BACKGROUND:Type 2 diabetes mellitus (T2DM) is the most common metabolic disorder and inflammation plays a key role in the pathogenesis of the disease. Ethnic variations have been linked with an increased prevalence of diabetes. This pilot study was designed to evaluate the inflammatory markers and demographic characteristics in T2DM patients among the ethnic group of Sikkim.MATERIALS AND METHODS:A total of 100 type 2 diabetic patients and 50 healthy controls were enrolled in the study. Demographic characteristics were recorded among the patients and inflammatory markers [high sensitivity C-reactive protein (hs-CRP) and interleukin-6 (IL-6)], body mass index (BMI), glycemic status, fasting blood sugar (FBS), and postprandial blood sugar (PPBS) level were compared between the cases and controls. Inflammatory markers were correlated with BMI, glycemic status, and duration of diabetes.RESULTS:Type 2 diabetes mellitus was found to be highest in between the age-group of 45 and 55 years with males being mostly affected. Among the ethnic group, Nepalis (57%) comprised the highest with diabetes. No significant correlation was found between inflammatory markers with BMI, glycemic status, and duration of diabetes.CONCLUSION:Our study showed that T2DM increases as age progresses with males being mostly affected. The Nepalese community showed a high proportion of the disease mostly living in the urban residences. Though we included patients on statin therapy, its specific roles and relationship were not in the scope of the current study, but further studies should be conducted at the community level to understand the role and relationship of statins among type 2 diabetic patients and the possibilities of ethnic differences in the levels of biomarkers due to variation in the body fat distribution.

Oxidative stress (OS) is an early event and at the same time also a consequence in the pathology of MetS. We investigated if oxidation markers of DNA, lipid and protein increased with an increase in the risk parameters of MetS. Participants (male:70, female:90 ≥ 20 yrs) were categorized based on the number of risk factors they had as 3 Risk, 2 Risk, 1 Risk and 0 Risk for MetS and were evaluated for various oxidation markers. Protein carbonyl and advanced oxidation protein product (protein oxidation marker) differed significantly between the four study group while malondialdehyde and hydroxynonenal (lipid peroxidation marker) did not. “8-OH dG” (DNA oxidation marker) differed significantly (P 0.05). Pairwise comparison for statistically significant markers(Protein oxidation markers and 8-OH dG), demonstrated that only 8-OH dG differed significantly between 0 Risk- 3 Risk (P< 0.012) but not between 0 Risk -2 Risk and 0 Risk-1 Risk. Oxidative stress markers of DNA, lipid and protein do not increase with an increase in the risk parameters of MetS. However, it is indeed high in MetS with 3 and more risk parameters. Presence of 2 or 1 Risk also increases OS compared to 0 Risk. There is oxidative stress damage in MetS to lipid and protein but DNA damage was of significant consequence.

BACKGROUND A successful isolation of Mesenchymal Stem Cells (MSCs) for its clinical application is very important. Isolation of MSCs from neonatal sources like human Umbilical Cord Wharton's Jelly (hUCWJ) have been reported in many studies and are considered primitive but isolation of MSCs from human umbilical cord blood (hUCB) is open to discussion. MATERIALS AND METHODS The MSCs from hUCWJ and hUCB were isolated by explant culture and Ficoll density gradient respectively. The media used for the two sources were Dulbecco's modified Eagles medium, GlutaMax and fetal bovine serum (FBS) for hUCWJ, and Iscove modified Dulbecco's medium and FBS for hUCB. Further, the isolated cells were studied as per criteria of International Society of Cellular Therapy to define the cells as MSCs. Finally, we compare the feasibility and ease of isolating the neonatal sources. RESULTS Unlike hUCWJ, defining hUCB cells to MSCs as per criteria is limited because the number of cells decreases once it reaches 50% confluence and after 2 weeks, they show differing morphology with an appearance of large dimension of multinucleated cells identifying them as osteoclast-like cells thus restricting further study. The cells derived from both the sources grew into long, spindle shaped cells with prominent nuclei but hUCB has limited proliferation and fulfilled only the first criterion to define as MSCs while the cells isolated from hUCWJ continue to grow for multiple passages and fulfil all the defining criteria. CONCLUSION Neonatal derived hUCWJ is more ideal source for isolation of MSCs than hUCB with the potential to use it in regenerative medicine.

Background: Human umbilical cord Wharton's jelly (hUCWJ) derived Mesenchymal stem cells (MSCs) is a non-controversial, easily available source of human tissue which has a close association with embryonic tissue. Explant culture technique of isolating MSCs do not require any proteolytic enzymes giving an edge when compared to enzymatic methods. Aims and Objective: This study was designed to develop a suitable protocol to optimize the number of primary cells by transferring the explants after a successful migration and before trypsinizing of MSCs from primary culture plate to new culture plates. Materials and Methods: For defining the isolated cells as MSCs, the cells were studied by colony forming assay, semiquantitative two steps reverse transcriptase polymerase chain reaction (RT-PCR) and International Society for Cellular Therapy (ISCT) criteria to determine the purity of the lineage. Results: The explant cultured cells fulfil the defining criteria by ISCT. Further, they form colony and RT-PCR confirmed the presence of thy-1, endoglin and β-actin. The incubated explant shows successful cells migration after transferring from one culture plate to another for five consecutive times. Further, it was seen that the migrated cells can be cultured for more than 18 passages. However, a reduction was seen in the migration of the cells after the 3rd transfer of the explants. Conclusion: This study found that the transferred explant has the potential to yield cells for 5 consecutive explant transfers. Further, this technique optimized the yield of primary cells and thereby reducing the quantity and frequency of sample collection, processing time and cost.

BACKGROUND & OBJECTIVES:Oxidative stress (OS) is associated with numerous components of metabolic syndrome (MetS). This study was aimed to investigate if hydrogen peroxide (H2O2) as the reactive oxygen species was capable of depicting OS in MetS, and If MetS patients showed DNA damage in the form of DNA strand breaks (DSB).METHODS:A total of 160 participants (90 males, 70 females) ≥20 yr of age were categorized into four groups based on the number of MetS risk parameters (n=40 in each group). Sugar and lipid profile, H2O2concentration in blood and DNA-strand breaks were measured.RESULTS:DSB was significantly more in those with MetS (n=40) than those without (n=120) whereas H2O2levels were the same in both the study groups. The number of DSB differed significantly between the control and 3 risk factor groups. DSB was also higher in groups with 2 and 1 risk factors compared to 0 risk but the difference was not significant. H2O2 level was higher in groups with 3, 2 and 1 risk factors compared to 0 risk group but the difference was not significant. The H2O2level correlated positively with triglyceride values but not with other MetS risk parameters. There was no significant correlation between DSB and MetS risk parameters.INTERPRETATION & CONCLUSIONS:Our findings showed a cumulative and synergistic effect of the risk factors of MetS on DSB. Individuals with three risk parameters had a greater effect on DNA damage than in those with two or one risk parameter. Although plasma H2O2level increased with an increase in the fat depots, use of H2O2to depict OS in MetS should be coupled with an adjunct and estimation of DSB in peripheral blood lymphocytes may be used as indicator of OS in MetS patients.

Single nucleotide polymorphism in the first intron of Collagen type I alpha 1 (COL1A1) gene which is the binding site of specificity protein 1 (Sp1) transcription factor associated with low bone mineral density and osteoporosis. To evaluate such genetic factors among the Sikkimese population, a total of 150 cases (75 men and 75 women) with primary osteopenia and osteoporosis and 150 healthy controls (75 men and 75 women) of age range between 35 and 65 years were enrolled in this study. The COL1A1 genotypes [SS, Ss and ss] were assessed by restriction enzyme [MscI] digestion of DNA after amplification by polymerase chain reaction. There, only 2.7% women and 1.3% men cases had restriction site with heterozygous genotype (Ss) and no homozygous genotype (ss) were detected. There was no statistically significant association between low bone mass and genotypes on analysis (χ2 = 1.014, P = 0.314; RR = 1.510) that, the Sp1 binding site polymorphism at the COLIA1 gene is very rare and has no contribution in the development low bone mineral density.

Sikkim has been reported to have the highest percentage of Diabetes Mellitus and Hypertension in the country. The study aimed to focus its precursor termed 'Metabolic Syndrome' (MetS) with special attention to its risk determinants as a measure to promote awareness in preventing the rise in number of these non communicable diseases in the state with only 6,10,577 inhabitants. Of 361 participants, 33.5% were diagnosed MetS as per the harmonized MetS criteria, 64.5% deemed non-MetS comprised of participants with "2 Risk", "1 Risk" and "0 Risk" for MetS, however not enough (3 or more) to be categorized as MetS. A "large WC + elevated BP + raised FBS"; "large WC + elevated BP" and "large WC" were the different types of risk combinations found frequently in the "3 Risk", "2 Risk" and "1 Risk" category respectively. MetS was most common among the females and highly found in 51-60 years of age. Ethnically "Bhutia/Lepcha/Tamang/Sherpa" were diagnosed with highest percentage of MetS followed by "Nepalese" and "Others". The chief contributing factor to compose MetS in the female population was a large WC. We conclude, Diabetes and Hypertension are indeed prevailing in Sikkim and that the presence of either an increased waist circumference or an elevated blood pressure mandates a check for the remaining four risk factors for MetS, rendering worthwhile to keep an account of MetS risk types by stratification to comprehend the influence of socio-cultural and ethno-geographical factors in causing MetS.

Osteoarthritis is characterized by the degeneration of articular cartilage. Cartilage metabolic markers have been explored as possible markers for osteoarthritis, and osteogenic protein -1 (OP-1) has emerged out to play a major role in cartilage repair. Oxidative stress has been implicated as a mediator of cartilage damage in patients with osteoarthritis. The aim of this study was to correlate the cartilage metabolic markers and antioxidants with the severity of knee osteoarthritis.

The thyroid dysfunction is one of the most common endocrine disorders. Sikkim lies in the severely iodine deficient zone. This was a hospital based cross-sectional study. The serum samples were used to check the levels of thyroid stimulating hormone (TSH), triiodothyronine (T3) and thyroxine (T4). Out of 674 patients with suspected thyroid dysfunction, 83% were Euthyroid followed by 10 % of patients having subclinical hypothyroidism. The incidence subclinical hyperthyroidism (1%) were lowest. Females were found to be maximum with thyroid disorders. The datas were represented as percentage and mean ± SD. Thyroid hormones were compared among the different thyroid disorder by One way analysis of variance (ANOVA). High incidence of thyroid dysfunction in females with subclinical hypothyroidism indicates that it still exists as a public health problem in Sikkim regardless of the implementation of iodized salt program since the last decade.

Role ofAmaranthusspinosusLinn.in indomethacin induced gastric ulcers was studied in rats. Amaranthusspinosus Linn. Leaves significantly reduced ulcer index induced byindomethacin. The plant leaves produced gastric anti secretory effect by decreasing gastric volume and acidity. It further increased gastric mucin which showedgastric cytoprotective effect. The plant prevented the increased lipid peroxidation during ulceration by indomethacin. Activities of the anti oxidant enzymes were enhanced during ulceration by this plant leaves. Results were comparable to that of ranitidine, a standard anti ulcer drug.Anti ulcerogenic activity of Amaranthusspinosus Linn. leaveswas, thus, mediated through anti oxidant defense mechanism.

INTRODUCTION:Osteoarthritis (OA) is a progressive degenerative disorder of the articular cartilage. Available diagnostic radiography has been poorly associated with the progress and severity of this clinical disease. As osteogenic protein-1 (OP-1) has been identified as a bone morphogenetic protein with a major role in cartilage repair, we aimed to evaluate its potential role in the diagnosis of OA.METHODS:This was an experimental study conducted at the Department of Biochemistry, Sikkim Manipal Institute of Medical Sciences, India. Polyclonal antibodies (i.e. anti-OP-1[f]) were raised against OP-1 in mice, and subsequently used in a sandwich enzyme-linked immunosorbent assay (ELISA) to detect the presence of OP-1 in the synovial fluids of 75 osteoarthritic patients. For the purpose of correlation, the radiographic assessments of the knees of the 75 patients were graded using the Kellgren-Lawrence scoring system.RESULT:The polyclonal antibody (i.e. anti-OP-1[f]) raised against OP-1 was able to detect the presence of OP-1 in the synovial fluids of all the osteoarthritic patients via sandwich ELISA. The level of the OP-1 was found to be much higher than the reference range and correlated positively with the severity of OA (r = 0.24; p = 0.04).CONCLUSION:Our study shows that the polyclonal antibody, anti OP-1(f), could be used for the immunodiagnosis of osteoarthritis via sandwich ELISA.

By different solvent extractions and chromatographic techniques an antibacterial constituent was isolated from leaves of Cassia alata Linn. Infra red spectroscopy, mass spectroscopy and nuclear magnetic resonance studies showed that the isolated compound was chemically 3,4 dihydroxy cinnamic acid. In vitro antibacterial activity of 3,4 dihydroxy cinnamic acid was studied against four Gram-positive and four Gram-negative bacteria using disc diffusion method. Minimum inhibitory concentration (MIC) of 3,4 dihydroxy cinnamic acid was also recorded against those bacteria by serial dilution technique. Kanamycin was used as positive control. Results showed that 3,4 dihydroxy cinnamic acid had antibacterial activity against the tested bacteria.

Background: Oxidative stress has been implicated as a mediator of cartilage damage in patient with Osteoarthritis (OA). Aim: To assess the antioxidant markers like superoxide dismutase (SOD) , glutathione peroxidase (GPx) and uric acid (UA) in the blood and cartilage metabolic markers like hyaluronic acid (HA) and keratan sulphate (KS) in the synovial fluid of osteoarthritic patients and to find the correlation between these parameters in the blood and synovial fluid. Methods: This was an observational study conducted in the department of Biochemistry, Sikkim Manipal Institute of Medical Sciences (SMIMS), Gangtok. 75 osteoarthritic patients and healthy volunteers were taken for the study. Blood levels of antioxidants (SOD, GPx and UA) were assayed in both osteoarthritic and healthy volunteers. Cartilage metabolic markers like HA and KS were assayed in the synovial fluid. Result: Osteoarthritic patients showed statistically significant increase in KS and decrease in HA level indicating cartilage damage. Decrease in SOD and GPx activities indicates increase in oxidative stress. The difference in UA levels in the two groups was not statistically significant. This study did not establish any significant positive/negative correlation between the antioxidants in the blood and cartilage metabolic markers in the synovial fluid of osteoarthritic patients.

BACKGROUND Age and oxidative stress have been implicated as the main causes of the cartilage damage in osteoarthritis (OA). The osteogenic protein-1(OP-1), a member of the bone morphogenetic family (BMP), plays a major role in cartilage repair. AIM To correlate the OP-1 in the synovial fluid with the age and antioxidants like superoxide dismutase (SOD), glutathione peroxidase (GPx) and uric acid (UA) in the blood and with the cartilage metabolic markers like hyaluronic acid (HA) and keratan sulphate (KS) in the synovial fluid of osteoarthritic patients. METHOD Seventy five osteoarthritic patients were taken up for the study. Heparinized blood samples were collected for the estimation of SOD, GPx and UA. Synovial fluid was aspirated for the estimation of HA, KS and OP-1. OP-1 was estimated by using a polyclonal antibody (anti-OP-1) which was produced against OP-1 in the mouse ascitic fluid by sandwich ELISA. RESULT Age and UA showed a significant correlation with OP-1. On the other hand, the correlations between OP-1 and the antioxidants (SOD and GPx) and the cartilage metabolic markers (HA and KS) were insignificant in osteoarthritic patients. CONCLUSION The study suggests about the important role of OP-1 in cartilage aging and degeneration, which may help us in understanding the potential for the therapeutic interventions in the treatment of cartilage degenerative processes.

Evidence of increased oxidative stress in patients of osteoarthritis in comparison with healthy control subjects was investigated by measuring the thiobarbituric acid reactive substances (TBARS), vitamin C, reduced glutathione (GSH) and the activities of superoxide dismutase (SOD), catalase and glutathione peroxidase (GPx) in erythrocytes. It was observed that osteoarthritis patients were more susceptible to oxidative damage than controls as evident from increased TBARS and decreased ascorbic acid, GSH, catalase and GPx in erythrocytes. Significant increase in SOD activity found in patients might be an adaptive response. With the understanding of the role of antioxidants in arthritis, it is becoming increasingly clear that these agents seem to be beneficial in osteoarthritis.

In view of association of diabetes mellitus and male infertility, present study was designed to investigate the functional status of hypothalamic pituitary gonadal (HPG) axis in diabetic men. Thirty-five diabetic men (BMI 22.24±0.21) in the age group 20–40 (30.6±4.7) years were selected. Twenty-five healthy men (BMI 23.85±0.25), in the same age group (29.5±4.8) served as control. Blood samples were collected for hormonal and biochemical estimations. Diabetic men had significantly low serum testosterone with low LH and FSH, hypercholesterolemia, hypertriglyceridemia, hypoalbuminemia and increased oxidative stress. Low serum testosterone in diabetic men was accompanied by low LH and FSH; the inability of the pituitary gland to respond appropriately to a decline in testosterone implying central effect of high serum glucose on the interaction between the nervous and endocrine system. Nutritional deficiency, increased oxidative stress and increased aromatase activity due to excessive body fat might have also contributed to low serum testosterone.

Paired samples of serum and hydrocele fluid of filarial patients associated with hydrocele were analysed for filarial antibody and antigen. Sera samples showed higher titers of filarial antibody and antigen compared to their corresponding hydrocele fluid samples. HFIgG isolated from hydrocele fluid was equally useful as FSIgG isolated from serum and detected filarial antigen in 23 out of 26 microfilaraemic sera, 7 out of 10 chronic sera and 3 out of 18 endemic normal sera by inhibition ELISA. Filarial antigen was isolated from hydrocele fluid. In inhibition ELISA antigen fraction, HFA-9 (20-25 kDa) isolated by sodium-dodecyl sulphate-polyacrylamide gel electrophoresis showed high reciprocal antigen titer of 2048. This active antigen fraction was evaluated for its diagnostic utility in comparison with Wuchereria bancrofti microfilariae excretory-secretory antigen (Wb mf ES Ag) in inhibition ELISA. Both antigen preparations detected filarial antigen in about 80% of microfilaraemic sera, 60% chronic sera and 20-30% of endemic normal sera. This study showed that antibody and antigen isolated from hydrocele fluid were equally sensitive as FSIgG and Wb mf ES Ag in the detection of filarial antigen by inhibition ELISA. Thus hydrocele fluid may be used as an alternative source for the isolation of antibody and antigen of immunodiagnostic importance.