Insights into the glioblastoma tumor microenvironment: current and emerging therapeutic approaches.

Glioblastoma (GB) is an intrusive and recurrent primary brain tumor with low survivability. The heterogeneity of the tumor microenvironment plays a crucial role in the stemness and proliferation of GB. The tumor microenvironment induces tumor heterogeneity of cancer cells by facilitating clonal evolution and promoting multidrug resistance, leading to cancer cell progression and metastasis. It also plays an important role in angiogenesis to nourish the hypoxic tumor environment. There is a strong interaction of neoplastic cells with their surrounding microenvironment that comprise several immune and non-immune cellular components. The tumor microenvironment is a complex network of immune components like microglia, macrophages, T cells, B cells, natural killer (NK) cells, dendritic cells and myeloid-derived suppressor cells, and non-immune components such as extracellular matrix, endothelial cells, astrocytes and neurons. The prognosis of GB is thus challenging, making it a difficult target for therapeutic interventions. The current therapeutic approaches target these regulators of tumor micro-environment through both generalized and personalized approaches. The review provides a summary of important milestones in GB research, factors regulating tumor microenvironment and promoting angiogenesis and potential therapeutic agents widely used for the treatment of GB patients.

The Effect of Difference in Word Order on Semantic Processing in Hindi-English Bilinguals.

Background: The typology of word order in Hindi (Subject-Object-Verb, SOV) differs from that of English (Subject-Verb-Object, SVO). Bilinguals whose two languages have conflicting word order provide a unique opportunity to understand how word order affects language processing. Earlier behavioural and event-related brain potential (ERP) studies with Spanish-Basque bilinguals showed longer reading times and more errors in the comprehension of OSV sentences than SOV sentences in Basque language, indicating that non-canonical word orders (OSV) were difficult to process than canonical word order (SOV). Purpose: This study was designed to explore how the difference in word order in Hindi and English languages affects N400 parameters in proficient Hindi-English bilinguals, using semantic congruity paradigm. Methods: Twenty-five proficient Hindi-English bilingual subjects were asked to silently read the congruent and incongruent sentences presented in one word at a time in both the languages. ERPs were recorded from midline frontal, central and parietal sites. Results: The mean amplitude of the N400 effect at the parietal sites in Hindi-English proficient bilinguals was larger for English than for Hindi but there was no significant difference in the N400 latencies. Conclusion: Hindi-English bilingual subjects processed SOV and SVO sentences with equal ease as evidenced by the N400 latencies. Higher amplitude of the N400 effect with English sentences indicate that placing 'Object' as the final word makes sentences more predictable than verb as the final word. Understanding the word order difference might help to unravel the neurophysiological mechanisms of language comprehension and may offer some insights in terms of functional advantage of a particular word order in bilinguals.

Reduced reward responsiveness in treatment resistant depression of middle-aged adults: Association with carotid artery stiffness and tetrahydrobiopterin.

Nearly one third of the population diagnosed with major depressive disorder (MDD) fail to respond to two or more antidepressant drugs of adequate dose and duration. This necessitates identification of confounding psychological and physiological factors that could contribute to treatment resistant depression (TRD). The present longitudinal study investigated the influence of behavioural inhibition system (BIS) and behavioural approach system (BAS) in treatment resistance. Further, the association of depression severity with physiological factors contributing to arterial stiffness was also investigated. Baseline data was acquired from 101 middle-aged (36-56 years) patients on immediate diagnosis with MDD using DSM-V criteria. Follow ups were conducted at 06 months and 12 months during treatment. Psychological assessment battery at baseline and follow ups comprised of Hamilton depression rating (HAM-D) for depression severity, WHODAS-2 and BIS-BAS score. Atherosclerosis and central arterial stiffness were measured by intima-media thickness of internal carotid artery and brachial-ankle pulse wave velocity. Physiological factors influencing central vascular function viz., body-mass index, estimated glomerular filtration rate, HbA1c, central systolic and diastolic blood pressure, heart rate and tetrahydrobiopterin were also investigated. Our results show lower reward responsiveness (BAS-RR) and higher BIS scores in TRD patients along with differentially higher intima-media thickness of left internal carotid artery. Higher depression severity at all stages of the study was correlated with lower tetrahydrobiopterin and BAS-RR scores. We, therefore, suggest that vascular depression resulting due to increased intima-media thickness of left carotid artery and lower tetrahydrobiopterin could be contributing factors for treatment resistance in middle-aged MDD patients.

Class switching of carbonic anhydrase isoforms mediates remyelination in CA3 hippocampal neurons during chronic hypoxia.

Chronic exposure to hypoxia results in cerebral white matter hyperintensities, increased P300 latency, delayed response and impairment in working memory. Despite burgeoning evidence on role of myelination in nerve conduction, the effect of chronic hypoxia on myelination of hippocampal neurons has been less studied. The present study provides novel evidence on alterations in myelination of hippocampal CA3 neurons following chronic hypoxic exposure. Sprague Dawley rats exposed to global hypobaric hypoxia simulating altitude of 25,000 ft showed progressive demyelination in CA3 hippocampal neurons on 14 days followed by remyelination on 21 and 28 days. The demyelination of CA3 neurons was associated with increased apoptosis of both oligodendrocyte precursor cells (OPCs) and mature oligodendrocytes (OLs), peroxidation of myelin lipids, and nitration induced reduced expression of Carbonic Anhydrase II (CAII). Prolonged hypoxic exposure of 21 and 28 days on the other hand resulted in peroxisome proliferator-activated receptor alpha (PPARα) induced upregulation of Carbonic Anhydrase IV (CAIV) expression in mature oligodendrocytes through iNOS mediated mechanisms along with reduction in lipid peroxidation and remyelination. Inhibition of carbonic anhydrase activity on the other hand prevented remyelination of CA3 neurons. Based on these findings we propose a novel iNOS mediated mechanism for regulation of myelination in hypoxic hippocampal neurons through class switching of carbonic anhydrases.

Conscious Abstinence from Smokeless Tobacco Evokes Higher Withdrawal Response and Impairs Cognitive Performance Independent of Sympathetic Response.

INTRODUCTION: High consumption of smokeless tobacco in adult Indian population increases the risk of developing oral cancers leading to high morbidity and mortality. Though the influence of abstinence from smoking on cognitive performance has been widely studied, the effect of smokeless tobacco on cognitive performance and its association with withdrawal symptoms is less understood. This study comparatively investigates the effect of short-term conscious abstinence and distraction during abstinence from smokeless tobacco consumption on the craving, withdrawal symptoms, sympathetic response, and cognitive performance in tobacco addicts. METHODS: Age, sex, education and socioeconomic status matched control (N = 15) and smokeless tobacco addicts (N = 60) were recruited from residential areas in Bhubaneswar for the study. Following randomization of the addicts, conscious abstinence (N = 30) was induced by informed abstinence from tobacco consumption for 8 hours, while distracted cessation (N = 30) was induced by involving the participants in a cognitively engaging task for 8 hours during uninformed tobacco abstinence. RESULTS: The results of the study show higher withdrawal symptoms and reduced cognitive performance in volunteers with conscious abstinence which was positively correlated. The decreased cognitive performance in conscious cessation was independent of tobacco-induced increase in the LF:HF ratio and cotinine concentration in saliva. CONCLUSION: While conscious abstinence results in higher withdrawal symptoms, distraction during abstinence lowers these symptoms. Inclusion of distraction sessions during cessation can, therefore, be a new element in tobacco control strategies.

Correction to: Epigenetic Regulation of SNAP25 Prevents Progressive Glutamate Excitotoxicty in Hypoxic CA3 Neurons.

The original version of this article unfortunately contained mistake.

Trans-Himalayan Phytococktail Confers Protection Against Hypobaric Hypoxia-Induced Hippocampal Neurodegeneration and Memory Impairment in Male Sprague Dawley Rats.

Background: Exposure to hypobaric hypoxia (HH) has been reported to cause neurodegeneration and memory impairment. Hippophae rhamnoides, Prunus armeniaca, and Rhodiola imbricata, the indigenous plants of Indian Trans-Himalaya are widely used in traditional Tibetan and Amchi system of medicine. These are rich sources of diverse bioactive metabolites having prophylactic and therapeutic uses against a wide array of neurodegenerative diseases. The objective of this study was to elucidate the prophylactic and neuroprotective efficacy of formulated phytococktail (PC) against simulated HH-induced neurodegeneration in male Sprague Dawley (SD) rats. Materials and Methods: A PC containing H. rhamnoides fruit pulp, P. armeniaca fruit pulp, and R. imbricata dry root extract (100:50:1) was formulated. The neuroprotective efficacy of PC was evaluated in male SD rats following exposure to 7 day HH at simulated altitude (25,000 ft, 282 mm Hg). Rats were divided into four groups viz., normoxia group (NOR), normoxic group treated with PC (NORPC), 7 day hypoxic group treated with vehicle (7DH), and 7 day hypoxic group treated with PC (7DHPC). Memory impairment and neuromorphological alterations were measured. Targeted protein expression was analyzed by immunoblotting study. Results: PC supplementation significantly reduced the oxidative stress markers during exposure to HH. Spatial memory impairment by HH was significantly ameliorated by PC. HH-induced augmented pyknosis, decreased dendritic arborization, and increased Hoechst-positive neurons in hippocampal CA3 region were significantly ameliorated by PC. Immunoblotting study showed upregulation of BDNF and TrkB expression by PC. PC also prevented the hippocampal neurodegeneration by activating the PI3K/AKT signaling pathway, which leads to GSK-3ß inactivation by its phosphorylation and alleviation of hippocampal Caspase3 expression leading to inhibition of apoptotic neuronal cell death. Conclusion: The present study advocates the potential role of PC as an effective neuroprotective supplement in preventing HH-induced neurodegeneration. Activation of the PI3K/Akt pathway through BDNF/TrkB interaction following PC supplementation after exposure to HH inhibits hippocampal neuronal apoptosis and memory impairment.

Methanolic root extract of Codonopsis clematidea prevents hypoxia induced procoagulant state by inhibition of GPIb receptor regulated Lyn kinase activation.

BACKGROUND: The prevalence of procoagulant state under prolonged hypoxic exposures and the complications and lack of specificity associated with use of existing anti-thrombotic agents have necessitated the search for safer and natural therapeutics. Codonopsis, a widely studied medicinal herb, has been reported to decrease whole blood viscosity but the bioactive ingredients involved, and their mechanism of action therein however remain to be investigated. PURPOSE: The present study aimed at evaluating the efficacy of C. clematidea root extract and mechanism of action of its bioactive constituent flavonoid, Kaempferol, in ameliorating hypobaric hypoxia induced procoagulant state. METHODS: Fingerprinting analysis of methanolic extract of C. clematidea root was performed by RP-HPLC. In vitro toxicity study was conducted using HUVEC cell line and in vivo acute and sub-acute toxicity were done according to OECD guidelines (section-4, number-420 and 407 respectively). Adult male Sprague-Dawley rats weighing 230-250 g were exposed to global hypoxia simulating an altitude of 7600 m (282 mmHg), in animal decompression chamber for 3, 7, 14 and 21 days for in vivo studies. Dose optimisation of the extract was done by quantification of Thromboxane A2 in the serum of hypoxic rats. C. clematidea root extract was also evaluated for its in vitro and in vivo antioxidant properties. Procoagulant changes were studied by biochemical plasma coagulation assays and expression analysis of the signalling molecules of the platelet activation cascade like vWF, platelet activation marker CD41, GpIb-IX-V (CD42), Lyn kinase, p-PI3K, p-ERK and p-PLCγ were conducted to investigate C. clematidea mediated signalling mechanisms. RESULTS: Methanolic extract of C. clematidea root showed improved antioxidant status and improvement in bleeding time and in vitro coagulation assays like pT, aPTT, INR. Decreased concentrations of D-Dimers along with that of platelet activation marker CD41 and serum concentration of Thromboxane A2 were observed in C. clematidea root extract supplemented hypoxic animals. Phosphorylation of Lyn kinase, was reduced despite increase in concentration of activating ligand vWF. CONCLUSION: C. clematidea root extract was effective in preventing hypoxia induced platelet activation and resultant procoagulant state by inhibiting Lyn kinase, a serine threonine kinase effector of vWF signalling cascade.

Hypoxia-mediated alteration in cholesterol oxidation and raft dynamics regulates BDNF signalling and neurodegeneration in hippocampus.

Brain-derived neurotrophic factor (BDNF) which is primarily associated with neuronal survivability, differentiation and synaptic plasticity has been reported to mediate neurodegeneration in hypoxia through its p75 Neurotrophin receptors (p75NTR). The molecular events promoting BDNF-mediated pro-death signalling in hypoxia, however, still remain an enigma. This study attempts towards deciphering the signalling cascades involved in alteration of BDNF isoforms and its cognate receptor subtypes leading to neurodegeneration in hypoxia. Adult Sprague-Dawley rats were exposed to global hypobaric hypoxia simulating an altitude of 7620 m at standard temperature and humidity. Chronic hypoxic exposure for 7 days resulted in higher expression of pro-BDNF and alteration in N-linked glycosylation in hippocampus along with increased expression of endoplasmic reticulum stress markers viz., glucose-regulated protein (Grp78), calnexin and changes in the endoplasmic reticulum morphology. Our findings reveal enriched expression of p75NTR in lipid rafts and higher expression of tyrosine receptor kinase ß (Trkß) in non-raft regions following hypoxic exposure. Further investigations on membrane properties revealed decline in membrane fluidity along with increased cholesterol oxidation resulting in reduced translocation of Trkß from non-raft to raft regions. Supplementation of vitamin E during hypoxic exposure on the other hand reduced cholesterol oxidation and increased translocation of Trkß from non-raft to raft regions and promoted neuronal survival. Hence, our findings suggest a novel mechanism of cholesterol oxidation-induced alteration in raft dynamics which is promotes p75 receptor-mediated death signalling in hippocampal neurons during chronic hypoxia.

Kaempferol Inhibits Extra-synaptic NMDAR-Mediated Downregulation of TRkß in Rat Hippocampus During Hypoxia.

Cognitive dysfunction on chronic exposure to hypobaric hypoxia has been attributed to a myriad of survival and degenerative factors. Downregulation of Trkß and compromised survival signaling has been ascribed as a major contributing factor for hypoxic neurodegeneration. The mechanisms leading to downregulation of Trkß in hypoxia, however, remain to be elucidated. The present study aimed at investigating the upstream signaling mechanisms leading to Trkß downregulation in hypoxia and the potential of Kaempferol in ameliorating these changes. Our results showed a duration-dependent increase in hypoxic neurodegeneration as measured by Fluoro-Jade C staining of hippocampal CA3 neurons. Protein expression studies revealed strong correlation of Trkß with NR1 and NR2b expression on exposure to hypoxic stress. Administration of Kaempferol during hypoxic stress revealed its neuroprotective effect and Morris Water Maze test also highlighted its efficacy in improving spatial learning and memory. Further elucidation of the signaling mechanisms using specific inhibitors and in vitro silencing experiments confirmed involvement of extra-synaptic N-methyl-d-aspartate receptor (NMDAR) i.e. NR2b receptor subunit in downregulation of Trkß under hypoxic conditions. ChIP assay showed involvement of E47 transcription factor in NR2b mediated Trkß downregulation. Selective inhibition of signaling intermediate MLK2 by CEP11004 and inhibition of extra-synaptic NMDAR during hypoxic stress prevented Trkß downregulation in the hippocampus of hypoxic rats. Administration of Kaempferol also inhibited phosphorylation of E47 and hypoxia-induced downregulation of Trkß. The present study establishes the role of extra-synaptic NMDAR in hypoxia-induced downregulation of Trkß and the efficacy of Kaempferol in inhibiting extra-synaptic NMDAR-mediated signaling.

High altitude with monotonous environment has significant impact on mood and cognitive performance of acclimatized lowlanders: Possible role of altered serum BDNF and plasma homocysteine level.

BACKGROUND: Monotony resulting due to the wilderness, sparse population and isolation from society could adversely affect human physiology and cause mood alterations. Thus, observations need to be conducted in order to elucidate the possible role of circulating biomarkers in inducing altered mood and cognitive performance following prolonged exposure to high altitude (HA) with persistent monotonous environment. OBJECTIVE: The present study aimed towards investigating the impact of monotonous environment in remote HA on mood and cognitive performance of human volunteers and its correlation with serum brain derived neurotrophic factor (BDNF) and plasma homocysteine level. METHODS: The present study was conducted on male lowlander participants who had normal social life prior to induction in HA environment. Baseline data was acquired at altitude ≤240 m mean sea level (MSL). Thereafter, the participants were inducted to an altitude of 4500-4800 m MSL. After acclimatization to HA, the participants were assigned as acclimatized low landers (ALL). Longitudinal follow up was conducted after 8 months of high altitude induction on acclimatized low landers (8ALL). Further, to study the effect of monotony, the participants were randomly assigned into different group sizes during their further stay of 4 months in HA viz. ≤5 (12ALL ≤ 5) and ≥10 (12ALL ≥ 10). Mood and cognitive performance of the participants were assessed by standard self-administered questionnaires. Serum BDNF and plasma homocysteine were estimated and their correlation with mood and cognition were determined. RESULTS: The findings showed significantly low serum BDNF in 12ALL ≤ 5 group when compared to baseline, 8ALL and 12AL ≥ 10 groups. Alleviated serum BDNF was associated with increased prevalence of mood alterations in HA with persistent monotonous environment. Participants of 12ALL ≥ 10 group showed significantly higher cognitive performance as compared to 12ALL ≤ 5 group which was associated with reduced plasma homocysteine level. LIMITATIONS: Total registered volunteers during baseline study were not available during the entire period of this study. The second limitation was exclusion of participants with medical history of severe head injuries, chronic diseases in family and extreme baseline serum profile. Third limitation of the study was to exclude the participants detected with MCI after 8 months of HA induction for negating the role of hypobaric hypoxia on mood and cognition. CONCLUSION: The study advocated that ALLs of 12ALL ≤ 5 group have increased prevalence of depressive trait and cognitive impairment which was correlated with reduced serum BDNF and augmented plasma homocysteine level as compared to participants of 12ALL ≥ 10 group having better social interaction with improved cognition and mood. The basic findings of the present study revealed that prolonged HA stay after physiological acclimatization should be regulated by proper social interaction involving normal group size to avoid detrimental effect of monotony and its significant impact on circulatory biomarkers.

Salidroside mediated stabilization of Bcl -xL prevents mitophagy in CA3 hippocampal neurons during hypoxia.

Chronic hypoxic stress results in deposition of lipofuscin granules in the CA3 region of hippocampal neurons which contributes to neurodegeneration and accelerated neuronal aging. Oxidative stress and mitophagy during hypoxia are crucial to cause aggregation of these lipofuscin granules in hypoxic neurons. Salidroside, a glucoside derivative of ß-Tyrosol, has been reported to protect hypoxic neurons through maintenance of mitochondrial activity. The present study is aimed at investigating the potential of Salidroside in preventing mitophagy during chronic hypoxia and identification of the molecular targets and underlying signaling mechanisms. In-silico analysis for interaction of salidroside with Bcl-xL was carried out using VLife MDS software. The prophylactic efficacy of Salidroside for amelioration of global hypoxia induced neuronal aging was studied in adult male Sprague-Dawley rats exposed to hypobaric hypoxia simulating an altitude of 7600 m for 21 days. Salidroside was supplemented at a daily dose of 25 mg kg-1b.w. p.o. during hypoxic exposure. Ultra-structural and immune-histological studies were conducted to study lipofuscin aggregation and mitophagy. In-silico findings on salidroside mediated stabilization of Bcl-xL were validated by investigating its effect on downstream signaling molecules involved in mitophagy. Administration of Salidroside reduced deposition of lipofuscin in hypoxic CA3 hippocampal neurons and prevented mitophagy. Salidroside stabilizes Bcl-xL in hypoxic neurons resulting in inhibition of PGAM5 phosphatase activity and maintenance of FUNDC1 in phosphorylated state. Salidroside mediated inhibition of pFUNDC1 dephosphorylation prevents FUNDC1-LC3 II interaction which is crucial for mitophagy. The present study demonstrates potential of Salidroside in preventing lipofuscin deposition during chronic hypoxic stress.

Differential responses of autonomic function in sea level residents, acclimatized lowlanders at >3500 m and Himalayan high altitude natives at >3500 m: A cross-sectional study.

We studied the differential responses of autonomic function in sea level residents (SLR), acclimatized lowlanders (ALH) in high altitude (HA) and HA natives (HAN) at >3500 m. Out of 771 male volunteers included in this cross-sectional study, SLR, ALH and HAN groups were comprised of 351, 307 and 113 volunteers, respectively. Our results showed persistent sympathetic dominance with significantly reduced (p < 0.05) parasympathetic response in ALH as compared to SLR and HAN populations. This may be attributed to significantly increased (p < 0.05) concentration of coronary risk factors and plasma catecholamines in ALH as compared to SLR and HAN. The ALH also showed significantly increased (p < 0.05) level of serum homocysteine as compared to SLR. The HAN exhibited no changes in autonomic function despite significantly elevated (p < 0.05) homocysteine level as compared to SLR. Our findings may have clinical relevance for assessment of susceptibility to cardiovascular risks in HA dwellers, native highlanders and patients with hypoxemia.

Prevalence and risk factors of hypertension in acclimatized lowlanders staying at high altitude for different durations.

Hypoxic exposure at high-altitude (HA) modulates blood pressure (BP). High prevalence of hypertension among native highlanders (NH) has been reported. However, information on prevalence and determinants of hypertension in acclimatized young lowlanders (ALL) staying at HA for different durations is sparse. We aimed to determine the prevalence of hypertension in ALL staying at HA for different durations and its association with cardiovascular risk factors. Male volunteers were categorized on the basis of their duration of stay at HA; Lowlanders (LL) (0 months; n = 151), ALL (1-24 months; n = 519) and NH (n = 103). ALL were sub grouped into ALL 1 (1-6 months; n = 165), ALL 2 (6-12 months; n = 181), and ALL 3 (12-24 months; n = 173). BP, sympathetic activity, arterial stiffness, lipid profile, and homocysteine were estimated. Regression analysis was performed to determine association of risk factors with hypertension. Prevalence of hypertension among ALL was highest with 17.53% followed by NH (11.6%) and LL (9.27%). Prevalence of hypertension in ALL sub group was in order ALL 1 < ALL 2 < ALL 3. Hypertension was significantly associated with sympathetic dominance (p < 0.001) in ALL 1. Hypertension in ALL 2 was associated with dyslipidemia (p < 0.01) while in ALL 3 hypertension was associated with hyperhomocysteinemia (hHCY, p < 0.001), arterial stiffness and dyslipidemia (p < 0.01). In conclusion, our report suggests higher prevalence of hypertension in ALL. The association of studied risk factors and hypertension in different ALL sub groups varied significantly. Our findings suggest the need for a differential clinical approach to control hypertension in ALL considering their duration of stay at HA.

Estrogen Receptor ß Mediated Neuroprotective Efficacy of Cicer microphyllum Seed Extract in Global Hypoxia.

Hypoxia induced oxidative stress and neurodegeneration in the hippocampus has been implicated for memory impairment in conditions like stroke, ischemia and hypobaric hypoxia. The present study, aimed at investigating the potential of ethanolic extract of Cicer microphyllum seeds (CSE) for amelioration of global hypoxia induced neurodegeneration in CA1 region of hippocampus. CSE supplementation considerably reduced neurodegeneration and dendritic atrophy in CA1 neurons along with improvement of memory in hypoxic rats. This effect of CSE was partly attributed to its antioxidant activity resulting in reduction of lipid peroxidation, protein oxidation and DNA damage during exposure to chronic hypoxia. CSE also promoted dendritic arborization through activation of estrogen receptor beta (ERß) and phosphorylation of extracellular signal regulated kinase (ERK1/2) which was independent of brain derived neurotrophic factor (BDNF) mediated signalling mechanisms. Extra nuclear activation of ERK1/2 by ERß resulted in phosphorylation of cyclic AMP response element binding protein (CREB) leading to increased expression of PSD-95.These molecular alterations translated to behavioural changes in CSE administered hypoxic animals that performed better in Morris Water Maze Task as compared to vehicle treated hypoxic animals. Toxicological studies show NOEAL > 2000 mg/kg b.w. for oral administration of CSE indicating its safety for consumption. Our findings not only suggest the neuroprotective potential of CSE in hypoxia but also provide evidence for involvement of estrogen receptor and pCREB mediated nootropic effect of the extract.

Acute and sub-acute toxicological evaluation of lyophilized Nymphaea x rubra Roxb. ex Andrews rhizome extract.

Nymphaea x rubra Roxb. ex Andrews (N. rubra) has been widely reported for immunomodulatory properties and treatment of piles, bleeding nose and dysentery in traditional medicinal systems. However, its in-vitro and in-vivo toxicity studies have never been investigated. So, the present study was designed to investigate in-vitro and in-vivo toxicity of methanolic extract of N. rubra rhizome in rats. In-vitro cytotoxicity studies were conducted for different doses of extract in N2a cell lines. For in-vivo toxicity studies, SD rats were divided into three groups and administered with 0, 300 and 2000 mg/kg b. w., p. o., of N. rubra extract respectively. In acute toxicity studies, female animals after extract administration animals were sacrificed for hematological profiling and gross necropsy. In sub-acute toxicity studies, both male and female animals were administered with extract daily for 14 days and were sacrificed for hematological, biochemical and histological examination. Body weight and food water intake was measured daily and animals were observed for visual toxic effects, behavioral changes and mortality. During in-vivo toxicity studies, none of the animals showed signs of toxicity and mortality during toxicity studies. The present findings suggest its safety and NOAEL of N. rubra rhizome extract to be > 2000 mg/kg b. w.

Epigenetic Regulation of SNAP25 Prevents Progressive Glutamate Excitotoxicty in Hypoxic CA3 Neurons.

Exposure to global hypoxia and ischemia has been reported to cause neurodegeneration in the hippocampus with CA3 neurons. This neuronal damage is progressive during the initial phase of exposure but maintains a plateau on prolonged exposure. The present study on Sprague Dawley rats aimed at understanding the underlying molecular and epigenetic mechanisms that lead to hypoxic adaptation of CA3 neurons on prolonged exposure to a global hypoxia. Our results show stagnancy in neurodegeneration in CA3 region beyond 14 days of chronic exposure to hypobaria simulating an altitude of 25,000 ft. Despite increased synaptosomal glutamate and higher expression of NR1 subunit of NMDA receptors, we observed decrease in post-synaptic density and accumulation of synaptic vesicles at the pre-synaptic terminals. Molecular investigations involving western blot and real-time PCR showed duration-dependent decrease in the expression of SNAP-25 resulting in reduced vesicular docking and synaptic remodeling. ChIP assays for epigenetic factors showed decreased expression of H3K9Ac and H3K14Ac resulting in SNAP-25 promoter silencing during prolonged hypoxia. Administration of sodium butyrate, a non-specific HDAC inhibitor, during 21 days hypoxic exposure prevented SNAP-25 downregulation but increased CA3 neurodegeneration. This epigenetic regulation of SNAP-25 promoter was independent of increased DNMT3b expression and promoter methylation. Our findings provide a novel insight into epigenetic factors-mediated synaptic remodeling to prevent excitotoxic neurodegeneration on prolonged exposure to global hypobaric hypoxia.

Effect of seabuckthorn seed oil in reducing cardiovascular risk factors: A longitudinal controlled trial on hypertensive subjects.

BACKGROUND & AIMS: The present study aimed at investigating whether dietary supplementation of seabuckthorn seed oil which is rich in omega fatty acids at an oral dose of 0.75 ml could affect cardiovascular risk factors and reduce hypertension and systolic blood pressure. METHODS: Toxicological evaluation and efficacy of seabuckthorn seed oil in reducing high fat diet induced dyslipidemia was initially conducted on adult male Sprague Dawley rats. 32 normal and 74 hypertensive and hypercholestrolemic human subjects participated in the randomized, controlled, double blind longitudinal study. Seabuckthorn seed oil or sunflower oil placebo was orally supplemented at a daily dose of 0.75 ml for 30 days. RESULTS: Supplementation of seabuckthorn seed oil at a daily dose of 0.75 ml for 30 days resulted in normalization of blood pressure in hypertensive subjects. Dietary supplementation of seabuckthorn seed oil markedly reduces cholesterol, oxy-LDL and triglycerides in hypercholesterolemic subjects though it's effect on subjects with normal blood pressure and cholesterol is less pronounced. Seabuckthorn seed oil supplementation also improves circulatory antioxidant status in both normal and hypertensive subjects. CONCLUSION: The present study demonstrates the efficacy of seabuckthorn seed oil in reducing dyslipidemia, cardiovascular risk factors and hypertension in human population which may be due to presence of omega 3, 6 and 9 fatty acids in the oil. The improvement in antioxidant status can be attributed to presence of beta carotene and vitamin E in seabuckthorn seed oil. The trial was registered with Clinical Trial Registry of India (Clinical trial registration number - CTRI/2015/11/006368).

Global hypoxia induced impairment in learning and spatial memory is associated with precocious hippocampal aging.

Both chronological aging and chronic hypoxia stress have been reported to cause degeneration of hippocampal CA3 neurons and spatial memory impairment through independent pathways. However, the possible occurrence of precocious biological aging on exposure to single episode of global hypoxia resulting in impairment of learning and memory remains to be established. The present study thus aimed at bridging this gap in existing literature on hypoxia induced biological aging. Male Sprague Dawley rats were exposed to simulated hypobaric hypoxia (25,000ft) for different durations and were compared with aged rats. Behavioral studies in Morris Water Maze showed decline in learning abilities of both chronologically aged as well as hypoxic rats as evident from increased latency and pathlength to reach target platform. These behavioral changes in rats exposed to global hypoxia were associated with deposition of lipofuscin and ultrastructural changes in the mitochondria of hippocampal neurons that serve as hallmarks of aging. A single episode of chronic hypobaric hypoxia exposure also resulted in the up-regulation of pro-aging protein, S100A9 and down regulation of Tau, SNAP25, APOE and Sod2 in the hippocampus similar to that in aged rats indicating hypoxia induced accelerated aging. The present study therefore provides evidence for role of biological aging of hippocampal neurons in hypoxia induced impairment of learning and memory.

Early mood behavioral changes following exposure to monotonous environment during isolation stress is associated with altered hippocampal synaptic plasticity in male rats.

Social isolation stress and its effect on mood have been well reported, but the effect of monotony (a state of repetition of events for a considerable period of time without variation) on mood and hippocampal synaptic plasticity needs to be addressed. Present study was conducted on male Sprague-Dawley rats. Singly housed (SH) rats were subjected to monotony stress by physical, visual and pheromonal separation in specially designed animal segregation chamber. Fluoxetine (a selective serotonin reuptake inhibitor) was administered orally. Behavioral assessment showed anxiety and depression like traits in SH group. Monotony stress exposure to SH group resulted in increased pyknosis, decreased apical dendritic arborization and increased asymmetric (excitatory) synapses with the corresponding decrease in the symmetric (inhibitory) synapses in the hippocampal CA3 region. Monotonous environment during isolation stress also decreased the serotonin level and reduced the expression of synaptophysin and pCREB in the hippocampus. Fluoxetine administration to singly housed rats resulted in amelioration of altered mood along with improvement in serotonin and decrease in excitatory synaptic density but no change in altered inhibitory synaptic density in the hippocampus. These findings suggest that monotony during isolation contributes to early impairment in mood state by altering hippocampal synaptic density and neuronal morphology.