MTHFR 677C>T polymorphism and the risk of breast cancer: evidence from an original study and pooled data for 28031 cases and 31880 controls

Background: Methylenetetrahydrofolate Reductase (MTHFR) acts at an important metabolic point in the regulation of cellular methylation reaction. It assists in the conversion of 5, 10-methylenetetrahydrofolate to 5-methyltetrahydrofolate. The latter aids in remethylation of homocysteine to de novo methionine that is required for DNA synthesis. The objective of this study was to examine the effect of MTHFR 677 C>T polymorphism on the risk of breast cancer in the Indian sub-continent. Methods and Results: We genotyped 677 C>T locus in 1096 individuals that were classified into cases (N = 588) and controls (N = 508). Genotype data were analyzed using chi-square test. No significant difference was observed in the distribution of genotypes between cases and controls in north Indian (P = 0.932), south Indian (P = 0.865), and pooled data (P = 0.680). To develop a consensus regarding the impact of 677C>T polymorphism on breast cancer risk, we also conducted a meta-analysis on 28031 cases and 31880 controls that were pooled from sixty one studies. The overall summary estimate upon meta-analysis suggested no significant correlation between the 677C>T substitution and breast cancer in the dominant model (Fixed effect model: OR = 0.97, P = 0.072, Random effects model: OR = 0.96, P = 0.084) or the recessive model (Fixed effect model: OR = 1.05, P = 0.089; Random effects model: OR = 1.08, P = 0.067). Conclusion: 677 C>T substitution does not affect breast cancer risk in the Indo-European and Dravidian populations of India. Analysis on pooled data further ruled out association between the 677 C>T polymorphism and breast cancer. Therefore, 677 C>T substitution does not appear to influence the risk of breast cancer

Strong impact of TGF-β1 gene polymorphisms on breast cancer risk in Indian women: a case-control and population-based study

Introduction: TGF-β1 is a multi-functional cytokine that plays an important role in breast carcinogenesis. Critical role of TGF-β1 signaling in breast cancer progression is well documented. Some TGF-β1 polymorphisms influence its expression; however, their impact on breast cancer risk is not clear. Methods: We analyzed 1222 samples in a candidate gene-based genetic association study on two distantly located and ethnically divergent case-control groups of Indian women, followed by a population-based genetic epidemiology study analyzing these polymorphisms in other Indian populations. The c.29C>T (Pro10Leu, rs1982073 or rs1800470) and c.74G>C (Arg25Pro, rs1800471) polymorphisms in the TGF-β1 gene were analyzed using direct DNA sequencing, and peripheral level of TGF-β1 were measured by ELISA. Results: c.29C>T substitution increased breast cancer risk, irrespective of ethnicity and menopausal status. On the other hand, c.74G>C substitution reduced breast cancer risk significantly in the north Indian group (p  =  0.0005) and only in the pre-menopausal women. The protective effect of c.74G>C polymorphism may be ethnicity-specific, as no association was seen in south Indian group. The polymorphic status of c.29C>T was comparable among Indo-Europeans, Dravidians and Tibeto-Burmans. Interestingly, we found that Tibeto-Burmans lack polymorphism at c.74G>C locus as true for the Chinese populations. However, the Brahmins of Nepal (Indo-Europeans) showed polymorphism in 2.08% of alleles. Mean TGF-β1 was significantly elevated in patients in comparison to controls (p<0.001). Conclusion: c.29C>T and c.74G>C polymorphisms in the TGF-β1 gene significantly affect breast cancer risk, which correlates with elevated TGF-β1 level in the patients. The c.29C>T locus is polymorphic across ethnically different populations, but c.74G>C locus is monomorphic in Tibeto-Burmans and polymorphic in other Indian populations

Association of IL-4 and IL-1RN (receptor antagonist) gene variants and the risk of type 2 diabetes mellitus: A study in the north Indian population

Background: Inflammation is a key event closely associated with the pathophysiology of type 2 diabetes mellitus (T2DM). Association of genetic polymorphisms of inflammatory cytokines with T2DM is largely unknown. Our objective was to investigate the relationship of polymorphism of IL-1RN and IL-4, two important biomarkers of inflammation, with the risk of T2DM. Setting and Design: We recruited 120 clinically diagnosed T2DM patients and 150 normal healthy controls for this study in order to evaluate the nature of polymorphisms of IL-1RN and IL-4. Materials and Methods: Genomic DNA was isolated from the blood of all subjects, and the variable number of tandem repeat (VNTR) polymorphisms of IL-1RN and IL-4 genes was identified by polymerase chain reaction. Statistical Analysis Used: Genotype distribution and allelic frequencies were compared between patients and control group. Means, as well as odds ratios (ORs) with 95% confidence intervals (CI), were calculated using SPSS software (version 11.5). Results: Our study revealed that distribution of both IL-4 and IL-1RN (VNTR) gene polymorphisms were significantly associated with T2DM subjects. We, however, failed to find any association of gene-gene (IL-4 and IL-1RN) interaction with T2DM. Conclusions: Both IL-4 and IL-1RN (VNTR) gene polymorphisms were significantly associated with T2DM subjects. This may suggest that the genetic polymorphisms of IL-4 and IL-1RN genes could serve as susceptibility indicators for T2DM in the Indian population, but the actual mechanism of these associations will require more elaborate investigations. Lack of association of gene-gene (IL-4 and IL-1RN) interaction with T2DM may indicate the independent nature of influence of both these genes on the risk of T2DM

CD3+, CD4+ & CD8+ tumour infiltrating lymphocytes (TILs) are predictors of favourable survival outcome in infiltrating ductal carcinoma of breast

Background & objectives: Tumour infiltrating lymphocytes (TILs) represent the host immune response against cancer cells associated with good or bad prognosis in different tumour types. This study was undertaken to evaluate the significance of CD3+, CD4+ and CD8+ TILs in breast cancer tissues in relation to clinico-pathological variables and survival outcome. Methods: Immunohistochemistry (IHC) was performed with antibodies against CD3, CD4 and CD8 antigens on formalin-fixed paraffin-embedded tissue sections of 150 breast cancer patients. Intratumoural and stromal TIL counting was performed semiquantitatively. Results: The higher CD3+, CD4+ and CD8+ intratumoural and stromal counts showed independent and direct association with good prognosis. The prognostic predictor value of intratumoural counts was higher than stromal counts. The independent associations of intratumoural and stromal counts became more prominent when adjusted with stage and grade, respectively. Among intratumoural counts, the high (++/+++) CD4+ count (OR=3.85, 95% CI=3.28-16.71, P<0.001) showed the highest survival followed by CD3+ (OR=2.70, 95% CI=1.76-8.30, P=0.001) and CD8+ (OR=2.58, 95% CI=1.55-5.86, p0 =0.001) the least when compared to respective low (+) counts. In contrast, among stromal counts, the high CD8+ count (OR=3.13, 95% CI=2.20-9.57, p0 <0.001) showed the highest survival followed by CD4+ (OR=3.02, 95% CI=2.07-8.89, p0 <0.001) and CD3+ (OR=2.45, 95% CI=1.53-6.73, p0 =0.002) the least. Interpretation & conclusions: Our results suggest that intratumoural CD4+ and stromal CD8+ counts by immunohistochemistry may serve as an independent prognosticator for favourable outcome in breast cancer

Does angiotensin-converting enzyme polymorphism have association with symptomatic benign prostatic hyperplasia?

Objectives : The aim of the study was to explore the putative significance of angiotensin-converting enzyme (ACE) insertion/deletion (I/D) polymorphism and its correlation with the lower urinary tract symptoms (LUTS) in benign prostatic hyperplasia (BPH). Materials and Methods : ACE I/D polymorphisms were determined by polymerase chain reaction (PCR) in 200 patients with moderate to severe LUTS due to BPH and 200 patients of same age group without the LUTS having normal size prostate. ACE levels were estimated by spectrophotometer method. Logistic regression models were used to determine the genetic effects using SPSS statistical software (version 12.0). Results and Conclusions : The distribution of genotypes along with allelic frequency and carriage rate did not significantly differ between study and control groups. This study suggests that I/D polymorphisms within the ACE gene are not associated with the presence of LUTS in BPH patients. Future studies in large cohorts are needed that may reveal the spectrum of cellular mechanism mediated by ACE relevant to pathophysiology of BPH and effect of ethnic differences

Esterification of carboxylic acids by acid activated Kaolinite clay

75-78Esterification of carboxylic acid was carried out by using acid activated kaolinite clay. This heterogeneous catalyst, activated kaolinite clay has been found to be a mild solid catalyst for the esterification of carboxylic acid in good yields. The catalyst is recoverable and can be recycled after activation

Vitamin D receptor (FokI, BsmI and TaqI) gene polymorphisms and type 2 diabetes mellitus : A North Indian study

Background : The vitamin D receptor (VDR) gene is a candidate gene for susceptibility to several diseases. Studies on association between VDR polymorphisms and risk of type 2 diabetes (T2DM) in different ethnic populations are yet inconclusive. Aims : This study was conducted to evaluate association between VDR polymorphisms and genetic susceptibility to T2DM in the north Indian population. Settings and Design : One hundred clinically diagnosed T2DM patients and 160 healthy controls from the north Indian population were recruited for genetic association study. Materials and Methods : Genomic DNA was extracted from blood and genotyped for the single nucleotide polymorphism SNPs of FokI (T/C) [rs2228570], BsmI (A/G) [rs1544410] and TaqI (C/T) [rs731236] by polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP) analysis. Statistical Analysis Used : Genotype distribution and allelic frequencies were compared between patients and controls. Mean values and odds ratios (ORs) with 95% confidence interval (CI) were calculated using SPSS software (version 15.0). Results : The genotype distribution, allele and haplotype frequencies of VDR polymorphism did not differ significantly between patients and controls. Mean age and waist-hip ratio of patients were found to be associated with VDR polymorphism. Combination studies showed FFBbtt increased the risk of T2DM in north Indians. Conclusions : Our data suggest that VDR gene polymorphism in combination of genotypes is associated with the risk of T2DM and thus requires further studies as a probable genetic risk marker for T2DM

Role of ethnic variations in TNF-&#945; and TNF-&#946; polymorphisms and risk of breast cancer in India

TNF-&#945; and -&#946;, the multi-functional pro-inflammatory cytokines, are known to play important roles in both tumor progression and destruction based on their concentrations. Growth factors and various stimuli such as cytokines regulate proliferation of the breast epithelial cells. Therefore, the polymorphisms in the genes encoding these signaling molecules could affect the risk of breast cancer. We have investigated selected genetic polymorphisms in TNF-&#945; promoter (rs1800629, −308 G&#62;A and rs361525, −238 G&#62;A) and TNF-&#946; intron 1 (rs909253, +252 A&#62;G) in ethnically two different case–control groups from India. The study included 200 cases and 200 controls from an Indo-European (North Indian) group, and 265 cases and 237 controls from a Dravidian (South Indian) group. Genotyping of a total of 902 individuals was done by direct DNA sequencing. None of the polymorphisms showed significant association with breast cancer in the Indo-European group; however, all the three polymorphisms showed strong association with breast cancer in the Dravidian group. Further, sub-group analysis in the Indo-European group showed no significant difference between pre-menopausal cases and controls or between post-menopausal cases and controls at any of the loci analyzed. However, all the polymorphisms in the Dravidian group were significantly associated with pre-menopausal but not with post-menopausal breast cancer. In conclusion, TNF-&#945; and -&#946; polymorphisms are strongly associated with breast cancer in the Dravidian but not in the Indo-European group

Electroporation of Mouse Follicles, Oocytes and Embryos without Manipulating Zona Pellucida

Electroporation is an effective technique of transfection, but its efficiency depends on the optimization of various parameters. In this study, a simplified and efficient method of gene manipulation was standardized through electroporation to introduce a recombinant green fluorescent protein (GFP) construct as well as RNA-inhibitors in intact mouse follicles, oocytes and early embryos, where various electroporation parameters like voltage, pulse number and pulse duration were standardized. Electroporated preantral follicles were cultured further in vitro to obtain mature oocytes and their viability was confirmed through the localization of a known oocyte maturation marker, ovastacin, which appeared to be similar to the in vivo-derived mature oocytes and thus proved the viability of the in vitro matured oocytes after electroporation. Standardized electroporation parameters, i.e., three pulses of 30 V for 1 millisecond at an interval of 10 s, were applied to manipulate the expression of mmu-miR-26a in preantral follicles through the electroporation of miR inhibitors and mimics. The TUNEL apoptosis assay confirmed the normal development of the electroporated embryos when compared to the normal embryos. Conclusively, for the first time, this study demonstrated the delivery of exogenous oligonucleotides into intact mouse follicles, oocytes and embryos without hampering their zona pellucida (ZP) and further development