Oleander attenuates hepatic inflammation in a TLR4-independent manner and by favorable modulation of hepatocellular global metabolome that supports cytoprotection.

ETHNOPHARMACOLOGICAL RELEVANCE: Nerium oleander is used to treat liver-associated chronic metabolic diseases in traditional medicinal systems across the globe. The hepatoprotective effects of oleander are mentioned in Indian and Chinese traditional medicinal literature. AIM OF THE STUDY: The present study aimed to investigate the cellular mechanisms behind the hepatoprotective effects of a non-toxic dose of oleander (NO). MATERIALS AND METHODS: The hepatoprotective effects of NO were tested against lipopolysaccharide (LPS)-treated HepG2 cells. Oxidative stress response was studied using cellular enzymatic assays, and gene expression was analyzed using qRT-PCR. HepG2 cells were pretreated with TAK-242 (pharmacological inhibitor of TLR4) to decipher the anti-inflammatory mechanisms of NO. Cell-free metabolites were analyzed using GCMS and were subjected to pathway enrichment analysis. RESULTS: NO reduced systemic inflammation, serum lipid peroxidation byproducts, and glucose without affecting serum transaminase levels and hepatic histopathological features. NO attenuated the inflammation-induced loss of antioxidant enzyme activities and mRNA expressions of toll-like receptor-4 (TLR4)/nuclear factor κß (NFκß)-dependent inflammatory genes. In TAK-242 pretreated cells, LPS was unable to induce inflammatory and oxidative responses. However, NO treatment in TAK-242 pretreated cells with LPS stimulation further reduced the signs of inflammation and improved hepatoprotective activities. A comparative analysis of the intracellular global metabolome from HepG2 cells with and without NO treatment indicated NO-mediated favorable modulation of intracellular metabolic pathways that support cytoprotective activities. CONCLUSION: NO protects HepG2 cells from LPS-induced oxidative and inflammatory injury. The hepatoprotective effects of NO are mediated by a TLR4-independent process and through a favorable modulation of the intracellular global metabolome that supports cytoprotection.

The antioxidant rich active principles of Clerodendrum sp. controls haloalkane xenobiotic induced hepatic damage in murine model.

Clerodendrum is a plant with potent antioxidant activity and has been frequently employed as a traditional remedy against bronchitis, asthma, liver and stomach disorders. Three species of genus Clerodendrum namely Clerodendrum indicum, C. colebrookianum and C. inerme (Syn. Volkameria inermis) were investigated for their possible activity against oxidative stress induced liver injury. Apart from generation of Reactive Oxygen Species (ROS) in the WRL-68 cell line (human hepatic cell line), in-vitro and in-vivo antioxidant assays were also assessed. Features of immune cell proliferation (MTT) were analyzed thoroughly. Gas Chromatography-Mass Spectrometry (GC-MS) and Fourier Transform Infrared Spectroscopy (FTIR) analyses have been performed to identify the active biological compounds. These active biological compounds were further subjected to molecular docking. The antioxidant activity of three Clerodendrum sp. was significantly high in DPPH, nitric oxide, hydroxyl radical and hydrogen peroxide etc. Biochemical parameters like catalase, superoxide dismutase (SOD) and reduced glutathione (GSH) were generated in excess due to CCl4 administration, which was ameliorated by treating with Clerodendrum extract. The phytochemical 24,25-Dihydroxyvitamin D shows excellent binding affinity in Autodock Vina. The present study provided convincing evidences that C. indicum and C. inerme showed good result but C. colebrookianum performed better by almost all means.

Local origin or external input: modern horse origin in East Asia.

BACKGROUND: Despite decades of research, the horse domestication scenario in East Asia remains poorly understood. RESULTS: The study identified 16 haplogroups with fine-scale phylogenetic resolution using mitochondrial genomes of 317 horse samples. The time to the most recent common ancestor of the 16 haplogroups ranges from [0.8-3.1] thousand years ago (KYA) to [7.9-27.1] KYA. With combined analyses of the mitochondrial control region for 35 extant Przewalski's horses, 3544 modern and 203 ancient horses across the world, researchers provide evidence for that East Asian prevalent haplogroups Q and R were indigenously domesticated or they were involved in numerous distinct genetic components from wild horses in the southern part of East Asia. These events of haplotypes Q and R occurred during 4.7 to 16.3 KYA and 2.1 to 11.5 KYA, respectively. The diffusion of preponderant European haplogroups L from west to East Asia is consistent with the external gene input. Furthermore, genetic differences were detected between northern East Asia and southern East Asia cohorts by Principal Component Analysis, Analysis of Molecular Variance test, the χ2 test and phylogeographic analyses. CONCLUSIONS: All results suggest a complex picture of horse domestication, as well as geographic pattern in East Asia. Both local origin and external input occurred in East Asia horse populations. And besides, there are at least two different domestication or hybridization centers in East Asia.

Correlative metabolomic fingerprinting and molecular docking studies of dermatological phytotherapeutics of South-Eastern Himalaya.

Viburnum erubescens Wall., Rhododendron arboretum Sm., Eurya japonica Thumb., Symplocos lucida (Thunb.) Siebold & Zucc, and Symplocos pyrifolia Wall. ex G. Don are extensively used by the native and ethnic populations of the South-Eastern Himalayan region for several dermatological conditions, yet their phytochemical composition remained largely unknown. Therefore, the aim of the study was to explore the therapeutically relevant volatile phytochemical compositions and study the molecular interactions against intracellular cytoregulatory transcription factors. Leaves of the five plants were subjected to Gas chromatography-Mass spectrometry (GCMS) post silylation derivation. The results were further analyzed using multivariate statistical methods such as Principal Component Analysis (PCA) and Hierarchical Cluster Analysis (HCA). A total of 115 compounds were identified in the five plants. Multivariate analysis revealed optimum metabolomic correlation between S. pyrifolia and S. lucida (0.876), whereas lowest correlation was found between E. japonica and V. erubescens (-0.242). Arbutin, ß-amyrin, betulin, ß-sitosterol and stigmasterol demonstrated highest interaction with the molecular targets. Collectively, the present study revealed the bioactive volatile phytochemicals responsible the therapeutic uses against diverse skin conditions.

Oleander Stem and Root Standardized Extracts Mitigate Acute Hyperglycaemia by Limiting Systemic Oxidative Stress Response in Diabetic Mice.

The extracts of different parts of Nerium oleander L. are used as antidiabetic remedy in the traditional medicinal systems of different parts of the world. Despite these uses in ethnomedicinal system, the antihyperglycemic potentials of oleander stem (NOSE) and root (NORE) extracts have not been pharmacologically evaluated. Therefore, we aimed at evaluating the antidiabetic ethnomedicinal claims of NOSE and NORE, primarily focusing on glucose homeostasis and associated metabolic implications. Alloxan-treated mice with hyperglycaemia (blood glucose >200 mg/dL) were treated with oleander 70% hydromethanolic extracts (200 mg/kg) for 20 consecutive days, and the results were compared with positive control glibenclamide. Blood glucose level was 52-65% lowered (P < 0.001) in oleander treated groups, which was otherwise 4.62 times higher in diabetic mice, compared to control. Insulin resistance was lowered 51-36% irrespective of any significant (P > 0.05) changes in insulin sensitivity throughout the treatments. Improved serum insulin remained associated with lowered glucose level (r P = -0.847 and -0.772; P < 0.01). Markers of hyperglycaemia-related hepatic glycogen, glycated haemoglobin (HbA1c), hyperlipidaemia, hepatic injury, and diabetic nephropathy were normalized as well. Improvement of systemic intrinsic antioxidant enzymes (catalase and peroxidase) were correlated (r P = -0.952 to -0.773; P < 0.01) with lower lipid peroxidation by-product malondialdehyde (MDA) in the circulation. Principal component analysis coupled with hierarchical cluster analysis represented shift in metabolic homeostasis in diabetic mice, which was further normalized by oleander and glibenclamide treatment. Additionally, molecular docking studies of the phenolic acids measured by HPLC with intracellular cytoprotective transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) revealed strong molecular interactions. The results collectively support the ethnomedicine antidiabetic claims of oleander stem and root and suggest that the oleander mediated elevation of systemic antioxidant status is likely responsible for the improved glycaemic control.

Amelioration of CCl4 induced liver injury in swiss albino mice by antioxidant rich leaf extract of Croton bonplandianus Baill.

The progress in industrialization has blessed mankind with a technologically superior lifestyle but poor management of industrial waste has in turn poisoned nature. One such chemical is carbon tetra chloride (CCl4), which is a potent environmental toxin emitted from chemical industries and its presence in the atmosphere is increasing at an alarming rate. Presence of CCl4 in human body is reported to cause liver damage through free radical mediated inflammatory processes. Kupffer cells present in the liver are potentially more sensitive to oxidative stress than hepatocytes. Kuffer cells produced tumor necrosis factor-α (TNF-α) in response to reactive oxygen species (ROS), that might further cause inflammation or apoptosis. In this study hepatoprotective capacity of antioxidant rich extract of Croton bonplandianus Baill. (CBL) was evaluated on CCl4 induced acute hepatotoxicity in murine model. Hydro-methanolic extract of C. bonplandianus leaf was used for evaluation of free radical scavenging activity. Liver cells of experimental mice were damaged using CCl4 and subsequently hepatoprotective potential of the plant extract was evaluated using series of in-vivo and in-vitro studies. In the hepatoprotective study, silymarin was used as a positive control. Antioxidant enzymes, pro-inflammatory markers, liver enzymatic and biochemical parameters were studied to evaluate hepatoprotective activity of Croton bonplandianus leaf extract. Free radical scavenging activity of CBL extract was also observed in WRL-68 cell line. The phytochemicals identified by GCMS analysis were scrutinized using in-silico molecular docking procedure. The results showed that CBL extract have potent free radical scavenging capacity. The biochemical parameters were over expressed due to CCl4 administration, which were significantly normalized by CBL extract treatment. This finding was also supported by histopathological evidences showing less hepatocellularnecrosis, inflammation and fibrosis in CBL and silymarin treated group, compared to CCl4 group. ROS generated due to H2O2 in WRL-68 cell line were normalize in the highest group (200 µg/ml) when compared with control and negative control (CCl4) group. After molecular docking analysis, it was observed that the compound α-amyrin present in the leaf extract of C. bonplandianus has better potentiality to protect hepatocellular damages than the standard drug Silymarin. The present study provided supportive evidence that CBL extract possesses potent hepatoprotective capacity by ameliorating haloalkane induced liver injury in the murine model. The antioxidant and anti-inflammatory activities also affirm the same. The synergistic effects of the phytochemicals present in CBL are to be credited for all the hepatoprotective activity claimed above.

Elevated levels of C-reactive protein and IL-6 among the antipsychotic medicating schizophrenia patients of Siliguri, West Bengal, India.

PURPOSE: Chronic, low-grade inflammation is a proposed etiological factor associated with schizophrenia. Thus, various studies have been conducted to understand the role of inflammatory process in schizophrenia by using inflammatory maker C-reactive protein (CRP) with conflicting findings. Inadvertently, studies of CRP among the Indian schizophrenia patients are very few. Therefore, the present study was undertaken to investigate the role of inflammatory process among Indian Bengalee schizophrenia patients of Siliguri, using the marker CRP and its stimulating cytokine interleukin-6 (IL-6). In addition, the study also intended to investigate the immunomodulatory effect of antipsychotic medication on serum levels of CRP and IL-6. MATERIALS AND METHODS: The serum levels of CRP and IL-6 were measured by Enzyme-Linked Immunosorbent Assay (ELISA) among 67 antipsychotic medicating, 28 psychotropic medication-free schizophrenia patients, and it was compared with 72 age, sex and ethnicity-matched controls. RESULTS: A significantly higher level of CRP and IL-6 were recorded among the antipsychotic medicating patients. Although CRP was found to be higher among the psychotropic medication-free patients than the controls, it was not found to be significant. However, a significantly higher level of IL-6 was observed in this group. CONCLUSIONS: The results provide the evidence for a possible immunomodulatory effect of antipsychotic drugs on CRP. Future investigations including the study of antipsychotics separately may help to understand the differential effects of individual antipsychotics on CRP level. Additional studies with a larger sample size of psychotropic medication-free patients may help to verify the role of inflammation in schizophrenia patients of this region.

Acacia nilotica leaf improves insulin resistance and hyperglycemia associated acute hepatic injury and nephrotoxicity by improving systemic antioxidant status in diabetic mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Acacia nilotica (L.) Delile is used as a traditional anti-diabetic remedy in Bangladesh, Pakistan, Egypt, Nigeria and is mentioned in Ayurveda as well. AIM: The objective of the study was to evaluate the ethnomedicinal claim of A. nilotica leaf (ANL) extract for its efficiency in ameliorating diabetic complications. MATERIALS AND METHODS: ANL was orally administrated (50 and 200mg/kg) to alloxanized mice (blood glucose > 200mg/dL) for 20d. Parameters of glucose metabolism, hepatotoxicity, hyperlipidemia and nephrotoxicity were measured with emphasis on elevated oxidative stress. ANL was chemically characterized using GC-MS. Further, docking studies were employed to predict molecular interactions. RESULTS: ANL lowered (65%, P< 0.001) systemic glucose load in diabetic mice, which was otherwise 398% higher than control. ANL lowered (35%) insulin resistance, without any significant effect on insulin sensitivity (P> 0.05). Anti-hyperglycemic properties of ANL was further supported by lowering of HbA1c (34%; P< 0.001) and improved glucose utilization (OGTT). Overall diabetic complications were mitigated as reflected by lowered hepatic (ALT, AST) and renal (creatinine, BUN) injury markers and normalization of dyslipidemia. Elevated systemic oxidative stress was lowered by increased catalase and peroxidase activities in liver, kidney and skeletal muscle, resulting in 32% decrease of serum MDA levels. Apart from high phenolic and flavonoid content, tocopherol, catechol and ß-sitosterol, identified in ANL, demonstrated substantial binding affinity with Nrf2 protein (5FNQ) reflecting possible crosstalk with intracellular antioxidant defense pathways. CONCLUSION: The present study revealed the potentials of A. nilotica to alleviate diabetes-related systemic complications by limiting oxidative stress which justified the ethnopharmacological antidiabetic claim.

A rapid and efficient DNA extraction protocol from fresh and frozen human blood samples.

BACKGROUND: Different methods available for extraction of human genomic DNA suffer from one or more drawbacks including low yield, compromised quality, cost, time consumption, use of toxic organic solvents, and many more. Herein, we aimed to develop a method to extract DNA from 500 µL of fresh or frozen human blood. METHODS: Five hundred microliters of fresh and frozen human blood samples were used for standardization of the extraction procedure. Absorbance at 260 and 280 nm, respectively, (A260 /A280 ) were estimated to check the quality and quantity of the extracted DNA sample. Qualitative assessment of the extracted DNA was checked by Polymerase Chain reaction and double digestion of the DNA sample. RESULTS: Our protocol resulted in average yield of 22±2.97 µg and 20.5±3.97 µg from 500 µL of fresh and frozen blood, respectively, which were comparable to many reference protocols and kits. CONCLUSION: Besides yielding bulk amount of DNA, our protocol is rapid, economical, and avoids toxic organic solvents such as Phenol. Due to unaffected quality, the DNA is suitable for downstream applications. The protocol may also be useful for pursuing basic molecular researches in laboratories having limited funds.

Variation in Phytochemical Composition Reveals Distinct Divergence of Aloe vera (L.) Burm.f. From Other Aloe Species: Rationale Behind Selective Preference of Aloe vera in Nutritional and Therapeutic Use.

In the present study, we have phytochemically characterized 5 different abundant Aloe species, including Aloe vera (L.) Burm.f., using silylation followed by Gas Chromatography-Mass Spectrometry technique and compared the data using multivariate statistical analysis. The results demonstrated clear distinction of the overall phytochemical profile of A vera, highlighted by its divergent spatial arrangement in the component plot. Lowest correlation of the phytochemical profiles were found between A vera and A aristata Haw. (-0.626), whereas highest correlation resided between A aristata and A aspera Haw. (0.899). Among the individual phytochemicals, palmitic acid was identified in highest abundance cumulatively, and carboxylic acids were the most predominant phytochemical species in all the Aloe species. Compared to A vera, linear correlation analysis revealed highest and lowest correlation with A aspera ( R2 = 0.9162) and A aristata ( R2 = 0.6745), respectively. Therefore, A vera demonstrated distinct spatial allocation, reflecting its greater phytochemical variability.

Radical Scavenging Activities of Lagerstroemia speciosa (L.) Pers. Petal Extracts and its hepato-protection in CCl4-intoxicated mice.

BACKGROUND: Lagerstroemia speciosa (L.) Pers. has medicinal importance. Bioactive phytochemicals isolated from different parts of L. speciosa, have revealed hypoglycemic, antibacterial, anti-inflammatory, antioxidant and hepato protective properties. Despite one report from Philippines detailing the use of L. speciosa as curative for fever and as well as diuretic, there is no experimental evidence about the hepatoprotective activity of the flower extracts. METHODS: Several spectroscopic methods, including GC-MS, were used to characterize phytochemicals present in the petal extract of L. speciosa. Ethanol extract of petals was evaluated for anti-oxidant and free radical scavenging properties by using methods related to hydrogen atom transfer, single electron transfer, reducing power, and metal chelation. This study has also revealed the in vitro antioxidant and in vivo hepatoprotective properties of petal extract against carbon tetra chloride (CCl4)-induced liver toxicity in Swiss albino mice. Hepatoprotection in CCl4 -intoxicated mice was studied with the aid of histology and different enzymatic and non-enzymatic markers of liver damage. Cytotoxicity tests were done using murein spleenocytes and cancareous cell lines, MCF7 and HepG2. RESULT: GCMS of the extract has revealed the presence of several potential antioxidant compounds, of them γ-Sitosterol and 1,2,3-Benzenetriol (Pyrogallol) were the predominant ones. The antioxidants activities of the flower-extract were significantly higher than curcumin (in terms of Nitric oxide scavenging activity; p = 0.0028) or ascorbic acid (in terms of 2,2-Diphenyl-1-Picrylhydrazyl (DPPH) assay; p = 0.0022). The damage control by the flower extract can be attributed to the reduction in lipid peroxidation and restoration of catalase activity. In vitro cytotoxicity tests have shown that the flower extract did not affect growth and survivability of the cell lines. It left beyond doubt that a flower of L. speciosa is a reservoir of antioxidant and hepatoprotective agents capable of reversing the damage inflicted by CCl4-intoxication. CONCLUSION: Results from the present study may be used in developing a potential hepato-protective health drink enriched with antioxidants from Lagerstroemia speciosa (L.) Pers.

Toxicological assessment of Diplazium esculentum on the reproductive functions of male Swiss albino mouse.

CONTEXT: Diplazium esculentum, a commonly consumed seasonal vegetable, has been reported to have some pathological effects in some animals. But, its effect on the male reproductive function has not yet been studied. OBJECTIVE: To investigate the effects of boiled D. esculentum (BDE), the form which human consumes, on male reproductive functions of Swiss albino mice. MATERIALS AND METHODS: Male (120 in no.) and female (80 in no.) Swiss albino mice (6-8 weeks of age) were fed orally with 80, 160 and 320 mg/kg bw of BDE within a span of 180 d. After the treatment, body weight, absolute- and relative-testis weight, relative-weight of other organs, their biochemical parameters, hypo-osmotic swelling test (HOST) of spermatozoa, testis histology and fertility and fecundity tests were performed to justify the toxic effects of D. esculentum on male reproductive functions. RESULTS: Significant dose- and time-dependent decreases were observed in body weight, absolute- and relative-testis weight, relative-weights of other organs and their biochemical parameters, percentage of live spermatozoa and percentage of fertility and fecundity in BDE fed mice. Significant decreases were observed in diameter, perimeter and area of the seminiferous tubules of mice treated for 180 d. The percentage of empty seminiferous tubules was increased significantly in BDE treated mice when compared to the controls. DISCUSSION AND CONCLUSION: These results suggest that the intake of D. esculentum, even after cooking, may induce infertility by altering the male reproductive function, and therefore, should be evaluated further as a potential antifertility agent.

Haloalkane induced hepatic insult in murine model: amelioration by Oleander through antioxidant and anti-inflammatory activities, an in vitro and in vivo study.

BACKGROUND: Nerium oleander L. (syn. Nerium indicum Mill, Nerium odorum Aiton) belongs to the family Apocynaceae. It is used for its anti-inflammatory, anti-diabetic, anti-cancer and hepatoprotective activities in traditional medicine. Previous pharmacognostic studies suggested that 70 % hydro-methanolic extracts of oleander possess potent free radical scavenging and anti-inflammatory activities, both of which are helpful against hepatotoxicity. METHODS: Hydro-methanolic extracts of oleander stem and root were evaluated for their hepatoprotective activities in acute CCl4 intoxicated mouse through in vitro and in vivo studies. Silymarin was used as positive reference. Antioxidant enzymes, pro-inflammatory markers and liver enzymatic and biochemical parameters were studied. The extracts were further chemically characterized using Fourier Transform Infrared (FTIR) spectroscopy and Gas chromatography-mass spectrometry (GC-MS). RESULTS: CCl4 toxicity caused fatty liver formation by increase of relative liver weight (32.53 g) compared to control group (16.08 g). The elevated liver enzymatic and biochemical parameters due to CCl4 toxicity were considerably normalized by the extracts treatment under both in vivo and in vitro models. Oleander stem (NOSE) and root (NORE) extracts increased the reduced hepatic catalase activity 27.37 and 25.25 %, whereas peroxidase activity was increased 18.19 and 22.78 %, respectively. The extent of lipid peroxidation was significantly (p < 0.01) lowered 20.76 % (NOSE) and 21.12 % (NORE) compared to CCl4 group. The levels of pro-inflammatory tumor necrosis factor-α (TNF-α) was lowered 71.33 % (NOSE) and 61.60 % (NORE). Histopathological study demonstrated substantial reduction of hepatocellular necrosis, fatty infiltration, sinusoidal dilation, bile duct proliferation, vascular congestion, leukocyte infiltration in the silymarin and extract treated groups. Furthermore, various bioactive compounds were identified in the extracts such as apocynin, tocopherol, squalene, vanillin, isoeugenol, amyrin, lupeol etc. CONCLUSION: The present study provided convincing evidence that oleander extracts possess potent hepatoprotective capacity which was primarily governed by its antioxidant and anti-inflammatory activities. The collegial bioactivities of the phytochemicals may be accredited behind the hepatoprotective activity of oleander.

Comparative phytochemical profiling of Clerodendrum infortunatum with some selected medicinal plants predominant in the Sub-Himalayan region of West Bengal.

BACKGROUND: The aim of the present study was to quantify the phytochemicals of Clerodendrum infortunatum root and correlate on the basis of phytochemicals present with other medicinal plants belonging to different families. METHODS: Various standard biochemical and spectrophotometric methods were employed to study the phytochemical status of the root of C. infortunatum. All the analysis was performed in multiple sets. Various statistical methods like principal component analysis, dendogram, Euclidean distance and correlation study were done to compare the phytochemical profile of C. infortunatum with different selected medicinal plants belonging to different families. RESULTS: Results indicated the presence of various phytochemicals like phenol, alkaloid, flavonoid, tannin, thiamine, riboflavin, etc. Among the estimated phytochemicals, phenol had the highest quantity (65.59±1.26 mg/g) followed by lipid (27.51±0.23 mg/g) and tannin (16.40±0.07 mg/100 g). The phytochemical contents of C. infortunatum are much higher than those of the other reference plants. CONCLUSIONS: It may be concluded from the present study that the root of C. infortunatum contains very high amount of phytochemicals which are chiefly responsible for various medicinal properties. Therefore, the presence of the bioactive compounds in high quantity may lead to the potent medicinal value of C. infortunatum root.

Assessment of haemolytic, cytotoxic and free radical scavenging activities of an underutilized fruit, Baccaurea ramiflora Lour. (Roxb.) Muell. Arg.

Baccaurea ramiflora Lour. (Roxb.) Muell. Arg. is an underutilized juicy fruit bearing plant found in sub-Himalayan area, South China, Indo-Burma region, etc. The fruit is considered to be nutritive, and in this study, we evaluated its antioxidant, haemolytic and cytotoxic properties. The juice was examined for the quenching activity of hydroxyl radical, nitric oxide, singlet oxygen, peroxynitrite, total antioxidant activity (TAA), erythrocyte membrane stabilizing activity (EMSA) along with quantification of phenolic and flavonoid contents and also tested for its potential activity as iron chelator, inhibitor of lipid peroxidation and total reducing power. Principal component analysis (PCA) and hierarchical cluster analysis (HCA) were also performed to correlate antioxidant capacities with the phenolic and flavonoid content. Haemolytic activity on murine erythrocyte and MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) cytotoxic test was performed on murine splenocytes, thymocytes, hepatocytes and peritoneal exudates macrophage to examine the cytotoxic effect of its juice. The result exhibited its potent free radical scavenging activity. In case of TAA, DPPH (2, 2-diphenyl-1-picrylhydrazyl), EMSA and lipid peroxidation, the fruit juice was found to have significant (P < 0.001) antioxidant capacity, which is evident from low IC50 (half maximal inhibitory concentration) value. Results obtained from haemolytic inhibition assay and MTT cytotoxic test confirms that the juice does not contain any cytotoxic effect and the fruit is safe for consumption. Fourier transform infrared (FTIR) spectra analysis exhibited high possibility of presence of flavonoid compounds in the juice.

Evaluation of anti-inflammatory activity and standardisation of hydro-methanol extract of underground tuber of Dioscorea alata.

Context The underground edible tuber of Dioscorea alata L. (Dioscoreaceae) is a functional food with high nutritive value and therapeutic potential. The tuber is known to possess anti-inflammatory properties in traditional medicine. Objective The present study explores the anti-inflammatory activity and standardisation of D. alata tuber hydromethanol extract. Materials and methods Hydromethanol extract (70%) of D. alata tuber was chemically characterised using HPLC and GC-MS techniques. Murine lymphocytes were cultured for 48 h with six different concentrations (0-80 µg/mL) of the extract. The expression of nitric oxide (NO), TNF-α, COX-1, COX-2, and PGE2 were evaluated using colorimetric and ELISA methods. Results Dioscorea alata extract inhibited the expression of NO and TNF-α with an IC50 value of 134.51 ± 6.75 and 113.30 ± 7.44 µg/mL, respectively. The IC50 values for inhibition of total COX, COX-1, COX-2 activities and PGE2 level were 41.96 ± 3.07, 141.41 ± 8.99, 32.50 ± 1.69, and 186.34 ± 15.36 µg/mL, respectively. Inhibition of PGE2 level and COX-2 activity was positively correlated (R(2) = 0.9393). Gallic acid (GA), 4-hydroxy benzoic acid (4HBA), syringic acid (SYA), p-coumaric acid (PCA), and myricetin (MY) were identified and quantified using HPLC. GC-MS analysis revealed the presence of 13 different phytocompounds such as hexadecanoic acid, methyl stearate, cinnamyl cinnamate, and squalene. Conclusion The D. alata extract significantly down-regulated the pro-inflammatory signals in a gradual manner compared with control (0 µg/mL). Different bioactive phytocompounds individually possessing anti-inflammatory activities contributed to the overall bioactivity of the D. alata tuber extract.

Environmental selection influences the diversity of TLR genes in ethnic Rajbanshi population of North Bengal Region of India.

BACKGROUND: Toll-like receptors are the mediators of the innate immune response to pathogens. In human, this gene family regulates the inflammatory pathways and is associated with the susceptibility to infection. SUBJECTS AND METHODS: The distribution and the diversity patterns of TLR genes in Rajbanshi population (n = 85) who are the inhabitants of the Northern part of West Bengal, have been investigated in the present study. PCR-SSP was done for all the ten TLR genes. We have also constructed the phylogenetic tree principal component analysis and genetic distance for all the four populations. RESULTS: It has been observed that in Rajbanshi population, the frequency of TLR8 (0.894) is higher and the frequency of TLR2 (0.176) is very low. Dendrogram based analysis, as well as the PCA plot, documented the closeness of Rajbanshi and Gurkha population. However, Rabha is distantly related to Rajbanshi population though evidences suggest their emergence from the same East-Asian lineage. Genetic distances between Rajbanshi-Gurkha and Rajbanshi-Muslim are very much smaller than that of Rajbanshi-Rabha populations. INTERPRETATIONS & CONCLUSIONS: Therefore, it may be concluded that Rajbanshi, Gurkha and Muslims are very much mixed populations and have genetic closeness due to exposure to similar environmental conditions. On the other hand, the Rabhas strictly follow the endogamy and are restricted to a particular region and therefore maintain considerable distances with the other three populations. The data showed some interesting observations which deviate the contemporary thought in respect to the population genetics.

Immunomodulatory activity of Nerium indicum through inhibition of nitric oxide and cyclooxygenase activity and modulation of TH1/TH2 cytokine balance in murine splenic lymphocytes.

Nerium indicum is a medicinal plant which is used in the treatment of wide range of illness in different ethnopharmacological practices. We have studied the immunomodulatory activity of the 70 % hydro-methanolic extract of N. indicum leaves (NILE) by studying plaque forming cell assay, haemagglutination titre, cell proliferation assay, inhibition of nitric oxide expression and cyclooxygenase activity, estimation of IL-2, IFN-γ, IL-4, IL-10, TNF-α and IgM levels. Furthermore, we have characterized NILE with Fourier Transform Infra-red (FTIR) spectroscopy. The results displayed that NILE possessed potent immunomodulatory activity by up-regulating IL-2, IFN-γ, IL-10 expression and down-regulating IL-4, TNF-α expression in vitro. NILE also stimulated the in vivo expression of immunoglobulin level in mouse. FTIR analysis revealed phytocompounds of diverse chemical nature present in NILE. Thus, the potent immunomodulatory activity of N. indicum may have serious implications in the treatment of inflammatory diseases in the future.

Study of genetic diversity of KIR and TLR in the Rabhas, an endogamous primitive tribe of India.

The Rabha tribe is a little known small endogamous population belonging to Indo-mongoloid group of north-eastern India. We have analyzed 16 KIR and 5 TLR gene polymorphisms in the Rabha population of northern West Bengal, India for the first time. The observed frequencies of the KIR genes (except framework and pseudogene loci) ranged between 0.26 (KIR2DS3) and 0.96 (KIR2DL1). Comparisons based on KIR polymorphism have revealed that although the Rabhas are of Indian origin the presence of mongoloid component in their gene pool cannot be denied. The frequencies of the 5 TLR genes ranged between 0.90 (TLR4) and 0.46 (TLR5). TLR variations found in the Rabhas may play a synergistic role in fighting against the bacterial invasions. Our results may contribute to the understanding of (1) genetic background and extent of genetic admixture in the Rabhas, (2) population migration events and (3) KIR-disease-TLR interactions.

Anti-inflammatory activity of Nerium indicum by inhibition of prostaglandin E2 in murine splenic lymphocytes.

OBJECTIVE: Nerium indicum Mill (syn. N. oleander L. and N. odorum Aiton; family: Apocynaceae) is a medicinal plant, used in the treatment of diverse ailments including various chronic inflammatory diseases in traditional medicine. We have previously demonstrated the immunomodulatory activity of a bioactive fraction of Nerium indicum leaf (NILE) by studying up-regulation of interleukin-2 (IL-2), IL-10, interferon-gamma and down regulation of IL-4, tumor necrosis factor-alpha (TNF-α), nitric oxide, cyclooxygenase-1 (COX-1) and COX-2 activities. Therefore, this study aimed to confirm the anti-inflammatory activity of NILE by inhibition of prostaglandin E2 (PGE2) activity in murine splenic lymphocytes in vitro. MATERIALS AND METHODS: Murine lymphocytes were isolated from spleen and stimulated with 5 ΅g/mL concanavalin A in RPMI-1640, supplemented with 50 U/mL penicillin, 50 U/mL streptomycin, 50 U/mL nystatin and 10% fetal bovine serum. Different concentrations (0-80 µg/mL) of NILE were added and the cells were cultured for 48 h. The culture supernatants were thereafter collected by centrifugation and assayed for expression of PGE2 level. The data were analyzed statistically. RESULTS: The results demonstrated a 2.26-fold inhibition of PGE2 level at 80 µg/mL of NILE. Half maximum inhibitory concentration (IC50) was calculated to be 44.95 ± 0.45 ΅g/mL. Linear correlation analysis of the dose-dependent PGE2 inhibition with other pro- and anti-inflammatory mediators demonstrated high inter-correlation between the parameters. CONCLUSIONS: Thus, the present study remains in accordance with our previous report and confirms the anti-inflammatory claim of N. indicum, mentioned in the traditional medicine.