30 years of American Society for Neural Therapy and Repair (ASNTR): A personal perspective at the intersection of science, politics, and culture.

The American Society for Neural Therapy and Repair (ASNTR) started 30 years ago in 1993 as the American Society for Neural Transplantation (ASNT), with an emphasis on neural transplantation. Through the years, the Society has been shaped as much by our expanding knowledge of neurodegenerative disorders and how to treat them as it has by politics and culture. What once felt like a leash on neuroscience research, has turned into an advantage as neural transplantation evolved into Neural Therapy and Repair. As a Co-Founder this brief commentary provides a personalized account of our research during the Society's years.

Effects of nutraceutical intervention on serum proteins in aged rats.

Aging is associated with many pathophysiological changes that could lead to the onset of degenerative disease. Some of the physiological changes that occur with aging include increased inflammation and decreased stem cell proliferation, leading to decreased capacity for tissue regeneration and loss of function. In previous studies, we and others have found nutraceutical intervention to ameliorate some of the deleterious effects associated with aging. In particular, we have previously shown that NT-020, a supplement composed of a proprietary blend of blueberries, green tea, vitamin D3, and carnosine, is able to rescue age-related cognitive deficits, impaired neurogenesis, and inflammation in rats. We have also previously demonstrated that stem cells cultured with old serum showed decreased proliferation; however, when stem cells were cultured in serum from old rats given a diet supplemented with NT-020, proliferation did not differ from that of cells cultured with serum from young rats. While it is clear that NT-020 is exerting a therapeutic, anti-aging effect, the mechanisms of action were yet to be fully elucidated.To that end, in the present study, we conducted a bioinformatics experiment to examine the rat proteome of serum from young and old control rats and young and old rats given a diet supplemented with NT-020. Serum from old rats showed an increase in some inflammatory and pro-aging factors while serum from old rats given a diet supplemented with NT-020 showed an increase in some anti-aging factors, most notably proteins associated with the complement system and autophagy. A number of immune functions that increase with age were shown to be downregulated with NT-020 treatment.

Retrospective Case Series of Traumatic Brain Injury and Post-Traumatic Stress Disorder Treated with Hyperbaric Oxygen Therapy.

Returning veterans are frequently diagnosed with traumatic brain injury (TBI) and post-traumatic stress disorder (PTSD). Considering a recent case-controlled study of hyperbaric oxygen therapy (HBOT) reporting a reduction in suicidal ideation, we investigated retrospectively three veterans with chronic TBI/PTSD symptoms who were treated with multiple rounds of HBOT with neurophysiological testing performed before and after treatment. Improvements were detected on parameters within neurocognitive domains, including reductions in suicide-related symptoms. These findings independently confirm that HBOT may be effective in treating specific symptoms of TBI/PTSD that are not currently addressed with existing therapeutic approaches.

Brazilian Jiu Jitsu Training for US Service Members and Veterans with Symptoms of PTSD.

INTRODUCTION: The United States has been actively involved in major armed conflicts over the last 15 years. As a result, a significant proportion of active duty service personnel and returning veterans have endured combat, putting them at risk for developing post-traumatic stress disorder (PTSD), a disabling disorder that may occur after exposure to a traumatic event. Current therapies often require long-term, time-intensive and costly commitment from the patient and have variable degrees of success. There remains an ongoing need for better therapies, including complementary medicine approaches that can effectively reduce PTSD symptoms. While anecdotal evidence suggests that routine practice of Brazilian Jiu Jitsu (BJJ) can reduce symptoms of PTSD, there have been no formal studies to address this. MATERIALS AND METHODS: This study was approved by the University of South Florida Institutional Review Board (#PRO00019430). Male US active duty service members and veterans from the Tampa area participated in a 5-month (40 sessions) BJJ training program. Before beginning and again midway through and upon completion of training the participants completed several validated self-report measures that addressed symptoms of PTSD and other co-morbid conditions. Effect size and 95% confidence intervals were determined using a within-person single-group pretest-posttest design. RESULTS: Study participants demonstrated clinically meaningful improvements in their PTSD symptoms as well as decreased symptoms of major depressive disorder, generalized anxiety and decreased alcohol use; effect sizes varied from 0.80 to 1.85. CONCLUSIONS: The results from this first-of-kind pilot study suggest that including BJJ as a complementary treatment to standard therapy for PTSD may be of value. It will be necessary to validate these promising results with a larger subject cohort and a more rigorous experimental design before routinely recommending this complementary therapy.

Plasma derived from human umbilical cord blood: Potential cell-additive or cell-substitute therapeutic for neurodegenerative diseases.

Limited efficacy of current therapeutic approaches for neurodegenerative disease has led to increased interest in alternative therapies. Cord blood plasma (CBP) derived from human umbilical cord blood (hUCB) may be a potential therapeutic. Benefits of CBP injection into rodent models of aging or ischaemic stroke have been demonstrated, though how benefits are elicited is still unclear. The present study evaluated various factors within the same samples of CBP and human adult blood plasma/sera (ABP/S). Also, autologous CBP effects vs. ABP/S or foetal bovine serum supplements on mononuclear cells from hUCB (MNC hUCB) in vitro were determined. Results showed significantly low concentrations of pro-inflammatory cytokines (IL-2, IL-6, IFN-γ, and TNF-α) and elevated chemokine IL-8 in CBP. Significantly higher levels of VEGF, G-CSF, EGF and FGF-basic growth factors were determined in CBP vs. ABP/S. Autologous CBP media supplements significantly increased MNC hUCB viability and decreased apoptotic cell activity. We are first to demonstrate the unique CBP composition of cytokines and growth factors within the same CBP samples derived from hUCB. Also, our novel finding that autologous CBP promoted MNC hUCB viability and reduced apoptotic cell death in vitro supports CBP's potential as a sole therapeutic or cell-additive agent in developing therapies for various neurodegenerative diseases.

Transplantation of human bone marrow stem cells into symptomatic ALS mice enhances structural and functional blood-spinal cord barrier repair.

Accumulating evidence shows alterations in the blood-brain barrier (BBB) and blood-spinal cord barrier (BSCB) in ALS patients and in animal models of disease, mainly by endothelial cell (EC) damage. Repair of the altered barrier in the CNS by replacement of ECs via cell transplantation may be a new therapeutic approach for ALS. Recently, we demonstrated positive effects towards BSCB repair by intravenous administration of unmodified human bone marrow CD34+ (hBM34+) cells at different doses into symptomatic ALS mice. However, particular benefits of these transplanted cells on microvascular integrity in symptomatic ALS mice are still unclear. The aim of the present study was to determine the structural and functional spinal cord capillary integrity in symptomatic ALS mice after intravenous administration of hBM34+ cells. The G93A mice at 13 weeks of age intravenously received one of three different cell doses (5 × 104, 5 × 105, or 1 × 106) and were euthanized at 17 weeks of age (4 weeks post-transplant). Control groups were media-treated and non-carrier mutant SOD1 gene mice. Capillary ultrastructural (electron microscopy), immunohistochemical (laminin and HuNu), and histological (myelin and capillary density) analyses were performed in the cervical and lumbar spinal cords. Capillary permeability in the spinal cords was determined by Evans Blue (EB) injection. Results showed significant restoration of ultrastructural capillary morphology, improvement of basement membrane integrity, enhancement of axonal myelin coherence, and stabilization of capillary density in the spinal cords primarily of ALS mice receiving the high dose of 1 × 106 cells. Moreover, substantial reduction of parenchymal EB levels was determined in these mice, confirming our previous results on capillary permeability. Additionally, transplanted cells were detected in blood smears of sacrificed late symptomatic mice by HuNu marker. Altogether, these results provide novel evidence that unmodified bone marrow hematopoietic stem cell treatment at optimal dose might be beneficial for structural and functional repair of the damaged BSCB in advanced stage of ALS, potentially resulting in delayed disease progression by increased motor neuron survival.

Suppressed acoustic startle response in traumatic brain injury masks post-traumatic stress disorder hyper-responsivity.

An exaggerated acoustic startle reflex (ASR) is a clinical indicator of anxiety disorders, such as post-traumatic stress disorder (PTSD). Given the prevalence of PTSD following traumatic brain injury (TBI), we studied the effects of TBI on ASR. Adult Sprague Dawley rats exposed to moderate controlled cortical impact injury model of TBI displayed suppression of ASR intensity and sensitivity. As patients with PTSD have been shown to display hyperactive startle responses, the present discrepant observation of TBI-induced suppression of ASR has clinical implications, in that the reduced, instead of elevated, startle response in patients with comorbid TBI/PTSD could be owing to a masking effect of TBI.

Hyperbaric oxygen therapy as a potential treatment for post-traumatic stress disorder associated with traumatic brain injury.

Traumatic brain injury (TBI) describes the presence of physical damage to the brain as a consequence of an insult and frequently possesses psychological and neurological symptoms depending on the severity of the injury. The recent increased military presence of US troops in Iraq and Afghanistan has coincided with greater use of improvised exploding devices, resulting in many returning soldiers suffering from some degree of TBI. A biphasic response is observed which is first directly injury-related, and second due to hypoxia, increased oxidative stress, and inflammation. A proportion of the returning soldiers also suffer from post-traumatic stress disorder (PTSD), and in some cases, this may be a consequence of TBI. Effective treatments are still being identified, and a possible therapeutic candidate is hyperbaric oxygen therapy (HBOT). Some clinical trials have been performed which suggest benefits with regard to survival and disease severity of TBI and/or PTSD, while several other studies do not see any improvement compared to a possibly poorly controlled sham. HBOT has been shown to reduce apoptosis, upregulate growth factors, promote antioxidant levels, and inhibit inflammatory cytokines in animal models, and hence, it is likely that HBOT could be advantageous in treating at least the secondary phase of TBI and PTSD. There is some evidence of a putative prophylactic or preconditioning benefit of HBOT exposure in animal models of brain injury, and the optimal time frame for treatment is yet to be determined. HBOT has potential side effects such as acute cerebral toxicity and more reactive oxygen species with long-term use, and therefore, optimizing exposure duration to maximize the reward and decrease the detrimental effects of HBOT is necessary. This review provides a summary of the current understanding of HBOT as well as suggests future directions including prophylactic use and chronic treatment.

Nutraceutical intervention reverses the negative effects of blood from aged rats on stem cells.

Aging is associated with a decline in function in many of the stem cell niches of the body. An emerging body of literature suggests that one of the reasons for this decline in function is due to cell non-autonomous influences on the niche from the body. For example, studies using the technique of parabiosis have demonstrated a negative influence of blood from aged mice on muscle satellite cells and neurogenesis in young mice. We examined if we could reverse this effect of aged serum on stem cell proliferation by treating aged rats with NT-020, a dietary supplement containing blueberry, green tea, vitamin D3, and carnosine that has been shown to increase neurogenesis in aged rats. Young and aged rats were administered either control NIH-31 diet or one supplemented with NT-020 for 28 days, and serum was collected upon euthanasia. The serum was used in cultures of both rat hippocampal neural progenitor cells (NPCs) and rat bone marrow-derived mesenchymal stem cells (MSCs). Serum from aged rats significantly reduced cell proliferation as measured by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 5-bromo-2'-deoxyuridine (BrdU) assays in both NPCs and MSCs. Serum from aged rats treated with NT-020 was not different from serum from young rats. Therefore, NT-020 rescued the effect of serum from aged rats to reduce stem cell proliferation.

Nutraceutical intervention improves older adults' cognitive functioning.

Interventions to improve the cognitive health of older adults are of critical importance. In the current study, we conducted a double-blind, placebo-controlled clinical trial using a pill-based nutraceutical (NT-020) that contained a proprietary formulation of blueberry, carnosine, green tea, vitamin D3, and Biovin to evaluate the impact on changes in multiple domains of cognitive functioning. One hundred and five cognitively intact adults aged 65-85 years of age (M=73.6 years) were randomized to receive NT-020 (n=52) or a placebo (n=53). Participants were tested with a battery of cognitive performance tests that were classified into six broad domains--episodic memory, processing speed, verbal ability, working memory, executive functioning, and complex speed at baseline and 2 months later. The results indicated that persons taking NT-020 improved significantly on two measures of processing speed across the 2-month test period in contrast to persons on the placebo whose performance did not change. None of the other cognitive ability measures were related to intervention group. The results also indicated that the NT-020 was well tolerated by older adults, and the presence of adverse events or symptoms did not differ between the NT-020 and placebo groups. Overall, the results of the current study were promising and suggest the potential for interventions like these to improve the cognitive health of older adults.

Optimized turmeric extract reduces ß-Amyloid and phosphorylated Tau protein burden in Alzheimer's transgenic mice.

In a previous in vitro study, the standardized turmeric extract, HSS-888, showed strong inhibition of Aß aggregation and secretion in vitro, indicating that HSS-888 might be therapeutically important. Therefore, in the present study, HSS-888 was evaluated in vivo using transgenic 'Alzheimer' mice (Tg2576) over-expressing Aß protein. Following a six-month prevention period where mice received extract HSS-888 (5mg/mouse/day), tetrahydrocurcumin (THC) or a control through ingestion of customized animal feed pellets (0.1% w/w treatment), HSS-888 significantly reduced brain levels of soluble (∼40%) and insoluble (∼20%) Aß as well as phosphorylated Tau protein (∼80%). In addition, primary cultures of microglia from these mice showed increased expression of the cytokines IL-4 and IL-2. In contrast, THC treatment only weakly reduced phosphorylated Tau protein and failed to significantly alter plaque burden and cytokine expression. The findings reveal that the optimized turmeric extract HSS-888 represents an important step in botanical based therapies for Alzheimer's disease by inhibiting or improving plaque burden, Tau phosphorylation, and microglial inflammation leading to neuronal toxicity.

Spirulina promotes stem cell genesis and protects against LPS induced declines in neural stem cell proliferation.

Adult stem cells are present in many tissues including, skin, muscle, adipose, bone marrow, and in the brain. Neuroinflammation has been shown to be a potent negative regulator of stem cell and progenitor cell proliferation in the neurogenic regions of the brain. Recently we demonstrated that decreasing a key neuroinflammatory cytokine IL-1beta in the hippocampus of aged rats reversed the age-related cognitive decline and increased neurogenesis in the age rats. We also have found that nutraceuticals have the potential to reduce neuroinflammation, and decrease oxidative stress. The objectives of this study were to determine if spirulina could protect the proliferative potential of hippocampal neural progenitor cells from an acute systemic inflammatory insult of lipopolysaccharide (LPS). To this end, young rats were fed for 30 days a control diet or a diet supplemented with 0.1% spirulina. On day 28 the rats were given a single i.p. injection of LPS (1 mg/kg). The following day the rats were injected with BrdU (50 mg/kg b.i.d. i.p.) and were sacrificed 24 hours after the first injection of BrdU. Quantification of the BrdU positive cells in the subgranular zone of the dentate gyrus demonstrated a decrease in proliferation of the stem/progenitor cells in the hippocampus as a result of the LPS insult. Furthermore, the diet supplemented with spirulina was able to negate the LPS induced decrease in stem/progenitor cell proliferation. In a second set of studies we examined the effects of spirulina either alone or in combination with a proprietary formulation (NT-020) of blueberry, green tea, vitamin D3 and carnosine on the function of bone marrow and CD34+ cells in vitro. Spirulina had small effects on its own and more than additive effects in combination with NT-020 to promote mitochondrial respiration and/or proliferation of these cells in culture. When examined on neural stem cells in culture spirulina increased proliferation at baseline and protected against the negative influence of TNFalpha to reduce neural stem cell proliferation. These results support the hypothesis that a diet enriched with spirulina and other nutraceuticals may help protect the stem/progenitor cells from insults.

Acute Treatment With Herbal Extracts Provides Neuroprotective Benefits in In Vitro and In Vivo Stroke Models, Characterized by Reduced Ischemic Cell Death and Maintenance of Motor and Neurological Functions.

The present study explored the prophylactic and restorative benefits of cacao and red sage using both in vitro and in vivo models of stroke. For the in vitro study, we initially exposed primary rat cells to the established oxygen-glucose deprivation (OGD) stroke model followed by reperfusion under normoxic conditions, then added different cacao and sage concentrations to the cell culture media. Trypan blue cell viability results revealed specific cacao and sage dosages exerted significant therapeutic effects against OGD-induced cell death compared to cultured cells treated with extract vehicle. We next embarked on testing the therapeutic effects of cacao and sage in an in vivo model of stroke when extract treatment commenced either prior to or after transient middle cerebral artery occlusion (MCAo). Significant reduction in ischemic cell death within the peri-infarct area coupled with better performance in routine motor and neurological tasks were demonstrated by stroke animals that received cacao or sage extracts prior to MCAo compared to those that received the extracts or vehicle after MCAo. In summary, the present results demonstrate that neuroprotective effects were afforded by plant extract treatment, and that the in vitro stroke paradigm approximates in vivo efficacy when considering prophylactic treatment for stroke.

Optimized turmeric extracts have potent anti-amyloidogenic effects.

Inhibition of beta-amyloid (A beta) accumulation and A beta fibril (fA beta) formation from A beta are attractive therapeutic targets for the treatment of Alzheimer's disease (AD). While previous studies have shown anti-amyloidogenic effects of curcumin in vitro and in vivo, no studies have examined optimized turmeric extracts enriched in curcuminoids or turmerones. Three standardized turmeric extracts, HSS-838, HSS-848, and HSS-888, were prepared with different chemical profiles to investigate their potential therapeutic benefits for AD. These extracts were fingerprinted by DART TOF-MS to reveal the significant chemical complexity. In addition four curcuminoids (curcumin, tetrahydrocurcumin, demethoxycurcumin and bisdemethoxycurcumin) were also examined. We measured the effects of the extracts and curcuminoids, on the aggregation of A beta by using a thioflavin T cell-free assay and the secretion of A beta from human neuronal cells (SweAPP N2A cells) in vitro. All three extracts and the curcuminoids showed dose-dependent inhibition of fA beta aggregation from A beta(1-42) in the cell-free assay, with IC(50) values of

From the basics to application of cell therapy, a steppingstone to the conquest of neurodegeneration: a meeting report.

The annual meeting of the American Society for Neural Therapy and Repair (ASNTR) showcases the latest research trends in neurodegenerative disease and the related medical regenerative science. The 2008 ASNTR meeting covered a variety of different topics ranging from basic research to exploration of currently unknown pathogenesis and mechanisms for specific neurodegenerative disease such as Parkinson's disease, Alzheimer's disease, or stroke. This included studies to characterize stem cells, such as neural stem cells, embryonic stem cells, bone marrow mesenchymal stem cells, and human umbilical cord blood cells, for transplantation and the conditions necessary to maximize the efficacy of endogenous and exogenous stem cells, such as isolation, purification, differentiation, and migration. Moreover, a number of studies looked at methods for more advanced application of transplantation of cells or specific factors, through tissue engineering or manipulation beyond simple injection. Finally, well-known or previously un-known dietary supplementation or pharmacological materials that can affect the nervous system positively or negatively, were also important topics.

Dietary supplementation exerts neuroprotective effects in ischemic stroke model.

This study examined whether dietary supplementation can be used to protect against ischemic stroke. Two groups of adult male Sprague-Dawley rats initially received NT-020, a proprietary formulation of blueberry, green tea, Vitamin D3, and carnosine (n = 8), or vehicle (n = 7). Dosing for NT-020 and vehicle consisted of daily oral administration (using a gavage) over a 2-week period. On day 14 following the last drug treatment, all animals underwent the stroke surgery using the transient 1-hour suture occlusion of middle cerebral artery (MCAo). To reveal the functional effects of NT-020, animals were subjected to established behavioral tests just prior to stroke surgery and again on day 14 post-stroke. ANOVA revealed significant treatment effects (p < 0.05), characterized by reductions of 11.8% and 24.4% in motor asymmetry and neurologic dysfunction, respectively, in NT-020-treated stroke animals compared to vehicle-treated stroke animals. Evaluation of cerebral infarction revealed a significant 75% decrement in mean glial scar area in the ischemic striatum of NT-020-treated stroke animals compared to that of vehicle-treated stroke animals (p < 0.0005). Quantitative analysis of subventricular zone's cell proliferative activity revealed at least a one-fold increment in the number of BrdU-positive cells in the NT-020-treated stroke brains compared to vehicle-treated stroke brains (p < 0.0005). Similarly, quantitative analysis of BrdU labeling in the ischemic striatal penumbra revealed at least a three-fold increase in the number of BrdU-positive cells in the NT-020-treated stroke brains compared to vehicle-treated stroke brains (p < 0.0001). In addition, widespread double labeling of cells with BrdU and doublecortin was detected in NT-020-treated stroke brains (intact side 17% and ischemic side 75%), which was significantly higher than those seen in vehicle-treated stroke brains (intact side 5% and ischemic side 13%) (p < 0.05). In contrast, only a small number of cells in NT-020-treated stroke brains double labeled with BrdU and GFAP (intact side 1% and ischemic side 2%), which was significantly lower than those vehicle-treated stroke brains (intact side 18% and ischemic side 35%) (p < 0.0001). Endogenous neurogenic factors were also significantly upregulated in the ischemic brains of NT-020-treated stroke animals. These data demonstrate the remarkable neuroprotective effects of NT-020 when given prior to stroke, possibly acting via its neurogenic potential.

Oxidative stress of neural, hematopoietic, and stem cells: protection by natural compounds.

During natural aging, adult stem cells are known to have a reduced restorative capacity and are more vulnerable to oxidative stress resulting in a reduced ability of the body to heal itself. We report here that the proprietary natural product formulation, NT020, previously found to promote proliferation of human hematopoietic stem cells, reduced oxidative stress-induced apoptosis of murine neurons and microglial cells in vitro. Furthermore, when taken orally for 2 weeks, cultured bone marrow stem cells from these mice exhibited a dose-related reduction of oxidative stress-induced apoptosis. This preclinical study demonstrates that NT020 can act to promote healing via an interaction with stem cell populations and forms the basis of conducting a clinical trial to determine if NT020 exhibits similar health promoting effects in humans when used as a dietary supplement.

Nutraceuticals synergistically promote proliferation of human stem cells.

A viable alternative to stem cell transplantation is to design approaches that stimulate endogenous stem cells to promote healing and regenerative medicine. Many natural compounds have been shown to promote healing; however, the effects of these compounds on stem cells have not been investigated. We report here the effects of several natural compounds on the proliferation of human bone marrow and human CD34(+) and CD133(+) cells. A dose-related effect of blueberry, green tea, catechin, carnosine, and vitamin D(3) was observed on proliferation with human bone marrow as compared with human granulocyte-macrophage colony-stimulating factor (hGM-CSF). We further show that combinations of nutrients produce a synergistic effect to promote proliferation of human hematopoietic progenitors. This demonstrates that nutrients can act to promote healing via an interaction with stem cell populations.