Publications by authors named "Syed F Ali"

195 Publications

Effects of acetyl L-carnitine on zebrafish embryos: Phenotypic and gene expression studies.

J Appl Toxicol 2021 Feb 20;41(2):256-264. Epub 2020 Jul 20.

Division of Neurotoxicology, National Center for Toxicological Research, United States Food and Drug Administration, Jefferson, AR, USA.

Acetyl L-carnitine (ALCAR), a dietary supplement and an antioxidant, plays a vital role in the bioenergetic process that produces ATP. Although there are reports on antioxidant toxicity, there is no information on the potential toxicity of ALCAR. Here, using zebrafish embryos, we explored whether ALCAR modulated ATP synthesis, generation of reactive oxygen species (ROS) and expression of specific genes related to major signaling pathways that control metabolism, growth, differentiation, apoptosis and oxidative stress. First, we show that ALCAR elicits a physiologic response, as ATP levels increased after ALCAR treatment. Simultaneously, an increase in the expression of ROS, a by-product of ATP synthesis, was observed in the ALCAR-treated embryos. Consistent with higher ROS expression, the level of cysteine, a precursor of glutathione, was significantly reduced. ALCAR did not have any drastic effect on overall development and heart rate. Polymerase chain reaction-based gene expression array analyses showed no significant change in the expression of 83 genes related to 10 major signaling pathways including: the transforming growth factor β (TGFβ), Wingless and Int-1 (Wnt), nuclear factor κ-light-chain-enhancer of activated B cells (NF-κB), Janus kinase/signal transducers and activators of transcription (JAK/STAT), p53, Notch, Hedgehog, Peroxisome proliferator-activated receptor (PPAR), oxidative stress, and hypoxia pathways. Our results show that the expression of 83 genes related to these major signaling pathways did not change significantly.
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http://dx.doi.org/10.1002/jat.4041DOI Listing
February 2021

Nifedipine toxicity is exacerbated by acetyl l-carnitine but alleviated by low-dose ketamine in zebrafish in vivo.

J Appl Toxicol 2020 02 9;40(2):257-269. Epub 2019 Oct 9.

Division of Neurotoxicology, US Food and Drug Administration, Jefferson, Arkansas.

Calcium channel blocker (CCB) poisoning is a common and sometimes life-threatening emergency. Our previous studies have shown that acetyl l-carnitine (ALCAR) prevents cardiotoxicity and developmental toxicity induced by verapamil, a CCB used to treat patients with hypertension. Here, we tested whether toxicities of nifedipine, a dihydropyridine CCB used to treat hypertension, can also be mitigated by co-treatment with ALCAR. In the zebrafish embryos at three different developmental stages, nifedipine induced developmental toxicity with pericardial sac edema in a dose-dependent manner, which were surprisingly exacerbated with ALCAR co-treatment. Even with low-dose nifedipine (5 μm), when the pericardial sac looked normal, ALCAR co-treatment showed pericardial sac edema. We hypothesized that toxicity by nifedipine, a vasodilator, may be prevented by ketamine, a known vasoconstrictor. Nifedipine toxicity in the embryos was effectively prevented by co-treatment with low (subanesthetic) doses (25-100 μm added to the water) of ketamine, although a high dose of ketamine (2 mm added to the water) partially prevented the toxicity.As expected of a CCB, nifedipine either in the presence or absence of ketamine-reduced metabolic reactive oxygen species (ROS), a downstream product of calcium signaling, in the rapidly developing digestive system. However, nifedipine induced ROS in the trunk region that showed significantly stunted growth indicating that the tissues under stress potentially produced pathologic ROS. To the best of our knowledge, these studies for the first time show that nifedipine and the dietary supplement ALCAR together induce adverse effects while providing evidence on the therapeutic efficacy of subanesthetic doses of ketamine against nifedipine toxicity in vivo.
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http://dx.doi.org/10.1002/jat.3901DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7015281PMC
February 2020

Stretch-Induced Deformation as a Model to Study Dopaminergic Dysfunction in Traumatic Brain Injury.

Neurochem Res 2019 Nov 16;44(11):2546-2555. Epub 2019 Sep 16.

Division of Neurotoxicology, National Center for Toxicological Research, 3900 NCTR Road, Jefferson, AR, 72079, USA.

Traumatic brain injury (TBI) is defined as damage to the brain that consequently disrupts normal function. Neuronal death, a hallmark of TBI, has been related to the development of neurodegenerative disorders like Parkinson's disease (PD), where loss of dopaminergic neurons and dopaminergic dysfunction are observed. To date, no in vitro model exists in which the dopaminergic damage observed in TBI is replicated. In this study, we evaluated the effects of in vitro simulated TBI on human dopaminergic neurons. To simulate TBI, neurons were subjected to 0%, 5%, 10%, 15%, 25% and 50% deformation. 24 h after injury, cell viability and apoptosis were determined by lactate dehydrogenase (LDH) release and DNA fragmentation, as well as ethidium homodimer and caspase 3/7 staining. Dopamine (DA) levels were determined by ELISA. Levels of tyrosine hydroxylase (TH) and DA transporter (DAT) were determined by western blot. Only 50% stretch increased LDH release and ethidium homodimer staining, suggesting the induction of necrosis. On the contrary, 25% and 50% stretch increased DNA fragmentation while 15%, 25% and 50% increased caspase 3/7 staining, suggesting that moderate and severe TBI promote apoptosis. Levels of intracellular DA decreased in a stretch-dependent manner with 15%, 25% and 50% stretch, which were related with a decrease in TH expression. Extracellular DA levels increased only at 50%. Levels of DAT remained unchanged regardless of treatment. These data support the use of stretch as a model to simulate TBI in vitro in human dopaminergic neurons, replicating the acute effects of TBI in the dopaminergic system.
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http://dx.doi.org/10.1007/s11064-019-02872-8DOI Listing
November 2019

Amyloid Beta 25-35 induces blood-brain barrier disruption in vitro.

Metab Brain Dis 2019 10 2;34(5):1365-1374. Epub 2019 Jul 2.

Division of Neurotoxicology, National Center for Toxicological Research/U.S. Food and Drug Administration, Jefferson, AR, 72079, USA.

The amyloid β-peptide (Aβ) is transported across the blood-brain barrier (BBB) by binding with the receptor for advanced glycation end products (RAGE). Previously, we demonstrated that the Aβ fraction 25-35 (Aβ) increases RAGE expression in the rat hippocampus, likely contributing to its neurotoxic effects. However, it is still debated if the interaction of Aβ with RAGE compromises the BBB function in Alzheimer' disease (AD). Here, we evaluated the effects of Aβ in an established in vitro model of the BBB. Rat brain microvascular endothelial cells (rBMVECs) were treated with 20 μM active Aβ or the inactive Aβ (control), for 24 h. Exposure to Aβ significantly decreased cell viability, increased cellular necrosis, and increased the production of reactive oxygen species (ROS), which triggered a decrease in the enzyme glutathione peroxidase when compared to the control condition. Aβ also increased BBB permeability by altering the expression of tight junction proteins (decreasing zonula occludens-1 and increasing occludin). Aβ induced monolayer disruption and cellular disarrangement of the BBB, with RAGE being highly expressed in the zones of disarrangement. Together, these data suggest that Aβ-induces toxicity by compromising the functionality and integrity of the BBB in vitro. Graphical abstract Aβ induces BBB dysfunction in vitro, wich is likely mediated by OS and ultimately leads to disruption of BBB integrity and cell death.
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http://dx.doi.org/10.1007/s11011-019-00447-8DOI Listing
October 2019

An Alternative In Vitro Method for Examining Nanoparticle-Induced Cytotoxicity.

Int J Toxicol 2019 Sep/Oct;38(5):385-394. Epub 2019 Jun 24.

Nanotechnology Core Facility, Office of Scientific Coordination, NCTR, FDA, Jefferson, AR, USA.

Conventional in vitro assays are often used as initial screens to identify potential toxic effects of nanoparticles (NPs). However, many NPs have shown interference with conventional in vitro assays, resulting in either false-positive or -negative outcomes. Here, we report an alternative method for the in vitro assessment of NP-induced cytotoxicity utilizing Fluoro-Jade C (FJ-C). To provide proof of concept and initial validation data, Ag-NPs and Au-NPs were tested in 3 different cell cultures including rat brain microvessel endothelial cells, mouse neural stem cells, and the human SH-SY5Y cell line. Conventional 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT) and lactate dehydrogenase (LDH) assays were run in parallel with the new method and served as references. The results demonstrate for the first time that FJ-C labeling can be a useful tool for assessing NP-induced cytotoxicity in vitro. Using these approaches, it was also demonstrated that removal of Ag-NPs-while keeping the Ag-ions that were released from the Ag-NPs in culture media-abolished the measured cytotoxicity, indicating that Ag-NPs rather than Ag-ions in solution contributed to the observed cytotoxic effects. Further, co-treatment of Ag-NPs with N-acetyl cysteine (NAC) prevented the observed cytotoxicity, suggesting a protective role of NAC in Ag-NP-induced cytotoxicity. Thus, this alternative in vitro assay is well suited for identify potential cytotoxicity associated with exposure to NPs.
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http://dx.doi.org/10.1177/1091581819859267DOI Listing
February 2020

Ketamine-induced attenuation of reactive oxygen species in zebrafish is prevented by acetyl l-carnitine in vivo.

Neurosci Lett 2019 07 9;706:36-42. Epub 2019 May 9.

Division of Neurotoxicology, National Center for Toxicological Research, U.S. Food and Drug Administration, 3900 NCTR Road, Jefferson, AR, 72079, USA. Electronic address:

Ketamine, an anesthetic, is a non-competitive antagonist of the calcium-permeable N-methyl-d-aspartate (NMDA) receptor. High concentrations of ketamine have been implicated in cardiotoxicity and neurotoxicity. Often, these toxicities are thought to be mediated by reactive oxygen species (ROS). However, findings to the contrary showing ketamine reducing ROS in mammalian cells and neurons in vitro, are emerging. Here, we determined the effects of ketamine on ROS levels in zebrafish larvae in vivo. Based on our earlier studies demonstrating reduction in ATP levels by ketamine, we hypothesized that as a calcium antagonist, ketamine would also prevent ROS generation, which is a by-product of ATP synthesis. To confirm that the detected ROS in a whole organism, such as the zebrafish larva, is specific, we used diphenyleneiodonium (DPI) that blocks ROS production by inhibiting the NADPH Oxidases (NOX). Upon 20 h exposure, DPI (5 and 10 μM) and ketamine at (1 and 2 mM) reduced ROS in the zebrafish larvae in vivo. Using acetyl l-carnitine (ALCAR), a dietary supplement, that induces mitochondrial ATP synthesis, we show elevated ROS generation with increasing ALCAR concentrations. Combined, ketamine and ALCAR counter-balanced ROS generation in the larvae suggesting that ketamine and ALCAR have opposing effects on mitochondrial metabolism, which may be key to maintaining ROS homeostasis in the larvae and affords ALCAR the ability to prevent ketamine toxicity. These results for the first time show ketamine's antioxidative and ALCAR's prooxidative effects in a live vertebrate.
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http://dx.doi.org/10.1016/j.neulet.2019.05.009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6556428PMC
July 2019

Characterization of Serum Exosomes from a Transgenic Mouse Model of Alzheimer's Disease.

Curr Alzheimer Res 2019 ;16(5):388-395

Division of Neurotoxicology National Center for Toxicological Research, 3900 NCTR Road, Jefferson, AR. 72079, United States.

Background: Alzheimer's Disease (AD) is the most common type of dementia characterized by amyloid plaques containing Amyloid Beta (Aβ) peptides and neurofibrillary tangles containing tau protein. In addition to neuronal loss, Cerebral Amyloid Angiopathy (CAA) commonly occurs in AD. CAA is characterized by Aβ deposition in brain microvessels. Recent studies have suggested that exosomes (cell-derived vesicles containing a diverse cargo) may be involved in the pathogenesis of AD.

Objective: Isolate and characterize brain-derived exosomes from a transgenic mouse model of AD that presents CAA.

Methods: Exosomes were isolated from serum obtained from 13-month-old wild type and AD transgenic female mice using an exosome precipitation solution. Characterization of exosomal proteins was performed by western blots and dot blots.

Results: Serum exosomes were increased in transgenic mice compared to wild types as determined by increased levels of the exosome markers flotillin and alix. High levels of neuronal markers were found in exosomes, without any difference any between the 2 groups. Markers for endothelial-derived exosomes were decreased in the transgenic model, while astrocytic-derived exosomes were increased. Exosome characterization showed increased levels of oligomeric Aβ and oligomeric and monomeric forms tau on the transgenic animals. Levels of amyloid precursor protein were also increased. In addition, pathological and phosphorylated forms of tau were detected, but no difference was observed between the groups.

Conclusion: These data suggest that monomeric and oligomeric forms of Aβ and tau are secreted into serum via brain exosomes, most likely derived from astrocytes in the transgenic mouse model of AD with CAA. Studies on the implication of this event in the propagation of AD are underway.
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http://dx.doi.org/10.2174/1567205016666190321155422DOI Listing
August 2020

Cytotoxicity profile of pristine graphene on brain microvascular endothelial cells.

J Appl Toxicol 2019 07 19;39(7):966-973. Epub 2019 Feb 19.

Division of Neurotoxicology, National Center for Toxicological Research, Jefferson, AR, USA.

Graphene-based nanomaterials hold the potential to be used in a wide variety of applications, including biomedical devices. Pristine graphene (PG) is an un-functionalized, defect-free type of graphene that could be used as a material for neural interfacing. However, the neurotoxic effects of PG, particularly to the blood-brain barrier (BBB), have not been fully studied. The BBB separates the brain tissue from the circulating substances in the blood and is essential to maintain the brain homeostasis. The principal components of the BBB are brain microvascular endothelial cells (BMVECs), which maintain a protectively low permeability due to the expression of tight junction proteins. Here we analyzed the effects of PG on BMVECs in an in vitro model of the BBB. BMVECs were treated with PG at 0, 10, 50 and 100 μg/mL for 24 hours and viability and functional analyses of BBB integrity were performed. PG increased lactate dehydrogenase release at 50 and 100 μg/mL, suggesting the induction of necrosis. Surprisingly, 2,3,-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino)-carbonyl]-2H-tetrazolium (XTT) conversion was increased at 10 and 50 μg/mL. In contrast, XTT conversion was decreased at 100 μg/mL, suggesting the induction of cell death. In addition, 100 μg/mL PG increased DNA fragmentation, suggesting induction of apoptosis. At the same time, 50 and 100 μg/mL of PG increased the endothelial permeability, which corresponded with a decrease in the expression of the tight junction protein occludin at 100 μg/mL. In conclusion, these results suggest that PG negatively affects the viability and function of the BBB endothelial cells in vitro.
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http://dx.doi.org/10.1002/jat.3786DOI Listing
July 2019

Rates, Characteristics, and Outcomes of Patients Transferred to Specialized Stroke Centers for Advanced Care.

Circ Cardiovasc Qual Outcomes 2018 09;11(9):e003359

Massachusetts General Hospital (L.S.), Harvard Medical School, Boston, MA.

Background While many patients are transferred to specialized stroke centers for advanced acute ischemic stroke (AIS) care, few studies have characterized these patients. We sought to determine variation in the rates and differences in the baseline characteristics and clinical outcomes between AIS cases presenting directly to stroke centers' front door versus Transfer-Ins from another hospital. Methods and Results We analyzed 970 390 AIS cases in the Get With The Guidelines-Stroke registry from January 2010 to March 2014 to compare hospitals with high Transfer-In rates (≥15%) versus those with low Transfer-In rates (<5%) and to compare the front-door versus Transfer-In patients admitted to those hospitals with high Transfer-In rates (high Transfer-In hospitals). Of 970 390 patients discharged from 1646 hospitals, 87% initially presented via the emergency department versus 13% were a Transfer-In from another hospital. High Transfer-In hospitals had a median 31% Transfer-In rate among all stroke discharges, were larger, had higher annual AIS volume and intravenous tPA (tissue-type plasminogen activator) rates, and were more often Midwest teaching hospitals and stroke centers. Compared with front-door, Transfer-In patients were younger, more often white, had higher median National Institutes of Health Stroke Scale scores, less often hypertension and previous stroke/transient ischemic attack, and higher in-hospital mortality (7.9% versus 4.9%; standardized difference, 12.4%). After multivariable adjustment, Transfer-In patients had higher in-hospital mortality and discharge modified Rankin scale. Conclusions There is significant regional variability in the transfer of patients with AIS. Because Transfer-In patients seem to have worse short-term outcomes, these patients have the potential to negatively influence institutional mortality rates and should be accounted for explicitly in hospital risk-profiling measures.
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http://dx.doi.org/10.1161/CIRCOUTCOMES.116.003359DOI Listing
September 2018

Evaluation of a score for the prehospital distinction between cerebrovascular disease and stroke mimic patients.

Int J Stroke 2019 06 10;14(4):400-408. Epub 2018 Oct 10.

1 Department of Neurology, Charité - Universitätsmedizin Berlin, Berlin, Germany.

Background: Patients with a sudden onset of focal neurological deficits consistent with stroke, who turn out to have alternative conditions, have been labeled stroke mimics.

Aims: We assessed a recently validated telemedicine-based stroke mimic score (TeleStroke mimic score; TM-score) and individual patient characteristics with regard to its discriminative value between cerebrovascular disease and stroke mimic patients in the in-person, pre-hospital setting.

Methods: We evaluated patients cared for in a mobile stroke unit in Berlin, Germany. We investigated whether the TM-score (comprising six parameters), Face Arm Speech Time test, and individual patient characteristics were able to differentiate cerebrovascular disease from stroke mimic patients.

Results: We included 423 patients (299 (70.7%) cerebrovascular disease and 124 (29.3%) stroke mimic) in the final analysis. A TM-score > 30 indicated a high probability of a cerebrovascular disease and a score ≤15 of a stroke mimic. The TM-score performed well to identify stroke mimics (area under the curve of 0.74 under receiver-operating characteristic curve analysis). The cerebrovascular disease patients were older (74.8 vs. 69.8 years, p = 0.001), had more often severe strokes (NIHSS > 14 25.8% vs. 11.3%, p = 0.001), presented more often with weakness of the face (70.9% vs. 42.7%, p = 0.001) or arm (60.9% vs. 33.9%, p = 0.001), dysarthria (59.5% vs. 40.3%, p < 0.001), history of atrial fibrillation (38.1% vs. 21.0%, p = 0.001), arterial hypertension (78.9% vs. 53.2%, p < 0.001), and less often with seizure (0.7% vs. 21.0%, p < 0.001).

Conclusions: The TM-score and certain patient characteristics can help paramedics and emergency physicians in the field to identify stroke mimic patients and select the most appropriate hospital destination.
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http://dx.doi.org/10.1177/1747493018806194DOI Listing
June 2019

Utilization of Hospice Care in Patients With Acute Ischemic Stroke.

Am J Hosp Palliat Care 2019 Jan 28;36(1):28-32. Epub 2018 Aug 28.

3 Division of Cerebrovascular and Neurocritical Care, Department of Neurology, Ohio State University, Columbus, OH, USA.

Background:: A significant percentage of terminally ill patients are discharged to hospice care following a devastating stroke.

Objective:: We sought to determine the factors associated with hospital discharge to hospice care in a large cohort of patients with stroke.

Methods:: Using the institutional Get With The Guidelines-Stroke database, all consecutive patients with acute ischemic stroke (AIS) who were alive at discharge, from January 2009 until July 2015, were analyzed. Univariate and multivariable statistical analyses were performed to determine the factors associated with discharge to hospice care.

Results:: Of 2446 patients with AIS, 3.4% died and were excluded of remaining 2363 patients, and 4.2% were discharged to hospice care. Univariate analysis identified patients who were discharged to hospice care to be older, caucasian, Medicare or private insurance, have atrial fibrillation, heart failure and less often had diabetes mellitus or smoked. Altered mentation at presentation and urinary tract infection were more common in patients discharged to hospice. On multivariable analysis, patients transferred to hospice care were older (odds ratio [OR]: 1.04, 95% confidence interval [CI]: 1.01-1.07; P < .001), had a high National Institute of Health Stroke Scale (NIHSS; OR: 1.15, 95% CI: 1.10-1.20; P < .001), and altered mental status at presentation (OR: 2.42, 95% CI: 1.29-4.55; P < .001).

Conclusion:: In our study, elderly patients with high NIHSS and altered mental status were identified as factors associated with transition to hospice care following AIS. Prospective studies on the optimal timing of initiation of these consults are needed.
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http://dx.doi.org/10.1177/1049909118796796DOI Listing
January 2019

Characterization of uniaxial high-speed stretch as an in vitro model of mild traumatic brain injury on the blood-brain barrier.

Neurosci Lett 2018 04 16;672:123-129. Epub 2018 Feb 16.

Division of Neurotoxicology, National Center for Toxicological Research, 3900 NCTR Road, Jefferson, AR, 72079, USA. Electronic address:

Traumatic brain injury (TBI) occurs when external mechanical forces induce brain damage as result of impact, penetration or rapid acceleration/deceleration that causes deformation of brain tissue. Depending on its severity, TBI can be classified as mild, moderate or severe and can lead to blood-brain barrier (BBB) dysfunction. In the present study, we evaluated the effects of uniaxial high-speed stretch (HSS) at 0, 5, 10 and 15% on a pure culture of primary rat brain endothelial cells as an in vitro model of TBI to the BBB. LDH release, viability and apoptosis analysis, expression of tight junction proteins and endothelial permeability were evaluated 24 h after a single stretch episode. HSS slightly increased cell death and apoptosis at 10 and 15%, while LDH release was increased only at 15% stretch. Occludin expression was increased at 10% stretch, while claudin-5 expression was increased at 5% stretch, which also decreased the endothelial permeability. In summary, 15% HSS induced low levels of cell death, consistent with mild TBI and very low percentages of HSS (5%) enhanced the BBB properties, promoting the formation of a stronger barrier. These data support the use of 15% HSS as valuable tool in the study of mild TBI to the BBB in vitro.
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http://dx.doi.org/10.1016/j.neulet.2018.02.019DOI Listing
April 2018

Validating the TeleStroke Mimic Score: A Prediction Rule for Identifying Stroke Mimics Evaluated Over Telestroke Networks.

Stroke 2018 03 26;49(3):688-692. Epub 2018 Jan 26.

From the Department of Neurology, Massachusetts General Hospital, Harvard Medical School, Boston (S.F.A., L.H.S.); Department of Neurology and Neurological Intensive Care, Staedtisches Klinikum München, TeleMedical Project for integrative Stroke Care, Munich, Germany (G.J.H.); Department of Neurology, Augusta University, GA (J.S., K.V.); Division of Vascular Neurology, University of Utah, Salt Lake City (J.J.M., L.W.S.); and Department of Neurology, University Regensburg, Germany (R.B.).

Background And Purpose: Up to 30% of acute stroke evaluations are deemed stroke mimics, and these are common in telestroke as well. We recently published a risk prediction score for use during telestroke encounters to differentiate stroke mimics from ischemic cerebrovascular disease derived and validated in the Partners TeleStroke Network. Using data from 3 distinct US and European telestroke networks, we sought to externally validate the TeleStroke Mimic (TM) score in a broader population.

Methods: We evaluated the TM score in 1930 telestroke consults from the University of Utah, Georgia Regents University, and the German TeleMedical Project for Integrative Stroke Care Network. We report the area under the curve in receiver-operating characteristic curve analysis with 95% confidence interval for our previously derived TM score in which lower TM scores correspond with a higher likelihood of being a stroke mimic.

Results: Based on final diagnosis at the end of the telestroke consultation, there were 630 of 1930 (32.6%) stroke mimics in the external validation cohort. All 6 variables included in the score were significantly different between patients with ischemic cerebrovascular disease versus stroke mimics. The TM score performed well (area under curve, 0.72; 95% confidence interval, 0.70-0.73; <0.001), similar to our prior external validation in the Partners National Telestroke Network.

Conclusions: The TM score's ability to predict the presence of a stroke mimic during telestroke consultation in these diverse cohorts was similar to its performance in our original cohort. Predictive decision-support tools like the TM score may help highlight key clinical differences between mimics and patients with stroke during complex, time-critical telestroke evaluations.
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http://dx.doi.org/10.1161/STROKEAHA.117.018758DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5829006PMC
March 2018

Mechanistic studies on ketamine-induced mitochondrial toxicity in zebrafish embryos.

Neurotoxicol Teratol 2018 Sep - Oct;69:63-72. Epub 2017 Dec 7.

Division of Neurotoxicology, National Center for Toxicological Research, U.S. Food and Drug Administration, 3900 NCTR Road, Jefferson, AR 72079, USA. Electronic address:

Ketamine, a phencyclidine derivative, is an antagonist of the Ca-permeable N-methyl-d-aspartate (NMDA)-type glutamate receptors. It is a pediatric anesthetic and has been implicated in developmental neurotoxicity. Ketamine has also been shown to deplete ATP in mammalian cells. Our previous studies showed that acetyl l-carnitine (ALCAR) prevented ketamine-induced cardiotoxicity and neurotoxicity in zebrafish embryos. Based on our finding that ALCAR's protective effect was blunted by oligomycin A, an inhibitor of ATP synthase, we further investigated the effects of ketamine and ALCAR on ATP levels, mitochondria and ATP synthase in zebrafish embryos. The results demonstrated that ketamine reduced ATP levels in the embryos but not in the presence of ALCAR. Ketamine reduced total mitochondrial protein levels and mitochondrial potential, which were prevented with ALCAR co-treatment. To determine the cause of ketamine-induced ATP deficiency, we explored the status of ATP synthase. The results showed that a subunit of ATP synthase, atp5α1, was transcriptionally down-regulated by ketamine, but not in the presence of ALCAR, although ketamine caused a significant upregulation in another ATP synthase subunit, atp5β and total ATP synthase protein levels. Most of the ATP generated by heart mitochondria are utilized for its contraction and relaxation. Ketamine-treated embryos showed abnormal heart structure, which was abolished with ALCAR co-treatment. This study offers evidence for a potential mechanism by which ketamine could cause ATP deficiency mediated by mitochondrial dysfunction.
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http://dx.doi.org/10.1016/j.ntt.2017.12.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5992061PMC
October 2019

Isolation and Culture of Brain Microvascular Endothelial Cells for In Vitro Blood-Brain Barrier Studies.

Methods Mol Biol 2018 ;1727:315-331

Division of Neurotoxicology, HFT-132, National Center for Toxicological Research/USFDA, Jefferson, AR, USA.

The blood-brain barrier (BBB) is essential to maintain the proper microenvironment for brain function. Although formed by different cell types, the endothelial cells (ECs) of the brain microvessels provide the BBB with its selective permeability. To study the BBB in vitro, EC lines as well as primary isolated ECs have been used. In this chapter, we will provide a detailed protocol on how to isolate and culture primary brain microvascular endothelial cells from different species for use as in vitro models of the BBB. When performed properly, this protocol will allow one to obtain a pure culture of brain microvascular endothelial cells with which to analyze the effects of therapeutic and toxic agents on BBB functions.
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http://dx.doi.org/10.1007/978-1-4939-7571-6_23DOI Listing
July 2018

Retraction notice to "RETRACTED: 6-OHDA-induced apoptosis and mitochondrial dysfunction are mediated by early modulation of intracellular signals and interaction of Nrf2 and NF-κB factors" [Toxicology (2013) 109 - 119].

Toxicology 2017 09;390:167

Laboratorio de Aminoácidos Excitadores, Instituto Nacional de Neurología y Neurocirugía - S.S.A., Mexico City, Mexico; Division of Neurotoxicology, National Center for Toxicological Research - FDA, Jefferson, AR, USA.

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http://dx.doi.org/10.1016/j.tox.2017.10.001DOI Listing
September 2017

Characterization of Biaxial Stretch as an In Vitro Model of Traumatic Brain Injury to the Blood-Brain Barrier.

Mol Neurobiol 2018 01;55(1):258-266

Neurochemistry Laboratory, Division of Neurotoxicology, National Center for Toxicological Research/FDA, Jefferson, AR, 72079, USA.

Traumatic brain injury (TBI) is one of the major causes of disability in the USA. It occurs when external mechanical forces induce brain damage that causes deformation of brain tissue. TBI is also associated with alterations of the blood-brain barrier (BBB). Using primary rat brain microvascular endothelial cells as an in vitro BBB model, the effects of biaxial stretch were characterized at 5, 10, 15, 25, and 50% deformation using a commercially available system. The results were compared to the effects of mild and moderate TBI in vivo, induced by the weight-drop method in mice. In vitro, live/dead cells, lactate dehydrogenase (LDH) release, caspase 3/7 staining, and tight junction (TJ) protein expression were evaluated 24 h after a single stretch episode. In vivo, Evans blue extravasation, serum levels of S100β, and TJ protein expression were evaluated. Stretch induced a deformation-dependent increase in LDH release, cell death, and activation of caspase 3/7, suggesting the induction of apoptosis. Interestingly, low magnitudes of deformation increased the expression of TJ proteins, likely in an attempt to compensate for stretch damage. High magnitudes of deformation decreased the expression of TJ proteins, suggesting that the damage was too severe to counteract. In vivo, mild TBI did not affect BBB permeability or the expression of TJ proteins. However, moderate TBI significantly increased BBB permeability and decreased the expression of these proteins, similar to the results obtained with a high magnitude deformation. These data support the use biaxial stretch as valuable tool in the study of TBI in vitro.
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http://dx.doi.org/10.1007/s12035-017-0738-5DOI Listing
January 2018

Monoaminergic toxicity induced by cathinone phthalimide: An in vitro study.

Neurosci Lett 2017 Aug 3;655:76-81. Epub 2017 Jul 3.

Neurochemistry Laboratory, Division of Neurotoxicology, National Center for Toxicological Research/FDA, 3900 NCTR Rd, HFT-132, Jefferson, AR, 72079, United States. Electronic address:

Bath salts, or synthetic cathinones, have cocaine-like or amphetamine-like properties and induce psychoactive effects via their capacity to modulate serotonin (5-HT) and dopamine (DA). Structurally distinct synthetic cathinones are continuously being generated to skirt existing drug laws. One example of these modified compounds is cathinone phthalimide (CP), which has already appeared on the global market. The lack of toxicological studies on the effects of CP on monoaminergic systems led to the development of the present study in order to generate an acute toxicity profile for CP, and to clarify whether it primarily affects both dopamine and serotonin, like the synthetic cathinones mephedrone and methylone, or primarily affects dopamine, like 3, 4-methylenedioxypyrovalerone (MDPV). For the first time, the toxicity profile of CP (10μM-1000μM) is reported. In pheochromocytoma cells, exposure to CP induced cell death, and altered mitochondrial function, as well as intracellular DA and 5-HT levels; at the same time, reduced glutathione (GSH) levels remained unaffected. This seems to indicate that CP functions like mephedrone or methylone. The role of CP metabolites, the effect of CP induced hyperthermia on neurotoxicity, and its ability to traverse the blood-brain barrier warrant further consideration.
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http://dx.doi.org/10.1016/j.neulet.2017.06.059DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6176714PMC
August 2017

Cyclosporine exacerbates ketamine toxicity in zebrafish: Mechanistic studies on drug-drug interaction.

J Appl Toxicol 2017 Dec 1;37(12):1438-1447. Epub 2017 Jun 1.

Division of Neurotoxicology, National Center for Toxicological Research, US Food and Drug Administration, Jefferson, AR, 72079, USA.

Cyclosporine A (CsA) is an immunosuppressive drug commonly used in organ transplant patients to prevent allograft rejections. Ketamine is a pediatric anesthetic that noncompetitively inhibits the calcium-permeable N-methyl-d-aspartic acid receptors. Adverse drug-drug interaction effects between ketamine and CsA have been reported in mammals and humans. However, the mechanism of such drug-drug interaction is unclear. We have previously reported adverse effects of combination drugs, such as verapamil/ketamine and shown the mechanism through intervention by other drugs in zebrafish embryos. Here, we show that ketamine and CsA in combination produce developmental toxicity even leading to lethality in zebrafish larvae when exposure began at 24 h post-fertilization (hpf), whereas CsA did not cause any toxicity on its own. We also demonstrate that acetyl l-carnitine (ALCAR) completely reversed the adverse effects. Both ketamine and CsA are CYP3A4 substrates. Although ketamine and CsA independently altered the expression of the hepatic marker CYP3A65, a zebrafish ortholog of human CYP3A4, both drugs together induced further increase in CYP3A65 expression. In the presence of ALCAR, however, CYP3A65 expression was normalized. ALCAR has been shown to prevent ketamine toxicity in mammal and zebrafish. In conclusion, CsA exacerbated ketamine toxicity and ALCAR reversed the effects. These results, providing evidence for the first time on the reversal of the adverse effects of CsA/ketamine interaction by ALCAR, would prove useful in addressing potential occurrences of such toxicities in humans. Published 2017. This article is a U.S. Government work and is in the public domain in the USA.
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http://dx.doi.org/10.1002/jat.3488DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5859923PMC
December 2017

The effects of physical exercise on nonmotor symptoms and on neuroimmune RAGE network in experimental parkinsonism.

J Appl Physiol (1985) 2017 Jul 6;123(1):161-171. Epub 2017 Apr 6.

Laboratory of Pharmacology and Experimental Therapeutics, Institute for Biomedical Imaging and Life Sciences, Faculty of Medicine, University of Coimbra, Coimbra, Portugal;

Parkinson's disease (PD) prodromal stages comprise neuropsychiatric perturbations that critically compromise a patient's quality of life. These nonmotor symptoms (NMS) are associated with exacerbated innate immunity, a hallmark of overt PD. Physical exercise (PE) has the potential to improve neuropsychiatric deficits and to modulate immune network including receptor for advanced glycation end products (RAGE) and Toll-like receptors (TLRs) in distinct pathological settings. Accordingly, the present study aimed to test the hypothesis that PE ) alleviates PD NMS and ) modulates neuroimmune RAGE network in experimental PD. Adult Wistar rats subjected to long-term mild treadmill were administered intranasally with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and probed for PD NMS before the onset of motor abnormalities. Twelve days after MPTP, neuroimmune RAGE network transcriptomics (real-time quantitative PCR) was analyzed in frontal cortex, hippocampus, and striatum. Untrained MPTP animals displayed habit-learning and motivational deficits without gross motor impairments (cued version of water-maze, splash, and open-field tests, respectively). A suppression of RAGE and neuroimmune-related genes was observed in frontal cortex on chemical and physical stressors (untrained MPTP: RAGE, TLR5 and -7, and p22 NADPH oxidase; saline-trained animals: RAGE, TLR1 and -5 to -11, TNF-α, IL-1β, and p22 NADPH oxidase), suggesting the recruitment of compensatory mechanisms to restrain innate inflammation. Notably, trained MPTP animals displayed normal cognitive/motivational performances. Additionally, these animals showed normal RAGE expression and neuroprotective PD-related gene upregulation in frontal cortex when compared with untrained MPTP animals. These findings corroborate PE efficacy in improving PD NMS and newly identify RAGE network as a neural substrate for exercise intervention. Additional research is warranted to unveil functional consequences of PE-induced modulation of RAGE/DJ-1 transcriptomics in PD premotor stages. This study newly shows that physical exercise (PE) corrects nonmotor symptoms of the intranasal 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) model of experimental parkinsonism. Additionally, we show that suppression of neuroimmune receptor for advanced glycation end products (RAGE) network occurs in frontal cortex on chemical (MPTP) and physical (PE) interventions. Finally, PE normalizes frontal cortical RAGE transcriptomics and upregulates the neuroprotective gene in the intranasal MPTP model of experimental parkinsonism.
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http://dx.doi.org/10.1152/japplphysiol.01120.2016DOI Listing
July 2017

Retraction notice toS-Allyl cysteine protects against 6-hydroxydopamine-induced neurotoxicity in the rat striatum: involvement of Nrf2 transcription factor activation and modulation of signaling kinase cascades [FRB 53/5 (2012) 1024 - 1040].

Free Radic Biol Med 2017 03;104:382

Laboratorio de Aminoácidos Excitadores, Instituto Nacional de Neurología y Neurocirugía - S.S.A., México City, Mexico; Departamento de Biología, Facultad de Química, Universidad Nacional Autónoma de México, México City, Mexico.

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http://dx.doi.org/10.1016/j.freeradbiomed.2016.12.044DOI Listing
March 2017

Baseline Predictors of Poor Outcome in Patients Too Good to Treat With Intravenous Thrombolysis.

Stroke 2016 12 10;47(12):2986-2992. Epub 2016 Nov 10.

From the Department of Neurology, Massachusetts General Hospital, Boston.

Background And Purpose: Several studies have reported poor outcomes in patients too good to treat with intravenous thrombolysis because of mild or rapidly improving symptoms. We sought to determine baseline clinical and imaging predictors of poor outcome in these patients.

Methods: Among 3950 consecutive stroke admissions (2009-2015) in our local Get With the Guidelines-Stroke database, 632 patients presented ≤4.5 hours and did not receive tissue-type plasminogen activator, with 380 of 632 (60.1%) being too good to treat. Univariate and multivariable analyses explored the clinical and imaging features associated with poor outcome (defined as not being discharged to home) in these 380 cases.

Results: Among these 380 cases, only 68% were discharged home; the other 25% to inpatient rehabilitation, 4% to a skilled nursing facility, and 3% expired or were discharged to hospice. Patients with poor outcome were older, were more often Hispanic, had more vascular risk factors, and had higher median National Institutes of Health Stroke Scale. Imaging characteristics associated with poor outcomes included large or multifocal infarction and poor collaterals. In multivariable analysis, only age, initial National Institutes of Health Stroke Scale, and infarct location were independently associated with poor outcome.

Conclusions: Approximately one third of patients deemed too good for intravenous tissue-type plasminogen activator are unable to be discharged directly to home. Given the current safety profile of intravenous tissue-type plasminogen activator, our results suggest that the concept of being too good to treat should be re-examined with an emphasis on the features associated with poor outcome identified in our study. If replicated, these findings could be incorporated into tissue-type plasminogen activator decision-making algorithms.
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http://dx.doi.org/10.1161/STROKEAHA.116.014871DOI Listing
December 2016

Protein Kinases and Parkinson's Disease.

Int J Mol Sci 2016 Sep 20;17(9). Epub 2016 Sep 20.

Department of Geriatrics, University of Arkansas for Medical Sciences, Little Rock, AR 72205, USA.

Currently, the lack of new drug candidates for the treatment of major neurological disorders such as Parkinson's disease has intensified the search for drugs that can be repurposed or repositioned for such treatment. Typically, the search focuses on drugs that have been approved and are used clinically for other indications. Kinase inhibitors represent a family of popular molecules for the treatment and prevention of various cancers, and have emerged as strong candidates for such repurposing because numerous serine/threonine and tyrosine kinases have been implicated in the pathobiology of Parkinson's disease. This review focuses on various kinase-dependent pathways associated with the expression of Parkinson's disease pathology, and evaluates how inhibitors of these pathways might play a major role as effective therapeutic molecules.
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http://dx.doi.org/10.3390/ijms17091585DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5037850PMC
September 2016

Acetazolamide Mitigates Astrocyte Cellular Edema Following Mild Traumatic Brain Injury.

Sci Rep 2016 09 14;6:33330. Epub 2016 Sep 14.

Department of Biomedical Engineering, University of Arkansas, Fayetteville AR 72701, USA.

Non-penetrating or mild traumatic brain injury (mTBI) is commonly experienced in accidents, the battlefield and in full-contact sports. Astrocyte cellular edema is one of the major factors that leads to high morbidity post-mTBI. Various studies have reported an upregulation of aquaporin-4 (AQP4), a water channel protein, following brain injury. AZA is an antiepileptic drug that has been shown to inhibit AQP4 expression and in this study we investigate the drug as a therapeutic to mitigate the extent of mTBI induced cellular edema. We hypothesized that mTBI-mediated astrocyte dysfunction, initiated by increased intracellular volume, could be reduced when treated with AZA. We tested our hypothesis in a three-dimensional in vitro astrocyte model of mTBI. Samples were subject to no stretch (control) or one high-speed stretch (mTBI) injury. AQP4 expression was significantly increased 24 hours after mTBI. mTBI resulted in a significant increase in the cell swelling within 30 min of mTBI, which was significantly reduced in the presence of AZA. Cell death and expression of S100B was significantly reduced when AZA was added shortly before mTBI stretch. Overall, our data point to occurrence of astrocyte swelling immediately following mTBI, and AZA as a promising treatment to mitigate downstream cellular mortality.
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http://dx.doi.org/10.1038/srep33330DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5022024PMC
September 2016

The role of surface chemistry in the cytotoxicity profile of graphene.

J Appl Toxicol 2017 04 4;37(4):462-470. Epub 2016 Sep 4.

Center of Integrative Nanotechnology Sciences, University of Arkansas at Little Rock, AR, 72204, USA.

Graphene and its derivative, because of their unique physical, electrical and chemical properties, are an important class of nanomaterials being proposed as foundational materials in nanomedicine as well as for a variety of industrial applications. A major limitation for graphene, when used in biomedical applications, is its poor solubility due to its rather hydrophobic nature. Therefore, chemical functionalities are commonly introduced to alter both its surface chemistry and biochemical activity. Here, we show that surface chemistry plays a major role in the toxicological profile of the graphene structures. To demonstrate this, we chemically increased the oxidation level of the pristine graphene and compared the corresponding toxicological effects along with those for the graphene oxide. X-ray photoelectron spectroscopy revealed that pristine graphene had the lowest amount of surface oxygen, while graphene oxide had the highest at 2.5% and 31%, respectively. Low and high oxygen functionalized graphene samples were found to have 6.6% and 24% surface oxygen, respectively. Our results showed a dose-dependent trend in the cytotoxicity profile, where pristine graphene was the most cytotoxic, with decreasing toxicity observed with increasing oxygen content. Increased surface oxygen also played a role in nanomaterial dispersion in water or cell culture medium over longer periods. It is likely that higher dispersity might result in graphene entering into cells as individual flakes ~1 nm thick rather than as more cytotoxic aggregates. In conclusion, changes in graphene's surface chemistry resulted in altered solubility and toxicity, suggesting that a generalized toxicity profile would be rather misleading. Copyright © 2016 John Wiley & Sons, Ltd.
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http://dx.doi.org/10.1002/jat.3379DOI Listing
April 2017

Corrigendum to 'The Janus faces of 3-hydroxykynurenine: Dual redox modulatory activity and lack of neurotoxicity in the rat striatum' [Brain Res. 1589 (2014) 1-14].

Brain Res 2016 10 7;1648(Pt A):525. Epub 2016 Jul 7.

Laboratorio de Aminoácidos Excitadores, Instituto Nacional de Neurología y Neurocirugía, SSA, Mexico City 14269, Mexico. Electronic address:

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http://dx.doi.org/10.1016/j.brainres.2016.07.004DOI Listing
October 2016

Methamphetamine, 3,4-methylenedioxymethamphetamine (MDMA) and 3,4-methylenedioxypyrovalerone (MDPV) induce differential cytotoxic effects in bovine brain microvessel endothelial cells.

Neurosci Lett 2016 08 16;629:125-130. Epub 2016 Jun 16.

Neurochemistry Laboratory, Division of Neurotoxicology, National Center for Toxicological Research/FDA, Jefferson, AR, USA. Electronic address:

Designer drugs such as synthetic psychostimulants are indicative of a worldwide problem of drug abuse and addiction. In addition to methamphetamine (METH), these drugs include 3,4-methylenedioxy-methamphetamine (MDMA) and commercial preparations of synthetic cathinones including 3,4-methylenedioxypyrovalerone (MDPV), typically referred to as "bath salts." These psychostimulants exert neurotoxic effects by altering monoamine systems in the brain. Additionally, METH and MDMA adversely affect the integrity of the blood-brain barrier (BBB): there are no current reports on the effects of MDPV on the BBB. The aim of this study was to compare the effects of METH, MDMA and MDPV on bovine brain microvessel endothelial cells (bBMVECs), an accepted in vitro model of the BBB. Confluent bBMVEC monolayers were treated with METH, MDMA and MDPV (0.5mM-2.5mM) for 24h. METH and MDMA increased lactate dehydrogenase release only at the highest concentration (2.5mM), whereas MDPV induced cytotoxicity at all concentrations. MDMA and METH decreased cellular proliferation only at 2.5mM, with similar effects observed after MDPV exposures starting at 1mM. Only MDPV increased reactive oxygen species production at all concentrations tested whereas all 3 drugs increased nitric oxide production. Morphological analysis revealed different patterns of compound-induced cell damage. METH induced vacuole formation at 1mM and disruption of the monolayer at 2.5mM. MDMA induced disruption of the endothelial monolayer from 1mM without vacuolization. On the other hand, MDPV induced monolayer disruption at doses ≥0.5mM without vacuole formation; at 2.5mM, the few remaining cells lacked endothelial morphology. These data suggest that even though these synthetic psychostimulants alter monoaminergic systems, they each induce BBB toxicity by different mechanisms with MDPV being the most toxic.
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http://dx.doi.org/10.1016/j.neulet.2016.06.029DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4983252PMC
August 2016

Regulation of striatal astrocytic receptor for advanced glycation end-products variants in an early stage of experimental Parkinson's disease.

J Neurochem 2016 08 13;138(4):598-609. Epub 2016 Jun 13.

Laboratory of Pharmacology and Experimental Therapeutics/IBILI, Faculty of Medicine, University of Coimbra, Coimbra, Portugal.

Convincing evidence indicates that advanced glycation end-products and danger-associated protein S100B play a role in Parkinson's disease (PD). These agents operate through the receptor for advanced glycation end-products (RAGE), which displays distinct isoforms playing protective/deleterious effects. However, the nature of RAGE variants has been overlooked in PD studies. Hence, we attempted to characterize RAGE regulation in early stages of PD striatal pathology. A neurotoxin-based rodent model of PD was used in this study, through administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) to C57BL/6 mice. Animals were killed 6 h post-MPTP to assess S100B/RAGE contents (RT-qPCR, ELISA) and RAGE isoform density (WB) and cellular distribution (immunohistochemistry). Dopaminergic and gliotic status were also mapped (HPLC-ED, WB, immunohistochemistry). At this preliminary stage of MPTP-induced PD in mice, RAGE inhibitory isoforms were increased whereas full-length RAGE was not affected. This putative cytoprotective RAGE phenotype paired an inflammatory and pro-oxidant setting fueling DAergic denervation. Increased RAGE inhibitory variants occur in astrocytes showing higher S100B density but no overt signs of hypertrophy or NF-κB activation, a canonical effector of RAGE. These findings expand our understanding of the toxic effect of MPTP on striatum and offer first in vivo evidence of RAGE being a responder in early stages of astrogliosis dynamics, supporting a protective rather tissue-destructive phenotype of RAGE in the initial phase of PD degeneration. These data lay the groundwork for future studies on the relevance of astrocytic RAGE in DAergic neuroprotection strategies. We report increased antagonistic RAGE variants paralleling S100B up-regulation in early stages of MPTP-induced astrogliosis dynamics . We propose that selective RAGE regulation reflects a self-protective mechanism to maintain low levels of RAGE ligands , preventing long-term inflammation and oxidative stress arising from sustained ligands/flRAGE activation . Understanding loss of RAGE protective response to stress may provide new therapeutic options to halt or slow down dopaminergic axonopathy and, ultimately, neuronal death .
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http://dx.doi.org/10.1111/jnc.13682DOI Listing
August 2016

Acetyl L-carnitine targets adenosine triphosphate synthase in protecting zebrafish embryos from toxicities induced by verapamil and ketamine: An in vivo assessment.

J Appl Toxicol 2017 02 18;37(2):192-200. Epub 2016 May 18.

Division of Neurotoxicology, National Center for Toxicological, Research, U.S. Food and Drug Administration, 3900 NCTR Road, Jefferson, AR, 72079, USA.

Verapamil is a Ca channel blocker and is highly prescribed as an anti-anginal, antiarrhythmic and antihypertensive drug. Ketamine, an antagonist of the Ca -permeable N-methyl-d-aspartate-type glutamate receptors, is a pediatric anesthetic. Previously we have shown that acetyl l-carnitine (ALCAR) reverses ketamine-induced attenuation of heart rate and neurotoxicity in zebrafish embryos. Here, we used 48 h post-fertilization zebrafish embryos that were exposed to relevant drugs for 2 or 4 h. Heart beat and overall development were monitored in vivo. In 48 h post-fertilization embryos, 2 mm ketamine reduced heart rate in a 2 or 4 h exposure and 0.5 mm ALCAR neutralized this effect. ALCAR could reverse ketamine's effect, possibly through a compensatory mechanism involving extracellular Ca entry through L-type Ca channels that ALCAR is known to activate. Hence, we used verapamil to block the L-type Ca channels. Verapamil was more potent in attenuating heart rate and inducing morphological defects in the embryos compared to ketamine at specific times of exposure. ALCAR reversed cardiotoxicity and developmental toxicity in the embryos exposed to verapamil or verapamil plus ketamine, even in the presence of 3,4,5-trimethoxybenzoic acid 8-(diethylamino)octyl ester, an inhibitor of intracellular Ca release suggesting that ALCAR acts via effectors downstream of Ca . In fact, ALCAR's protective effect was blunted by oligomycin A, an inhibitor of adenosine triphosphate synthase that acts downstream of Ca during adenosine triphosphate generation. We have identified, for the first time, using in vivo studies, a downstream effector of ALCAR that is critical in abrogating ketamine- and verapamil-induced developmental toxicities. Published 2016. This article is a U.S. Government work and is in the public domain in the USA.
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http://dx.doi.org/10.1002/jat.3340DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5115992PMC
February 2017

Fucoidan Extracted from Hijiki Protects Brain Microvessel Endothelial Cells Against Diesel Exhaust Particle Exposure-Induced Disruption.

J Med Food 2016 May;19(5):466-71

1 Department of Preventive Medicine, College of Medicine and Medical Research Institute, Chungbuk National University , Cheongju, Korea.

This study was performed to evaluate the protective effects of fucoidan against the decreased function of primary cultured bovine brain microvessel endothelial cells (BBMECs) after exposure to diesel exhaust particles (DEPs). BBMECs were extracted from bovine brains and cultured until confluent. To evaluate the function of BBMECs, we performed a permeability test using cell-by-cell equipment and by Western blot analysis for zonular occludens-1 (ZO-1), which is a tight junction protein of BMECs, and evaluated oxidative stress in BBMECs using the DCFH-DA assay and the CUPRAC-BCS assay. The increased oxidative stress in BBMECs following DEP exposure was suppressed by fucoidan. In addition, permeability of BBMECs induced by DEP exposure was decreased by fucoidan treatment. Our results showed that fucoidan protects against BBMEC disruption induced by DEP exposure. This study provides evidence that fucoidan might protect the central nervous system (CNS) against DEP exposure.
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http://dx.doi.org/10.1089/jmf.2015.3635DOI Listing
May 2016