Surinder Singh

SNP–SNP interactions within APOE gene influence plasma lipids in postmenopausal osteoporosis

Biological revelations have gradually started clearing the complex relationships of bones with lipids. Studies have shown that lipid proWles are related to bone mass, bone fragility and fracture risk [1, 2]. It has been observed that oxidized lipids inhibit mineralization of bone, induce osteoblastic diVerentiation in vascular cells, and hyperlipidemia reduces bone density in mice [3]. To understand the genetic link between lipids and osteoporosis risk, apolipoprotein E gene (APOE) is a potential candidate because of its vital contribution to both lipid and vitamin K metabolism. APOE is located on the chromosome 19q13.2 and has three codominant alleles i.e. APOE2, APOE3 and APOE4. It serves as a ligand for receptor-mediated uptake of lipoprotein particles, which are the main carriers of vitamin K, and availability of vitamin K is an essential step for the carboxylation of glutamic acid residues of osteocalcin, an important bone protein. A study has shown that decrease in bone mineral density (BMD) in hemodialysis patients is correlated with hyperlipidemia, and the risk of fractures in such patients varies according to APOE genotypes [4]. Some preliminary Wndings suggest that APOE contributes to the variation in cholesterol increase with menopause [5], and APOE-E4-negative subjects respond better to hormone replacement therapy (HRT) for lowering lipid levels than APOE-E4-positive subjects [6]. However, in what way, genomic changes within APOE inXuence lipids for the risk of osteoporosis remains unclear. The impact of individual APOE SNP or allelic variant on lipid levels may vary if another nearby SNP is also participating or in linkage disequilibrium with another functional SNP. Studies comprising single SNP would have overlooked coordinated eVects of such SNPs. The present study explored the SNP–SNP interactions of four pertinent APOE SNPs (rs440446, rs769450, rs429358 and rs7412) as the genetic mediators of lipids in postmenopausal osteoporotic women of Punjab Province of Northwest India. Out of 366 randomly selected postmenopausal women who were not using HRT and were aged between 45 and 70 (Mean age 53.2 § 9.2), 154 osteoporotic women (71-lumbar spine and 83-femoral neck cases) were enrolled after veriWcation of the fractures from original radiographs and validation by DEXA (dual energy X-ray absorptiometry) unless they met the exclusion criteria: hyperlipidemia, irregular cycles or premature ovarian failure (menopause < 38 years), women with surgical menopause, any systemic disease or receiving medications known to inXuence calcium metabolism. DEXA veriWed population-based 83 postmenopausal women were enrolled as controls, according to a random-digit invitation from residents of the same geographic suburban and rural areas from Northwest India with three main exclusion criteria: hyperlipidemia, any fracture before or after menopause or any anti-osteoporotic treatment. Detailed information on age, medical history, years since menopause and gynecological history was obtained from these subjects. Height and weight were measured, and BMI (kg/m) was calculated. In the present study, none of the subjects were found to be alcohol users or smokers. Bone mineral density was measured by DEXA using Hologic QRR 4500 (Hologic Inc. Waltham, MA, M. Singh (&) · P. Singh Molecular Genetics Laboratory, Department of Human Biology, Punjabi University, Patiala, Punjab 147002, India e-mail: singh_m12@rediffmail.com

A susceptible haplotype within APOE gene influences BMD and intensifies the osteoporosis risk in postmenopausal women of Northwest India

BACKGROUNDnThe association of apolipoprotein E (APOE) genotypes with bone mineral density (BMD) and risk of osteoporosis have remained unclear. The influence of APOE gene polymorphisms on BMD as genetic mediators of osteoporosis risk needs to be explored in Indian postmenopausal females where this disease is rising rampantly.nnnMETHODS AND RESULTSnThe present study investigated the role and relevance of four pertinent APOE single nucleotide polymorphisms: 5UTR G/C (rs440446), Int2 G/A (rs769450), Exon4 T/C (rs429358), Exon4C/T (rs7412) in DEXA verified 133 osteoporotic, 57 osteopenic and 83 normal postmenopausal females of India, who were not taking hormone replacement therapy. Minor allele frequencies of rs440446 and rs429358 were higher in osteoporotic females (0.31, 0.18) than osteopenic (0.29, 0.15) and females having normal bone mass (0.16, 0.07). Disease association analysis revealed a susceptibility haplotype CGTC (in order of rs440446, rs769450, rs429358, rs7412) and the carriers of this haplotype has higher risk of osteopenia (OR 3.53, 95% CI 1.21-11.0, P=0.017) and osteoporosis (OR 3.61, 95% CI 1.53-9.48, P=0.002) after adjusting the confounding effect of age, BMI and years since menopause. Females who possess either one copy or two copies of the haplotype have lesser BMD values of lumbar spine (0.88 and 0.85 g/cm(2)) and femoral neck (0.84 and 0.82 g/cm(2)) than those females who possess zero copy (0.9 and 0.87 g/cm(2), respectively).nnnCONCLUSIONSnThe present study exposed a susceptibility haplotype CGTC, within APOE gene, which was found to be associated with BMD and risk of osteopenia and osteoporosis in postmenopausal females of India.

A haplotype derived from the common variants at the −1997G/T and Sp1 binding site of the COL1A1 gene influences risk of postmenopausal osteoporosis in India

The aim of the present study was to investigate the association between Collagen 1 alpha 1 (COL1A1) polymorphism and osteoporosis in DEXA verified 349 (145 osteoporotic, 87 osteopenic and 117 normal) postmenopausal women of India, who were not taking hormone replacement therapy. Two single-nucleotide polymorphisms (SNPs), that is, −1997G/T (rs1107946) and +1245G/T (rs1800012, Sp1) of the COL1A1 gene, were analyzed. Minor allele frequencies of rs1107946 and rs1800012 were 0.15 and 0.20 in osteoporotic women, 0.18 and 0.18 in osteopenic and 0.20 and 0.17 in women having normal bone mass. An allele dose effect with BMD of lumbar spine has been exhibited by major allele G of rs1107946 (GG: 0.86xa0g/cm2, GT: 0.91xa0g/cm2 and TT: 0.93xa0g/cm2) and minor allele T of rs1800012 (GG: 0.91xa0g/cm2, GT: 0.87xa0g/cm2 and TT: 0.81xa0g/cm2). Disease association analysis revealed a haplotype GT that confers approximately threefold higher risk of osteoporosis in the carriers (OR 3.12, 95% CI 1.24–8.88, Pxa0=xa00.008) after adjusting the confounding effect of age, BMI and years since menopause. These results suggest that GT haplotype of COL1A1 gene is associated with a higher risk of postmenopausal osteoporosis in Northwest Indian women.

Vitamin D receptor (VDR) gene polymorphism influences the risk of osteoporosis in postmenopausal women of Northwest India

SummaryThe influence of VDR gene for the risk of osteoporosis has remained inconclusive. VDR gene polymorphism in relation to BMD in postmenopausal women of Northwest India revealed a susceptibility haplotype AGT. Possession of this haplotype exacerbates the risk of osteoporosis by 2.8 times, which manifests in recessive mode of inheritance.PurposeThe purpose of this study is to understand the influence of coordinated effect of various single nucleotide polymorphisms (SNPs) within vitamin D receptor (VDR) gene for the risk of osteoporosis, which has remained undefined so far.MethodsFour pertinent SNPs of VDR gene, i.e., rs2228570, rs1544410, rs17879735, and rs731236 were examined with polymerase chain reaction–restriction fragment length polymorphism in dual energy X-ray absorptiometry verified 188 osteoporotics, 115 osteopenics, and 147 normal postmenopausal women of Northwest India.ResultsMinor allele ‘T’ of rs2228570 showed significant influence for the risk of osteoporosis (OR 1.60, 95%CI 1.16–2.20, Pu2009=u20090.004) and also in dominant (OR 2.32, 95%CI 1.47–3.64, Pu2009=u20090.0006) and additive model (OR 2.41, 95%CI 1.49–3.87, Pu2009=u20090.0006) after Bonferroni correction. Minor allele (T) of rs2228570 showed an allele dose effect with BMD of L1-L4 (Pu2009=u20090.009) and FN (Pu2009=u20090.036). Disease association analysis exposed a susceptibility haplotype AGT which influences the risk of osteopenia (OR 2.04, 95%CI 1.03–4.08, Pu2009=u20090.036) and osteoporosis (OR 2.90, 95%CI 1.61–5.38, Pu2009=u20090.00005) after adjusting the effects of age, BMI and years since menopause. This haplotype is significantly associated with BMDs at lumbar spine (Pu2009=u20090.0001) and femoral neck (Pu2009=u20090.016).ConclusionIn-depth analysis of this haplotype with other methods of Wald statistics and Akaike information criterion confirmed that carriers of each unit of this haplotype AGT increases the risk of osteoporosis by a factor of 2.80u2009±u20090.34 (βu2009±u2009SE) which manifests (Pu2009=u20090.1u2009×u200910−6) in its recessive mode of inheritance.

A susceptibility haplotype within the endothelial nitric oxide synthase gene influences bone mineral density in hypertensive women

The influence of the coordinated effect of various single-nucleotide polymorphisms (SNPs) within the endothelial nitric oxide synthase (eNOS) gene on the risk of osteoporosis in hypertension has remained undetermined. Four pertinent SNPs of the eNOS gene, rs2070774, rs1799983, rs1800780 and rs3918181, were examined for the risk of osteoporosis in 313 hypertensive postmenopausal women in Northwest India. All the hypertensive women were verified with dual energy X-ray absorptiometry and categorized as 150 with osteoporosis and 163 without osteoporosis. The minor allele (T) of rs1799983 exerts a statistically significant risk for osteoporosis both in dominant [odds ratio (OR) 3.71, 95xa0% confidence interval (CI) 2.12–6.49, Pxa0<xa00.001] and recessive mode (OR 5.75, 95xa0% CI 1.24–26.69, Pxa0=xa00.036) after Bonferroni correction. Bone mineral density (BMD) values (corrected for the effects of risk variables) according to eNOS SNP genotypes revealed a significant association with rs1799983 at both the lumbar spine (Pxa0=xa00.001) and femoral neck (Pxa0=xa00.023). Risk association analyses revealed a susceptibility haplotype TTAG which influences the risk of osteoporosis (OR 2.02, 95xa0% CI 1.05–3.39, Pxa0=xa00.042) in hypertension after adjusting for the effects of risk factors. Furthermore, this haplotype was significantly associated with BMD at the lumbar spine (Pxa0=xa00.029) and femoral neck (Pxa0=xa00.021) in a dose-dependent manner. The results suggest that possession of the TTAG haplotype of the eNOS gene may increase the risk of osteoporosis two-fold in hypertensive postmenopausal women in Northwest India.

Extraction titrimetric determination of copper.

Lead xanthate undergoes a quantitative, stoichiometric and fast exchange reaction with copper(II), even near the equivalence point, making it a suitable reagent for an extraction titration of copper. Many other metal ions do not participate in exchange reactions with lead xanthate. making this method selective for copper.