Assessment of potential biomarkers of atherosclerosis in Indian patients with type 2 diabetes mellitus.

BACKGROUND & OBJECTIVES: : Various biological markers of subclinical atherosclerosis have been proposed to predict cardiovascular events in patients with diabetes mellitus (DM). However, there are only a few clinical studies assessing the role of invasive biomarkers [CD-36, peroxisome proliferator-activated receptor gamma (PPAR-γ) and YKL-40] in Indian patients with type 2 DM (T2DM). Hence, the present study was conducted to assess protein levels and gene expression of CD-36, PPAR-γ and YKL-40 in patients with T2DM and compare that with hypertensive and healthy controls. METHODS: : All the participants were subjected to medical history, anthropometric measurements and biochemical and biomarker (ELISA and real-time polymerase chain reaction) estimations. The study groups consisted of patients with T2DM (>5 yr) with hypertension (n=55), patients with T2DM (<2 yr) without hypertension (n=28), hypertensive controls (n=31) and healthy controls (n=30). RESULTS: : Gene expressions of YKL-40 and CD36 were significantly higher in patients with T2DM (>5 yr) with hypertension compared to healthy controls (P=0.006). In addition, a significant increase in serum levels of sCD36, PPAR-γ and YKL-40 was observed in patients with T2DM (>5 yr) with hypertension compared to healthy controls (P< 0.05). Serum levels as well as gene expression of CD36 showed significant correlation with serum levels as well as gene expression of PPAR-γ (ρ=0.45 and ρ=0.51; P< 0.001), respectively. INTERPRETATION & CONCLUSIONS: : CD36 and YKL-40 may be potential inflammatory biomarkers for early onset of atherosclerosis in patients with T2DM.

Study of oxidative stress in peripheral blood of Indian vitiligo patients.

BACKGROUND: Vitiligo is an acquired skin disease that involves the interplay of complex genetic, immunological, neural and self-destructive mechanisms in its pathogenesis. According to autocytotoxic hypothesis, oxidative stress has been suggested to be the initial pathogenic event in melanocyte degeneration. OBJECTIVES: The aim of our investigation was to evaluate the role of oxidative stress by studying the role of catalase (CAT) in the destruction of melanocytes in patients with vitiligo and compare the same in healthy normal controls. MATERIALS AND METHODS: We determined the serum catalase enzyme by ELISA method. The catalase activity was studied in two groups, Group I-localized vitiligo: (i) active stage, (ii) static or inactive stage and Group II-generalized vitiligo: (i) active stage, (ii) static or inactive stage patients, and the levels were compared with healthy controls. RESULTS: Group I active stage patients showed significant difference in the catalase levels with a P < 0.044 when compared with healthy controls, whereas Group II static stage patients did not show any significant difference (P < 0.095) although the catalase activity was increased. CONCLUSION: Our study could not explain the cause of melanocyte damage in patients in the active stage of the disease. The increase in the oxidative stress as detected by catalase activity was more significant in Group I active disease than Group II active disease patients although the levels were higher than the healthy normals. This is the first study conducted on active and static stage of vitilgo in India. It is possible that the number of compounds of hydrogen peroxide produced is not balanced by the production of catalase in the body.

Regulation of adiponectin secretion in human subcutaneous and omental adipose tissue: effects of pioglitazone and endothelin-1: a pilot study.

OBJECTIVE: The study was designed to test the effect of anti-diabetic agent pioglitazone and Endothelin-1 (ET-1) on adiponectin secretion from human adipose tissue in depot dependent manner. METHODS: Subcutaneous adipose tissue (SAT) and omental adipose tissues (OAT) were obtained from 19 subjects, including 6 non-obese controls, 7 obese and 6 obese T2DM patients. Adipose tissue was treated with pioglitazone and ET1. Adiponectin secreted into the culture medium after treatment at different time interval (0, 24, 48, 96 hours) was determined by ELISA and normalized for cellular DNA content. RESULTS: Basal adiponectin secretion from both the depots significantly associated with serum adiponectin, BMI, waist and HOMA-IR. Though no depot-specific difference was found in adiponectin secretion from SAT and OAT in our population, significant reduction in adiponectin secretion was observed in SAT of obese and T2DM patients compared to controls. Responsiveness to pioglitazone treatment was more in SAT, while ET1 inhibits adiponectin secretion in OAT. CONCLUSION: These data suggest that, SAT, appears to be major contributor to regulation of adiponectin in circulation. Pioglitazone stimulate adiponectin secretion in SAT compared to OAT in diabetic patients while ET-1 inhibiting adiponectin secretion in OAT of diabetic patients. We need to focus on mechanism underlying these regulatory agents mediated stimulation or inhibition of adiponectin secretion in human adipose tissue.

Increased prevalence of subclinical atherosclerosis in rheumatoid arthritis patients of Indian descent.

BACKGROUND: Studies have shown that rheumatoid arthritis (RA) patients are two to five times more likely to develop premature cardiovascular disease, thus shortening their life expectancy by five to 10 years. This risk has risen to approximately 12.6% in the urban population and 7.4% in the rural population of India. The Framingham risk score (FRS) identifies patients at increased cardiovascular risk and helps determine the need for preventive interventions. An investigation of the patients' coronary arteries and coronary artery calcification (CAC) - a measure of atherosclerotic plaque - has been found to be a strong predictor of cardiovascular disease. OBJECTIVE: To identify important biological markers for easy and non-invasive identification of cardiovascular disease in RA patients, and to investigate whether there is a relationship between the FRS and coronary artery atherosclerosis in RA patients. METHODS: The present study included 43 established RA patients and 50 healthy individuals (controls). Traditional and nontraditional risk factors were studied and compared with the control group. Insulin resistance was assessed using the homeostasis model of assessment of insulin resistance (HOMA-IR) and the homeostasis model of assessment of beta cell function. The FRS and the 10-year cardiovascular risk were compared between RA patients and controls. The presence of CAC was determined using electron-beam computed tomography, and the association between the FRS and CAC was examined. RESULTS: Significant differences in body mass index, waist circumference, rheumatoid factors (immunoglobulin [Ig]G, IgM and IgA) and inflammatory markers - C-reactive protein and erythrocyte sedimentation rate - were noted. There was significant correlation between HOMA-IR and body mass index, hypertension and C-reactive protein, but no correlation was seen with the homeostasis model of assessment of beta cell function. Significant differences were observed in the nontraditional biomarkers in RA patients, thus supporting their importance. Calcium deposition was observed in only seven RA patients. CONCLUSIONS: RA patients with increased C-reactive protein levels and erythrocyte sedimentation rates showed an increase in serum insulin levels and significant differences in HOMA-IR, thus indicating insulin resistance, which could lead to underlying progression of artherosclerosis. Significant differences were observed in the nontraditional risk factors, which could be chosen as biomarkers for endothelial dysfunction. There was a significant correlation between calcium score and the FRS in seven patients, suggestive of an underlying risk of atherosclerosis.

Genetic variations in nalp1 mRNA expressions in human vitiligo.

INTRODUCTION: Vitiligo is an acquired autoimmune disease of unknown etiology showing depigmentation of the skin due to the absence of melanocytes. Familial vitiligo suggests a genetic origin to this disease. Chromosome 17 was recently demonstrated to harbor the gene coding for NALP1. PATIENTS AND METHODS: A total of 18 patients of vitiligo were selected on the basis of clinical history. Group 1 (N=8) showing segmental or localized vitiligo with one or two macules on the body. Group 2 (N=10) with generalized or whole body vitiligo. A control group of 10 healthy individuals were selected from our laboratory persons with no history or any infections or skin disease. NALP1 gene expression was studied using RT-PCR assay and the bands quantitated as intensity using volume as measurement and comparison of results was done using SPSS 16 version for statistical analysis. NALP1 gene expression was observed in vitiligo patients with different intensities. RESULTS: Greater reduction in the intensity was seen in Group I, which was inversely proportional to the volume of the band. The intensity of the NALP1 and the GAPDH gene expression was more in Group 2 patients than that shown by Group 1. CONCLUSION: This study shows expression of NALP1 gene in patients as well as normals. NALP1 is widely expressed at low levels but is expressed at high levels in immune cells, particularly T cells and Langerhans cells, in which different patterns are seen that are consistent with the particular involvement of NALP1 in skin autoimmunity.

Role played by Th2 type cytokines in IgE mediated allergy and asthma.

OBJECTIVE: Recent evidence suggest that allergen type 2 helper T cells (Th2) play a triggering role in the activation/recruitment of IgE antibody producing B cells, mast cells and eosinophils. Reduced microbial exposure in early life is responsible for a shift of Th1/Th2 balance in the immune system towards the pre-allergic Th2 response. The Th1 predominantly produce IFNgamma and delayed type hypersensitivity while Th2 secrete IL-4, IL-5, IL-6, IL-13 and regulate B cell and eosinophil mediated responses. To assess regulatory changes in the immune system, in patients with allergy and asthma, we studied the cytokine profile in serum in comparison with normal healthy controls. PATIENTS AND METHODS: A total of 170 patients with various allergies and asthmatic conditions were studied, for cytokines in the serum by ELISA using kits from Immunotech, and analyzed to identify the triggering factors or main contributors towards allergy and asthma. RESULTS: Our study showed increase in the levels of IL-4, IL-5 and IL-6 in all groups which were non- significant. But the levels of IL-10, IL-13 and TNF alpha were highly significant. Besides, we found correlation of GM-CSF with IL-10. Significant correlation with different cytokines was observed. Most of these patients showed increase in IgE levels. CONCLUSIONS: This study gives a better understanding of how cytokines are the mediators of balance of Th1 and Th2 immune responses and IgE synthesis is controlled by cytokines. Further studies will eventually lead to improved treatment strategies in the clinical management of IgE mediated allergy.

Detection of Viral Citrullinated Peptide Antibodies Directed Against EBV or VCP: In Early Rheumatoid Arthritis Patients of Indian Origin.

AIM: Study was undertaken to analyze the frequency of anti-viral citrullinated peptide (anti-VCP) antibodies in sera from patients with early rheumatoid arthritis (ERA). MATERIALS AND METHODS: Viral citrullinated peptide (VCP) and Epstein-Barr nuclear antigen (EBNA-1) peptide were commercially prepared and antibodies to these were determined in 25 patients of ERA, 40 disease control patients constituting 25 rheumatoid arthritis (RA), 7 systemic lupus erythematosus (SLE), 2 scleroderma, 1 spondyloarthritis (SpA), 1 juvenile rheumatoid arthritis (JRA), 1 osteoarthritis (OA), 1 psoriatic arthritis (PsA), 1 undifferentiated arthritis (UA), and 1 gout and 25 healthy controls (HCs) were taken for comparison. In-house ELISA was established for both the antibodies while cyclic citrullinated peptide (CCP) antibody was detected by commercial ELISA kit. RESULTS: Significant increase in VCP antibody by ERA and disease controls than healthy normal was observed. VCP IgM antibody was significantly increased in RA patients than HC. The presence of VCP antibody signifies a good marker for ERA. We observed significant difference in the VCP IgG and IgM antibody when compared to EBNA-1. In-house ELISA established for EBNA-1 and VCP antibodies showed low sensitivity but 96% specificity. CONCLUSIONS: We observed that sera from early RA patients reacted to the deiminated protein encoded by Epstain Barr Virus (EBV). Thus a possible role of virus in inducing an anti-citrullinated peptide antibody (ACPA) response reveals viral etiology in this disease.

Association of adiposity, inflammation and atherosclerosis: the role of adipocytokines and CRP in Asian Indian subjects.

BACKGROUND: To reveal the exact link between adipose tissue, inflammation, and cardiovascular disease (CVD), we studied the association of C-reactive protein (CRP) with insulin resistance and adipocytokines in Asian Indian subjects. METHODS: Forty-one controls, 40 obese, and 53 type 2 diabetes (T2DM) patients (total 134) were recruited. Enzyme-linked immunoassay (ELISA) technique was used to determine serum CRP and adipocytokine concentrations. Serum insulin was measured by radioimmune assay, and insulin resistance index was calculated by the homeostasis model assessment (HOMA). Association of CRP with different adipocytokines and insulin resistance was assessed with univariate regression analysis. RESULTS: Serum leptin, resistin, and CRP levels were significantly increased and adiponectin levels were significantly reduced in obese subjects. In T2DM patients, CRP levels were increased and adiponectin levels were significantly decreased but no difference in leptin and resistin levels were found compared to controls. An important finding of this study was the significantly reduced levels of leptin, adiponectin, and resistin in nonobese T2DM patients compared to their BMI-matched controls. CRP in all subjects showed a significant correlation with obesity parameters like BMI (P < 0.001), waist circumference (P < 0.01), body fat percentage (P < 0.01), HOMA-IR (P < 0.001), leptin (P < 0.05), and resistin (P < 0.01). CONCLUSIONS: The association of CRP with insulin resistance, adipocytokines, and resistin reveals close links between inflammation, CVD, and adipose tissue. These findings provide an exciting therapeutic opportunity in cardiovascular disease by targeting various proinflammatory cascades in adipocytes.

Elevated levels of secretory immunoglobulin A (sIgA) in urinary tract infections.

OBJECTIVE: Urinary secretory IgA (sIgA), an immunoglobulin synthesized locally in mucosal surface is an important immunological defense in preventing bacterial adherence to periurethral epithelia and uroepithelia. Therefore attempts were made to measure secretory IgA in the urine of children with urinary tract infections (UTI), by using sIgA specific ELISA. METHODS: Fresh or unprocessed urine samples from healthy donors (children and adults N=10 each), 68 children and 17 Adults with UTI were tested for the presence of sIgA. RESULTS: The level of sIgA in 10 healthy normal children was 2.7 +/- 0.94 ug/ml and that in 10 healthy adults was 5.2 +/- 0.73 ug/ml. Children with UTI showed highly elevated levels of sIgA amounting to 279 +/- 80 ug/ml (p<0.001). It was interesting to note that 96% of children and 76% of adults with UTI had sIgA level significantly above that of +/- 2SD of respective controls (287 +/- 99 and 80 +/- 48 ug/ml respectively). On culturing the urine obtained from these children the colonies identified were E. coil about 46%, Klebsiella about 24% and Pseudomonas about 24%. The sIgA antibody from urine samples assessed by indirect immunoflourescense. specifically reacted with the respective organism. CONCLUSION: Taken together the results show that the presence of sIgA not only correlated with the UTI in children as well in adults but sIgA seems to be directed to the infective agent and can also be used to identify the type of infection. Thus measurement of urine antibody levels may provide an alternative marker of host responses to infection, which can be used either as a simple screening test or could be useful to assist alongwith other tests in establishing a diagnosis.