We still do not fully understand why certain people with COVID-19 are asymptomatic or experience only mild symptoms. What differentiates those who fight off the SARS-CoV-2 infection without any treatments from others who suffer severe symptoms, and even die from the disease?
So far, we know that underlying health conditions (obesity, diabetes, heart disease, and lung problems to name a few), gender (men are more at risk than women) and age (over 65) increases vulnerability to COVID-19. But, what about genetic make-up? When exposed to the virus, are some people at higher risk of infection and developing symptoms that are more serious simply because they carry or lack one or several specific genes? This question can be posed for each infectious disease, not just COVID-19. Yet, given the huge prevalence of COVID-19, rigorously pursuing the answer could have a major impact on human health.
Why do so many COVID-19 patients exhibit neurological disorders?
In one of our previous blogs, we explained that proteins called spikes protrude from the surface of SARS-CoV-2 particles. This spike protein (S protein) has two subunits, S1 and S2. S1 contains the receptor-binding domain (RBD), which binds to the angiotensin-converting enzyme 2 (ACE2) receptors on human host cells. The S protein conveys the virus to receptors on the host cell surface, helping the virus efficiently invade the human cell to produce more viruses.
However, reports published in Science last month suggest that the differences observed in various tissues infected with COVID-19 may be related to another host factors. The research shows that the membrane protein neuropilin-1 (NRP1), which promotes SARS-CoV-2 entry, is another host factor for the coronavirus infection. Moreover, pathological analysis of olfactory epithelium cells obtained from human COVID-19 autopsies revealed that SARS-CoV-2 infected NRP1-positive cells. Within the olfactory epithelium, NRP1 was also observed in cells positive for oligodendrocyte transcription factor 2, which is mostly expressed by olfactory neuronal progenitors. That means the SARS-CoV-2 virus not only enters pulmonary and pharyngeal epithelium but also directly infects the brain, particularly through the nasal cavity, thus better explaining why so many infected patients suffer from neurological disorders, including loss of smell (anosmia) and taste (ageusia). Moreover, understanding the role of NRP1 gene expression in SARS-CoV-2 infection may provide potential targets for future antiviral treatments.
What about the influence of genetics in immune response?
The highly complex human immune response involves many genes. Scientists look for links between specific genetic markers and risk for and severity of disease. Investigating the interplay between human genetics and COVID-19, a study published in the New England Journal of Medicine identified a gene cluster called 3p21.31 on chromosome 3 as a risk locus for respiratory failure after infection with SARS-CoV-2. A study by Regeneron scientists also emphasized the positive link between the 3p21.31 locus and the severity of COVID-19. Another striking paper published in Nature demonstrated that this modified gene cluster is inherited from Neanderthals and carried by about 50% of people in south Asia and about 16% in Europe. In theory, the absence of this particular gene cluster in Africa might in part account for the decreased severity of COVID-19 cases across the continent. Of course, COVID-19 severity depends on many factors, but it is interesting that some genetic variations may also influence the characteristics of the current pandemic.
In a paper published in Science, researchers studied type I Interferons (IFNs)-based immunity, investigating whether defects in this immune defense might account for life-threatening COVID-19. The scientists found that indeed those individuals who lack functional IFNs signaling are more susceptible to severe COVID-19.
In fact, they identified two mechanisms that jeopardize IFNs functioning. First, 3.5 % of patients with life-threatening COVID-19 pneumonia had genetic defects in the genes controlling the type I interferon pathway. Secondly, and this is the first report in virology of this kind, at least 10.5% with severe COVID-19 pneumonia yielded type I interferon autoantibodies that existed before the infection. Autoantibodies are misguided antibodies that attack the immune system instead of the virus. Researchers did not find those autoantibodies in patients who were asymptomatic or experienced milder infections, or in healthy people. Patients with either modified genes or autoantibodies lacked an effective type I interferon-dependent immune response. These findings further underscore the importance of investigating in-depth the interferon pathway to help identify individuals at higher risk for severe SARS-CoV-2 infections.
In summary, the susceptibility to and severity of any infection is the result of interactions between viral- and host-related factors. Therefore, studying human genomics can help us understand the transmission, pathogenesis, susceptibility and severity of the infectious disease, as well as potentially lead to new, advanced COVID-19 diagnostics and treatments. Such studies must be evidence-based and conducted with caution to avoid misunderstandings and stigmatization of groups of patients.
Christian Brèchot, MD, PhD Senior Associate Dean for Research in Global Affairs, USF Health Morsani College of Medicine
Associate Vice President for International Partnerships and Innovation, USF
Professor, Department of Internal Medicine
President, Global Virus Network
Linman Li, MBA, MPH, PMP, CPH
Director, USF- GVN Center
USF Health Morsani College of Medicine
Vice President, Global Virus Network
Despite the progress made in managing asthma and chronic obstructive pulmonary disease (COPD), poorly controlled symptoms for both respiratory diseases can lead to severe shortness of breath, hospitalizations or even death.
“Only about 50 percent of asthmatics, and an even lower percentage of people with COPD, achieve adequate control of lung inflammation and airway constriction with currently available medications,” said Stephen Liggett, MD, vice dean for research at the University of South Florida Morsani College of Medicine and a USF Health professor of medicine, molecular pharmacology and physiology, and biomedical engineering. “So, we’re clearly missing something from our drug armamentarium to help all these patients.”
Dr. Liggett’s laboratory has discovered several subtypes of bitter taste receptors (TAS2Rs) — G protein-coupled receptors expressed on human smooth airway muscle cells deep inside the lungs. In asthma and COPD, tightening of smooth muscles surrounding bronchial tubes narrows the airway and reduces air flow, and Dr Liggett’s lab found that these taste receptors open the airway when activated. They are now looking for new drugs to treat asthma and other obstructive lung diseases by targeting smooth muscle TAS2Rs to open constricted airways.
A promising bronchodilator agonist rises to the top
In a preclinical study published Nov. 5 in ACS Pharmacology and Translational Science, Dr. Liggett and colleagues identified and characterized 18 new compounds (agonists) that activate bitter taste receptor subtype TAS2R5 to promote relaxation (dilation) of human airway smooth muscle cells. The cross-disciplinary team found 1,10 phenanthroline-5,6-dione (T5-8 for short) to be the most promising of several lead compounds (drug candidates). T5-8 was 1,000 times more potent than some of the other compounds tested, and it demonstrated marked effectiveness in human airway smooth muscle cells grown in the laboratory.
For this drug discovery project, Dr. Liggett’s laboratory collaborated with Jim Leahy, PhD, professor and chair of chemistry at the USF College of Arts and Sciences, and Steven An, PhD, professor of pharmacology at the Rutgers Robert Wood Johnson Medical School.
In an extensive screening conducted previously, another research group identified only one compound that would bind to and specifically activate the TASR5 bitter taste receptor – although apparently with limited effectiveness. Using this particular agonist (called T5-1 in the paper) as a starting point, the team relied on their collective disciplines to devise new activators, aiming for a much better drug profile for administration to humans.
USF Health’s Stephen Liggett, MD
“The two key questions we asked were: ‘Is it possible to find a more potent agonist that activates this receptor?’ and ‘Is it feasible to deliver by inhalation given the potencies that we find?’” said Dr. Liggett, the paper’s senior author. “T5-8 was the bronchodilator agonist that worked best. There were a few others that were very good as well, so we now have multiple potential new drugs to carry out the next steps.”
The researchers developed screening techniques to determine just how potent and effective the 18 compounds were. A biochemical test assessed how well these new agonists activated TAS2R5 in airway smooth muscle cells isolated from non-asthmatic human donor lungs. Then, the researchers validated the effect on airway smooth muscle relaxation using a technique known as magnetic twisting cytometry, pioneered by Dr An.
“Team science” solves a structural problem
“The biggest challenge we faced was not having a 3-D crystal structure of TAS2R5, so we had no idea exactly how agonist T5-1 fit into this mysterious bitter taste receptor,” Dr. Liggett said. “By merging our strength in receptors, pharmacology, physiology, and drug development, our team was able to make the breakthrough.”
T5-8 was superior to all the other bronchodilator agonists screened, exhibiting a maximum relaxation response (50%) substantially greater than that of albuterol (27%). Albuterol belongs to the only class of direct bronchodilators (beta-2 agonists) available to treat wheezing and shortness of breath caused by asthma and COPD. However, this drug or its derivatives, often prescribed as a rescue inhaler, does not work for all patients and overuse has been linked to increased hospitalizations, Dr. Liggett said. “Having two distinct classes of drugs that work in different ways to open the airways would be an important step to help patients optimally control their symptoms.”
The ACS Pharmacology paper highlights the importance of translational research in bridging the gap between laboratory discoveries and new therapies to improve human health, he added. “This study yielded a drug discovery that successfully meets most of the criteria needed to advance the compound toward its first trial as a potential first-in-class bronchodilator targeting airway receptor TAS2R5.”
Mounting studies indicate that the nicotine and other chemicals delivered by vaping, while generally less toxic than conventional cigarettes, can damage the lungs and heart. “But so far there has been no clear understanding about what happens when the vaporized flavoring molecules in flavored vaping products, after being inhaled, enter the bloodstream and reach the heart,” said the study’s principal investigator Sami Noujaim, PhD, an associate professor of molecular pharmacology and physiology at the USF Health Morsani College of Medicine.
In their study published Nov. 20 in the American Journal of Physiology- Heart and Circulatory Physiology, Dr. Noujaim and colleagues report on a series of experiments assessing the toxicity of vape flavorings in cardiac cells and in young mice. “The flavored electronic nicotine delivery systems widely popular among teens and young adults are not harm-free,” Dr. Noujaim said. “Altogether, our findings in the cells and mice indicate that vaping does interfere with the normal functioning of the heart and can potentially lead to cardiac rhythm disturbances.”
Dr. Noujaim’s laboratory is among the first beginning to investigate the potential cardiotoxic effects of the many flavoring chemicals added to the e-liquids in electronic nicotine delivery systems, or ENDS. He recently received a five-year, $2.2-million grant from the NIH’s National Institute of Environmental Health Sciences to carry out this laboratory research. Commonly called e-cigarettes, ENDS include different products such as vape pens, mods, and pods.
Sami Noujaim, PhD, of USF Health Molecular Pharmacology and Physiology, is investigating the potential cardiotoxicity of flavoring chemicals added to vaping e-liquids using preclinical models, including “cardiac cells in a dish.” | Photo by Allison Long, USF Health Communications
Vaping involves inhaling an aerosol created by heating an e-liquid containing nicotine, solvents such as propylene glycol and vegetable glycerin, and flavorings. The vaping device’s battery-powered heat converts this e-liquid into a smoke-like aerosolized mixture (e-vapor). Manufacturers tout e-cigarettes as a tool to help quit smoking, but evidence of their effectiveness for smoking cessation is limited, and they are not FDA approved for this use. E-cigarettes contain the same highly addictive nicotine found in tobacco products, yet many teens and young adults assume they are safe.
Among the USF Health study key findings:
In mouse cardiac muscle cells (HL-1 cells), the researchers tested the toxicity of three different, popular flavors of e-liquid: fruit flavor, cinnamon, and vanilla custard. All three were toxic to HL-1 cells exposed to e-vapor bubbled into the laboratory dish where the cells were cultured.
Cardiac cells derived from human pluripotent stem cells were exposed to three distinct e-vapors. The first e-vapor containing only solvent interfered with the electrical activity and beating rate of cardiac cells in the dish. A second e-vapor with nicotine added to the solvent increased the toxic effects on these cells. The third e-vapor comprised of nicotine, solvent, and vanilla custard flavoring (the flavor previously identified as most toxic) augmented damage to the spontaneously beating cells even more. “This experiment told us that the flavoring chemicals added to vaping devices can increase harm beyond what the nicotine alone can do,” Dr. Noujaim said.
Healthy young mice implanted with tiny electrocardiogram devices were exposed to 60 puffs of vanilla-flavored e-vapor five days a week, for 10 weeks. Heart rate variability (HRV) – that is, fluctuations in the time interval between successive heartbeats – decreased in these test mice compared to the control mice that inhaled only puffs of air under the same regimen. A sophisticated analysis by the USF Health researchers showed that vaping interfered with normal HRV in the mice by disrupting the autonomic nervous system’s control of heart rate (the acceleration and slowing down of heartbeats), Dr. Noujaim said.
Finally, mice exposed to vaping were more prone to an abnormal and dangerous heart rhythm disturbance known as ventricular tachycardia compared to control mice.
Whether the mouse findings will translate to people is unknown. Dr. Noujaim emphasizes that more preclinical and human studies are needed to further determine the safety profile of flavored ENDS and their long-term health effects.
A partial government ban on flavored e-cigarettes aimed at stopping young people from vaping focused on enforcement against flavored e-cigarettes with pre-filled cartridges, like those produced by industry leader JUUL. However, teens quickly switched to newer disposable e-cigarettes still sold in a staggering assortment of youth-appealing fruity and dessert-like flavors.
“Our research matters because regulation of the vaping industry is a work in progress,” Dr. Noujaim said. “The FDA needs input from the scientific community about all the possible risks of vaping in order to effectively regulate electronic nicotine delivery systems and protect the public’s health. At USF Health we will continue to examine how vaping may adversely affect cardiac health.”
Short-term inflammation is one of the body’s key defense mechanisms to help repair injury and fight infection. But low-level inflammation that does not subside has been linked to many common chronic conditions, including cardiovascular diseases such as atherosclerosis, atrial fibrillation and heart failure.
Ganesh Halade, PhD, an associate professor of cardiovascular sciences at the USF Health Morsani College of Medicine, investigates the safe clearance of acute inflammation – and what happens at the molecular and cellular levels when initially beneficial inflammation becomes harmful to the heart. His team at the USF Health Heart Institute works on bridging the gap between the immune-responsive metabolism of fat and cardiac health by more clearly defining two distinct but simultaneous processes: the inflammatory response and how inflammation is safely cleared, or resolved.
In particular, Dr. Halade’s laboratory focuses on discovering ways to prevent, delay or treat the unresolved inflammation after a heart attack, which plays a key role in the pathology leading to heart failure. Their goal is to contribute to individualized therapies that may account for possible sex, racial/ethnic or age-related physiological differences in heart failure, a leading cause of hospitalizations and deaths worldwide.
Ganesh Halade, PhD, associate professor of cardiovascular sciences, joined the USF Health Heart Institute in February 2020.
Heart failure — a progressively debilitating condition in which weakened or stiff heart muscle cannot pump enough blood to meet the body’s demand for nutrients and oxygen — has become a growing public health problem, fueled in part by an aging population and obesity epidemic. About 6.2 million adults in the U.S. suffer heart failure, according to the Centers for Disease Control and Prevention. Nearly half will die within five years of diagnosis, and the annual cost for health care, medications and missed work is estimated at more than $30 million.
“Although several treatments and devices exist to help manage heart failure, the challenge remains the growth of metabolic risk factors like obesity, diabetes, hypertension and aging that amplify heart failure – and inflammation underlies all these conditions,” Dr. Halade said. “We’re in the early stages of understanding how the inflammatory response becomes chronic, or unresolved” after heart attack-induced injury.
Honing in on “the roots”
Dr. Halade’s late father, a farmer in Nashik close to Mumbai, India, emphasized to his young son that if he wanted to make a difference in life to “look to the roots, rather than the fruits.”
That philosophy drives Dr. Halade’s research endeavors. “We focus on the root causes of inflammation so that we can successfully treat the chronic inflammation that leads to heart failure,” he said.
Dr. Halade (center) with his research team, postdoctoral fellow Bochra Tourki, PhD, (left) and research associate Vasundhara Kain, PhD, (right).
When a blocked coronary artery triggers a heart attack, inflammation caused by the tissue injury has two overlapping phases, Dr. Halade explains. During the inflammatory response, over-activated leukocytes (white blood cells of the immune system) rush from the spleen to the heart to remove dead cardiac tissue and start repairs. In the resolving phase, acute inflammation is cleared with the help of macrophages (another type of immune cell) that arrive to further repair the damage, and form a stable scar. Both timely responses are governed by coordinated ‘get in’ and ‘get out’ signals to leukocytes infiltrating the site of heart muscle injured by the heart attack. If the leukocytes do not receive a ‘get out’ signal, the sustained presence of inflammation impairs cardiac repair and eventually leads to heart failure.
Clinical trials of several anti-inflammatory therapies so far have failed to show benefit in heart failure patients. Dr. Halade suggests that the investigational compounds intended to suppress inflammation very early in the cardiovascular disease process likely disrupt the tight control of immune-responsive signaling needed for timely resolution of inflammation.
“The inflammatory response and its resolution are two sides of the same coin – and they roll together. Blocking one side will affect the other,” he explained. “So, we don’t want to block the ‘get in signal’ needed to promote the early, ‘good’ inflammation. We want to accelerate the ‘get out’ signal to immune cells, so that as soon as repair of cardiac injury is done the acute inflammation leaves without becoming chronic.”
Dr. Halade views a high-resolution image (below) of a normally beating heart.
Connecting dysfunctional inflammation control and heart failure
A class of immune-system molecules orchestrates the resolution of tissue inflammation, an active process essential for advancing cardiac healing after a heart attack. These specialized proresolving mediators, or SPMs, are signaling molecules that form when fatty acids metabolize in response to immune activation of leukocytes.
Dr. Halade’s work is helping uncover new details on how heart failure-inducing inflammation may be limited (without promoting immunosuppression) – either by administering pharmacological SPMs, or activating enzymes that help stimulate the body’s own SPMs.
Over the last two years, he has published significant findings in several leading journals (papers summarized below) making the connections between fatty acids, inflammation control, and heart failure. Among Dr. Halade’s study collaborators is Charles Serhan, PhD, of Harvard Medical School, a pioneer in the emerging field of inflammation resolution.
Science Signaling: This study followed the time course of inflammation and its resolution in a mouse heart attack model. The research showed for the first time that the active inflammation-resolving phase coincided with the acute inflammatory response facilitating cardiac repair after a heart attack. Among other factors, the researchers looked at types and amounts of SPMs, and the expression of enzymes that synthesize SPMs, both in the spleen and at the injured site of the heart. Macrophages, a type of white blood cell, are needed to generate SPMs as opposed to other immune cells, they reported.
Dr. Halade’s laboratory focuses on discovering ways to prevent, delay or treat the unresolved inflammation after a heart attack, which plays a key role in the pathology leading to heart failure.
Journal of the American Heart Association: The preclinical study discovered male-female cardiac repair differences in heart failure survival after heart attack, including improved recovery of cardiac function and greater survival of acute and chronic heart failure in female mice. Females generated higher levels of a particular fatty acid-derived signaling molecule (EET; epoxyeicosatrienoic acids) known to facilitate healing after a heart attack.
ESC Heart Failure: The researchers profiled bioactive lipids (inflammatory biomarkers) in blood samples from hospitalized Black and White patients soon after a severe heart attack. They found a potent SPM signature (resolvin E1) was significantly lower in Black men and women than in Whites. The study concluded bioactive lipids are key for the diagnosis and treatment of cardiac repair after heart attack to delay heart failure.
The FASEB Journal: Halade and colleagues identified a mouse model to study heart failure with preserved ejection fraction (HFpeF), a common form of heart failure linked to age-related obesity. Using this unique model of obese aging, they defined how the deficiency of a single resolution receptor triggers obesity in mice at an early age, which can give rise to many of the molecular and cellular processes ultimately leading to HFpEF.
Vasundhara Kain (seated) and Bochra Tourki, look at slides for a paper on age-related obesity and heart failure.
Insight into potential inflammation-resolving therapies
As they learn more about the metabolic and immune-responsive signals that control acute cardiac inflammation, researchers hope to harness the capacity of fatty acid-derived bioactive molecules to prevent, diagnose and treat heart failure, Dr. Halade said. SPMs are derived primarily from omega-3 fats in our diet – the polyunsaturated “good” fats in foods like salmon, avocados, almonds, and walnuts.
Some evidence indicates that omega 3-rich diets and/or SPM supplements, as well as getting enough exercise and quality sleep may help prevent the unresolved inflammation leading to heart failure, Dr. Halade said. If SPMs are not produced due to risk factors like obesity or aging, or because enzymes required to metabolize fatty acids are deficient, then drugs specifically designed to facilitate cardiac repair and calm inflammation might delay or treat heart failure, he added. Distinctive biochemical signatures acquired by analyzing SPMs or other metabolites might even be used to help diagnose heart failure or predict which treatments will work best for certain patients.
Dr. Halade joined USF Health this February from the University of Alabama at Birmingham, where he was a faculty member since 2013. He received his PhD in pharmacology from the University of Mumbai Institute of Chemical Technology in 2007. He completed two postdoctoral fellowships at the University of Texas Health Science Center in San Antonio. The first fellowship focused on nutritional immunology. The second was conducted with mentor Merry Lindsey, PhD, to examine the effects of obesity on post-heart attack cardiac structure and function.
Foods rich in omega-3 fatty acids (including salmon, walnuts and avocados), as well as enough exercise and quality sleep, may help prevent unresolved inflammation contributing to cardiovascular disease.
Dr. Halade’s research is supported by funding from the NIH’s National Heart, Lung and Blood Institute. In 2018, he received American Physiological Society Research Career Enhancement Award to train in lipidomics at the RIKEN Center for Integrative Medical Sciences in Japan.
His inflammation resolution research has been recognized with two awards for studies published in the American Journal of Physiology-Heart and Circulatory. An Article Impact Award 2020 was conferred this March by the American Physiological Society for Dr. Halade’s work defining the impact of the cancer drug doxorubicin on the heart and spleen. He also received a 2017 Best Paper Award from the Unbound Science Foundation. Dr. Halade is associate editor for the American Journal of Physiology-Heart and Circulatory and for Scientific Reports, and serves on the editorial boards of several other high-impact journals in cardiovascular sciences.
At left: Beneficial resolution of inflammation following cardiac repair. At right: Risk factors like aging, obesity and some medications can contribute to unresolved (chronic) inflammation, which impairs cardiac repair and can lead to heart failure. [Graphic courtesy of Ganesh Halade]
Some things you may not know about Dr. Halade
As an undergraduate student in India, Dr. Halade won the gold medal in fencing at a statewide collegiate competition.
To help promote a heart healthy lifestyle, he enjoys recreational bicycling and gardening in his backyard, where he grows vegetables and chiles.
Halade lives in Tampa with his wife Dipti, an information technology engineer, and their son Arav, 13.
Top: Sources of inflammation include injury (like damage from a heart attack), infection (viruses, bacteria or other pathogens), and factors associated with lifestyle (such as poor diet and lack of exercise). Below: Ways to help prevent unresolved cardiac inflammation associated with lifestyle. [Graphics courtesy of Ganesh Halade]
-Photos by Anne DeLotto Baier and Allison Long, Video by Allison Long, USF Health Communications and Marketing
Clinical trial shows first 3D printed nasal swabs work as well as commercial swabs for COVID-19 diagnostic testing
The device invented by USF Health doctors, teaming with Tampa General Hospital, Northwell Health and Formlabs, has been used worldwide to address critical shortages of test kit swabs
Seeking a solution to an unprecedented demand for nasal swabs at their own institution and others, USF Health researchers in the Departments of Radiology and Infectious Diseases reached out to colleagues at Northwell Health, New York’s largest health care provider, and leading 3D-printer manufacturer Formlabs. Working around the clock, this multidisciplinary team rapidly designed, tested and produced a 3D printed nasal swab prototype as a replacement for commercially-made flocked nasal swabs. Bench testing (24-hour, 3-day, and leeching) using respiratory syncytial virus as a proxy for SARS-CoV-2, as well as local clinical validation of the final prototype (fabricated with FDA-approved nontoxic, surgical grade materials), was successfully completed in mid-March 2020.
The paper’s first author Summer Decker, PhD, directs the USF Health Radiology-TGH Division of 3D Clinical Applications, which creates and prints 3D anatomical models for surgeons and other clinicians and designs medical devices.
Although USF Health held a provisional patent on the concept and design of the new 3D printed swab, they freely shared the information with hospitals, clinics, governments and international agencies experiencing supply chain shortages. Since the first batches of 3D printed swabs were processed, tens of millions of the USF Health-invented devices have been used in 22 countries, said lead author Summer Decker, PhD, an associate professor of radiology at the USF Health Morsani College of Medicine. Dr. Decker directs the USF Health Radiology-TGH Division of 3D Clinical Applications, a group with expertise in creating and printing 3D anatomical models for surgeons and other clinicians as well as designing medical devices.
“In the midst of a pandemic, our team of experts representing academic medicine, health care delivery systems, and the medical device industry put aside boundaries to quickly work together toward a common purpose,” Dr. Decker said. “It’s rewarding that the novel design for a 3D swab we created has been adopted around the world, equipping more providers to diagnose COVID-19 and hopefully help prevent its spread.”
The gold standard for diagnosing respiratory infections is to look for viral genetic material found in mucosal fluid collected with a long, slender swab inserted into the patient’s nose and back of the throat. The nasal swab is put into a plastic tube with chemicals that stabilize the sample until the virus-specific genetic material can be extracted and amplified by polymerase chain reaction (PCR) in a diagnostics laboratory. Conventional swabs feature a bushy tip coated with nylon flock; the USF Health doctors designed a tip with a 3D printed textured pattern able to capture a sufficient sample for COVID testing while keeping patient safety and comfort in mind.
Kami Kim, MD, infectious diseases division director at USF Health Morsani College of Medicine, led the multisite clinical trial comparing the performance of commercial nasal swabs with the 3D-printed alternative.
The clinical trial fully tested the safety and effectiveness of this 3D printed swab in 291 symptomatic adults undergoing COVID-19 screening at the TGH, Northwell Health and Thomas Jefferson University Hospital sites. The 3D printed nasal swab was compared to the standard synthetic nasal swab across three SARS-CoV-2 testing platforms FDA-authorized for emergency use — a modified version of the Center for Disease Control and Prevention’s real-time reverse transcriptase PCR diagnostic panel, and two commercial molecular diagnostic tests.
“This trial provided the first rigorous head-to-head comparison to make sure that the 3D swab performed as well as the standard,” said principal investigator Kami Kim, MD, professor and division director for infectious disease at the USF Health Morsani College of Medicine. “Across all three platforms used in our study, we demonstrated that the commercial swab and the 3D printed swab were comparable for accurate detection of COVID-19 infection.”
For both swabs, the only adverse patient reaction documented during the trial was a few instances of slight nasal bleeding. The cost of materials per 3D printed nasal swab ranges from 26-to 46-cents, while commercial swabs cost about $1 each, the authors reported.
Given the ongoing need for widespread COVID-19 testing, the study authors concluded that 3D printing technology offers a viable, cost-efficient option to address swab supply shortages, particularly when local hospitals or other clinical sites already have 3D printing labs equipped to print and process the devices.
The 3D printed nasal swabs were specifically designed for patients using FDA-approved surgical grade material.
Frank Rybicki, MD, PhD, vice chair of operations and quality at the University of Cincinnati College of Medicine’s Department of Radiology, wrote a commentary on 3D printing in medicine to accompany the Clinical Infectious Diseases paper. The article frames the contributions of Decker et. al. in the context of the larger 3D manufacturing community.
“Among all parts 3D printed during COVID-19, nasopharyngeal swabs have received the most attention, with participants ranging from humanitarians to charlatans,” Dr. Rybicki wrote in his summary. “The authors should be congratulated for staying on the right side of the curve, and for their perseverance, leadership, scientific rigor, and good will.”
-Photos and Video by Allison Long, USF Health Communications and Marketing
Compounds halt SARS-CoV-2 replication by targeting key viral enzyme
A University of Arizona-University of South Florida team identified and analyzed four promising antiviral drug candidates in the preclinical study
The most promising drug candidates – including the FDA-approved hepatitis C medication boceprevir and an investigational veterinary antiviral drug known as GC-376 – target the SARS-CoV-2 main protease (Mpro), an enzyme that cuts out proteins from a long strand that the virus produces when it invades a human cell. Without Mpro, the virus cannot replicate and infect new cells. This enzyme had already been validated as an antiviral drug target for the original SARS and MERS, both genetically similar to SARS-CoV-2.
“With a rapidly emerging infectious disease like COVID-19, we don’t have time to develop new antiviral drugs from scratch,” said Yu Chen, PhD, USF Health associate professor of molecular medicine and a coauthor of the Cell Research paper. “A lot of good drug candidates are already out there as a starting point. But, with new information from studies like ours and current technology, we can help design even better (repurposed) drugs much faster.”
Before the pandemic, Dr. Chen applied his expertise in structure-based drug design to help develop inhibitors (drug compounds) that target bacterial enzymes causing resistance to certain commonly prescribed antibiotics such as penicillin. Now his laboratory focuses its advanced techniques, including X-ray crystallography and molecular docking, on looking for ways to stop SARS-CoV-2.
Using 3D computer modeling, Michael Sacco (left), a doctoral student in the Department of Molecular Medicine, worked with Dr. Chen to determine the interactions between antiviral drug candidate GC-376 and COVID-19’s main protease.
Mpro represents an attractive target for drug development against COVID-19 because of the enzyme’s essential role in the life cycle of the coronavirus and the absence of a similar protease in humans, Dr. Chen said. Since people do not have the enzyme, drugs targeting this protein are less likely to cause side effects, he explained.
The four leading drug candidates identified by the University of Arizona-USF Health team as the best (most potent and specific) for fighting COVID-19 are described below. These inhibitors rose to the top after screening more than 50 existing protease compounds for potential repurposing:
Boceprevir, a drug to treat Hepatitis C, is the only one of the four compounds already approved by the FDA. Its effective dose, safety profile, formulation and how the body processes the drug (pharmacokinetics) are already known, which would greatly speed up the steps needed to get boceprevir to clinical trials for COVID-19, Dr. Chen said.
GC-376, an investigational veterinary drug for a deadly strain of coronavirus in cats, which causes feline infectious peritonitis. This agent was the most potent inhibitor of the Mpro enzyme in biochemical tests, Dr. Chen said, but before human trials could begin it would need to be tested in animal models of SARS-CoV-2. Dr. Chen and his doctoral student Michael Sacco determined the X-ray crystal structure of GC-376 bound by Mpro, and characterized molecular interactions between the compound and viral enzyme using 3D computer modeling.
Calpain inhibitors II and XII, cysteine inhibitors investigated in the past for cancer, neurodegenerative diseases and other conditions, also showed strong antiviral activity. Their ability to dually inhibit both Mpro and calpain/cathepsin protease suggests these compounds may include the added benefit of suppressing drug resistance, the researchers report.
All four compounds were superior to other Mpro inhibitors previously identified as suitable to clinically evaluate for treating SARS-CoV-2, Dr. Chen said.
Michael Sacco looks at COVID-19 viral protein crystals under a microscope.
A promising drug candidate – one that kills or impairs the virus without destroying healthy cells — fits snugly, into the unique shape of viral protein receptor’s “binding pocket.” GC-376 worked particularly well at conforming to (complementing) the shape of targeted Mpro enzyme binding sites, Dr. Chen said. Using a lock (binding pocket, or receptor) and key (drug) analogy, “GC-376 was by far the key with the best, or tightest, fit,” he added. “Our modeling shows how the inhibitor can mimic the original peptide substrate when it binds to the active site on the surface of the SARS-CoV-2 main protease.”
Instead of promoting the activity of viral enzyme, like the substrate normally does, the inhibitor significantly decreases the activity of the enzyme that helps SARS-CoV-2 make copies of itself.
Visualizing 3-D interactions between the antiviral compounds and the viral protein provides a clearer understanding of how the Mpro complex works and, in the long-term, can lead to the design of new COVID-19 drugs, Dr. Chen said. In the meantime, he added, researchers focus on getting targeted antiviral treatments to the frontlines more quickly by tweaking existing coronavirus drug candidates to improve their stability and performance.
Two viral protein images generated by Yu Chen, University of South Florida Health, using X-ray crystallography. Above: The protein dimer (one molecule is blue and the other orange) shows the overall structure of the COVID-19 virus’s main protease (Mpro), the researchers’ drug target. Below: Three configurations of active sites where inhibitor GC-376 binds with the Mpro viral enzyme, as depicted by 3D computer modeling.
Dr. Chen worked with lead investigator Jun Wang, PhD, UA assistant professor of pharmacology and toxicology, on the study. The work was supported in part by grants from the National Institutes of Health.
-Photos by Torie Doll, USF Health Communications and Marketing
Responding to the urgent need for solutions to a potentially deadly emerging infectious disease, USF Health has jumpstarted a variety of COVID 19-related research projects.
The work may contribute to worldwide efforts to accelerate the discovery and validation of new technologies to diagnose and prevent the spread of SARS-CoV-2, as well as finding potential new treatments for COVID-19, the respiratory disease caused by the new coronavirus.
The new studies are being conducted by scientists across disciplines including biochemistry, infectious diseases and international medicine, medical engineering, nursing, pharmacy, public health, structural biology and virology. The ambitious research efforts include the joint clinical trials launched last month by USF Health and Tampa General Hospital.
“Many fundamental questions remain about this newest coronavirus, including how it functions, the ability of antibodies to convey immunity, and whether genetic differences in certain populations affect their susceptibility to COVID-19 infection or severity of the illness,” said Stephen Liggett, MD, associate vice president of the USF Health Office of Research and vice dean for research in the Morsani College of Medicine. “Our faculty and student researchers have been quick to mobilize their talent and resources, because they want to do whatever they can to find answers — both to help fight this pandemic and to prepare for future outbreaks.”
Some of the COVID-19 related projects build upon knowledge and insights USF Health scientists have acquired using advanced technologies to study the underlying molecular and cellular biology of other viruses and pathogens, including respiratory syncytial virus and HIV.
Among the more than 60 new research projects underway, or being scaled up, are:
3D printed nasal swabs for testing: This multidisciplinary project was recently cited in Nature. At the request of USF Health Senior Vice President Dr. Charles J. Lockwood, USF Health Morsani College of Medicine faculty in partnership with faculty at Northwell Health (New York, NY), developed, bench lab-tested, and clinically validated 3D printable flocked nasopharyngeal (NP) swabs to obtain viral RNA samples for COVID-19 testing. The NP swabs were designed with FDA-cleared software and produced on FDA-cleared 3D printers using FDA-cleared surgical grade material that can withstand high-temperature sterilization. The swabs are comparable to commercially available, conventional NP swabs composed of nylon or polyester to obtain enough viral particles to reliably diagnose COVID-19. The FDA has cleared the use of 3D printed swabs made with medical/dental grade materials. USF Health and Northwell Health hold the provisional patent for this technology but are sharing the print file with institutions that have the FDA-cleared technology and materials to print their own swabs. The 3D formula has been provided to medical centers throughout the U.S. and to other countries.
Antibody tests and immunity: The reliability of new tests that detect blood markers of SARS-CoV-2 infection (antibodies) varies. This collaborative research with Tampa General Hospital’s laboratory, will compare different combinations of antibody tests and analyze which best determines whether a person infected with the virus develops a protective immune response. Knowing whether, and when, antibodies convey immunity could help hospitals and other health care employers decide whom among their medical staff might safely return to work. It can also help researchers recalculate a more accurate fatality rate in the general population by providing a broader picture of COVID-19 infections (including in those with mild or no symptoms). Principal investigator: Kami Kim, MD
Susceptibility in different ethnic backgrounds: African Americans and Hispanics have double the infection rates and deaths from COVID-19, and in some disease hotspots they are among those disproportionately affected by pre-existing cardiovascular conditions like hypertension and heart failure. This project seeks to answer key questions about COVID-19 related racial/ethnic disparities, including whether socioeconomic differences alone account for differences in infection rates and cardiovascular complications, or if cellular-level or other physiological factors contribute to worse disease outcomes. Principal investigator: Thomas McDonald, MD, USF Health Heart Institute, MCOM.
Protective antibodies and vaccines: People previously exposed to SARS CoV-2 make immune responses to viral proteins. This public health-medicine team project aims to find parts of the viral proteins, called epitopes, that are recognized by antibodies. Some epitopes are associated with strong protective antibody responses that neutralize the virus while others are not. This Morsani College of Medicine-College of Public Health study will help identify specific epitopes that lead to strong neutralizing antibodies so that researchers can develop more effective vaccines. The information will also help in screening donor plasma that can be best used for treatment of critically ill COVID-19 patients. Principal investigator: Kami Kim, with co-investigators Michael Teng, PhD; John Adams, PhD; and Thomas Unnasch, PhD
Portable biomedical testing system: This project explores whether a mobile phone version of the Enzyme Linked Immunosorbent Assay (ELISA), technology patented by USF, could be used to accurately measure antibodies (proteins) made in response to the COVID-19 virus. Laboratory-based ELISA is the gold standard in biochemical analysis of proteins, but the blood test requires big, expensive machines for its complex steps, including incubation and reading. The lightweight Mobile Enzyme Linked Immunosorbent Assay (MELISA) device may allow patients to cost-effectively obtain point-of-care antibody testing and results at a clinic or even a remote area. Anna Pyayt, PhD, principal investigator, College of Engineering (affiliated with Department of Medical Engineering).
Targeting viral replication– This study applies advanced techniques, including X-ray crystallography and molecular docking, to identify new or existing drugs that prevent viral replication by inhibiting SARS-CoV-2 main protease, or by blocking entry of the virus into human cells. Principal investigator: Yu Chen, PhD, Department of Molecular Medicine, MCOM.
Hana Totary-Jain, PhD, an associate professor of molecular pharmacology and physiology in the USF Health Morsani College of Medicine, was senior author of the study published in Scientific Reports.
The placenta, an organ which attaches to the lining of the uterus during pregnancy, supplies maternal oxygen and nutrients to the growing fetus. Abnormal formation and growth of the placenta is considered an underlying cause of various pregnancy complications such as miscarriages, stillbirth, preeclampsia and fetal growth restriction. Yet, much remains to be learned about molecular mechanisms regulating development of this blood-vessel rich organ so vital to the health of a pregnant woman and her developing fetus.
During the first trimester, fetal-derived placental cells known as trophoblasts invade the maternal uterine lining and modify its blood vessels to allow oxygenated blood to flow from the mother to fetus. However, trophoblast invasion requires tight regulation of EMT. If inadequate, trophoblast invasion is too shallow to adequately remodel the maternal blood vessels, and adverse pregnancy outcomes can occur. Conversely, excess EMT can cause exaggerated trophoblast invasion through the uterine wall leading to placenta accreta, a condition that can cause hemorrhage and often requires hysterectomy at delivery.
The USF Health researchers used a powerful genome editing technology called CRISPR (shorthand for “CRISPR-dCas9) to activate all of the chromosome 19 microRNA cluster (known as C19MC), so they could study the gene’s function in early pregnancy. C19MC — one of the largest microRNA gene clusters in the human genome — is normally turned off but becomes expressed only in the placenta, embryonic stem cells and certain cancers.
Dr. Totary-Jain discusses the molecular aspects of placenta development and pregnancy complications with research collaborator Umit Kayisli, PhD, a professor of obstetrics and gynecology at USF Health.
In their cell model study, published Feb. 20 in Scientific Reports, a Nature research journal, the USF Health team showed that robust activation of C19MC inhibited EMT gene expression, which has been shown to reduce trophoblast invasion.
But when trophoblast-like cells were exposed to hypoxia – a lack of oxygen similar to that occurring in early placental development — C19MC expression was significantly reduced, the researchers found. The loss of C19MC function causes differentiation of trophoblasts from stem-like epithelial cells into mesenchymal-like cells that can migrate and invade much like metastatic tumors. This EMT process helps explain trophoblast invasion and early placental formation.
“We were the first to use CRISPR to efficiently activate the entire gene, not just a few regions of this huge gene, in human cell lines,” said the paper’s senior author Hana Totary-Jain, PhD, an associate professor in the Department of Molecular Pharmacology and Physiology, USF Health Morsani College of Medicine. “Our study indicates C19MC plays a key role in regulating many genes important in early implantation and placental development and function. The regulation of these genes is critical for proper fetal growth.”
Above: Chromosome 19 microRNA cluster (stained purple) expressed in first-trimester placenta. Below: In preparation for pregnancy, fetal trophoblast cells (brown) from which the placenta arises invade maternal decidual cells (pink) in the uterus lining. | Images courtesy of Hana Totary-Jain, originally published in Scientific Reports: doi.org/10.1038/s41598-020-59812-8
“You need EMT, but at some point the process needs to cease to prevent adverse pregnancy outcomes,” Dr. Totary-Jain said. “You really need a balance between not enough invasion and too much invasion, and C19MC is important in maintaining that balance.”
“The USF Health study offers new insight into how trophoblasts interact with the maternal uterine environment to become more invasive or less invasive in the formation of the placenta,” said coauthor Umit Kayisli, PhD, a USF Health professor of Obstetrics and Gynecology. “More research on microRNA expression and how it inhibits EMT may help us better understand the causes and potential prevention of preeclampsia and fetal growth restriction, which account for 5-to-10 percent of all pregnancy complications as well as spontaneous preterm births.”
Investigating the effects of altered C19MC expression on cell differentiation and trophoblast invasion has implications not only for a better understanding of normal and abnormal placental development, but also for cancer and stem cell research, Dr. Totary-Jain added.
Dr. Totary-Jain and Dr. Kayisli
-Photos by Freddie Coleman, USF Health Communications and Marketing
Gopal Thinakaran pursues genetic clues to Alzheimer’s disease pathways
The USF Health neurobiologist focuses on understanding genetic risk factors that may offer new therapy targets to delay or protect against age-related cognitive decline
The steep rise in the number of Americans dying of Alzheimer’s disease – up 145 percent between 2000 and 2017— is not just reflected in unprecedented statistics driven by an aging population. This neurodegenerative disease that relentlessly diminishes the mind is borne by those living more years in a state of disability and dependence before dying — and the family members who care for them.
No treatments exist to cure or slow the progression of Alzheimer’ disease, the major form of dementia afflicting an estimated 5.8 million Americans.
“The goal of our research is to reduce the (brain) pathology leading to Alzheimer’s disease, identify targeted treatments to delay the onset of disease and protect cognitive function,” said Gopal Thinakaran, PhD, professor of molecular medicine and associate dean for neuroscience research at the USF Health Morsani College of Medicine. “Finding ways to extend cognitive function so that an older person is still able to continue their daily activities or recognize a loved one – even for five more years – would greatly benefit both those suffering from Alzheimer’s and their families or other caregivers.”
Gopal Thinakaran, PhD (center), who holds the Bagnor Endowed Chair in Alzheimer’s Research, with his research team at the USF Health Byrd Alzheimer’s Center.
Dr. Thinakaran, an internationally recognized Alzheimer’s disease researcher, joined the University of South Florida from the University of Chicago in August to help accelerate the interdisciplinary work of the USF Health Neuroscience Institute. That includes recruiting a critical mass of basic scientists who can complement the university’s ongoing Alzheimer’s research while also expanding efforts to translate laboratory findings into new therapies for other neurodegenerative disorders, including Parkinson’s disease, ataxias, ALS, and multiple sclerosis.
In addition to his leadership role, Dr. Thinakaran oversees a laboratory at the Byrd Alzheimer’s Center where he uses cutting-edge cell biology techniques and mouse models to study the molecular and cellular processes underlying Alzheimer’s disease. His research is supported by more than $6.1 million in grants from the National Institutes of Health (NIH), National Institute on Aging.
With normal brain aging, people experience minor lapses of memory (i.e., forgetting where their keys were left, or the name of someone just met) and some reduced speed in processing information. But disruptions in attention, memory, language, thinking and decision-making that interfere with daily life are signs of dementia.
In addition to overseeing his own laboratory research, Dr. Thinakaran holds Morsani College of Medicine leadership roles as associate dean of neuroscience research and Neuroscience Research Institute associate director of research.
Dr. Thinakaran’s lab pursues findings on relatively new genes identified through genome-wide association studies to gain insights into the mechanisms of late-onset Alzheimer’s disease, which affects people age 65 or older and accounts for the overwhelming majority of cases. Recently, the group has been investigating the role of bridging integrator 1 (BIN1), the second most common genetic risk factor for late-onset Alzheimer’s (exceeded only by APOE). Approximately 40% of people with Alzheimer’s have one of three variations in the BIN1 gene – a glitch in a single DNA building block (nucleotide) that heightens their risk for the disease, Dr. Thinakaran said.
Pursuing A Common Risk Factor For Late-Onset Alzheimer’s
BIN1, expressed in all the body’s cells, has been shown to play a role in suppressing tumors and in muscle development — but little is known about what the protein does in the brain. Dr. Thinakaran was among the first to embrace the challenge of pursuing how BIN1 contributes to Alzheimer’s disease risk at a time when most researchers focused on amyloid and tau, two proteins considered the primary drivers of Alzheimer’s pathology.
Now, his team and a few others across the country probe what goes wrong in Alzheimer’s patients who carry the BIN1 risk allele. They have already confirmed that BIN1 is present both in the brain’s nerve cells (neurons) and its non-neuronal cells, such as oligodendrocytes and microglia.
Biochemist Melike Yuksel, PhD, a postdoctoral scholar in Dr. Thinakaran’s lab.
A healthy human brain contains tens of billions of neurons that process and transmit chemical messages (neurotransmitters) across a tiny gap between neurons called a synapse. Alzheimer’s disease severely disrupts this synaptic communication, eventually killing cells throughout the brain and leading to a steep decline in memory and other signs of dementia.
“The single biggest correlation with cognitive decline is the loss of these synaptic communication centers between neurons,” Dr. Thinakaran said, adding that individuals most susceptible to developing full-blown Alzheimer’s in later life are those who lose the most synapses.
In a study posted online last year as a Cell Reports Sneak Peak manuscript and since accepted for publication, Dr. Thinakaran and colleagues demonstrated for the first time that the loss of BIN1 expression impaired spatial learning and memory associated with remembering where things are located. The researchers used an Alzheimer’s disease “knockout” mouse model in which neuronal BIN1 expression was inactivated in the hippocampus, a brain region involved with higher cognitive functions.
Discovering A Defect In Brain Cell Communication
A lack of BIN1 leads to a defect in the transmission of neurotransmitters needed to activate the brain cell communication that allows us to think and behave, the researchers found. Further analysis distinguished that BIN1 primarily locates on neurons that send neurotransmitters across the synapse (presynaptic sites) rather than residing on those neurons that receive the neurotransmitter messages (postsynaptic sites). The BIN1 deficiency was also associated with reduced synapse density; a back-up of docked vesicles, the tiny bubble-like carriers that transfer neurotransmitters from presynaptic to postsynaptic neurons; and likely slower release of the neurotransmitters from their vesicles.
“Our findings so far that BIN1 localizes right at the point of (presynaptic) communication and may be precisely regulating neurotransmitter vesicle release brings us much closer to understanding how BIN1 could exert its function as a risk factor (for Alzheimer’s disease),” Dr. Thinakaran said. “We suspect it helps control how efficiently neurons communicate.”
Peering into the brain, one synapse at a time. Electron micrograph depicting selected region of a mouse brain hippocampus, the brain area responsible for learning and memory. A single synapse is marked with the yellow outline. The human brain is estimated to have trillions of these synapses, which transmit information from one neuron to the next.| Image courtesy of Gopal Thinakaran, PhD
Antibody-stained mouse brain with Alzheimer’s disease β-amyloid deposits. The amyloid precursor proteins within healthy nerve cells and swollen neuronal processes are depicted in blue. The late-onset Alzheimer’s risk factor BIN1 is shown in green, and a marker for brain glial cells responsible for neuroinflammation is shown in magenta.| Image courtesy of Gopal Thinakaran, PhD
Dr. Thinakaran’s team also became interested in investigating whether BIN1 risk variants can interfere with the protective capacity of glia (cells supporting neurons) to mount a full inflammatory response needed to clear toxins from the brain. His USF Health group will work with researchers at Emory University to further investigate why the absence of BIN1 may impair the brain’s removal of abnormal beta-amyloid protein associated with Alzheimer’s disease.
Exploring The Type 2 Diabetes Connection
Collaborating with a coprincipal investigator at the University of Kentucky, Dr. Thinakaran explores the molecular link between type 2 diabetes and Alzheimer’s disease progression. An Alzheimer’s mouse model created by the Thinakaran lab allows researchers to turn on, or switch off, production of the human hormone amylin in the pancreas.
Amylin is secreted by the pancreas at higher levels, along with insulin, as diabetes begins to develop. Small amounts of this excess amylin migrate from pancreatic cells into the bloodstream and can cross the blood-brain barrier, especially in older brains where the protective barrier becomes leakier. The amylin then mixes with the brain’s beta-amyloid, which eventually builds into the sticky amyloid plaques that are a hallmark of Alzheimer’s pathology. The researchers will test in their preclinical model whether this brain amylin elevates the risk for Alzheimer’s disease, and if reducing amylin in peripheral circulation can help prevent or slow damage to cognition.
Dr. Thinakaran with biological scientist Stanislau (Stas) Smirnou.
Scientists are still trying to figure out why some people remain cognitively resilient throughout life despite having neuropathology that would otherwise cause dementia. On the horizon, Dr. Thinakaran said, integrating large databases of gene expression and individual cell types will help scientists drill deeper into what specific inflammatory, metabolic and neural circuit changes shift a normally aging brain to one in which the abilities to remember, think and reason abnormally accelerate.
At the same time, data on genetics and environment/lifestyle (including diet, physical and mental exercises, sleep patterns and uncontrolled cardiovascular risk factors such as hypertension, diabetes and high cholesterol) is being collected both for patients in various stages of Alzheimer’s disease and for older adults with healthy cognitive function. “Bridging these two sets of data will be extremely valuable in understanding what confers higher risk and delineating what can keep our brains healthy as we age,” Dr. Thinakaran said.
Fascinated By A Field With Unprecedented Challenges
Dr. Thinakaran holds a PhD in molecular biology and genetics from the University of Guelph in Canada. He completed a postdoctoral research fellowship in neuropathology and was an assistant professor of pathology at Johns Hopkins University School of Medicine. Before joining USF Health, he was a professor of neurobiology at the University of Chicago, where he built one of the country’s leading laboratories investigating pathways responsible for Alzheimer’s disease pathology and neuronal dysfunction.
Known as an accomplished scientist and thought leader who does not hesitate to tackle uncharted territory, Dr. Thinakaran studied muscle differentiation as a PhD student. But, he soon realized that muscle research had advanced to a stage where it was unlikely he could make much of an impact. At that time (early 1990s) Alzheimer’s disease research was just gaining momentum in molecular and cellular biology and posing unprecedented challenges, he said.
Biological scientist Xiaolin Zhang, MS
Once Dr. Thinakaran’s interest in Alzheimer’s was sparked during his postdoctoral training at Johns Hopkins, he seized the opportunity to pursue the emerging area of neuroscience research. “In many ways the brain and its complexity as we age is the final frontier in understanding human behavior. We’re continuing to learn every day the basics of how this organ system works, and what goes wrong when it doesn’t,” he said. “It’s a field that still has great opportunities for the next generation of young minds to make a difference.”
Dr. Thinakaran has authored more than 140 peer-reviewed publications. He is associate editor for the journals Molecular Neurodegeneration and Genes and Diseases and an editorial board member for Neurodegenerative Diseases and for Current Alzheimer Research. He serves on several scientific review/advisory committees for federal, private and public institutions. Dr. Thinakaran has received numerous awards, including the Alzheimer’s Association prestigious Zenith Fellows Award supporting senior scientists pursuing new ideas to advance Alzheimer’s and dementia research.
Some Things You May Not Know About Dr. Thinakaran
Dr. Thinakaran combines his artistic talents of drawing and painting with his research. Andy Warhol-like microscopic art he created won a competition to be featured as the program cover for a brain research symposium at the University of Chicago. The multicolor montage of images depicts a mouse brain section (hippocampus) stained to visualize β-secretase, an enzyme critical for generating the hallmark Alzheimer’s disease β-amyloid pathology.
He is married to neurophysiologist Angèle Parent, PhD, associate professor of molecular medicine at the Byrd Alzheimer’s Center. They have three children: Abigaël, a freshman and aspiring neuroscientist at the University of Chicago; Daphné, 14; and Cédric, 12.
Dr. Thinakaran enjoys cooking authentic South Indian food and other international dishes with his family.
This microscopic brain art created by Dr. Thinakaran was featured on the program cover of a University of Chicago brain research symposium.
-Video by Allison Long, and photos by Freddie Coleman, USF Health Communications and Marketing
USF Health cell biologist studies role of capillaries in tissue health and disease
The laboratory of Dr. George Davis grows three-dimensional “blood vessel networks in a dish” under defined, serum-free conditions
Capillaries are our smallest, yet most abundant, blood vessels.
With walls barely as thick as a single red blood cell, they form exquisitely branching networks, spanning a total surface area of 1,000 square miles. Connecting with arteries and veins, capillaries exchange oxygen, nutrients and waste between the bloodstream and tissues throughout the body.
Healthy communication, or molecular signaling, inside and outside capillaries appears to play a critical role in promoting healthy tissues such as the heart, lungs and liver. Conversely, many diseases arise from abnormalities in blood vessels that fail to communicate properly with tissues, especially those requiring a lot of oxygen to work properly.
“I believe the root cause of many patient treatment failures is a lack of understanding of the underlying basis of disease,” said Dr. Davis, professor of molecular physiology and pharmacology and member of the college’s USF Health Heart Institute. “My philosophy is that understanding the fundamental biology of capillaries in their normal state will lead to answers about what goes wrong in diseases,” including coronary artery disease, stroke, diabetes, and malignant cancers.
“If blood vessel formation is altered or begins to break down, we should be able to find a way to pharmacologically fix that.”
George E. Davis, MD, PhD, professor of molecular physiology and pharmacology, is a member of the USF Health Heart Institute.
Growing Blood Vessel Networks “In A Dish”
Dr. Davis joined USF Health in June 2018 from the University of Missouri-Columbia School of Medicine, where he was an investigator at the Dalton Cardiovascular Research Center. “What really attracted me to USF Health was having the opportunity to be part of a new heart institute being built around world-class research that interfaces with clinicians and biomedical engineering,” he said.
To delve into the complexity of capillary formation, Dr. Davis grows three-dimensional “blood vessel networks in dish” under serum-free, defined conditions to reduce variability. “The (cell culture) system we’ve developed over the years is really quite powerful, because it allows us to study molecular signaling — but also helps us in trying to understand what genes are regulated when vessels form,” he said.
His in vitro research primarily centers on two types of human cells – endothelial cells, which line the inner surface of capillaries, and pericytes, cells recruited to the outer surface of developing capillaries to help fortify the endothelial-lined tubes. He investigates the molecular “cross-talk” between these cells that controls how capillary networks arise and mature to support adjacent tissues.
Dr. Davis’ laboratory is known for discovering the combination of five growth factors (SCF, IL-3, SDF-1α, FGF-2 and insulin) needed to create viable human capillary networks in culture. Vascular endothelial growth factor (VEGF), considered to be a primary driver of blood vessel formation, surprisingly was not in mix; it did not directly stimulate the assembly of capillary networks.
Biological scientist Gretchen Koller works in Dr. Daviss laboratory.
Healthy Capillaries As “Disease Suppressors”
His group defined at least two steps in capillary formation, validated by experiments. First, upstream priming by VEGF “wakes up” the capillary cells to respond to the five growth factors required for vessel assembly. Second, this now activated group of downstream factors promotes capillary tube formation and branching.
The Davis laboratory also recently proposed that healthy capillaries within tissues may be “disease suppressors,” since communication between endothelial cell-and pericyte-derived growth factors can inhibit basic disease mechanisms. These underlying pathological mechanisms include blood-clotting (thrombosis), inflammation, excessive fibrous connective tissue (fibrosis), inadequate blood supply (ischemia), and transformation of normal cells into cancer cells (carcinogenesis).
For example, Dr. Davis said, dysfunctional capillaries are a hallmark of diabetes — particularly for a complication known as diabetic retinopathy where pericytes drop off the tiny blood vessels in light-sensitive tissue at the back of the eye. This loss of pericytes, coupled with capillary breakdown leading to bleeding events, can lead to vision loss.
Even as they continue to uncover fundamental details about how capillary networks normally take shape, Dr. Davis’ team has reached a point where they can start applying what they’ve learned to attack, possibly even prevent, diseases.
“We want to take a model of capillary formation and create a diabetic-type state to see if we can mimic any of the changes in pericytes and the associated vessels observed in diabetes,” he said.
Endothelial cell-lined tubes (red) with associated pericytes (green). Both cell types co-assemble to create the capillary networks vital to the health of tissues throughout the body.
Applications For Tissue Regeneration
Besides helping identify potential treatments to repair dysfunctional capillaries or to promote their disease suppression capacities, the research may have applications for tissue regeneration. Engineering functional tissue to repair or replace a damaged heart, lung, kidney or other organs requires robust capillary networks.
“The biggest problem in creating these tissue engineered constructs has been a failure in the vasculature,” Dr. Davis said. “It’s critical to understand specifically what makes capillaries form and stabilize to sustain healthy tissue, so (when the construct does not work) it’s possible the growth factors added may not be the right ones.
“We’re working to figure out, in our defined system, which growth factors do what, and when they act.”
And the USF Health work in angiogenesis, the growth of new blood vessels, can provide insight into the control of tumor cell migration and invasion into distant tissues (metastasis). “If we can better understand the nuts and bolts of how to make a capillary network, we’ll gain a better sense of what makes the tumor microenvironment so abnormal,” Dr. Davis said.
Dr. Davis earned dual MD and PhD degrees from the University of California, San Diego, in 1986, and completed a medical staff fellowship (anatomical pathology residency) at the National Cancer Institute’s Laboratory of Pathology in Bethesda, MD.
Biological scientist Kalia Aguera.
Dr. Davis’ research has been continuously funded by the National Institutes of Health (NIH) for more than 20 years. Working with a UT Southwestern colleague specializing in mouse genetic models of blood vessel development, he is co-principal investigator for a four-year, $1 million grant from the NIH’s National Health, Lung and Blood Institute. The project focuses on the role of VEGF-dependent signaling molecules in controlling downstream growth factors that prompt blood vessel formation.
Dr. Davis was among a group of scientists world-wide to develop the first consensus guidelines for use and interpretation of angiogenesis assays, published last year in the journal Angiogenesis. He has authored more than 140 peer-reviewed publications, and served as a member and chair of the NIH Cardiovascular Differentiation and Development Study Section.
Some Things You May Not Know About Dr. Davis
Drafted as an outfielder by a well-known baseball scout, he played after high school in a San Francisco Giants minor league team for a year before entering college. His perspective as a physician-scientist was shaped by intense determination that enabled him to excel at a team sport, and early medical training that included learning about a broad range of diseases and treating patients. “In science,” Dr. Davis said, “it’s critical to focus on the details, but you also need to understand the overall playing field, the big picture.”
Davis and his wife Nancy have four grown children: a daughter, age 22, and three sons, ages 24 to 37.
-Photos by Allison Long, USF Health Communications and Marketing; Microscopic image of capillary co-assembly courtesy of George Davis laboratory
David Kang probes brain changes in aging that tip the balance toward dementia
His team searches beyond the hallmark Alzheimer’s disease proteins for alternative treatments
In his laboratory at the USF Health Byrd Alzheimer’s Center, neuroscientist David Kang, PhD, focuses on how different types of proteins damage the brain when they accumulate there. In the case of Alzheimer’s disease, decades of good science has zeroed in on amyloid and tau, as the two types of hallmark proteins driving the disease process that ultimately kills brain cells.
Dr. Kang and his team investigate molecular pathways leading to the formation large, sticky amyloid plaques between brain cells, and to the tau neurofibrillary tangles inside brain cells –including the interplay between the two proteins. But, he is quick to point out that amyloid and tau are “not the full story” in the quest to understand how normally aging brains go bad.
“Our goal is to understand as much of the entire Alzheimer’s disease process as possible and then target specific molecules that are either overactive or underactive, which is part of the drug discovery program we’re working on,” said Dr. Kang, professor of molecular medicine and director of basic research for the Byrd Alzheimer’s Center, which anchors the USF Health Neuroscience Institute.
Neuroscientist David Kang, PhD, (third from left) stands with his team in his laboratory at the Byrd Alzheimer’s Center, which anchors the USF Health Neuroscience Institute.
Attacking dementia from different angles
Dr. Kang’s group takes a multifaceted approach to studying the biological brain changes that impair thinking and memory in people with Alzheimer’s, the most common type of dementia, as well as Lewy body, vascular and frontotemporal dementias.
That includes examining how damaged mitochondria, the energy-producing power plants of the cell, contribute to pathology in all neurodegenerative diseases. “Sick mitochondria leak a lot of toxins that do widespread damage to neurons and other cells,” Dr. Kang said.
Dr. Kang’s team was the first to identify how mutations of a gene, called CHCHD10, which contributes to both frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS), cause both mitochondrial dysfunction and protein pathology called TDP-43. Their findings on the newly identified mitochondrial link to both neurodegenerative diseases were published in Nature Communications in 2017.
The role of selective degradation in ridding cells of abnormal proteins, old or damaged organelles (including mitochondria) and other debris is another key line of research pursued by Dr. Kang and colleagues.
A single stained nerve cell | Microscopic image courtesy of Kang lab.
“We believe something more fundamental is going wrong in the brain during the aging process to tip the balance toward Alzheimer’s disease – beyond what we call proteinopathy” or deposits of malformed proteins like toxic amyloid and tau, said Dr. Kang, whose work is bolstered by nearly $8 million in grant funding from the National Institutes of Health (NIH), the Veterans Administration (VA merit awards) and the Florida Department of Health.
“I think one of the fundamental things happening is that the (cellular) plumbing system isn’t working to clear out all the accumulating junk,” he said. “That’s why we’re looking at the protective clearance mechanisms (autophagy and mitophagy) that would normally quickly remove misfolded proteins and dysfunctional mitochondria.”
Unfortunately, pharmaceutical trials to date have yielded no effective treatments for Alzheimer’s disease, the sixth leading cause of death in the U.S. Most clinical studies have centered on developing medications to block or destroy the amyloid protein plaque formation, and a few have targeted the tau-containing neurofibrillary tangles. The five Alzheimer’s drugs currently available may provide temporary relief of symptoms, such as memory loss and confusion. But, they do not prevent or delay the mind-robbing disease as toxic proteins continue to build up and dismantle the brain’s communication network.
Lesson learned: The critical importance of intervening earlier
Some scientists argue that the “amyloid hypothesis” approach is not working. Dr. Kang is among those who maintain that amyloid plays a key role in initiating the disease process that leads to brain atrophy in Alzheimer’s – but that amyloid accumulation happens very early, as much as 10 to 20 years before people experience memory problems or other signs of dementia.
Early detection and treatment are key, Dr. Kang says, because as protein plaques and other lesions continue to accumulate in the brain, reversing the damage may not be possible.
“One reason we’ve been disappointed in the clinical trials is because so far they have primarily targeted patients who are already symptomatic,” Dr. Kang said. “Over the last decade we’ve learned that by the time someone is diagnosed with early Alzheimer’s disease, or even mild cognitive impairment, the brain has degenerated a lot. And once those nerve cells are gone they do not, for the most part, regenerate… The amyloid cascade has run its course.”
As protein plaques and other lesions continue to accumulate, becoming apparent with MRI imaging, reversing the damage may not be possible. So, for anti-amyloid therapies – or even those targeting downstream tau – to work, patients at risk of Alzheimer’s need to be identified and treated very early, Dr. Kang said.
USF Health is recruiting healthy older adults with no signs of memory problems for a few prevention trials. A pair of Generation Program studies will test the effectiveness of investigational anti-Alzheimer’s drugs on those at high genetic risk for the disease before symptoms start. And, the NIH-sponsored Preventing Alzheimer’s with Cognitive Training (PACT) study is examining whether a specific type of computerized brain training can reduce the risk of mild cognitive impairment and dementias like Alzheimer’s disease in those age 65 and older.
To accelerate early intervention initiatives, more definitive tests are needed to pinpoint biomarkers that will predict Alzheimer’s disease development in genetically susceptible people. Dr. Kang is hopeful about the prospects. His own team investigates how exosomes, in particular the lipid vesicles that shuttle proteins and other molecules from the brain into the circulating bloodstream, might be isolated and used to detect people at risk of proteinopathy.
“I think within the next five years, some type of diagnostic blood test will be available that can accurately identify people with early Alzheimer’s brain pathology, but not yet experiencing symptoms,” he said.
Graduate research assistant Yan Yan, a member of Dr. Kang’s research team, works at a cell culture hood.
Searching for alternative treatment targets
Meanwhile, Dr. Kang’s laboratory continues searching for other treatment targets in addition to amyloid and tau — including the enzyme SSH1, which regulates the internal infrastructure of nerve cells, called the actin cytoskeleton. SSHI, also known as slingshot, is needed for amyloid activation of cofilin, a protein identified by the USF Health neuroscientists in a recent study published in Communications Biology as a possible early culprit in the tauopathy process.
“Cofilin is overactive in the brains of Alzheimer’s patients so if we can inhibit cofilin by targeting slingshot, it may lead to a promising treatment,” Dr. Kang said.
Ultimately, as with other complex chronic diseases, Alzheimer’s may not be eliminated by a single silver-bullet cure. Rather, Dr. Kang said, a combination of approaches will likely be needed to successfully combat the neurodegenerative disorder, which afflicts 5.8 million Americans.
“I think prevention through healthy living is definitely key, because brain aging is modifiable based on things like your diet as well as physical activity and brain exercises,” he said. “Also, we need to focus on earlier diagnosis, before people become symptomatic, and develop next-generation drugs that can attack the disease on multiple fronts.”
Xingyu Zhao, PhD, a research associate in the Department of Molecular Medicine, is among the scientists in Dr. Kang’s laboratory studying the basic biology of the aging brain.
Fascinated by how the brain works — and malfunctions
Dr. Kang came to USF Health in 2012 after nearly 20 years as a brain researcher at the University of California San Diego, where he earned M.S. and PhD degrees in neurosciences and completed NIH National Research Service Award fellowships in the neuroplasticity of aging.
As an undergraduate Dr. Kang switched from studying engineering to a dual major in science/psychology. He began focusing on neurosciences in graduate school, he said, because tackling how the brain works and malfunctions was fascinating and always challenged him.
“With every small step forward, we learn something else about the basic biology of the aging brain,” said Dr. Kang, “It’s not just helpful in discovering what therapeutic approaches may work best against Alzheimer’s disease – we’re also learning more about other neurodegenerative conditions affecting the brain.”
In addition to leading day-to-day research operations at the Byrd Center and helping to recruit new Alzheimer’s investigators, Dr. Kang holds the Mary and Louis Fleming Endowed Chair in Alzheimer’s Research and serves as a research neurobiologist at the James A. Haley Veterans Haley Veterans’ Hospital.
He has authored more than 50 peer-reviewed journal articles on brain aging and Alzheimer’s disease research. A member of the NIH Clinical Neuroscience and Neurodegeneration Study Section since 2016, he has served on multiple national and international editorial boards, scientific panels and advisory boards.
Dr. Kang sits next to a computer monitor depicting stained microscopic images — a single neuron (far left) and the two hallmark pathological proteins for Alzheimer’s disease, tau tangles (center) and amyloid plaques (right).
Some things you may not know about Dr. Kang
His parents were Presbyterian missionaries in Africa, so he spent nine years of his early life (third through 10th grade) in Nigeria.
Dr. Kang practices intermittent fasting, often forgoing breakfast and eating only within an 8-hour window. Animal studies indicate the practice may contribute to lifespan and brain health by improving cellular repair through the process of autophagy, he said. “Autophagy really kicks your cells’ plumbing system into gear to clear out all the waste.”
-Video and photos by Allison Long, USF Health Communications and Marketing
USF Health scientist part of urgent global effort to rid Southeast Asia of malaria
What Dr. Liwang Cui’s team learns in the laboratory and the field can help curb the spread of this increasingly multidrug-resistant, mosquito-borne disease
USF Health molecular parasitologist Liwang Cui, PhD, co-leads one of 11 international centers of excellence for malaria research funded by the NIH’s National Institute of Allergy and Infectious Diseases.
While some of the largest declines in malaria deaths since 2010 have been reported worldwide, the battle continues against the parasite’s resistance to artemisinin-based antimalarial drugs and an emerging threat of mosquito insecticide resistance.
USF Health molecular parasitologist Liwang Cui, PhD, leads a team that is part of an ambitious effort to eliminate malaria from the Greater Mekong subregion (GMS) in Southeast Asia — six countries bound together by the Mekong River — by 2030. Malaria spreads to people through the bites of female Anopheles mosquitoes (malaria vectors), which inject Plasmodium parasites into the bloodstream. The GMS eradication effort targets both Plasmodium falciparum, the most lethal form of human malaria, and Plasmodium vivax, a less virulent species but the most widespread globally.
Public health officials want to avert a crisis that could arise if the artemisinin-resistant malaria parasites so prevalent in the GMS reach India and then sub-Saharan Africa, where the greatest international burden of malaria lies.
Dr. Cui’s team conducts malaria field studies focused primarily along the border areas of six countries comprising Southeast Asia’s Greater Mekong subregion.
Collectively these researchers — working both in Dr. Cui’s laboratory at USF and international field study sites — possess diverse expertise, including malaria parasite developmental biology, epidemiology, vector biology, genomics, epigenetics, host-parasite interactions and drug resistance. The Cui laboratory strengthens USF’s growing cadre of medical and public health investigators committed to taking the university’s global infectious diseases research to the next level.
Dr. Cui (center) with team members who joined him at USF from Pennsylvania State University. Their native countries include Cameroon, China, India, Thailand, Uganda and the United States.
And, while malaria is not endemic to the United States, the advances they make have implications closer to home. Travelers who live in areas with no malaria transmission, and therefore acquire no malaria immunity, are among those most susceptible to the severest form of the disease.
“With all vector-borne diseases, our goal is to learn more about the mechanisms of infection, to try to prevent multidrug resistance, and to eradicate the disease before it comes to our doorstep,” Dr. Cui said.
“International travel is so easy now. Today in Florida — tomorrow in Africa, Asia or South America… A few years ago, no one considered that the Zika virus would reach this country, but now the National Institutes of Health is funding Zika research to avoid further outbreaks.”
Postdoctoral scholar Amuza Lucky, PhD, lifts a container of malaria parasites cryopreserved in liquid nitrogen.
The six GMS countries include China, the region’s more economically advanced, which has invested significantly in malaria surveillance, diagnosis, treatment and prevention, particularly in the Yunnan Province counties near its border. Meanwhile hotbeds of the mosquito-borne disease remain in bordering Myanmar, an economically depressed GMS country plagued for decades by ongoing civil conflicts.
“Mosquitoes do not need passports to cross borders, so it is critical to prevent disease introduction and reintroduction at international borders” Dr. Cui said.
Even though China and Thailand have little or almost no malaria now, mosquitoes harboring the parasite on the Myanmar side of the borders, along with highly mobile and displaced ethnic minority populations, could potentially undermine the entire region’s recent progress in combatting malaria, he explained.
After reviving the malaria parasites from cold storage, researchers manipulate the genome of parasites, including those isolated from patients. They use the latest molecular techniques to study gene functions and learn more about drug-resistant malaria.
Dr. Cui is principal co-investigator for the Southeast Asia Malaria Research Center, a network of institutions in the U.S., Thailand, Myanmar and China where internationally recognized investigators conduct coordinated malaria research and education projects.
One of 11 international centers of excellence for malaria research funded by the National Institute of Allergy and Infectious Diseases (NIAID), the Southeast Asia center (with USF as the lead institution) is currently supported by a six-year, $9.2 million NIAID grant. Its interdisciplinary researchers seek to better define how mobile human populations, parasite drug and insecticide resistance, and mosquito biology contribute to ongoing malaria transmission along international borders, so that more effective control measures can be developed and strategically deployed.
They remain focused on the overarching goal — achieving the World Health Organization’s (WHO) plan to eradicate the life-threatening malaria parasite P. falciparum in the Greater Mekong subregion by 2025, and to make the region malaria free by 2030.
Research technician Xiaolian Li
Led by China, the political will to eliminate malaria throughout the region is strong, Dr. Cui said, but the daunting challenges to be overcome require a multipronged approach. Among the challenges:
– Multidrug resistance: The malaria parasite P. falciparum has become increasingly resistant to artemisinin or its derivatives and to several partner antimalarial drugs given in combination with artemisinins. Resistance first reported in 2008 along Cambodia-Thailand border areas has spread to parts of other countries, including Laos, Myanmar and Vietnam. This means the drugs take longer to work, and urgency has risen to find new cost-effective compounds to delay drug resistance and halt malaria’s spread.
– Less insecticide effectiveness: Insecticides that reduce the number of mosquitoes have been shown to make a big difference in the incidence of malaria cases. But, growing mosquito resistance to insectides used in long-lasting bed nets and sprayed indoors has become an emerging threat. In Thailand, Dr. Cui said, resistance to pyrethroids, one of only two WHO-approved insecticides to treat bed nets, is now emerging.
Magnified P. falciparum malaria parasites (dark purple) inside human red blood cells stained blue.
– Barriers to care: Malaria distribution is geographically uneven. Vulnerable populations living near borders and remote rural areas lacking access to the latest diagnostics and treatment bear a disproportionate burden of the disease.
– Fake drugs: Falsified and substandard antimalarials present another growing concern, according to WHO. A 2017 survey reported that nearly 28 percent of the antimalarial medicines sold by the private sector in Myanmar were still monotherapy (a single drug) — not the artemisinin-based combination therapy (ACT) recommended as the first-line treatment for P. falciparum to provide an adequate cure rate. Drug resistance can develop when the concentration of antimalarials in the blood is lacking – at least two drugs (a fast-acting artemisinin and second longer-lasting ingredient) are needed to completely clear malaria parasites, Dr. Cui said.
Beakers containing the media used to feed malaria parasites.
While the challenges complicate elimination efforts, Dr. Cui’s group and others are fighting back with the latest technology. They employ drones to help map topography and learn more about potential mosquito breeding habitats (ponds, puddles of water) in areas difficult reach by land. They use advanced genetic tools to help track the cross-border movement of malaria parasites and to look for new ways to attack all stages in the complex life cycle of Plasmodium, which can readily mutate to avoid immune system destruction and evolve resistance to drugs.
Dr. Cui is the lead investigator for a $1-million R01 grant from NIAID to identify molecular markers that can help manage P. falciparum malaria’s artemisinin resistance with targeted control measures.
Dr. Cui (green shirt) oversees a team surveying mosquito larvae at a remote field site near the China-Myanmar border, where malaria is endemic.
As he works with his team at the epicenter of drug-resistant malaria in Southeast Asia — whether collecting blood samples used to isolate malaria parasites for genomic sequencing or surveying mosquito larvae sites in rural areas near the China-Myanmar border — Dr. Cui is reminded of the real-world costs of malaria, both in human lives and socioeconomic productivity.
“Malaria is a preventable and treatable,” he said, “so why are we still seeing more than 200 million cases a year worldwide, and nearly half a million, mostly kids, die from this disease?” he said. “When you see the suffering of the people in villages and (refugee) camps it really touches your heart and helps drive your work toward malaria control and elimination.”
USF Preeminence funding helped recruit internationally-recognized malaria researcher Liwang Cui and his team to Tampa.
Dr. Cui received one PhD degree in biology from Moldova Agricultural University (former USSR) and a second in molecular virology from the University of Kentucky; he conducted a postdoctoral fellowship in entomology and molecular parasitology at Walter Reed Army Institute of Research, Washington, DC. Before coming to USF, he spent 18 years as a professor in Penn State’s Department of Entomology.
Dr. Cui’s malaria research related to parasites, mosquito vectors and human hosts has been continuously funded by the NIH since 2001, and he was awarded two training grants from the NIH Fogarty International Center and one WHO grant for tropical disease research. He has authored more than 240 peer-reviewed publications, including five book chapters. He currently serves as the academic editor of several scientific journals.
Some things you may not know about Dr. Cui:
While he has worked in some very remote areas where malaria remains rampant, Dr. Cui takes plenty of precautions to avoid infection. During his years of field studies in Southeast Asia, he has experienced the effects of civil war and even an earthquake – but he has not contracted malaria.
Some of his most productive scientific research writing is done late at night and the early hours of the morning, fueled by a pot of freshly brewed tea.
One of Dr. Cui’s first introductions to Florida wildlife came during an evening jog around the New Tampa subdivision where he lives with wife Rosabel and their two children: Stephen, 3, and Sophia, 9. He stopped short before nearly running over a 6-foot alligator lying in his path – then took a photo with his smartphone from a safe distance.
Close-up of the Anopheles mosquito that carries the malaria parasite.
Some late-stage malaria parasites growing inside human red blood cells.
Written by Anne Delotto Baier. Photos by Torie Doll, USF Health Communications and Marketing. Field site photos courtesy of Dr. Liwang Cui.
Sea squirt’s microbiome offers clues to frontline immune defense, gut health
National Science Foundation research by USF Health’s Larry Dishaw has relevance for debilitating digestive disorders like inflammatory bowel disease
Supported by a four-year, $867,581 grant from the National Science Foundation (NSF), Dr. Dishaw’s immune research explores the populations of microbes living together in the gut – trillions of bacteria, viruses, fungi and even some parasites – collectively known as the gut microbiome. His team is also interested in defining how disruption of a stable gut microbiome relates to the onset of inflammatory bowel disease, an autoimmune disorder affecting 1.6 million Americans.
Microbiologist Larry Dishaw, PhD, associate professor of pediatrics at the USF Health Morsani College of Medicine.
“Many factors are at play in regulating the microbiome,” said Dr. Dishaw, an associate professor of pediatrics at USF Health Morsani College of Medicine. “And a lot of researchers around the world are interested in how the microbiome colonizes in animals, how this microbial community achieves stability, how it can shift out of balance, and how it is relevant to protecting human health.”
One Of Closest Evolutionary Invertebrate Relatives Of Humans
Ciona intestinalis, an invertebrate marine animal resembling a spongy plant, spends its life attached to underwater structures like boat hulls and pier pilings where it continually siphons in water through one end, filtering out plankton and algae to eat and then squirting water and waste out the other end.
If you look under the microscope at the translucent juvenile sea squirts cultivated by Dr. Dishaw’s lab you can see a gut (with a stomach and intestinal compartment) similar to the digestive tract of vertebrates. Despite its primitive appearance, the sea creature – a protochordate — is considered one of the closest evolutionary invertebrate relatives of humans.
Unlike humans, sea squirts rely their entire lives solely on naturally present innate immunity for host defense. They lack the other more evolved arm of immunity know as adaptive immunity, which differs from one animal species to the next and activates in response to specific foreign invaders.
The adult sea squirt Ciona intestinalis.
That, Dr. Dishaw says, is what helps make the sea creature a good model for investigating the evolution of the host immune system and its role in creating and maintaining a well-balanced gut microbiome.
“When humans are born, they do not develop adaptive immunity until ages 3 to 5… so the initial process of colonizing bacteria and other microbes that make up their microbiomes is all mediated by innate immunity,” he said.
With a simpler model system like the sea squirt, the researchers can look at innate immunity in isolation. They can monitor how the immune system responds when encountering microbes for the first time, how microbes initially “choose” where to colonize in the gut, and how the animal maintains homeostasis – that is, a healthy balance of gut microbes needed to digest food and absorb nutrients throughout its life.
Ultimately, the not-so-lowly sea squirt may provide better insight into gut defense evolution and define ways that beneficial (non-pathogenic) bacteria may help prevent the overgrowth of disease-causing (pathogenic) bacteria.
Tipping The “Good” Microbe – “Bad” Microbe Balance
Dr. Dishaw’s NSF grant builds upon the work of his mentor Gary Litman, PhD, USF professor emeritus of pediatrics, whose lab discovered the genes for a family of variable region-containing chitin-binding proteins, or VCBPs, made and secreted by the gut wall in a different protochordate, the ancient fish-like organism amphioxus. These immune proteins appear to help regulate how bacteria and fungi interact and grow such that intestinal barrier function is enhanced, Dr. Dishaw said. The VCBPs also seem to influence the production or release of phages, viruses that infect and destroy specific gut bacteria, he added.
Dr. Dishaw with some laboratory team members. From left: Michael Schepps, undergraduate research assistant; Zachary Graham, lab technician; and Julie Voelschow, undergraduate research assistant. Not pictured are Ojas Natarajan, PhD, postdoctoral scientist; and Celine Atkinson, graduate student.
“The goal of our project is to find out what happens when we muck with that system. When the VCBPs don’t bind fungi or bacteria correctly, how is the physiological fitness of the animal affected?” Dr. Dishaw said. “We believe these immune effectors can shape the ecology of the gut microbiome in ways that promote (or deter) health. And we think the sea squirt model can help us understand how that happens.”
The researchers can rear germ-free juvenile Ciona, then introduce into their water whatever microbes they choose. Because the sea squirt filters water constantly, they can quickly track where in this controlled environment the newly introduced gut microbe populations settle and thrive as well as examine the role host-microbe interactions play in developing a frontline immune defense system.
In a series of experiments published last year in Open Biology, the USF researchers induced colitis-like inflammation and damage in sea squirts by exposing their guts to the chemical dextran sulfate sodium (DSS). They showed that DSS altered the production and settlement of the secreted immune molecule that binds bacteria.
The laboratory grows its own algae, cultured in beakers shown on the windowsill, to feed to the juvenile sea squirts.
Most invertebrates, including sea squirts, defend their gut walls against potential microbial attack and prevent infection with mucous rich in chitin. The researchers found that pretreatment with microparticles of chitin, a fibrous substance prevalent in Ciona’s epithelium-associated gut mucous, protected the animal from the colitis-like effects of subsequent DSS exposure.
Ciona, which permits the study of innate immunity in isolation, “may help us determine how innate immunity modulates recovery from colitis and the re-establishment of (gut microbiome) homeostasis,” the study authors concluded.
Promising Power Of Phages To Treat Drug-Resistant Infections
Dr. Dishaw is also co-investigator for a National Institutes of Health grant led by the USF College of Nursing’s Maureen Groer, PhD. The project is investigating the link between the gut microbiome of premature infants and their health as they age.
Lab tech Zachary Graham and Dr. Dishaw look over culture plates of bacterial biofilms stained with crystal violet. The bacteria were recovered from the gut of the sea squirts.
“We think early colonization of microbes is critically important in establishing lifelong gut microbiome features,” Dr. Dishaw said. “So, early life events that shape the (evolving) microbiome — like spending 4 to 6 weeks in a neonatal intensive care unit where the infant receives multiple antibiotics to treat or prevent infections — could translate into long-term changes in health.”
The ultimate goal of microbiome research is to come up with effective treatments to reset the equilibrium of a microbial community gone awry – whether by diet, stress, pathogens, or even the medications used to fight disease.
Fecal microbiota transplants, which insert a healthy donor’s fecal matter into a recipient’s colon to reconstitute a stable gut microbiome, have already become a relatively common treatment for Clostridium difficile (C. diff) infection, a debilitating gastrointestinal disease resistant to many antibiotics.
The crystal violet-stained cultures allow researchers to estimate the amount of biofilm formed during stationary growth of the bacteria.
In an age of growing multidrug resistance, Dr. Dishaw believes that phage therapy also offers promise as an alternative or supplement to antibiotics for patients suffering from recurrent, difficult-to-treat infections. In another NSF grant with co-principal investigator Mya Breitbart of the USF College of Marine Sciences, he is colonizing a variety of bacteria in the sea squirts in an effort to test how phages – the viruses that infect bacteria – can be used to selectively eliminate harmful gut bacteria.
Phages, which specifically target single types of bacteria, might be harnessed to chase pathogens out of the microbial community without having to rely on antibiotics, which can wipe out both harmful and beneficial bacteria, Dr. Dishaw said. “In theory, you could use a C diff phage with high precision to target and kill only C. diff bacteria.”
Dr. Dishaw also participates in a clinical study with the USF Health Department of Pediatrics and Johns Hopkins All Children’s Hospital to characterize the gut microbiomes of patients with primary immune deficiencies. The research may provide a better understanding of how different aspects of immunity regulate the gut microbiome.
Microscopic image of a juvenile sea squirt shows its gut colonized by fluorescently labeled bacteria.
Written by Anne Delotto Baier, Photos and Video by Torie M. Doll, USF Health Communications and Marketing
Infectious diseases expert looks for new ways to combat resilient parasites
USF Health’s Dr. Kami Kim probes the epigenetics of two global parasitic infections, malaria and toxoplasmosis
While an undergraduate at Harvard University, Kami Kim, MD, participated in a research thesis project exploring leukemia’s resistance to chemotherapy and the effectiveness of combination drugs in combatting it. While she was excited to help figure out (and publish) a mechanism, she recalls that she signed on for this laboratory research primarily “to help in get into medical school.”
Her interest in research intensified in medical school in early 1980s at the beginning of the domestic AIDS era, about the same time tuberculosis cases were exploding and malaria, once considered virtually eliminated as a major public health threat, began to re-emerge globally.
Kami Kim, MD, a USF Health professor of infectious disease, with her multidisciplinary laboratory research team, which includes expertise from the medicine, public health, mathematics and statistics.
Undergraduate laboratory work that sparked a lifelong passion for research
Basic science and clinical infectious diseases expertise
After clinical training as an infectious diseases fellow at the University of California San Francisco (UCSF) – and witnessing firsthand the devastating consequences of acquired immunodeficiency syndrome – Dr. Kim returned to laboratory research, with an emphasis on parasitic infectious diseases. Meanwhile, she continued to see patients as an attending academic physician at some of the nation’s best hospitals in San Francisco and New York City.
That blend of rigorous clinical and basic science expertise makes Dr. Kim one of the first of several high-profile, energetic recruits who will help take USF Health’s global infectious disease research to the next level.
Dr. Kim came to USF Health in November 2017 from Albert Einstein College of Medicine in New York City, where she was a professor of medicine, microbiology and immunology, and pathology. In addition to her laboratory research at USF, she consults monthly on infectious diseases cases at Tampa General Hospital. At Einstein, she directed the infectious diseases section of the Center for Epigenomics and helped launch and led the National Institutes of Health-funded Geographic Medicine and Emerging Infections Training Program, which supports interdisciplinary training in translational research for pre-doctoral students, post-doctoral research fellows and clinical fellows.
A plaque formation of the intracellular parasite Toxoplasma gondii
Seeking solutions to life-threatening global parasitic diseases
Dr. Kim’s USF Health research team, working out of a laboratory in the university’s research park, focuses on two major areas — malaria and toxoplasmosis. The world’s most dangerous parasitic disease, malaria claims more than 2 million victims and 445,000 deaths yearly, primarily in sub-Saharan Africa. Toxoplasmosis, often asymptomatic, can be life-threatening to babies born to women infected during pregnancy and people with weakened immune systems.
Toxoplasmosis project: Combining advanced techniques from genetics, cell biology and proteomics, the researchers investigate the ways that epigenetics – the interface of genetics and environmental factors – regulate development of chronic infection by the cat-borne gondii parasite. They seek to understand how this pervasive parasite switches back and forth between a rapidly dividing acute stage destructive to healthy tissue (tachyzoite) and a chronic, or dormant, stage, where bradyzoite forms within pseudocysts remain invisible to the immune system. Dr. Kim collaborates with other leading Toxoplasma experts: Distinguished USF Health Professor Michael White, PhD, a long-time colleague, as well as investigators at Indiana University, Pennsylvania State University and Albert Einstein College of Medicine.
Malaria project: In the hot, wet regions of Africa, mosquitoes are ubiquitous and children exposed to malaria from birth may contract the infection several time a year. The overwhelming majority of clinical cases are uncomplicated, with flu-like symptoms of fever and malaise that typically resolve. Researchers are trying to determine why a small percentage of individuals, in particular certain children, are more likely to develop severe malaria with coma and death (cerebral malaria) or long-term neurological complications such as seizures and cognitive and behavioral problems. In particular, the USF team is assessing specific biomarkers, or genetic predispositions, and parasite or host factors that may help predict disease development or its outcomes.
Dr. Kim discusses research correlating HIV co-infection with cerebral malaria
Research instructor Iset Vera
Both research initiatives harness the latest genomic technology to better understand how immunity works within the framework of host-parasite interactions – all with the aim of devising better or first-time treatments.
Valuable insights into cerebral malaria, future therapies
With collaborators from the Blantyre Malaria Project, based in Malawi, Africa, Dr. Kim published a high-profile paper in mBIO in 2015 reporting for the first time that children co-infected with HIV were much more likely than those who were not to die from severe malaria. Autopsies of the children who died from cerebral malaria indicated that those with HIV had brain blood vessels more clogged with white blood cells and platelets than those of children with malaria alone. HIV appeared to rev up brain inflammation that could lead to death.
In another study, published in Cell Host & Microbe in 2017, Dr. Kim and colleagues used neuroimaging, parasite transcript profiling and laboratory blood profiles to develop machine-learning models of malarial retinopathy and brain swelling. The researchers found that the interaction of high parasite biomass, low platelet levels and certain parasite protein variants that bind to the endothelial protein C receptor (EPCR) play a pivotal role in fatal cases of malaria. Their findings added strength to the rationale that anti-inflammatory and anticoagulant treatments counteracting the breakdown of endothelium may benefit those with severe malaria.
“We still don’t entirely know why some of these kids get super sick and have complications requiring hospitalization,” Dr. Kim said. “If we could figure that out we could save lives, reduce complications and use limited healthcare dollars more effectively in these under-resourced countries.”
The “Goldilocks” theory of immunity
When it comes to infectious diseases, too much of a good thing may make you sick. Dr. Kim calls it the “Goldilocks” theory of immunity – not too much (overactive immune system) and not too little (under-responsive immune system).
For instance, “for someone with malaria the right amount of immunity might not be just the right amount if they already also have tuberculosis,” Dr. Kim said. “What we’re realizing now with the human immune response to parasites or other foreign invaders (pathogens) is that you have to get the balance just right, so you get rid of the pathogen without damaging the human host.”
Otherwise, she added, even after the pathogen is eliminated, long-term complications like a damaging autoimmune inflammatory condition may linger.
Rigorously studying the dynamics of host-parasite interaction – including how parasites hijack the epigenome, which adjusts specific genes in response to signals from the outside world such as diet and stress — is critical to bridging the gap between discovery and effective treatments for different subgroups of infected patients.
“Both the pathogen and the infected host are duking it out to see which one wins, so figuring out what’s happening on both sides is really important to understanding immunity – how our body fights off disease,” Dr. Kim said. “Using genomic information to tell us who’s most susceptible to certain conditions will likely help us to tailor therapies to the individual, or perhaps to know who needs to be vaccinated.”
Dr. Kim with Li-Min Ting, PhD, an assistant professor in the Department of Internal Medicine’s Division of Infectious Disease
Striking the right balance of immunity
Potential applications for other diseases
Within their complex life cycles, both malaria and toxoplasma parasites have dormant forms that the human immune system can’t identify and kill, and antimicrobial drugs can’t touch. For malaria, this silent form lurks in the liver. For Toxoplasma, cysts can settle quietly into the infected person’s brain and muscle tissue without replicating, sometimes for years, until weakened immunity reactivates the disease.
Dr. Kim and other researchers continue to look for new ways to combat chronic infection by parasites.
“Normally when treating a disease you think of killing the form that makes a person clinically symptomatic,” she said, “but with both malaria and Toxoplasma if you can kill the biologically silent form, which is absolutely essential for the disease to continue, you’re accomplishing the same thing.”
Although Dr. Kim’s group targets specific problems underlying malaria and toxoplasmosis, such immune research may have broad applications for understanding and treating other conditions. For instance, atherosclerosis has been linked to the release of molecules from the immune system that can cause inflammation, blood vessel injury and plaque instability leading to heart attacks and stroke.
A T. gondii plaque assay
“Even though drug companies, because of financial return on investment, aren’t necessarily willing to invest in research on malaria host factors,” Dr. Kim said, “they are really interested in stroke and cardiovascular disease. And the big players in the kind of inflammation seen in these two major diseases are platelets and monocytes” – the same inflammatory culprits implicated in cerebral malaria.
While more research is needed, perhaps statin and antiplatelet drugs already approved for another indication could be effective in helping combat malaria,” she said. “It’s entirely possible by better understanding what’s a good immune response to malaria in one situation and bad in another will lead to insights that can be used to develop treatments for other diseases, or insight into what’s protective in another disease.”
Pursuing new approaches to outsmart elusive pathogens
Dr. Kim received her MD degree from the Columbia College of Physicians and Surgeons in New York City. She completed her residency in medicine at Columbia-Presbyterian Medical Center, a clinical fellowship in infectious diseases at UC San Francisco, and two postdoctoral research fellowships – one in parasitology at San Francisco General Hospital and a second in microbiology and immunology at Stanford University.
Dr. Kim is a fellow of the Infectious Diseases Society of America and the American Academy of Microbiology. She is also an elected member of American Society for Clinical Investigation and the Association of American Physicians, national honor societies for physician-scientists. A recipient of the Burroughs Wellcome Fund (BWF) New Investigator Award in Molecular Parasitology early in her career, she has served on the BWF Postdoctoral Research Enrichment Program’s scientific advisory board since 2014. She is a member of the NIH Pathogenic Eukaryotes Study Section.
The cutting-edge tools of computational genomics contribute to Dr. Kim’s work
Throughout much of her career, Dr. Kim’s research has been funded by the National Institute of Allergy and Infectious Diseases. She holds several patents, and was one of the first investigators to develop techniques to genetically manipulate T. gondii. She is the co-editor and currently preparing the third edition of Toxoplasma gondii, the Model Apicomplexan: Perspectives and Methods, a textbook widely considered the seminal source for scientists and physicians working with this parasite.
Dr. Kim said she was attracted to USF because of the university’s upward national trajectory and USF Health leadership’s commitment to building translational research and pursuing innovative approaches and excellence in all its academic missions. She is a member of the USF-wide Genomics Program.
“I enjoy being part of a USF clinical community that excels in the treatment of infectious diseases and working with physicians and scientists who do parasitology research,” she said. “You constantly have to think outside the box and come up with clever strategies – because we’re dealing with pathogens that do not behave like they are supposed to.”
Toxplasma parasite strategy for survival
Dr. Kim, one of the first investigators to develop techniques to genetically manipulate Toxoplasma gondii, co-edits a textbook widely considered the seminal source for scientists and physicians working with this parasite
Some things you might not know about Dr. Kim
Korean was her first language, and she also speaks Spanish.
She enjoys food, arts and crafts, and travel. Countries she has visited include Korea, Malawi, South Africa, France, Japan, and Brazil.
Kim is married to Thomas McDonald, MD, USF Health professor of cardiovascular sciences and director of the new Cardiogenetics Clinic. They have two sons who are both mathematicians, one in a mathematics PhD program and the other in college. They met in the cardiac intensive care unit at Columbia Presbyterian Medical Center when Dr. McDonald was a resident and Dr. Kim was rounding as a medical student.
Dr. Kim’s career as an infectious disease physician-scientist bridges basic research and clinical practice
Written by Anne Delotto Baier, Photos and Video by Torie M. Doll, USF Health Communications and Marketing
USF Health cardiologist studies genetic predisposition to sudden cardiac death
In the laboratory and the clinic, Dr. Thomas McDonald focuses on inherited heart diseases that can lead to potentially deadly heart rhythm disturbances.
Sudden cardiac death most often makes the news when athletes in peak physical condition collapse and die while exercising or competing. This spring, Zeke Upshaw, 26, a basketball player for the Grand Rapids Drive, a G-league affiliate of the Detroit Pistons, collapsed face-down on home court during the final minute of a game and later died at the hospital. A medical examiner ruled that he had suffered sudden cardiac death.
Most of the 200,000 to 450,000 sudden cardiac deaths each year in United Start are caused by heart rhythm disturbances provoked by certain strenuous activities, prescription medications, recreational drugs, or other triggers. “Sometimes it just happens in your sleep. The most severe and earliest form would be sudden infant death, or SIDS,” Dr. McDonald said.
Physician-scientist Thomas McDonald, MD, a professor in the USF Health Department of Cardiovascular Sciences and member of the USF Health Heart Institute, with his laboratory team.
Dr. McDonald was recruited to the USF Health Heart Institute in October 2017 from Albert Einstein College of Medicine in New York City, where he was a professor of both cardiology and molecular pharmacology. He also co-directed the thriving Montefiore-Einstein Clinic for CardioGenetics, the first such interdisciplinary clinic in metropolitan New York for families at risk of sudden cardiac death from arrhythmias.
At USF Health his laboratory continues to focus on the fundamental causes of heart conditions passed from one generation to the next — and what can be done to help prevent disease and its consequences. The hereditary conditions he studies include those affecting the heart’s electrical system to cause arrhythmias, like long QT syndrome and Brugada syndrome, and those affecting heart muscle, such as hypertrophic cardiomyopathy and dilated cardiomyopathy. While rare, these conditions can substantially increase an individual’s risk for sudden cardiac death and devastate families.
In long QT syndrome, the heart takes longer than normal to recharge between beats. This electrical disturbance, called a prolonged QT interval, can often be seen on an electrocardiogram (ECG) like the one pictured here.
Dr. McDonald has also started a USF Health Cardiogenetics Clinic, modeled after the Montefiore-Einstein center he co-founded, to evaluate and treat families in which members succumb to unexplained sudden cardiac death or SIDS, or where suspicion of an underlying, hereditary heart rhythm disturbance exists.
His work bridging the laboratory and clinic has implications for a much larger population than people with relatively rare inherited cardiac disorders. Dr. McDonald points to growing evidence of the interplay between genetics and environmental factors like diet, exercise and stress.
“By studying these rare or uncommon cardiac diseases,” he said, “we may uncover more generalizable biochemical pathways that could be influenced to harm the heart given the wrong environment — even in genetically unaffected families.”
Studying uncommon (inherited) heart diseases to gain better insight into more common ones.
Dr. McDonald lifts a container including pluripotent stem cells from storage in liquid oxygen. Alexander Bertalovitz, PhD, (right) an assistant professor of cardiovascular sciences who helps manage the cardiogenetics laboratory, followed Dr. McDonald to USF Health from Albert Einstein College of Medicine in New York City.
Pinpointing The Meaning Of Genetic Variants Of “Unknown Significance”
Dr. McDonald analyzes genetic changes, or mutations, which may lead to malfunctioning of ion channels that create electrical signals in the heart.
His team has spent the last few years characterizing the function of 1,000 different mutations found in cardiac ion channel genes associated with hereditary rhythm conditions such as long QT syndrome and Brugada syndrome. The researchers recreate the genetic variations in a cellular model and use automated electrophysiology techniques to analyze how the mutations affect the ion channel’s ability to correctly generate each heartbeat. All these variations have been cited in published scientific literature; however, it is still largely unclear which ones truly increase the risk of abnormal rhythms leading to palpitations, seizures, fainting or sudden death – and which are benign.
“Our ultimate goal is to work with other investigators to create a NIH-curated public database that physicians and genetic counselors could access to find out whether a genetic variant is likely, or unlikely, to cause a potentially life-threatening heart rhythm disturbance in a patient or their family members,” Dr. McDonald said.
As genetic testing is becomes more common, a growing challenge is that lab reports of people referred for DNA sequence testing often come back listing many “variants of unknown significance,” Dr. McDonald said. “That drives physicians and patients crazy because they don’t know what that means… what do they do with that information?”
Dr. McDonald with Jiajia Yang, a PhD student in the Department of Molecular Pharmacology and Physiology.
An important step toward improving the guidance that doctors offer individuals with inherited heart disorders would be the ability to more precisely distinguish between disease-causing mutations and mutations with little or no harmful physiological effects through a resource like a scientifically validated database, he added.
Recommended treatment options for long QT are life-long and vary, including regular cardiac monitoring, taking medication such as beta blockers, restricting strenuous sports activities, or sometimes implanting pacemakers or defibrillators to help control abnormal heartbeats. So, for example, if DNA testing of a child or young adult revealed a long QT genetic variation characterized as having little risk of leading to sudden cardiac death, prescribing beta blockers and routine cardiac monitoring might be the best preventive therapy – avoiding the long-term management and small, but real, lifetime risk of complications from an implantable device.
Dr. McDonald comments on the focus of his laboratory’s research on genetic variations.
Opening Tampa Bay Region’s First CardioGenetics Clinic
The twice-monthly Cardiogenetics Clinic, which opened in March, is held at USF Health Cardiology’s Armenia Avenue location. The new clinic is staffed by a team with the expertise to address the diverse medical, psychological, social and ethical issues arising when evaluating genetic heart conditions that predispose patients to sudden cardiac death.
Dr. McDonald — with certified genetic counselor Melissa Racobaldo (far left) and clinical geneticist Christopher Griffith, MD — leads a comprehensive discussion of family medical history with a patient and his mother referred to the USF Health Cardiogenetics Clinic.
“When I arrived there was no formal cardiogenetics program in the greater metropolitan area of Tampa Bay where 4 million people live — so the prospect of building one from scratch was very attractive,” said Dr. McDonald, who specializes in adult cardiology. He leads the clinic working with USF Health faculty members Christopher Griffith, MD, assistant professor of pediatrics and a clinical geneticist; and Melissa Racobaldo, a genetic counselor; as well as Gary Stapleton, MD, a pediatric interventional cardiologist from Johns Hopkins All Children’s Hospital. USF College of Public Health students specializing in genetic counseling are expected to join the clinic in coming months.
Many with congenital cardiac conditions have no signs or symptoms. Patients and their families referred to the clinic typically have experienced a history of arrhythmias or other cardiac events, or suffered the unexpected death of a loved one.
During the initial visit, families meet with team members for a cardiac history and examination, review of medical records and/or autopsy reports, and baseline tests that include an electrocardiogram and echocardiogram. Based on the family’s medical history, a tree-like chart known as the DNA pedigree is created to identify familial genetic patterns and sudden unexpected deaths linked to cardiac disorders. “The most important genetic test is still a complete family history,” Dr. McDonald said.
Certified genetic counselor Melissa Racobaldo consults with Dr. McDonald about a patient.
DNA testing is usually only recommended when the team discerns that the pattern of cardiac-related events is highly likely to be genetic rather than environmental. For instance, a family history indicating that a few relatives died from heart disease in their 80s might be considered environmental.
If a mutation is found in one of the genes known to be associated with a dangerous cardiac arrhythmia, then the first (affected) patient who got tested receives immediate counseling by a cardiologist, and genetic testing and counseling will be offered to all at-risk relatives. Different heart rhythm genetic mutations have different effects on ion channels, so individualized remedies are required.
“Genetics is still quite complex to most people, so we try to make our explanations understandable and not so scary,” said Dr. McDonald, who has co-authored several articles on how patients are affected by cardiogenetic testing, including in the journals Qualitative Health Research and Personalized Medicine.
“Our dominant message is ‘we’re here to provide information, which gives you knowledge, and knowledge gives you power to manage your life and to help the next generation.”
A pedigree, which depicts the relationship between individuals and relevant facts about their medical histories, can be used to help understand the transmission of genes within the family. Dr. McDonald points to a square indicating the presence of a particular genetic trait in a male.
The Cardiogenetics Clinic will offer patients access to the latest clinical trials for new drugs or devices. Dr. McDonald was recently named USF site lead investigator for a world-wide Phase 3 study testing the effect on walking endurance of an investigational medication for patients with dilated cardiomyopathy caused by a rare genetic mutation. This form of heart disease, in which inadequate pumping of blood causes the heart to become weaker, can lead to heart failure.
“Heart In A Dish” As A Drug Screening Tool
Interested in drug discovery for inherited heart diseases lacking effective medications, Dr. McDonald’s lab has begun collecting blood cells from USF Health cardiomyopathy patients who provide informed consent.
The adult blood cells can be genetically reprogrammed into induced pluripotent stem cells (iPSCs) with the potential to develop into any cell type in the body, including heart cells. The goal is to model the early stages of inherited heart disease with patient-specific cells grown in a petri dish, working out at a molecular level how the disease does its damage to heart muscle.
On the horizon: Modeling inherited heart diseases using pluripotent stem cells
Dr. McDonald and Maliheh Najari Beidokhti, PhD, a postdoctoral associate in the Department of Cardiovascular Sciences.
“Once you do that,” Dr. McDonald said, “you can use the ‘heart disease in a cell culture dish’ to screen any number of drugs or chemical compounds for their potential therapeutic benefit.”
Dr. McDonald is also collaborating with colleagues in the USF Health Department of Neurology to look at rare genetic mutations for nervous system diseases, such as certain types of muscular dystrophy and ataxias, which can lead to severe heart damage,
Dr. McDonald received his bachelor’s degree in zoology from USF in 1977 and MD degree from the University of Florida. He completed a residency in medicine and research fellowship in cardiology at Columbia-Presbyterian Medical Center in New York City. At Stanford University School of Medicine, he conducted fellowships in clinical cardiology and interventional cardiology, as well as a postdoctoral research fellowship. He spent 22 years as a faculty member at Albert Einstein College of Medicine before joining the USF Health Morsani College of Medicine last fall.
Continuously funded throughout his career by the NIH or the American Heart Association (AHA), Dr. McDonald has authored more than 70 peer-reviewed publications. Among his many high-impact papers was a 2013 article published in FASEB. The NIH-supported study was among the first to report that synonymous (silent) changes in DNA traditionally considered neutral may adversely affect the processing speed and efficiency of ion channels associated with the heart arrhythmia syndrome Long QT and alter disease severity.
Dr. McDonald has served on multiple study sections of the NIH and AHA. He was elected in 2011 as an AHA Fellow-Basic Cardiovascular Sciences Council.
Dr. McDonald says his clinical practice helps inform and complement the translational science he conducts in the laboratory.
Some Things You May Not Know About Dr. McDonald
During high school, he worked one year as a head cook for a restaurant in Winter Park, Fla., before entering college. “It made me realize that hard work is important, but also motivated me to study so I could make a living by using my head more than my hands.”
Wife Kami Kim, MD, also a USF Health physician-scientist, is a professor with joint appointments in the Department of Internal Medicine and in the Department of Global Health. They met in the cardiac intensive care unit at Columbia Presbyterian Medical Center when Dr. McDonald was a resident and Dr. Kim was rounding as a medical student. Their two sons, both studying theoretical math, are Clayton, 24, a PhD student at Boston College, and Vaughan, 20, starting his junior year at Harvard University.
McDonald enjoys bicycling, Japanese cooking, and nearly exclusively reads fiction – “it’s another window on the human condition.” His two favorite books are One Hundred Years of Solitude, an acclaimed novel by Nobel Prize-winning Latin-American author Gabriel García Márquez, and Infinite Jest, a literary bestseller and unconventional comedy by David Foster Wallace.
Written by Anne Delotto Baier, Photos and Video by Torie M. Doll, USF Health Communications and Marketing
USF neuroscientist probes how different states of tau drive brain cell damage
Research by Laura Blair’s team seeking to untangle tau may lead to targeted treatments for Alzheimer’s, Parkinson’s and other neurodegenerative diseases
As an USF undergraduate student conducting research in chemist Bill Baker’s laboratory, she had the chance to work at the USF Health Byrd Alzheimer’s Institute with Chad Dickey, PhD, an accomplished and creative NIH-funded neuroscientist who was the first to find that proteins involved in learning and memory were selectively impaired in a mouse model of Alzheimer’s disease.
Blair jumped at the opportunity. Dr. Dickey’s National Institutes of Health (NIH) work was focusing on defects in the removal of damaged proteins from cells, with promising studies of the key role “chaperone proteins” play in regulating brain cell function in Alzheimer’s disease and other neurodegenerative disorders.
“I was excited to be part of the translational work he was doing – seeking to understand how the cell’s natural defense, these chaperone proteins, might be harnessed to regulate (abnormal) misfolding proteins in order to help fix or prevent neurodegenerative disease,” Dr. Blair said.
That was nine years ago.
Dr. Blair pulls up microscopic images of stained neurons with doctoral student Lindsey Shelton, right. The stain is used to determine if treatments administered to mice — for instance, overexpressing cochaperone protein Aha1 or human enzyme CyP40 — change how toxic tau becomes to nerve cells.
Dr. Laura Blair describes the focus of her laboratory’s research.
Carrying on a scientific legacy
Since then, Dr. Blair received her doctorate in medical sciences at USF in 2014 (she also earned bachelor’s and master’s degrees here) and continued to work in Dr. Dickey’s laboratory as a postdoctoral fellow. When Dr. Dickey passed away last year at the age 40 following a courageous battle with cancer, Dr. Blair and colleague John Koren, PhD, assumed the leadership of Dickey’s team with heavy hearts. But they push forward with their mentor’s same dedication to seeking answers and sense of urgency to “publish, publish, publish.”
“Chad was always larger than life and one of the biggest influences in my life, so losing him has been surreal,” said Blair, who keeps Dr. Dickey’s nameplate on the office she now occupies. “It’s hard, but we will continue the work he started and carry on his scientific legacy.”
“He was a tremendous mentor who taught all of us how to think about a problem and ways to come up with solutions.” And she said, smiling at the memory of Dr. Dickey’s unrelenting drive to find solutions, “he taught us there’s no sense in waiting until tomorrow for an experiment that could have been started yesterday.”
Dr. Blair with Jeremy Baker, doctoral candidate in the Department of Molecular Medicine who was the lead author for a recent PLOS Biology paper reporting that a human enzyme can reduce neurotoxic amyloids in a mouse model of dementia.
Dr. Blair’s laboratory studies how chaperone proteins drive different states of the tau protein associated with Alzheimer’s disease and more than a dozen other tauopathies, including Parkinson’s disease and traumatic brain injury. Just as their name suggests, “chaperone” proteins escort other proteins in the cell to the places they need to be, ensure these proteins do not interact with others that could be bad influences, and see to it that proteins degrade, “or are put to bed, so to speak,” when the time is right, Dr. Blair said. Under normal circumstances, chaperone proteins help ensure tau proteins are properly folded to maintain the healthy structure of nerve cells.
Pursuing how chaperones drive different states of tau
In particular, the USF researchers explore how various chaperone proteins interact, for better or worse, with various forms of tau – ranging from soluble tau protein that can spread from one brain cell to another to the aggregated, misfolded species of tau tangles inside brain cells. (Sticky plaques of B-amyloid protein and toxic tangles of tau protein both accumulate in patients with Alzheimer’s disease, but recent studies suggest that tau deposits may be more closely linked to the actual death of neurons leading to memory loss and dementia.)
“If we can understand which states of tau are worse in terms of driving neurotoxicity,” Dr. Blair said, “maybe we can begin to shift tau into a better state so we can help delay disease progression, if not stop it in its tracks.”
Dr. Blair comments on differences in Alzheimer’s disease.
Dr. Blair and colleague John Koren, PhD, assistant professor (far left), co-manage a research team focusing on how chaperone proteins drive different states of the tau protein associated with Alzheimer’s disease and more than a dozen other tauopathies.
With a support of a new five-year, $1.5-million R01 grant from the National Institute on Aging, Dr. Blair, along with co-principal investigators Paula Bickford, PhD, of the USF Center of Excellence for Aging and Brain Repair, and Vladimir Urvesky of the Department of Molecular Medicine, is looking at a family of energy-independent, small heat shock proteins known to prevent harmful tau aggregation. In an earlier paper published several years ago in the Journal of Neuroscience, Drs. Blair, Dickey and colleagues reported that high levels of one of these chaperone proteins, Hsp27, reduced tau accumulation in neurons and rescued learning and memory in a mouse model for Alzheimer’s disease.
Dr. Blair is also principal investigator of a second, five-year $1.36-million grant from the National Institute of Mental Health (the continuation of a grant originally awarded to Dr. Dickey) to develop a treatment blocking the effect of a stress-related protein genetically linked to depression, anxiety and other behaviors associated with post-traumatic stress disorder (PTSD). The grant builds upon earlier USF research, showing that as levels of this stress-related protein, known as FKBP51, increase in the brain with age it partners with chaperone protein Hsp90 to make tau more deadly to brain cells involved in memory formation. Using a new mouse model genetically engineered to overexpress FKBP51 and exhibit symptoms like those seen in humans with PTSD, Dr. Blair’s team will test various treatments on mice exposed to early-life environmental stresses.
Dr. Blair and Shelton, lead author of a USF-led paper published this September in Proceedings of the National Academy of Sciences demonstrating that Hsp90 cochaperone Aha1 boosted production of toxic tau aggregates in a mouse model of neurodegenerative disease.
The ultimate goal of all this tau regulation research is to discover and commercialize targeted treatments that work, whether that’s drugs that inhibit or activate chaperone proteins, or gene therapies, or a combination. Currently no FDA-approved medications for Alzheimer’s disease specifically target beta amyloid or tau; they only help improve symptoms for some patients for a limited time.
A single-bullet therapeutic approach to a disease like Alzheimer’s that affects diffuse areas of the brain is unlikely, Dr. Blair said. “The multi-treatment option is probably going to be the most effective, but it’s difficult to address that until we have individual treatments moving forward.”
“Working toward treatments to help slow and prevent these devastating neurodegenerative diseases is what keeps our laboratory so motivated and determined.”
On working in a building that bridges research and clinical care
Slides, containing stained nerve cells from mouse brain tissue, are used by the researchers to evaluate neuronal health following various treatments.
Identifying potential treatments for neurodegenerative diseases
Building on Dr. Dickey’s work with chaperone proteins, the USF researchers continue to make significant progress in identifying potential targets to help slow or prevent neurodegenerative disease progression.
The team recently identified cochaperone protein Aha1, which binds to and stimulates the activity of heat shock protein Hsp90, as one promising therapeutic target. Hsp90 regulates the folding, degradation and accumulation of tau. In a study published this September in Proceedings of the National Academy of Sciences, the researchers demonstrated that Hsp90 cochaperone Aha1 boosted production of toxic tau aggregates in a mouse model of neurodegenerative disease and led to neuron loss and memory impairment.
The researchers also found that inhibiting Aha1 prevented the dramatic accumulation of tau in cultured cells. “We think Aha1 inhibitors offer promise for effects similar to Hsp90 inhibitors with less side effects,” Dr. Blair said.
Dr. Blair and Shelton work at an automated stereotactic injector station. The equipment helps the researchers determine the effects of gene therapy in the brain.
This June, in the journal PLoS Biology, Dr. Blair (principal investigator), Jeremy Baker (lead author) and colleagues reported for the first time that a naturally-occurring human enzyme – called cyclophilin 40 or CyP40 – could break apart clumps of tau in a mouse model of dementia. The USF led study found that CyP40 reduced the amount of aggregated tau, converting it into a more soluble and less toxic form of amyloid. In a mouse model of an Alzheimer’s-like disease, experimental expression of CyP40 preserved brain neurons and rescued cognitive deficits. The same enzyme also disaggregated alpha-synuclein, an aggregate associated with Parkinson’s disease.
Exactly how CyP40 can untangle clumps of tau and alpha-synuclein is not yet clear, but, Blair said “our finding suggests that CyP40, or one of the more than 40 other proteins with similar activity, may have a role to play in treating neurodegenerative diseases.”
Dr. Blair points to the hippocampus on the map of a mouse brain. An area critical for learning and memory, the hippocampus is especially vulnerable to damage at early stages of Alzheimer’s disease.
What Dr. Blair hopes she can impart to students as a mentor
Some things you might not know about Dr. Blair:
Blair and her husband Tom belong to a local swing dancing community, where they enjoy dancing the Lindy Hop and Balboa to ‘40s jazz.
Blair is a mother to 7-year-old Oliver, and a stepmother to 17-year-old Laney, who is a high school senior. They also have two dogs, a Shih Tzu named Oreo and a Shih Tzu-Yorkshire terrier mix named Pumpkin.
Participating in the Great American Teach-in two years ago, Dr. Blair spoke to her son’s kindergarten class about her career as a neuroscientist, including working on brain teaser puzzles and discussing a video of a preclinical neurobehavioral test with the young students. But, she said, the kids were particularly impressed with the squishy plastic “stress-reliever brains” she brought along.
Research associate Leo Breydo loads protein into a gel.
Cultured cells are harvested for analysis.
–Written by Anne Delotto Baier, Photos by Sandra C. Roa and Eric Younghans, University Communications and Marketing
Genetics research may help tailor more precise therapies for asthma, heart failure
Studies led by USF’s Dr. Stephen Liggett shed light on genetic variability of adrenergic receptors and how they might best be used to treat disease
Dr. Stephen Liggett, who leads the research enterprise for the Morasani College of Medicine and for USF Health, also oversees a genomics laboratory working on NIH-funded studies. Behind him is a radioligand binding machine used to determine the number of receptors in each cell.
While significant progress has been made managing asthma over the last two decades, about half of all asthmatics achieve optimal control of this chronic inflammatory disease using currently available medications. Similarly, only about 50 percent of patients with congestive heart failure, which occurs when the heart is too weak to pump enough blood to meet the body’s needs, have an average life expectancy of more than five years.
More still needs to be known at the molecular level about these common diseases to identify potential new targets for drug therapies, said Stephen B. Liggett, MD, associate vice president for research at USF Health, vice dean for research at the Morsani College of Medicine, and professor of internal medicine and molecular pharmacology and physiology.
What ties these two diseases together are the receptors on cardiac muscle and on smooth muscle of the airways. Dr. Liggett’s laboratory helps shed light on the genetic variability of adrenergic receptors and on how these receptors can best be used for treatment. The genetic studies have been particularly useful in developing the concept of pharmacogenetics, a tailoring of therapy based on an individual’s genetic makeup, for heart failure and asthma.
“Twenty years ago we had a handful of medicines for high blood pressure, and today we don’t use any of them. Now, we have a whole new group of more effective (antihypertensive) drugs with much fewer side effects,” he said. “And, I’m sure that one day, we’ll have more tools in our toolbox to better treat heart failure and asthma – drugs that work better for subgroups of people as defined by their genetic makeup and environmental exposures.”
Dr. Liggett comments on some of his laboratory’s contributions to the field over his career.
Mining a “superfamily” of receptors for better drug targets
Dr. Liggett leads a USF team that studies the genetic, molecular biology, structure and function of G-coupled protein receptors, or GPCRs, the largest family of human proteins. More than 800 GPCRs have been discovered within cell membranes in the human body, Dr. Liggett said, and one or more of these receptors plays a role in virtually everything the body does, including controlling thoughts in the brain, sight and smell, uterine contraction and relaxation, blood pressure, cardiac, lung and kidney function, to name just a few.
Consequently, malfunctions of GPCR signaling pathways are implicated in many chronic diseases including asthma and cardiovascular diseases. Already this “superfamily” of receptors accounts for nearly half the targets of all prescribed drugs. But, a deeper understanding of the dynamics of the GPCR signaling network and how it maintains a healthy cell or responds to pathogens could lead to the design of drugs that more precisely target diseases with greater effectiveness and fewer side effects.
Dr. Liggett began his work with GPCRs in 1988 as a Howard Hughes Institute postdoctoral research fellow in the Duke University Medical Center laboratory of mentor Robert Lefkowitz, MD. Dr. Lefkowitz was awarded the 2012 Nobel Prize in Chemistry with Brian Kobilka, MD, for groundbreaking discoveries revealing the inner workings of GPCRs.
Building upon his interest and advanced training in pulmonary and critical care medicine, Dr. Liggett began early in his career to concentrate on one of the classes of GPCRs known as adrenergic receptors, which are stimulated by the hormone epinephrine and the neurotransmitter noepinephrine. They are involved in increasing the rate and force of contraction of the heart, as well as constriction and dilation of blood vessels throughout the body and of airways in the lung. For the last 28 years, he has been continuously funded by the National Institutes of Health (NIH) to study the molecular basis of beta-adrenergic receptors in asthma.
Biological scientist Ashley Goss
Dr. Liggett is the principal investigator of a four-year, $1.12-million R01 grant from the NIH’s National Heart, Blood and Lung Institute (NHBLI) that seeks to understand how beta-adrenergic signaling is regulated to influence the development and treatment of asthma. Over his career, he has also been awarded millions of dollars in NIH funding to explore the role of genetic variations of GPCRs in heart failure, including whether those variations may alter how effectively drugs work in individual patients.
Bitter taste receptors in a new place
Dr. Liggett is also currently a project principal investigator for a five-year, $2-million NHBLI P01 grant examining how airway smooth muscle bitter taste receptors might be applied as new treatments for asthma and chronic obstructive pulmonary disease.
Using a genomics-based method that Dr. Liggett pioneered, his team had previously identified bitter taste receptors, initially thought only to exist on the tongue, deep inside the lung at the airway smooth muscle and demonstrated they act to open the airway. “When activated, they appear far superior to the beta-agonists commonly prescribed to patients to open their airways during an asthma attack,” said Dr. Liggett, who published the discovery and the need for alternatives to current bronchodilators in Nature Medicine and other journals.
Overall, discoveries emerging from Dr. Liggett’s research have yielded more than 250 peer-reviewed papers, many highly cited and appearing in top journals such as Nature Medicine, Science, Proceedings of the National Academy of Sciences, and the New England Journal of Medicine. His work has been cited by other papers more than 26,000 times. He also holds 18 patents detailing potential new targets for drug therapy or genetic variations of known drug targets and how they might be used to predict response to medications and customize treatment.
The serendipity of finding bitter taste receptors on smooth airway muscle in the lungs
Laboratory assistant Hiwot Zewdie
Among some of his laboratory’s major findings:
– While at the University of Maryland, Dr. Liggett’s team worked with colleagues at the University of Wisconsin-Madison to sequence for the first time the entire genomes (more than 100 different strains) of all known rhinoviruses, a frequent cause of respiratory infections including the common cold. The groundbreaking work, published on the cover of Science, provided a powerful framework for large-scale, genome-based epidemiological studies and the design of antiviral agents or vaccines to combat rhinoviruses. “I originally suggested sequencing 10 strains, and then my collaborator asked why not do them all,” he said. “This made the difference between a mediocre proof-of-concept paper and a full article in Science. I learned that it is important to think big if you want to make a real difference”
– Discovered and characterized genetic variations that may predict which patients with congestive heart failure respond best to a life-saving beta-blocker drug. These landmark studies occurred over several years and were published in Nature Medicine twice, and the Proceedings of the National Academy of Sciences three times. “This is a good example of the progression of an idea over time, where every year or so an unexpected turn of events occurred, and new insight was gained,” he said.
– While at the University of Cincinnati, Dr. Liggett, working with colleagues at Washington University and Thomas Jefferson University, found that a genetic variation of an enzyme, which inhibits beta-adrenergic receptor signaling, confers “genetic beta-blockade” in cardiac muscle and protects against early death in African Americans with heart failure. The findings, published in Nature Medicine, provided insight into individual variations in disease outcomes. Another key study from Cincinnati revealed that a certain combination of genetic variants within a single gene conferred low vs. excellent responses to inhaled beta-agonists in treating asthma. These combinations, called haplotypes, had never been identified in GPCRs. The work was published in Proceedings of the National Academy of Sciences.
Dr. Liggett’s groundbreaking research sequencing all known human rhinoviruses, a frequent cause of respiratory infections, was featured on the April 3, 2009 cover of the journal Science.
Advancing outside his field of study
Dr. Liggett joined USF Health in 2012 from the University of Maryland School of Medicine in Baltimore, where he was associate dean for interdisciplinary research and professor of medicine and physiology. He received his MD degree at the University of Miami and completed both a residency in internal medicine and fellowship in pulmonary diseases and critical care medicine at Washington University School of Medicine and Barnes Hospital in St. Louis, MO.
Within two years, he advanced from a postdoctoral research fellowship in Dr. Lefkowitz’s laboratory at Duke to tenured associate professor and director of pulmonary and critical care medicine at the University of Cincinnati College of Medicine. By the time he left Cincinnati for the University of Maryland in 2005, he held an endowed chair in medicine and directed the university’s Cardiopulmonary Research Center.
Though he had no significant wet-lab experience, Dr. Liggett was fascinated by the emerging science called “molecular biology” and was undeterred from branching into a field of study in which he had no formal training.
He secured a position as assistant professor at Duke following his fellowship there, and figured out how to sequence adrenergic receptor genes from a patient’s blood. While routine now, such genetic testing had not been done previously. He unexpectedly kept finding multiple variations (called polymorphisms or mutations) in genes coding for the same receptors, so he sought out the advice of some classic geneticists. At the time, Dr. Liggett said, their traditional thought was modeled after diseases like cystic fibrosis — if a person had the genetic mutation they developed the disease, if the mutation was absent they did not.
“There was no consideration for common genetic variants and how they might affect disease risk, progression, or response to treatment. It simply was not in their thought process,” Dr. Liggett said. He was told “it’s probably nothing and don’t quit your day job.” He did not take their advice.
Some advice Dr. Liggett would give to emerging young scientists
Assistant professor Donghwa Kim, PhD
Instead, he returned to the laboratory to sequence and clone receptors from many different populations with asthma and heart failure, showing that the receptor genes did indeed differ from one individual to another, generally with several common “versions.” His team also created “humanized” mice expressing the human genes for asthma and heart failure so they could begin to understand the physiology of the receptors. They began to find that some genetic alterations increased receptor function, some decreased the drug’s affinity to bind (responsiveness) to a receptor, and still others altered how the receptor was regulated. And, through NIH-supported clinical trials, the researchers correlated outcomes observed in patients undergoing drug therapies with the genetic variations uncovered in the laboratory.
“If there’s a lesson to be learned here by young investigators, I’d say it’s that you can collect information from experts in the field, but you need to use your gut to ultimately decide on whether to pursue a line of research or not,” Dr. Liggett said.
Personalized medicine challenge: Common diseases, multiple genetic variations
Realizing personalized medicine’s full potential will require a better understanding of how environmental variables – including diet, exercise, the gastrointestinal microbiome (gut bacteria) and toxin exposure – combine with genetic variations to affect disease and its treatment, he said. “Personalized medicine faces its greatest challenges in the common diseases like asthma, atherosclerotic heart disease and heart failure, because they involve multiple variations in multiple genes that interact with the environment to give you a disease – and also provide a set-up for unique ways to treat the disease.”
Biological scientist Maria Castano
Dr. Liggett was one of the first physicians recruited for what would become the USF Health Heart Institute. He recalls that he still had the letter of offer in his pocket when he stood before the Hillsborough County Commission in 2012 to help USF Health leadership pitch the need for a cardiovascular institute to include a focus on genomics-based personalized medicine. The county joined the state in funding the project, and Dr. Liggett was instrumental in the early planning stages of the Heart Institute before the arrival of its founding director Dr. Samuel Wickline. The institute is now under construction in downtown Tampa as part of the new Morsani College of Medicine facility, a key anchor of Water Street Tampa. Already, 21 of the 31 institute’s biomedical scientists who will investigate the root causes of heart and vascular diseases with the aim of finding new ways to detect, treat and prevent them, have been recruited.
“There’s an excitement here and philosophy of excellence that’s rewarding to see,” Dr. Liggett said. “We have a strategic plan in place, including moving ahead to expand research in cardiovascular disease, infectious disease and the microbiome, and the neurosciences. Our departments are recruiting at a good pace, and the faculty we’re bringing in all have NIH funding and are highly collaborative.”
Dr. Liggett is an elected fellow of the American Association for the Advancement of Science – one of only five Morsani College of Medicine faculty members to receive that prestigious honor. He is also an elected Fellow of the National Academy of Inventors and the American College of Chest Physicians. Last year, he was one of 30 scientists nationwide selected to join The Research Exemplar Project – recognition of his outstanding reputation as a leader whose high-impact, federally-funded research yields novel and reproducible results.
Over his career, he has served on several NIH study sections and on the editorial board of high-impact journals relevant to fundamental biochemistry as well as heart and lung diseases. He is currently editor-in-chief of the Journal of Personalized Medicine.
The potential of new treatments for asthma and heart failure
Dr. Liggett holds 18 patents detailing potential new targets for drug therapy or genetic variations of known drug targets, which might be used to predict response to medications and customize treatment.
Some things you may not know about Dr. Liggett:
He has asthma, which helps motivate his research toward finding better treatments for this common lung disease affecting one in 12 people in the United States.
Restores vintage cars, primarily DeLoreans. Although he recently finished bringing a funky lime green 1974 Volkswagen Thing back to life, and over the holidays restored a 1973 VW camper.
Lives with wife Julie on the beach in Treasure Island, where they enjoy surfing, paddle boarding, and photography.
Has three children – Elliott, an engineer at NASA’s Jet Propulsion Laboratory at Cal Tech in Pasadena, CA; Grace, who recently completed her master’s degree in public health at USF; and Mara, an undergraduate student studying social work at Florida Atlantic University, and two step-children — Madison, an undergraduate at the University of Florida, and Tripp, a senior at St. Petersburg Catholic High School. He also has three grandchildren, ages 2 to 9.
– Written by Anne Delotto Baier, photos by Sandra C. Roa, and audio clips by Eric Younghans, University Communications and Marketing
Dr. Mack Wu studies molecular control of ischemia-reperfusion injury, leaky gut
USF Health researcher Mack Wu, MD, studies what happens when the microvascular endothelial barrier controlling blood-tissue exchange is compromised during ischemia-reperfusion injury, a condition that can lead to irreversible tissue damage. He also investigates the molecular control of gut permeability, also known as “leaky gut,” in tissue injuries caused by trauma and severe burns.
His group’s work has broad implications for a variety of conditions including stroke, heart attack, thrombosis, sepsis, trauma or other inflammatory diseases associated with microvascular injury.
Mack Wu, MD, is a professor of surgery and molecular medicine at USF Health Morsani College of Medicine and a research physiologist at James A. Haley Veterans’ Hospital. On the monitor next to him are images of microvessels in the small intestine injected with fluorescent dye.
The closely connected endothelial cells lining the interior of blood vessel walls play a critical role in limiting the how much fluid, proteins and small molecules cross the wall of the tiny blood vessels, or microvessels. However when this protective endothelial barrier is damaged, excessive amounts of blood fluid, proteins and molecules leak outside the microvessels into nearby body tissue – a process known as microvascular hyperpermeability. If this breech of endothelial barrier is associated with a body-wide inflammatory response, it can trigger a chain of events leading to edema (swelling), shock from severe blood and fluid loss (hypovolemic shock), and ultimately multiple organ failure.
Pinpointing Potential Solutions For Ischemia-Reperfusion Injury
Previous research by Dr. Wu’s laboratory and other groups discovered that ischemia-reperfusion injury can cause endothelial barrier damage leading to vascular hyperpermeability, or abnormally leaky blood vessels.
Ischemia-reperfusion injury is typically associated with conditions like organ transplantation, stroke, heart attack, or cardiopulmonary bypass where blood supply to a vital organ is temporarily cut off (ischemia), resulting in oxygen deprivation. For instance, a period of ischemia occurs while a donor organ is transported to a recipient in the operating room, or when a clot interrupts blood circulation to the brain. When blood supply is re-established with new blood returned to the previously oxygen-deprived area (reperfusion), tissue injury can worsen because the reperfusion itself causes inflammation and oxidative damage rather than restoring normal function. It its severest form, ischemia-reperfusion injury can result in multiple organ failure, or even death.
“I believe endothelial barrier injury is one of the key elements of ischemia-reperfusion injury, so my group is trying to find out which molecule is ultimately responsible for the endothelial barrier damage,” said Dr. Wu, a professor of surgery and molecular medicine at USF Health Morsani College of Medicine and a research physiologist at James A. Haley Veterans’ Hospital.
Dr. Wu with some members of his laboratory team. From left, Rebecca Eitnier, research assistant; Shimin Zhang, Department of Molecular Medicine graduate student; Ricci Haines, research associate; and Fang Wang, research assistant.
With the support of a $1.49-million, four-year R01 grant from the National Heart, Lung and Blood Institute, Dr. Wu’s team is zeroing in on a molecule known as focal adhesion kinase, or FAK, an enzyme that may play a role in weakening the microvascular endothelial barrier during ischemia-reperfusion injury. Using cell models and a newly developed mouse model in which the endothelial-specific gene for FAK is knocked out, the USF researchers are testing whether selectively inhibiting FAK activity can rescue the endothelial barrier from such injury.
The work is critical because no FDA-approved treatment exists to prevent tissue damage following reperfusion. Identifying a new mechanism for the injury would provide potential targets for drug development, Dr. Wu said. So for instance, he said, after an initial stroke a new intravenously administered drug selectively targeting endothelial cells in the brain’s microvessels might stop further harmful swelling of the brain caused by stroke.
Defining Molecular Control Of “Leaky Gut” In Severe Burn Trauma
A second grant from the U.S. Department of Veterans Affairs funds Dr. Wu’s studies to define the underlying molecular mechanisms of leaky guts induced by traumatic injury associated with thermal (fire, scald or chemical) burns. Massive burn trauma is a significant cause of injury and death in American soldiers. With a $960,000 VA Merit Award, Dr. Wu focuses on how intestinal epithelial barrier damage happens during severe burns, with the aim of developing targeted therapies to prevent posttraumatic complications. In particular, he is working to determine the pathways by which the protein palmitoylation in gut epithelial cells are stimulated by burn injury.
Epithelial cells line the interior of the small intestines, and after severe burn injury, this protective epithelial barrier commonly breaks down, causing bacteria and toxins to flow from the intestine into the circulating blood. The result of this abnormal epithelial permeability, or “leaky gut,” can be deadly if sepsis ensues – a bacterial infection in the bloodstream sets up a body-wide inflammatory response leading to multiple organ failure.
While the role gut barrier failure plays in posttraumatic complications is well recognized, its cellular and molecular mechanisms remain poorly understood. Currently, pushing IV fluids to help prevent hypovolemic shock and administering antibiotics and anti-inflammatories are the only therapies, mostly supportive, Dr. Wu said.
“More effective early therapeutic interventions to prevent leaky gut and systemic inflammatory response will be key to preventing sepsis,” he added, whether in soldiers with trauma or VA patients with inflammatory bowel diseases.
From Industry To Academia
Dr. Wu joined USF Health and the Haley VA Hospital in 2011. He came from Sacramento, Calif, where he was an associate professor of surgery at the University of California at Davis School of Medicine and a research physiologist at Sacramento VA Medical Center. Previously, Dr. Wu was a faculty member in the Department of Medical Physiology at Texas A&M University Health Science Center. He screened pharmaceutical compounds as a toxicologist in a biotechnology laboratory before joining Texas A&M, moving from industry to academia in 1995.
Dr. Wu received his MD degree from Second Military Hospital in Shanghai, China, and conducted an internship at Shanghai Second Hospital.
One of his earliest and most highly cited studies, published in the American Journal of Physiology (1996), was first to report nitric oxide’s role in contributing to cardiovascular injury. The study showed an increase in nitric oxide induces vascular endothelial growth factor (VEGF) to promote leakage in tiny coronary veins.
Another more recent study in Shock (2012) provided direct evidence that thermal burn injury causes intestinal barrier disruption and inflammation characterized by intestinal mucosal permeability (leakage) and an infiltration of immune system cells known as neutrophils.
Something You May Not Know About Dr. Wu:
He loves deep-sea fishing. Dr. Wu has fished for sharks off the Golf coast of Texas, rockfish off the Pacific coast of California, and grouper off the west coast of Florida.
Dr. Wu is a member of the USF Health Heart Institute. His team’s work has broad implications for a variety of conditions including stroke, heart attack, thrombosis, sepsis, trauma or other inflammatory diseases associated with microvascular injury.
– Written by Anne Delotto Baier, photos by Eric Younghans, USF Health Communications and Marketing
Seeking to understand how the heart short circuits, Sami Noujaim looks for new drugs to fix atrial fibrillation
Within the last three years, USF biomedical scientist Sami Noujaim, PhD, lost his older brother to sudden cardiac death and his 80-year-old father was diagnosed with atrial fibrillation.
The experiences gave Dr. Noujaim a new appreciation for his research on understanding how normal and abnormal electrical impulses are generated in the heart. The Cardiac Electrophysiology Research Laboratory he directs focuses on finding more effective drugs to treat atrial fibrillation, the most common irregular heart rhythm and a condition for which prevalence rises markedly after age 65.
Sami Noujaim, PhD, directs the Cardiac Electrophysiology Research Laboratory in the USF Health Department of Molecular Pharmacology and Physiology.
“When life throws something like that at you, the work you do takes on a more personal tone, a sense of mission. I realized that neither myself nor my loved ones are immune from the cardiovascular diseases I’m studying,” said Dr. Noujaim, an assistant professor in the Morsani College of Medicine’s Department of Molecular Pharmacology and Physiology. “We are all at risk.”
Dr. Sami Noujaim describes the focus of his laboratory’s research.
Searching For Noninvasive Solutions To Atrial Fibrillation
Atrial fibrillation is a problem with the cardiac electrical circuitry that controls the rate and rhythm of the heartbeat. The condition affects about 9 percent of the U.S. population age 65 or older, according to the Centers for Disease Control and Prevention, and can lead to potentially deadly complications such as stroke and heart failure. While signs of atrial fibrillation may include heart palpitations, fatigue, lightheadedness, dizziness and shortness of breath, some people experience no noticeable symptoms.
Treatment and management options include lifestyle changes, medications to help control heart rate and rhythm and reduce the risk of blood clots, controlled electrical shock (cardioversion to reset heart rhythm), invasive procedures (catheter ablation) and surgical implantation of pacemakers. However, a major challenge is that atrial fibrillation frequently recurs after normal heart rhythm (sinus rhythm) is restored.
“The existing treatments can be good, but there is a lot of room for improvement, so we are focusing on contributing to noninvasive treatment options,” Dr. Noujaim said. “If we could help physicians get patients with atrial fibrillation to long-term normal sinus rhythm, and perhaps increase the ability to take them off blood thinners, it would be a significant improvement.”
Dr. Noujaim’s laboratory uses specialized equipment to measure the electrical activity of heart muscle cells and image what he describes as “atrial fibrillation in a dish.”
Closing In On A Pathway Linking Aging And Afib
Dr. Noujaim currently works with colleagues at USF and other institutions, including Tufts University, the University of Michigan, and Northeastern University, to investigate how age-related changes in specific potassium ion channels known as GIRK may trigger a cascade of molecular events leading to atrial fibrillation. His research, supported by a five-year $2.14 million RO1 grant from the National Heart, Lung and Blood Institute employs techniques including structural biology, molecular simulations, and cellular and whole organ electrophysiology.
The researchers hypothesize that in aging-associated atrial fibrillation, the condition may arise when a biochemical pathway controlling the GIRK postassium channels begins behaving abnormally, in part because of structural and metabolic cardiovascular changes that occur with aging. High blood pressure, diabetes, and coronary artery disease– among the most common risk factors for atrial fibrillation – become more common as people grow older.
Mohammed Alhadidy (left), a biomedical graduate student, and Bojjibabu Chidipi, PhD, a postdoctoral scholar in Molecular Pharmacology and Physiology, record the electrical signals from several cells using a multichannel automated patch clamp.
The significance of investigating the relationship between aging and atrial fibrillation.
Designing A “Perfect Plug” Using An Antimalarial Drug Model
“Right now, we are trying to design a perfect plug using the antimalarial drug chloroquine as a model,” Dr. Noujaim said, “We have evidence that if we block that specific type of potassium channels we will be able to stop atrial fibrillation or at least reduce its occurrence.”
Earlier work by Dr. Noujaim and others, including a study reported in the journal FASEB, demonstrated that the antimalarial drug chloroquine was effective in blocking the GIRK potassium channels and suggested a new path for discovering antiarrhythmic drugs.
Dr. Noujaim continues using cellular and animal models to pinpoint how and where the chloroquine molecule interacts with the potassium channel – with the aim of discovering a “plug” that works even better than the antimalarial drug. At the same time, he has reached out to USF Health Department of Cardiovascular Sciences to begin applying the laboratory findings to the clinic.
Working with cardiologists Bengt Herweg, MD, and Dany Sayad, MD, Dr. Noujaim recently gained approval for a pilot study to enroll 40 adults without heart damage whose atrial fibrillation has persisted more than one week and less one year. The team will test the effectiveness of chloroquine in restoring and maintaining normal heart rhythm in patients with atrial fibrillation.
The laboratory uses techniques including structural biology, molecular simulations, and cellular and whole organ electrophysiology to conduct its NIH-funded research.
Collaborating With Clinicians On New Treatment Options
Dr. Sayad, initially surprised at Dr. Noujaim’s proposal to try chloroquine, said the strength of the preclinical data convinced him of the antimalarial drug’s potential as another antiarrhythmic option. “Treating atrial fibrillation can be especially challenging in older patients, who experience a higher recurrence of atrial fibrillation (following cardioversion) and more failure on drugs used to regulate heart rhythm,” he said.
Clinicians help biomedical scientists like Dr. Noujaim frame and focus their studies to make the research more relevant to challenges faced in treating patients, such as maintaining sinus rhythm once a normal heartbeat has been restored.
“It does not mean that the fundamental, basic science questions are not important,” Dr. Noujaim said. “To the contrary, those questions are at the heart of every single experiment we do; however, we must always think about the big picture and why we are asking those questions. And that always goes back to the clinic.”
In previous electrophysiology experiments, Dr. Noujaim and colleagues helped better define the contribution of the nervous system within the heart, otherwise known as the intrinsic cardiac ganglia, to normal and abnormal heart rhythm. Using both mouse models and patients with atrial fibrillation, the study shed light on how nerves emerging from these cardiac ganglia regulate activity of the sinus node, the heart’s natural pacemaker. The study appeared in Cardiovascular Research in 2013.
Dr. Noujaim with members of his research team, from left, Bojjibabu Chidipi, Mohammed Alhadidy and laboratory manager Michelle Reiser.
Dr. Noujaim comments on the importance of a clinical perspective to frame biomedical research.
Impressed By USF’s Biomedical Research Opportunities
Dr. Noujaim came to USF in 2015 from the Molecular Cardiology Research Institute at Tufts University School of Medicine. He received his PhD in pharmacology, with distinction, from SUNY Upstate Medical University in Syracuse, NY, followed by a year of postdoctoral training there. He then completed a three-year postdoctoral fellowship, supported by the American Heart Association, and the National Institutes of Health, at the Center for Arrhythmia Research, University of Michigan, in Ann Arbor, MI.
Dr. Noujaim said he was attracted to USF by the opportunity to be part of an emerging preeminent university committed to establishing a cardiovascular institute bridging top biomedical research and clinical care.
“The opportunities that the University of South Florida is providing for scientists are equal or greater than those at any other major academic medical center,” he said. “It was also striking to me that, in a place the size of USF, the USF Health leadership is so actively engaged in research, with their own laboratories and grants. That’s not what you would see in a lot of places, and as a biomedical scientist it makes me feel that the leadership here really values research.”
Dr. Noujaim is passionate about the cardiovascular research his laboratory conducts, which he says has taken on a greater sense of mission since his own family’s experience with sudden cardiac death and atrial fibrillation.
His take on the benefit of brainstorming with scientists in other disciplines.
Some Things You May Not Know About Dr. Noujaim:
Born in Lebanon, he moved to the United States after graduating from high school.
He routinely swims laps in an indoor pool.
He enjoys experimenting with cooking, specializing in inventing new dishes by combining ingredients he finds in his refrigerator. “I’ve discovered by trial and error that no matter how bad what I cook really is, adding a tablespoon of soy sauce makes it alright,” he said.
His first scientific experiment as a college student volunteering in Boston’s Beth Israel Deaconess Medical Center laboratory was unforgettable. He fainted while his blood was being drawn so he could use it to help study blood platelet activation and aggregation. Click on video below to find out more.
– Written by Anne Delotto Baier, Photos by Sandra C. Roa and Eric Younghans
Pioneering nanotechnology research has applications for cardiovascular disease
The USF Health Heart Institute’s new leader Dr. Samuel Wickline arrives with an impressive NIH portfolio and strong track record of entrepreneurial research.
The founding director of the USF Health Heart Institute has a passion for innovation, translational medicine and entrepreneurship.
Samuel A. Wickline, MD, has parlayed his expertise in harnessing nanotechnology for molecular imaging and targeted treatments into an impressive $1-million portfolio of National Institutes of Health awards, multiple patents and four start-up biotechnology companies.
“We’ve developed nanostructures that can carry drugs or exist as therapeutic agents themselves against various types of inflammatory diseases, including, cancer, cardiovascular disease, arthritis and even infectious diseases like HIV,” said Dr. Wickline, who arrived at USF Health last month from the Washington University School of Medicine in St. Louis.
Dr. Samuel Wickline talks about his vision for the USF Health Heart Institute.
At Washington University, Dr. Wickline, a cardiologist, most recently was J. Russell Hornsby Professor in Biomedical Sciences and a professor of medicine with additional appointments in biomedical engineering, physics, and cell biology and physiology.
“I like the challenge of building things,” he said.
In St. Louis, he built a 29-year career as an accomplished physician-scientist keenly interested in translating basic science discoveries into practical applications to benefit patients. He served as chief of cardiology at Jewish Hospital, developed one of the first cardiac MRI training and research programs in the country, helped establish Washington University’s first graduate program in biomedical engineering, and led a university consortium that works with academic and industry partners to develop medical applications for nanotechnology.
At USF, there will be no shortage of challenging opportunities to build.
Building the USF Health Heart Institute
A major part of Dr. Wickline’s new job is helping to design, build and equip the Heart Institute. Most importantly, he will staff the state-of-the-art facility with a critical interdisciplinary mix of top biomedical scientists (including immunologists, molecular biologists, cell physiologists and genomics experts), who investigate the root causes of heart and vascular disease with the aim of finding new ways to detect, treat and prevent them. The Heart Institute will be co-located with new Morsani College of Medicine in downtown Tampa; construction on the combined facility is expected to begin later this year.
“I have been impressed by the energy and commitment here at the University of South Florida to invest substantial resources in a heart institute,” Dr. Wickline said. “I believe we have a lot to offer in terms of bench-to-bedside research that could solve some of the major cardiovascular problems” like atherosclerosis or heart failure.
“We want to put together a program that supplies the appropriate core facilities to attract the best and brightest researchers to this cardiovascular institute.”
Cardiovascular disease is the leading cause of death in the United States and worldwide, so exploring potential new treatment options is critical. One of the Heart Institute’s driving themes will be advancing concepts and findings that prove promising in the laboratory into projects commercialized for clinical use, Dr. Wickline said.
“Our goal is to make a difference in the lives of patients,” he said. “Innovation is not just about having a new idea, it’s about having a useful idea.”
Dr. Wickline also serves as associate dean for cardiovascular research and a professor of cardiovascular sciences at the Morsani College of Medicine. He holds the Tampa General Endowed Chair for Cardiovascular Research created last year with a gift from USF’s primary teaching hospital.
With Washington University colleague Hua Pan, PhD, a biomedical engineer and expert in molecular biology, Dr. Wickline is re-building his group at USF. Dr. Pan was recently recruited to USF as an assistant professor of medicine to continue her collaborations with Dr. Wickline.
An example of Dr. Wickline’s group using nanotechnology to help combat atherosclerosis.
Dr. Wickline’s lab focuses on building nanoparticles to deliver drugs or other therapeutic agents to specific cell types, or targets.
Designing nanoparticles to “kill the messenger”
Dr. Wickline’s lab focuses on building nanoparticles – shaped like spheres or plates, but 10 to 50 times smaller than a red blood cell – to deliver drugs or other therapeutic agents through the bloodstream to specific cell types, or targets. These tiny carrier systems can effectively deliver a sizeable dosage directly to a targeted tissue, yet only require small amounts of the treatment in the circulation to reduce the risk of harmful side effects.
Some types of nanoparticles can carry image-enhancing agents that allow researchers to quantify where the illuminated particles travel, serving as beacons to specific molecules of interest, and enabling one to determine whether a therapeutic agent has penetrated its targeted site, Dr. Wickline said.
Dr. Wickline also is known for designing nanoparticles derived from a component of bee venom called melittin. While bee venom itself is toxic, Dr. Wickline’s laboratory has detoxified the molecule and modified its structure to produce a formula that allows the nanoparticles to carry small interfering (siRNA), also known as “silencing RNA,” or other types of synthetic DNA or RNA strand.
Among other functions, siRNA can be used to inhibit the genes that lead to the production of toxic proteins. Many in the nanotechnology research and development community are working to make siRNA treatment feasible as what Dr. Wickline calls “a message killer,” but the challenges have been daunting.
“The big challenge in the field of siRNA, and many companies have failed at this, is how to get the nanostructure to the cells so that the siRNA can do what it’s supposed – hit its target and kill the messenger — without being destroyed along the way, or having harmful side effects,” Dr. Wickline said. “We figured out how to engineer into a simple peptide all of the complex functionality that allows that to happen.”
Dr. Wickline comments on how being a physician adds perspective to the science he conducts.
Different targets, same delivery vehicle
In a recent series of experiments in mice, Dr. Wickline and colleagues have shown that silencing RNA messages delivered by nanoparticle to a specific type of immune cell known as a macrophage – a “big eater” of fat – actually shrinks plaques that accumulate inside the walls of the arteries during atherosclerosis, one of the main causes of cardiovascular disease. The build-up of atherosclerotic plaques with fat-laden macrophages narrows, weakens and hardens arteries, eventually reducing the amount of oxygen-rich blood delivered to vital organs.
This type of plaque-inhibiting nanotherapy could be useful in aggressive forms of atherosclerosis where patients have intractable chest pain or after an acute heart attack or stroke to prevent a secondary cardiac event, Dr. Wickline said.
In another study, Washington University School of Medicine researchers investigated the potential of the siRNA nanoparticle designed by co-investigators Dr. Pan and Dr. Wickline in treating the inflammation that may lead to osteoarthritis, a degenerative joint disease that is a major cause of disability in the aging population. The nanoparticles — injected directly into injured joints in mice to suppress the activity of the molecule NF-κB — reduced local inflammation immediately following injury and reduced the destruction of cartilage. The findings were reported September 2016 in the Proceedings of the National Academy of Sciences.
Previously, Dr. Wickline said, the Washington University group had shown that nanoparticles delivered through the bloodstream inhibited inflammation in a mouse model of rheumatoid arthritis. And, another laboratory at the University of Kentucky is studying whether locally injected siRNA nanoparticles can quell the bacterial inflammation that can lead to a serious gum disease known as periodontitis. Other collaborating labs are using these nanoparticles in pancreatic, colon, and ovarian cancers with good effects.
“The specific targets in these cases may be different, but the nice thing about this kind of delivery system for RNA interference is that the delivery agent itself, the nanostructures, are the same,” Dr. Wickline said. “All we have to do is change out a little bit of the genetic material that targets the messages and we’re set up to go after another disease. So it’s completely modular and nontoxic.”
The St. Louis-based biotechnology company Trasir Therapeutics is developing these peptide-based nanocarriers for silencing RNA to treat diseases with multiple mechanisms of inflammation. Dr. Wickline co-founded the company in 2014 and continues to serve as its chief scientific officer.
Dr. Wicklne: “Innovation is not just about having a new idea, it’s about having a useful idea.”
Inhibiting chronic inflammation without getting rid of beneficial immune responses.
In essence, Dr. Wickline said, he is interested in suppressing chronic inflammation, without disrupting the beneficial functions of surveillance by which the immune system recognizes and destroys invading pathogens or potential cancer cells.
“If you can inhibit the ongoing inflammation associated with (inappropriate) immune system response, you inhibit the positive feedback cycle of more inflammation, more plaques, more damage and more danger,” he said. “If you can cool off inflammation by using a message killer that says (to macrophages) ‘don’t come here, don’t eat fat, don’t make a blood clot’ – that’s what we think could be a game changer.”
Another NIH grant has funded collaborative work to develop an image-based nanoparticle that detects where in a compromised blood vessel too much blood clotting (hypercoagulation) occurs, and delivers potent anti-clotting agent only to that site. Formation of abnormal blood clots can trigger a heart attack when a clot blocks an artery that leads to heart muscle, or a stroke when a clot obstructs an artery supplying blood to the brain.
Because this site-specific nanotherapy targets only areas of active clotting, it may provide a safer, more effective approach against cardiac conditions like atrial fibrillation and acute heart attack than existing anticoagulant drugs such as warfarin and newer blood thinners like Xarelto® (rivaroxoban) or Eliquis® (apixiban), all which work systemically and come with raised risk for serious bleeding, Dr. Wickline said.
In a study published last year in the journal Arteriosclerosis, Thrombosis, and Vascular Biology, Dr. Wickline and colleagues found that nanoparticles delivering a potent inhibitor of thrombin, a coagulant protein in blood that plays a role in inflammation, not only reduced clotting risk but also rapidly healed blood vessel endothelial barriers damaged during plaque growth.
The preclinical work showed the experimental treatment “is actually an anti-atherosclerotic drug as well as an anti-clotting drug, so there are many potential applications,” Dr. Wickline said.
Dr. Wickline received his MD degree from the University of Hawaii School of Medicine. He completed a residency in internal medicine, followed by clinical and research fellowships in cardiology at Barnes Hospital and Washington University, where he joined the medical school faculty in 1987.
He has authored more than 300 peer-reviewed papers and holds numerous U.S. patents. Dr. Wickline is a fellow of the American College of Cardiology and the American Heart Association, and a 2014 recipient of the Washington University Chancellor’s Award for Innovation and Entrepreneurship.
Some things you may not know about Dr. Wickline
Hisfirst scientific experiment was conducted at age 21, before entering medical school, when he worked in a research laboratory in Hawaii run by a cardiovascular surgeon interested in techniques to best support the heart during cardiopulmonary bypass surgery. Dr. Wickline figured out why one particular way of perfusing blood through the heart pump machine could dangerously compromise oxygenated blood flow to the brain. The results were published in the Annals of Thoracic Surgery. One of Dr. Wickline’s first studies in medical school, published in the prestigious journal Circulation, described a method for determining the size of a heart attack by recording electrical signals, called vector cardiograms.
He grew up in St. Petersburg, Fla., and did some trick water skiing as a teen. “I learned how to ski backwards.” After being “landlocked” in Missouri for 37 years following medical school in Hawaii, he said he’s glad for the opportunity to get back to get back to the beach and water sports.
He likes to play the ukulele.
– Story by Anne Delotto Baier, photos and video by Sandra C. Roa