New study teams researchers from College of Arts and Sciences, USF Health

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Jan. 9, 2019 — The ketogenic diet has proven successful in helping people lose weight and improve their overall health, including those with epilepsy. The low-carb diet transitions the body from burning sugar to burning fat and ketones for energy. New research suggests that increasing blood ketones by using ketogenic supplementation can reduce seizures without dietary restriction.

A study published this month in Physiological Reports finds supplementing a normal, carbohydrate-rich diet with specific ketogenic agents may significantly delay tonic-clonic seizures caused by exposure to high levels of oxygen.

Oxygen toxicity is a complication that can arise following hyperbaric oxygen therapy, an FDA approved treatment used to manage various medical conditions, such as carbon monoxide poisoning, air/gas embolism and diabetic wounds. It involves inhaling pure oxygen in a pressurized or hyperbaric chamber.

“Exposure to high-pressure oxygen is also a danger to recreational, technical and military scuba divers, including Navy SEAL divers, as a seizure manifesting underwater can be lethal,” said lead author Csilla Ari D’Agostino, PhD, research assistant professor in the Department of Psychology at the University of South Florida College of Arts and Sciences. “As a scuba diver, I am very excited about the implications of these findings, since during 20 years of diving I have heard many stories about the dangers of being exposed to high partial pressure of oxygen and it is something that always has to be considered when planning a dive.”

An experienced scuba diver, Csilla Ari D’Agostino, PhD (pictured here), says the ketogenic supplement findings also have implications for recreational, technical and military scuba divers, who are at risk for lethal seizures induced by exposure to high-pressure oxygen.

Dr. Ari D’Agostino and her team, including collaborators from the USF Health Morsani College of Medicine’s Department of Molecular Pharmacology and Physiology and the USF College of Pharmacy, used a small hyperbaric environmental chamber to test the effects of ketogenic agents on rats. The animals were freely fed standard rodent chow, consisting of 70% carbohydrates. They also received different ketogenic supplements one hour before being exposed to pure oxygen in the chamber, which was pressurized to simulate technical and military dive operations. These conditions were maintained until physical symptoms of a seizure were observed.

The team found that the most effective supplement was a combination of ketone ester and medium-chain triglyceride oil in delaying the onset of hyperbaric oxygen-induced seizures. The latency to seizures was delayed by 219 percent in that group and the seizure severity was significantly reduced as well with ketone supplementation. The neuroprotective effects of ketone supplements were associated with an elevation of blood ketone levels.

The study was conducted using the Hyperbaric Biomedical Research Laboratory at the USF Health Morsani College of Medicine.

The findings show that boosting the level of blood ketones by specific ketone supplements produce therapeutic ketosis, which, in turn, may provide increased resistance to seizures induced by extreme levels of hyperbaric oxygen. Importantly, the findings support that this neuroprotective effect may not require strict adherence to dietary restrictions and can be achieved through supplementation alone.

The study was supported in part by grants from the federal Office of Naval Research.

Some of the same gases in the air we breathe to stay alive can become harmful, even deadly, at increased atmospheric pressure.

“Oxygen becomes toxic and nitrogen starts to act like a narcotic that will anesthetize you in some of these high pressure, or hyperbaric, environments encountered by the military in deep-sea diving or submarine operations,” said Jay Dean, PhD, professor of molecular pharmacology and physiology at the USF Health Morsani College of Medicine.

USF Health’s Jay Dean, PhD, is one of the world’s leading experts in hyperbaric neurophysiology.

  Listen to Dr. Dean talk about the hyperbaric lab.

Dr. Dean, one of the world’s leading experts in hyperbaric neurophysiology, has attracted more than $4 million in external funding from the Office of Naval Research (ONR) Undersea Medicine Program, since joining USF in 2006.

The USF Hyperbaric Biomedical Research Laboratory he established and directs houses various styles of pressure chambers, which mimic the environmental conditions challenging divers who breathe pure oxygen as they swim deeper and longer. The largest, at 3.2 tons, is specially designed for use with an atomic force microscope and patch clamping apparatus to help researchers determine how gases with different solubility affect brain cell function.

To date, Dr. Dean and his USF colleague, Dominic D’Agostino, PhD, have adapted electrophysiology, radiotelemetry and various types of microscopy techniques for use under hyperbaric pressures, including fluorescence, confocal and atomic force microscopy.

“Atomic force microscopes are common, but not atomic force microscopes placed under hyperbaric pressure,” Dr. Dean said. “In our lab, we’ve been able to apply very powerful research tools to unique conditions.”

Dr. Dean’s lab will soon send its second graduate student to work as a physiologist at the Navy’s medical research center. “At USF, we are helping train the next generation of undersea medicine experts in the novel techniques needed to study human performance under extreme conditions.”

The USF Hyperbaric Biomedical Research Laboratory houses various pressure chambers, including a 3.2-ton one specially designed for use with an atomic force microscope, which mimic the extreme environmental conditions challenging deep-sea divers.

Shedding light on role of oxygen toxicity in seizures

Dr. Dean started his career studying the effects of carbon dioxide on the neural control of breathing and cardiovascular function. His collaborations with the Department of Defense and Undersea Medicine program shifted his primary focus to the role of oxygen toxicity in seizures as well as the toxic effects of carbon dioxide retention.

Recently, Dr. Dean’s team expanded the scope of their hyperbaric neurosciences research by probing the cellular mechanisms of nitrogen narcosis, a major factor limiting divers’ safety and performance. This new research direction was supported by a transfer of $700,000 of equipment to USF from the Navy’s Experimental Diving Unit in Panama City, FL. In addition, Dr. D’Agostino’s team, housed with Dr. Dean in the Hyperbaric Biomedical Research Lab, has broadened the work on hyperoxia to include studies that may lead to non-toxic cancer therapies combining dietary supplements and hyperbaric oxygen.

Over the last decade Dr. Dean’s research has helped shed light on the role of hyperbaric oxygen toxicity in triggering seizures. The condition can be a life-threatening by-product of breathing too much oxygen at high ambient pressures that impacts deep-sea divers as they swim deeper and longer.

Deep-sea divers can be at risk for oxygen toxicity seizures, a life-threatening condition caused by breathing too much oxygen at high ambient pressures. – U.S. Dept. of Defense photo

Navy SEALs are especially at risk because they wear a closed circuit rebreather, to mitigate the narcotic and other debilitating effects of nitrogen and carbon dioxide breathed under increasing ocean pressure. The special device filters out these gases in such a way that bubbles do not appear on the water’s surface – useful in helping avoid enemy detection. However, the additional stealth comes at a cost. The ratio of oxygen the divers breathe greatly increases the deeper they plunge (essentially becoming pure oxygen) and, when combined with physical exertion and mission stress, can lead to nausea, dizziness, seizures, and even coma or death – all symptoms of oxygen toxicity.

A possible countermeasure, anti-seizure sedatives, requires high doses that could impair warfighters’ mental and physical performance.

Without a reliable way to treat oxygen toxicity or predict which divers are more prone to seizures than others, the Navy takes rigorous precautions to restrict all divers to no more than 10 minutes in 50 feet of seawater.

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Applications extend beyond undersea medicine

“This risk of central nervous system oxygen toxicity limits oxygen’s use — not only in diving operations, but also its clinical applications in hyperbaric oxygen therapy,” Dr. Dean said.

Hyperbaric oxygen therapy, which increases blood oxygen to temporarily restore blood gases and tissue function, can help treat unhealed wounds, burns, crushing injuries, decompression sickness, carbon monoxide poisoning, and other medical conditions. The therapeutic benefit might be maximized if the doses of hyperbaric oxygen administered could be boosted without the risk of CNS oxygen toxicity.

In their search to find solutions, Dr. Dean and colleagues analyze the response of individual brain cells to the powerful effects of oxygen and other gases under altered pressure. In the laboratory’s hyperbaric chambers, they measure changes in brain cell membranes and electrical activity, and the damage of oxygen-induced free radicals.

The researchers also monitor physiological changes in the breathing and heart rate of normal rats moving about in a chamber mimicking the environment of an increasingly deep dive. An electroencephalogram (EEG) shows electrical signals in the brain in real time, indicating the hyperexcitability that precedes and peaks with oxygen toxicity seizures.

An intracellular recording of the electrical signaling by a brain cell (middle trace) in a rodent brain slice that is stimulated by hyperbaric oxygen (top trace).

Promising discoveries to predict, delay seizures

The USF group made what could be a key discovery – the breathing rate of the rats exposed to pure oxygen increases several minutes before a seizure starts. “This may be a biomarker – an early physiological signal that predicts the impending seizure,” said Dr. Dean, who was principal investigator for the study published in the Journal of Applied Physiology.

If this early-predictor hypothesis bears out in larger animal models, he said, the next step would be to work with the Navy to devise and test a mask-fitted with a device designed to monitor divers’ breathing underwater. The ultimate aim: preventing oxygen-induced seizures to safely allow Navy SEALs to dive deeper and longer.

Another of the laboratory’s major findings evolved from an idea by Dr. D’Agostino, Dr. Dean’s former postdoctoral fellow who is now an associate professor, to harness the power of ketones, natural compounds produced by the body when it burns fat during periods of fasting or calorie restriction.

They’ve focused on better understanding how the ketogenic diet — a special low-carbohydrate, high-fat diet that elevates blood ketones and alters brain metabolism — produces anticonvulsive and neuroprotective effects. The diet has been successfully used to treat drug-resistant epilepsy or other seizure disorders.

Dr. Dean with laboratory colleagues Angela Poff, PhD, research associate, and Dominic D’Agostino, PhD, associate professor, all members of the Department of Molecular Pharmacology and Physiology.

  Dr. Dean comments on the team’s two approaches to studying oxygen toxicity seizures.

Working with collaborators in academia and industry, USF continues to develop and test naturally derived and synthetic supplements that will more rapidly mimic the therapeutic effects of ketosis without the problems associated with adhering to the ketogenic diet.

In a first of its kind study, Dr. D’Agostino and Dr. Dean tested whether feeding laboratory rats a ketone ester and placing them in the hyperbaric chamber simulating underwater conditions could delay oxygen toxicity seizures. It worked. Their study was published in the American Journal of Physiology: Regulatory, Integrative and Comparative Physiology. They also hold a patent on the use of the USF-developed ketone ester, a highly efficient fuel for the brain, to prevent CNS oxygen toxicity.

More research is needed, but the experiments pave the way for a ketone supplement that would allow Navy SEALs to dive longer while protecting them against seizures, Dr. Dean said. “If what we’ve observed in rat model experiments holds true in humans, the Navy diver should be able to increase the amount of time spent at a depth of 50 feet of seawater (10 minutes) by 600 percent… which means that the divers could get more work done with fewer dives.”

The USF Hyperbaric Laboratory will be included in an upcoming independent documentary on nitrogen narcosis, which features Sherri Ferguson, a colleague of Dr. Dean’s from Simon Fraser University in British Columbia, who studies the health effects of narcosis in deep-sea divers.

Unmatched expertise in hyperbaric cellular electrophysiology

Earlier this year, Sherri Ferguson, director of the Environmental Medicine and Physiology Unit at Simon Fraser University in British Columbia, visited Dr. Dean’s lab to observe and collaborate on some experiments investigating brain cell response to nitrogen under pressure. Ferguson, helping to make a documentary on nitrogen narcosis and its health effects in deep-sea divers, brought along the independent filmmaker who included an interview with Dr. Dean in the piece.

When breathed beneath the ocean’s depths, nitrogen can create state of mental impairment similar to the intoxicating effect of alcohol.

Ferguson recently visited Dr. Dean’s lab to collaborate on some experiments investigating brain cell response to nitrogen under pressure. “Dr. Dean has the only cellular hyperbaric electrophysiology lab of its kind in North America,” she said.

Ferguson says she was attracted to the USF Hyperbaric Biomedical Research Laboratory by Dr. Dean’s development of continuous intracellular recordings measuring how mammalian neurons behave under varying gas and pressure conditions.

“Dr. Dean has the only cellular hyperbaric electrophysiology lab of its kind in North America. His expertise in this field is unmatched, so I was excited to learn from him,” she said. “To leave him out of a documentary on cellular mechanisms of narcosis would not accurately reflect where the research is today and where it is going in the future.”

A microscopic image of neurons hyper-excited by exposure to pure oxygen under high pressure in the hyperbaric chamber.

Something you might not know about Dr. Dean:

He has spent 33 years researching the physiological problems of flight encountered by World War II pilots and their crews, who flew at high altitudes in unpressurized aircraft and suffered hypoxia from lack of oxygen and decompression sickness from low pressure.

Dr. Dean is writing a book on advances by the Allies in aviation physiology research during the war and has presented on this topic across the United States. His impressive collection of historical documents, manuscripts, films and other artifacts from Wright Field Aeromedical Laboratory (Dayton, Ohio) and several universities and medical centers documents the pioneering work on oxygen equipment, G-suits, high-altitude parachute escape, explosive decompression and development of the first pressurized airplanes.

This photo from the Mayo Historical Unit Archives shows the team of researchers from the Wright Field Aero Med Lab and Mayo Aero Med Unit before the aircraft Nemisis, a B-17E testing platform used during World War II, takes off for a study of the opening shock of a parachute at high altitude. Physiologists trained a 145-pound St. Bernard dog, Major, (lower right) to parachute — simulating the jump of a man. Major wore protective clothing and an oxygen mask during his descent.

Listen to Dr. Dean’s recent presentation on WWII aeromedical research at the Institute of Human and Machine Cognition lecture series.

Photos by Katy Hennig, USF Health Communications

The mouse model study combined a ketogenic diet and supplements with hyperbaric oxygen therapy 

Tampa, FL (June 10, 2015) — A team of researchers from the Hyperbaric Biomedical Research Laboratory at the University of South Florida (USF) has doubled survival time in an aggressive metastatic cancer model using a novel combination of non-toxic dietary and hyperbaric oxygen therapies.

The study, “Non-toxic metabolic management of metastatic cancer in VM mice: Novel combination of ketogenic diet, ketone supplementation, and hyperbaric oxygen therapy,” was published online today in PLOS ONE.

Led by principal investigator Dominic D’Agostino, PhD, assistant professor in the Department of Molecular Pharmacology and Physiology at the USF Health Morsani College of Medicine, the recently published research shows the beneficial effects of using ketone supplements in conjunction with a non-toxic therapeutic regimen developed previously by the team.  Ketones are produced when the body begins burning fat instead of carbohydrates for energy.

Principal investigator Dominic D’Agostino, PhD, and research associate Angela Poff, PhD, measure tumor growth in the mice receiving the investigational treatment in USF’s Hyperbaric Biomedical Research Laboratory

The research group previously published a study in PLOS ONE demonstrating the anti-cancer effects of therapeutic ketosis induced by the high-fat, low-carbohydrate ketogenic diet (KD) combined with hyperbaric oxygen therapy (HBOT), which involves breathing high-pressure oxygen.  Inducing therapeutic ketosis solely with the ketogenic diet can be difficult, however, so the USF researchers created novel metabolic agents that induce ketosis without dietary restriction.  These ketone supplements slowed cancer growth on their own, and further enhanced the combined therapeutic effects of KD and HBOT.

In the recent USF study, mice with advanced metastatic cancer were fed either a standard high-carbohydrate diet or a carbohydrate-restricted ketogenic diet with ketone supplements and HBOT.  Therapeutic ketosis causes the body to shift from using glucose to fatty acids and ketones bodies for energy.

Normal healthy cells readily adapt to using ketone bodies for fuel, but most cancer cells lack this metabolic flexibility. Solid tumors also have areas of low oxygen, which promote tumor growth and metastatic spread.  HBOT involves breathing 100 percent oxygen at elevated barometric pressure, saturating the tumors with oxygen.  When administered properly, both ketosis and HBOT are non-toxic and may even protect healthy tissues while simultaneously damaging cancer cells.

Animals receiving the combination of KD, ketone supplements, and HBOT lived 103 percent longer than mice fed a standard high-carbohydrate diet. The researchers believe their study demonstrates the potential of these non-toxic therapies to contribute to current cancer treatment regimens and significantly improve the outcome of patients with advanced metastatic cancer.

Researchers at USF and elsewhere are investigating the potential benefits of the physiological state of therapeutic ketosis for several major diseases. The USF team believes these novel ketone supplements may be effective in other disorders besides cancer and have ongoing studies to test their potential use in wound healing, epilepsy, amyotrophic lateral sclerosis (ALS), Alzheimer’s disease, glucose transporter type 1 (GLUT1) deficiency syndrome, and exercise performance.

The cancer study, funded by a charitable donation from Scivation Inc., was inspired by the research of Professor Thomas Seyfried of Boston College.  Dr. Seyfried has advanced the theory that cancer is a metabolic disease, leading to the development of new strategies to treat and prevent cancer.  The USF researchers are currently collaborating with other scientists to explore options for establishing human clinical trials.

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Media contact:
Anne DeLotto Baier, USF Health Communications & Marketing
abaier@health.usf.edu or (813)974-3303

The mouse model study by USF researchers combined ketogenic diet and hyperbaric oxygen therapy

Tampa, FL (June 5, 2013) — A combination of nontoxic dietary and hyperbaric oxygen therapies effectively increased survival time in a mouse model of aggressive metastatic cancer, a  research team from the Hyperbaric Biomedical Research Laboratory at the University of South Florida has found.

The study, “The Ketogenic Diet and Hyperbaric Oxygen Therapy Prolong Survival in Mice with Systemic Metastatic Cancer,” was published online today in PLOS ONE. 

Led by Dominic D’Agostino, PhD, principal investigator  in the Department of Molecular Pharmacology and Physiology at the USF Health Morsani College of Medicine, the research shows the effects of combining two nontoxic adjuvant cancer therapies, the ketogenic diet and hyperbaric oxygen therapy, in a mouse model of late-stage, metastatic cancer. 

“Our study demonstrates the potential of these cost-effective, nontoxic therapies to contribute to current cancer treatment regimens and significantly improve the outcome of patients with advanced metastatic cancer,” D’Agostino said.

Study lead author Angela Poff, a doctoral student in the Department of Molecular Pharmacology and Physiology, is shown here in the USF Hyperbaric Biomedical Research Laboratory.

Metastasis, the spreading of cancer from the primary tumor to distant spots, is responsible for over 90 percent of cancer-related deaths in humans.  A lack of available therapies effective against metastatic disease remains the largest obstacle in finding a cure for cancer. 

In the study, mice with advanced metastatic cancer were fed either a standard high carbohydrate diet or carbohydrate-restricted ketogenic diet.  Mice on both diets also received hyperbaric oxygen therapy, which uses a special chamber to increase the amount of oxygen in the tissues.

The ketogenic diet forces a physiological shift in substrate utilization from glucose to fatty acids and ketone bodies for energy.  Normal healthy cells readily adapt to using ketone bodies for fuel, but cancer cells lack this metabolic flexibility, and thus become selectively vulnerable to reduced glucose availability.  Solid tumors also have areas of low oxygen, which promotes tumor growth and metastatic spread.  

Hyperbaric oxygen therapy involves breathing 100 percent oxygen at elevated barometric pressure, saturating the tumors with oxygen.   When administered properly, both the ketogenic diet and hyperbaric oxygen therapy are non-toxic and may even protect healthy tissues while simultaneously damaging cancer cells, D’Agostino said. 

While both therapies slowed disease progression independently, animals receiving the combined ketogenic diet and hyperbaric oxygen therapy lived 78 percent longer than mice fed a standard high-carbohydrate diet.

The research, funded by a charitable donation from Scivation, was inspired by the research of Professor Thomas Seyfried of Boston College.  Dr. Seyfried has advanced the theory that cancer is a metabolic disease, inspiring the development of metabolic strategies to treat and prevent cancer. 

D`Agostino`s team is currently collaborating with Dr. Seyfried and other scientists to secure funding and develop protocols for establishing human clinical trials.

Article Citation:
“The Ketogenic Diet and Hyperbaric Oxygen Therapy Act Synergistically to Prolong Survival in Mice with Systemic Metastatic Cancer;” A.M. Poff, C. Ari, T.N. Seyfried and D.P. D’Agostino; PLOS ONE, June 5, 2013: http://dx.plos.org/10.1371/journal.pone.0065522

Media contact:
Anne DeLotto Baier, USF Health Communications
abaier@health.usf.edu or (813) 9745-3303