A new University of South Florida preclinical study finds that the regenerative cell therapy boosts motor nerve cell survival by repairing the blood-spinal cord barrier

TAMPA, Fla. (April 2, 2019) — Transplantation of human bone marrow-derived endothelial progenitor cells (EPCs) into mice mimicking symptoms of amyotrophic lateral sclerosis (ALS) helped more motor neurons survive and slowed disease progression by repairing damage to the blood-spinal cord barrier (BSCB), University of South Florida researchers report.

The study was published March 27 in Scientific Reports, one of the Nature journals. The findings contribute to a growing body of work exploring cell therapy approaches to barrier repair in ALS and other neurodegenerative diseases.

Human bone marrow-derived endothelial progenitor cells in vitro

The progressive degeneration of nerve cells that control muscle movement (motor neurons) eventually leads to total paralysis and death from ALS. Each day, an average of 15 Americans are diagnosed with the disease, according to the ALS Association.

Damage to the barrier between the blood circulatory system and the central nervous system has been recognized as a key factor in the development of ALS. A breach in this protective wall opens the brain and spinal cord to immune/inflammatory cells and other potentially harmful substances circulating in peripheral blood. The cascade of biochemical events leading to ALS includes alterations of endothelial cells lining the inner surface of tiny blood vessels near damaged spinal cord motor neurons.

This latest study by lead author Svitlana Garbuzova-Davis, PhD, and colleagues at the USF Health Morsani College of Medicine’s Center of Excellence for Aging & Brain Repair, builds upon a previous study showing that human bone marrow-derived stem cells improved motor functions and nervous system conditions in symptomatic ALS mice by advancing barrier repair. However, in that earlier USF study the beneficial effect was delayed until several weeks after cell transplant and some severely damaged capillaries were detected even after a high-dose treatment. So in this study, the researchers tested whether human EPCs – cells harvested from bone marrow but more genetically similar to vascular endothelial cells than undifferentiated stem cells – would provide even better BSCB restoration.

Svitlana Garbuzova-Davis, PhD

ALS mice were intravenously administered a dose of human bone-marrow derived EPCs.  Four weeks after transplant, the results of the active cell treatment was compared against findings from two other groups of mice:  ALS mice receiving a media (saline) treatment and untreated healthy mice.

The symptomatic ALS mice receiving EPC treatments demonstrated significantly improved motor function, increased motor neuron survival and slower disease progression than their symptomatic counterparts injected with media. The researchers suggest that these benefits leading to BSCB repair may have been promoted by widespread attachment of EPCs to capillaries in the spinal cord. To support this proposal, they point to evidence of substantially restored capillaries, less capillary leakage, and re-establishment of structural support cells (perivascular astrocytes) that play a role in helping form a protective barrier in the spinal cord and brain.

Further research is needed to clearly define the mechanisms of EPC barrier repair.  But, the study authors conclude: “From a translational viewpoint, the initiation of cell treatment at the symptomatic disease stage offered robust restoration of BSCB integrity and shows promise as a future clinical therapy for ALS.”

The USF study was supported by a grant from the National Institute of Neurological Disorders and Stroke.

Article citation:
Svitlana Garbuzova-Davis, Crupa Kurien, Edward Haller, David J. Eve, Stephanie Navarro, George Steiner, Ajay Mahendrasah, Surafuale Hailu, Mohammed Khatib, Kayla J. Boccio, Cesario V. Borlongan, Harry R. Van Loveren, Stanley H. Appel and Paul R. Sanberg. Human Bone Marrow Endothelial Progenitor Cell Transplantation into Symptomatic ALS Mice Delays Disease Progression and Increases Motor Neuron Survival by Repairing Blood-Spinal Cord Barrier, Scientific Reports, March 27, 2019. https://doi.org/10.1038/s41598-019-41747-4.

ALS mice improved with stem cell therapy; first step for science in finding better treatment  

TAMPA, Fla. (May 12, 2017) – Researchers at the University of South Florida show in a new study that bone marrow stem cell transplants helped improve motor functions and nervous system conditions in mice with the disease amyotrophic lateral sclerosis (ALS) by repairing damage to the  blood-spinal cord barrier.

In a study recently published in the journal Scientific Reports, researchers in USF’s Center of Excellence for Aging and Brain Repair say the results of their experiment are an early step in pursuing stem cells for potential repair of the blood-spinal cord barrier, which has been identified as key in the development of ALS. USF Health Professor Svitlana Garbuzova-Davis, PhD, led the project.

Previous studies in development of various therapeutic approaches for ALS typically used pre-symptomatic mice. This is the first study advancing barrier repair that treats symptomatic mice, which more closely mirrors conditions for human patients, Dr. Garbuzova-Davis said.

Svitlana Garbuzova-Davis, PhD, led the study.

Using stem cells harvested from human bone marrow, researchers transplanted cells into mice modeling ALS and already showing disease symptoms. The transplanted stem cells differentiated and attached to vascular walls of many capillaries, beginning the process of blood-spinal cord barrier repair.

The stem cell treatment delayed the progression of the disease and led to improved motor function in the mice, as well as increased motor neuron cell survival, the study reported.

ALS is a progressive neurodegenerative disease that affects neuronal cells in the brain and the spinal cord, which send signals to control muscles throughout the body. The progressive degeneration of motor neuron cells leads to death from ALS. More than 6,000 Americans each year are diagnosed with the disease.

Because stem cells have the ability to develop into many different cell types in the body, researchers at USF’s Center of Excellence for Aging and Brain Repair, Department of Neurosurgery and Brain Repair have focused on using stem cells to restore function lost through neurodegenerative disorders or injuries.

Damage to the barrier between the blood circulatory system and the central nervous system has been recently recognized as a factor in ALS development, leading researchers to work on targeting the barrier for repair as a potential strategy for ALS therapy.

In this study, the ALS mice were given intravenous treatments of one of three different doses of the bone marrow stem cells. Four weeks after treatment, the scientists determined improved motor function and enhanced motor neuron survival. The mice receiving the higher doses of stem cells fared better in the study, the researcher noted.

The transplanted stem cells had differentiated into endothelial cells – which form the inner lining of a blood vessel, providing a barrier between blood and spinal cord tissue – and attached to capillaries in the spinal cord. Furthermore, the researchers observed reductions in activated glial cells, which contribute to inflammatory processes in ALS.

USF Health Morsani College of Medicine researchers Crupa Kurien, Avery Thomson, Dimitri Falco, Sohaib Ahmad, Joseph Staffetti, George Steiner, Sophia Abraham, Greeshma James, Ajay Mahendrasah, Paul R. Sanberg and Cesario V. Borlongan joined in the project. The study was funded by the National Institutes of Health, National Institute of Neurological Disorders and Stroke.

Read the full study here.

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Stroke’s long-term effects on blood-spinal cord barrier can lead to ‘an increasingly toxic environment’ in spinal cord and ‘significant input on disease pathology’

Tampa, Fla. (June 14, 2016) – A team of researchers at the University of South Florida investigating the short and long-term effects of ischemic stroke in a rodent model has found that stroke can cause long-term damage to the blood-spinal cord barrier (BSCB), creating a “toxic environment” in the spinal cord that might leave stroke survivors susceptible to motor dysfunction and disease pathology.

The paper describing their study was recently published online and will appear in an upcoming issue of Journal of Neuropathology and Experimental Neurology.

“This study, carried out using laboratory rats modeling stroke, demonstrated that ischemic stroke — in both its subacute and chronic stages — damages the BSCB in a variety of ways, creating a toxic environment in the spinal cord that can lead to further disability and exacerbate disease pathology,” said study lead author Svitlana Garbuzova-Davis, PhD, associate professor in USF’s Center of Excellence for Aging and Brain Repair, Department of Neurosurgery and Brain Repair. “The aim of our study was to evaluate post-stroke BSCB condition that might lead to the development of more effective therapies for stroke survivors.”

Svitlana Garbuzova-Davis, PhD

The BSCB provides a specialized protective ‘microenvironment’ for neural cells in the spinal cord. Substantial vascular damage is a major pathologic feature of both subacute and chronic stroke caused by an extended period of microvascular permeability after the BSCB loses integrity. Damage to the BSCB, explained the researchers, plays a fundamental role in the development of several pathological conditions, including abnormal motor function.

The researchers, who evaluated the BSCB in test animals at seven and 30 days after stroke modeling, found that ischemic stroke damaged the gray and white matter in the cervical spinal cord on both sides of the spinal column, based on analysis of electron microscope images. Among the effects were damage to neural cells called ‘astrocytes,’ loss of motor neurons, reduced integrity of a tight junction protein between barrier cells, and swollen axons with damaged myelin in ascending and descending tracts connecting to the brain.

They also found stroke-associated ‘upregulation’ of Beclin-1 in endothelial cells composing the BSCB. Beclin-1, explained the researchers, helps induce autophagy, an activity associated with removal of various intracellular components. They also observed a decrease in LC3B, an essential autophagy protein, at a later stage post-stroke. These observations of Beclin-1 and LC3B suggest an impaired post-stroke autophagy process in spinal cord capillaries, inducing endothelial cell degeneration.

These stroke-related alterations in the cervical spinal cord indicate pervasive and long-lasting BSCB damage that would severely affect spinal cord function, wrote the researchers, adding that the widespread microvascular impairment in the gray and white matter of the cervical spinal cord aggravated motor neuron deterioration and had the potential to cause motor dysfunction.

“Because our investigations on the post-stroke microvascular alterations, including BSCB damage, have just begun, many questions remain,” said senior author Cesario Borlongan, PhD, professor and director of the USF Center of Excellence for Aging and Brain Repair. “Specifically, the protein expression responsible for endothelial cell degeneration and tight junction damage we identified in this study needs to be confirmed through further tests. Also, behavioral tests of motor function in post-stroke animals in correlation with BSCB damage are needed. These questions and others will be addressed in our future studies.”

Paul R. Sanberg, PhD, DSc, Distinguished University Professor, a co-author of the paper, concluded that “these novel data showing BSCB damage in subacute and chronic ischemic stroke may lead to development of new therapeutic approaches for patients with ischemic cerebral infarction.”

Article citation:
Blood-Spinal Cord Barrier Alterations in Subacute and Chronic Stages of a Rat Model of Focal Cerebral Ischemia. Svitlana Garbuzova-Davis; Edward Haller; Naoki Tajiri; Avery Thomson; Jennifer Barretta; Stephanie N. Williams; Eithan D. Haim; Hua Qin; Aric Frisina-Deyo; Jerry V. Abraham; Paul R. Sanberg; Harry Van Loveren; Cesario V. Borlongan. Journal of Neuropathology & Experimental Neurology 2016; doi: 10.1093/jnen/nlw040.

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USF Health’s mission is to envision and implement the future of health. It is the partnership of the USF Health Morsani College of Medicine, the College of Nursing, the College of Public Health, the College of Pharmacy, the School of Physical Therapy and Rehabilitation Sciences, the Biomedical Sciences Graduate and Postdoctoral Programs, and the USF Physicians Group. The University of South Florida is a Top 50 research university in total research expenditures among both public and private institutions nationwide, according to the National Science Foundation. For more information, visit www.health.usf.edu

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Findings suggest that blood-brain barrier integrity suffers days after ischemic stroke, leading to serious complications; repair of this protective barrier might prevent them

Tampa, FL (May 20, 2013) — While the effects of acute stroke have been widely studied, brain damage during the subacute phase of stroke has been a neglected area of research. Now, a new study by the University of South Florida  reports that within a week of a stroke caused by a blood clot in one side of the brain, the opposite side of the brain shows signs of microvascular injury.  

Stroke is a leading cause of death and disability in the United States, and increases the risk for dementia.

 “Approximately 80 percent of strokes are ischemic strokes, in which the blood supply to the brain is restricted, causing a shortage of oxygen,” said study lead author Svitlana Garbuzova-Davis, PhD, associate professor in the USF Department of Neurosurgery and Brain Repair. “Minutes after ischemic stroke, there are serious effects within the brain at both the molecular and cellular levels.  One understudied aspect has been the effect of ischemic stroke on the competence of the blood-brain barrier and subsequent related events in remote brain areas.”

USF Health neuroscientist Svitlana Garbuzova-Davis, PhD, was lead author of the study.

Using a rat model, researchers at USF Health investigated the subacute phase of ischemic stroke and found deficits in the microvascular integrity in the brain hemisphere opposite to where the initial stroke injury occured.

The study was published in the May 10, 2013 issue of PLOS One.

The USF team found that “diachisis,” a term used to describe certain brain deficits remote from primary insult, can occur during the subacute phase of ischemic stroke. The research discovered diachisis is closely related to a breakdown of the blood-brain barrier, which separates circulating blood from brain tissue.

In the subacute phase of an ischemic stroke, when the stroke-induced disturbances in the brain occur in remote brain microvessels, several areas of the brain are affected by a variety of injuries, including neuronal swelling and diminished myelin in brain structures. The researchers suggest that recognizing the significance of microvascular damage could make the blood-brain barrier (BBB) a therapeutic “target” for future neuroprotective strategies for stroke patients.

The mechanisms of BBB permeability at different phases of stroke are poorly understood.  While there have been investigations of BBB integrity and processes in ischemic stroke, the researchers said, most examinations have been limited to the phase immediately after stroke, known as acute stroke.  Their interest was in determining microvascular integrity in the brain hemisphere opposite to an initial stroke injury at the subacute phase.

Accordingly, this study using rats with surgically-simulated strokes was designed to investigate the effect of ischemic stroke on the BBB in the subacute phase, and the effects of a compromised BBB upon various brain regions, some distant from the stroke site.

“The aim of this study was to characterize subacute diachisis in rats modeled with ischemic stroke,” said co-author Cesar Borlongan, PhD, professor and vice chairman for research in the Department of Neurosurgery and Brain Repair and director of the USF Center for Aging and Brain Repair.  “Our specific focus was on analyzing the condition of the BBB and the processes in the areas of the brain not directly affected by ischemia. BBB competence in subacute diachisis is uncertain and needed to be studied.”

Their findings suggest that damage to the BBB, and subsequent vascular leakage as the BBB becomes more permeable, plays a major role in subacute diachisis. 

The increasing BBB permeability hours after the simulated stroke, and finding that the BBB “remained open” seven days post-stroke, were significant findings, said Dr. Garbuzova-Davis, who is also a researcher in USF Center for Aging and Brain Repair. “Since increased BBB permeability is often associated with brain swelling, BBB leakage may be a serious and life-threatening complication of ischemic stroke.”

Another significant aspect was the finding that autophagy — a mechanism involving cell degradation of unnecessary or dysfunctional cellular components —plays a role in the subacute phase of ischemia.  Study results showed that accumulation of numerous autophagosomes within endothelial cells in microvessels of both initially damaged and non-injured brain areas might be closely associated with BBB damage.  Autophagy is a complex but normal process usually aimed at “self-removing” damaged cell components to promote cell survival. It was unclear, however, whether the role of autophagy in subacute post-ischemia was promoting cell survival or cell death.

More than 30 percent of patients who survive strokes develop dementia within two years, the researchers noted.

“Although dementia is complex, vascular damage in post-stroke patients is a significant risk factor, depending on the severity, volume and site of the stroke,” said study co-author Dr. Paul Sanberg, USF senior vice president for research and innovation. “Ischemic stroke might initiate neurodegenerative dementia, particularly in the aging population.”

The researchers conclude that repair of the BBB following ischemic stroke could potentially prevent further degradation of surviving neurons.

“Recognizing that the BBB is a therapeutic target is important for developing neuroprotective strategies,” they said.

In addition to researchers from USF, researchers from the Ribeirao Preto School of Medicine, University of Sao Paulo, Sao Paulo, Brazil, contributed to the study.

The study was supported by the National Institutes of Health (1RO1NS071956-01A1) and the James and Esther King Biomedical Research Program (1KG01-33966).

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The Department of Neurosurgery and Brain Repair has strong and ongoing commitments to  neurosurgical education both at the residency and fellowship level and also to providing ongoing educational efforts for practicing physicians. Multiple avenues of research under investigation in the department include conducting clinical research in the fields of complex spinal disorders, epilepsy, brain tumor treatment, and stroke care, among others. In the departmental laboratories, studies of spinal biomechanics, neurodegenerative disorders, and microsurgical techniques are also underway. We are trying to improve the treatments available for our neurosurgical patients and neurological patients throughout the world. http://health.usf.edu/medicine/neurosurgery/index.htm

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The mission of the Center of Excellence for Aging and Brain Repair is to develop new therapeutic strategies to promote repair and regeneration of aging and diseased brain. Building on a foundation of excellence in basic and clinical research, the center focuses on translating innovative ideas into industrial partnerships and educational and clinical services to address key needs of the community and those suffering from brain injury and disease. http://health.usf.edu/nocms/medicine/neurosurgery/ceabr/