INS VNTR Polymorphisms Research Articles

Insulin gene VNTR class III allele is a risk factor for insulin resistance in Kashmiri women with polycystic ovary syndrome

PurposePCOS is a complex heterogeneous multifactorial endocrine disorder characterized by anovulation, hyperandrogenism and polycystic ovaries. PCOS affects 5–10% women of reproductive age is also suggested to increase the risk of insulin resistance, cardiovascular disorders and T2DM. The aim of this study was to assess the possible association of INS VNTR polymorphism with PCOS and their phenotype-genotype interactions in Kashmiri women with PCOS. MethodsA total of 349 subjects including 249 cases and 100 age-matched healthy controls were recruited in the study. DNA was extracted from peripheral venous blood from all subjects and association analysis was done by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique. Genotype-phenotype correlation analysis was performed to evaluate the association of INS VNTR polymorphism with phenotypic features. ResultsAllele frequency (OR = 0.86, C.I. 0.48–1.54, P = .61) and genotype distribution (P = .79, χ2 = 0.46) showed no significant association between INS VNTR and PCOS. The dominant (OR = 1.21, χ2 = 0.026, P = .871), recessive (OR = 0.83, χ2 = 0.35, P = .549) and heterozygote vs. homozygote (OR = 1.25, χ2 = 0.45, P = .50) genotype model analysis further supported this finding. Genotype-phenotype correlation analysis showed class III allele carrier (I/III + III/III genotype) was significantly associated with increased weight (P = .044), BMI (P = .021) insulin (F) (P = .049) and HOMA IR (P = .02) in recessive model in PCOS women. ConclusionsINS VNTR polymorphism does not increase the risk of PCOS. However, INS VNTR class III allele is associated with elevated BMI and HOMA IR, suggesting its role in the pathogenesis of insulin resistance and obesity in Kashmiri women with PCOS.

Association of INS VNTR polymorphisms with polycystic ovary syndrome (PCOS) in south Indian population

Association of INS VNTR polymorphisms with polycystic ovary syndrome (PCOS) in south Indian population

P055 : INS-VNTR POLYMORPHISM AND GENE EXPRESSION IN TYPE 1, TYPE 2 AND GESTATIONAL IN BRAZILIAN DIABETES MELLITUS PATIENTS

Besides HLA class II alleles, the variable number of tandem repeat polymorphism located at 5’ of the insulin gene (INS-VNTR) at chromosome 11p15.5 is the second most important susceptibility region to type 1 diabetes (T1D). INS-VNTR alleles (ACAGGGGTGTGGGG) is divided into class I (30–60 repeats), class II (60–120) and class III (120–160). INS-VNTR class I allele transcribe lower thymic INS than class III allele favoring escape of auto-reactive thymocytes from negative selection. Since insulin resistance is a major shared feature among diabetes patients and since there no studies evaluating these markers in the major types of the disease, we characterized the INS-VNTR polymorphisms (348patients/339controls) and the differential mRNA/microRNA (40 patients) expression in T1D, gestational (GDM) and type 2 (T2D) Brazilian diabetic patients. INS-VNTR was performed using −23HphI PCR-RFLP, and gene expression profiles were performed using Agilent microarrays. INS-VNTR I:I genotype was associated with susceptibility, whereas the III:III genotype conferred protection against T1D development, and I:III genotype was associated to T2D. The multivariate analyses results showed that the I:I genotype was associated with age and dyslipidemia only in T1D patients. Considering all types of diabetes, the differential gene expression profiles (mRNA/miRNA) for INS-VNTR I:I genotype were distinct from those observed for the III:III genotype. The miRNA expression profile according to INS-VNTR polymorphism yielded 31 differentially expressed transcripts. Networks of functional miRNA target, presenting possible INS interactions, showed 6 upregulated ( I:I genotype), 10 upregulated (I:III genotype), and 6 upregulated and 8 downregulated for III:III genotype. Conclusion: This is the first study of INS-VNTR polymorphisms evaluating the three major types of diabetes in the same population group. Moreover, the INS-VNTR genotypes influenced gene and miRNAs expression profiles.

Lack of association of INS VNTR polymorphism with polycystic ovary syndrome: a meta-analysis

An association between the INS VNTR polymorphisms and polycystic ovary syndrome (PCOS) susceptibility has been reported in previous studies, but the results were inconsistent. This study was conducted to explore this association using meta-analysis. PubMed, Embase, and China National Knowledge Infrastructure (CNKI) were searched according to predefined criteria for all relevant studies published up to August 2013. Four genetic models, together with odds ratios (ORs) and 95 % confidence intervals (CI), were calculated. Subgroup analyses were performed by ethnicity, anovulatory PCOS, and Hardy-Weinberg equilibrium (HWE) in the controls. In total, 13 case-control studies, including 1,767 cases and 4,108 controls, were included. No significant association was detected in overall population in all models (III/III vs. I/I: OR = 1.200, 95%CI = 0.866-1.664, P=0.277; I/III vs. I/I: OR = 1.041, 95%CI = 0.880-1.232, P=0.637; III/III + I/III vs. I/I: OR = 1.191, 95%CI = 0.912-1.554, P=0.199; III/III vs. I/III + I/I: OR = 1.100, 95%CI = 0.816-1.484, P=0.531), the same as in Caucasian and Asian populations. When the studies were limited to conform to HWE, the results remained persistent and robust. The anovulation subgroup showed significantly elevated risk in the I/III vs. I/I (OR = 1.460, 95%CI = 1.017-2.095, P=0.040). This meta-analysis revealed no significant association between INS VNTR polymorphisms and the risk of PCOS in the overall population, while it supported that variance may be associated with susceptibility to PCOS with anovulation. Further confirmation is needed from more well-designed and larger studies.

Influence of Polymorphisms in Genes Encoding for Insulin-Like Growth Factor (IGF)-I, Insulin, and IGF-Binding Protein (IGFBP)-3 on IGF-I, IGF-II, and IGFBP-3 Levels in Umbilical Cord Plasma

Background/Aims: In postnatal life, polymorphisms in the promoter region of IGFBP3 were associated with insulin-like growth factor binding protein (IGFBP)-3 plasma levels. Whether these associations exist in utero has not been studied yet. Polymorphisms in the IGF1 promoter (polymorphic CA-repeat) and the insulin gene variable number tandem repeats locus (INS VNTR) are further polymorphisms of interest, because associations with birth weight have been reported. We aimed to investigate associations between polymorphisms in the promoter regions of IGF1 (wild type 192 bp), IGFBP3 (rs2854744; rs13241830), and INS VNTR (rs689) with cord plasma levels of IGF-I, IGF-II, and IGFBP-3. Methods: We measured IGF-I, IGF-II, and IGFBP-3 concentrations in cord blood from 677 neonates and genotyped the selected polymorphisms. Results: Carriers of the minor allele of both polymorphisms in the IGFBP3 gene had, on average, 4–5% lower IGFBP-3 levels per copy of the respective minor allele (p = 0.002 and p = 0.028) when compared to wild type carriers. The IGF1 promoter and the INS VNTR polymorphisms were not associated with IGF-I, IGF-II, or IGFBP-3 levels. Conclusions: Our data show associations of cord plasma IGFBP-3 levels and the IGFBP3 gene variants but not of IGF1 promoter and INS VNTR polymorphisms with IGF-I, IGF-II, or IGFBP-3 levels in utero.

Association between INS-VNTR polymorphism and polycystic ovary syndrome in a Korean population

Polycystic ovary syndrome (PCOS) is a common disorder in women of reproductive ages. But its etiology is not fully understood yet. Variability in the number of tandem repeats of the insulin gene (INS-VNTR) is known to associate with PCOS, and it is associated with an increased risk of diabetes mellitus and other cardiovascular diseases. The aim of our study was to analyze an association between the INS-VNTR polymorphism and PCOS in a Korean population. The −23/Hph I polymorphism was used as a surrogate marker for INS-VNTR polymorphism and a total of 218 PCOS patient and 141 control DNAs were analyzed by restriction fragment length polymorphism method. Statistical analysis of genotyping results were performed using HapAnalyzer. χ2 test and logistic regression were used to analyze the association between two groups. A p value <0.05 was considered statistically significant. The frequencies of A/A and A/T genotypes indicated a similar change between PCOS patients and controls. In conclusion, there was no association between PCOS and INS-VNTR polymorphism (p = 0.0544, odds ratio = 1.69). Our present data demonstrate that INS-VNTR polymorphism is not related with PCOS in Korean women. Thus, it is suggested that INS-VNTR polymorphism is not a key factor in the etiology and the pathogenesis of PCOS in a Korean population.

No association of the insulin gene VNTR polymorphism with polycystic ovary syndrome in a Han Chinese population

BackgroundPolycystic ovary syndrome (PCOS) is a common endocrine disorder associated with an increased risk of type II diabetes mellitus. The results of previous research about the association of the VNTR polymorphism in 5-prime flanking region of the insulin (INS) gene with PCOS have been inconsistent. The present study was to investigate the association of the INS-VNTR polymorphism with PCOS in a Han Chinese population.MethodsThe -23/HphI polymorphism as a surrogate marker of the INS-VNTR length polymorphism was genotyped by polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP) in 216 PCOS patients and 192 non-PCOS women as a control group. Allelic and genotypic frequencies were compared between patients and controls, and these results were analyzed in respect to clinical test data.ResultsNo significant differences were observed between the cases and controls groups either in allele (P = 0.996) or genotype (P = 0.802) frequencies of INS-VNTR polymorphism; Regarding anthropometric data and hormone levels, there were no significant differences between INS-VNTR genotypes in the PCOS group, as well as in the non-PCOS group.ConclusionThe present study demonstrated for the first time that the INS-VNTR polymorphism is not a key risk factor for sporadic PCOS in the Han Chinese women. Further studies are needed to give a global view of this polymorphism in pathogenesis of PCOS in a large-scale sample, family-based association design or well-defined subgroups of PCOS.

VNTR polymorphism of the insulin gene and childhood overweight in a general population.

The VNTR polymorphism 5' of the insulin gene has been related to obesity in a previous study on children with early onset of severe obesity. Our purpose was to analyze the association between this polymorphism and adiposity variability in an unselected population of children and adolescents in northern France. In 293 nuclear families from the Fleurbaix Laventie Ville Santé study, we genotyped the INS VNTR polymorphism in 431 children and adolescents (8 to 18 years of age) and their parents. Overweight was defined according to the international definition in both children and adults. A transmission disequilibrium test in families with an overweight offspring was performed. The prevalence of overweight was compared according to genotype. The effect of the genotype on BMI and waist circumference was tested with a linear regression model, adjusting for age, gender, and Tanner stage. There was an undertransmission of class III alleles from heterozygous parents to their overweight offspring (p < 0.002). Overweight was associated with class I alleles in children and adolescents (12% I/I, I/III vs. 3% III/III; p < 0.08). Those with a class III/III genotype had a 1 kg/m(2) lower mean BMI (p = 0.04) and 3 cm lower waist circumference (p = 0.02) than those bearing one or two class I alleles. No association of adiposity or obesity with class I alleles was found in parents. INS VNTR polymorphism seems to contribute to differences in adiposity level in the general population of children and adolescents.

Polymorphism of INS VNTR is associated with glutamic acid decarboxylase antibodies and postprandial C-peptide in patients with diabetes onset after 35 years of age.

Variability in the number of tandem repeats of the insulin gene (INS VNTR) is probably involved in the genetic regulation of insulin secretion. The aim of this study was to investigate the association of INS VNTR polymorphism with the presence of glutamic acid decarboxylase antibodies (GADA) and C-peptide levels in patients with the onset of diabetes after 35 years of age. We investigated 117 patients, median of age 63 (range 40-83) years, median of diabetes duration 8 (range 1-30) years; 31 GADA-positive and 86 GADA-negative subjects. INS VNTR polymorphism was typed indirectly using - 23HphI (T/A) polymorphism, which is in complete linkage disequilibrium with INS VNTR. The I/I, I/III and III/III genotypes were found in 22 (71 %), 8 (26 %), 1 (3 %) GADA-positive individuals and in 39 (45 %), 35 (41 %), 12 (14 %) GADA-negative individuals, respectively. The Class I allele and the genotype I/I were significantly associated with the presence of GADA (OR=2.72, CI 95 %=1.29-5.73 and OR=2.95, CI 95 %=1.22-7.13). The presence of Class III allele was significantly associated with a higher level of postprandial C-peptide in GADA-positive subjects, even when regarding the duration of diabetes. Our results of INS VNTR polymorphism in patients with the onset of diabetes after 35 years of age confirm the association of Class I INS VNTR with autoimmune diabetes and the protective effect of Class III INS VNTR on the insulin secretion in GADA-positive subjects.

Association of insulin gene VNTR polymorphism with polycystic ovary syndrome.

Variability in the number of tandem repeats of the insulin gene (INS VNTR) is known to influence several phenotypes, including polycystic ovary syndrome (PCOS), diabetes mellitus type 1, diabetes mellitus type 2, and birth weight. The presence of the class III allele of INS VNTR has been reported to be protective in diabetes mellitus type 1, but in contrary it increases the disease risk of PCOS and diabetes mellitus type 2. PCOS is a very common endocrinopathy in women of reproductive age. The etiology of PCOS is uncertain, but family history of this syndrome suggests a major genetic cause. The aim of this pilot study was to investigate the possible association of INS VNTR polymorphism with PCOS in Czech women. In PCOS, significantly higher WHR, BMI, G(0), G(180), I(30), Cp(0), Cp(30), Cp(60), AUC-I, AUC-Cp, and insulinogenic index and significantly lower AUC-G/AUC-I were found. No significant differences in INS VNTR genotype, phenotype, or allele frequencies were found between PCOS and controls. In spite of several differences in anthropometric and biochemical parameters (abdominal fat localization, increased beta-cell function, and lower insulin sensitivity in PCOS women), no effect of INS VNTR polymorphism was found on insulin secretion, insulin action, or any other screened parameter.

Nonisotopic, PCR-based method for 5' insulin gene VNTR allele class assignment.

The identification of genes that contribute to the susceptibility to polygenic human diseases is a laborious task that requires the analysis of multiple polymorphic markers in search of variants putatively associated with the disease phenotype. Besides, it is often necessary to study large cohorts of affected and unaffected individuals to achieve acceptable levels of significance. Once preliminary evidence of association has been obtained, a confirmation of results in independent samples from different populations is also required. If this type of research is to be cost effective, genotyping methods must be straightforward, easily scalable to large numbers of samples and results must be obtained in a short time. A clear example of a polygenic disease is insulin-dependent (type 1) diabetes mellitus, an autoimmune disorder in which T-cell-mediated destruction of the insulin-producing pancreatic beta cell occurs, and is currently the object of intense genetic investigation in the search for susceptibility genes. During the last few years, several groups have focused their attention towards the allelic variation at the polymorphic region situated 5' upstream of the human insulin gene (INS) on chromosome 11p15.5, which arises from a variable number of tandem repetitions (VNTR) of a 14–15 bp oligonucleotide with the consensus sequence 5' -ACAGGGGT(G/C)(T/C)GGGG-3'. Based on Southern blot analysis (2), these VNTR alleles fall into three distinctive size classes according to the number of repeats of the oligonucleotide (classes I, II and III average 570, 1200 and 2200 bp, respectively). It has been shown that in Caucasians, class I alleles are positively associated with type I diabetes, class II alleles are rare and class III alleles are dominantly protective. More recently, a PCR-based method has been used for class I allele subtyping, and allele-specific effects on disease susceptibility have been described (3,8). However, all reported methods for PCR amplification of the INS-VNTR polymorphism are limited to the analysis of class I alleles and fail to amplify either class II or class III alleles (3,5,6) probably because of (i) the increased length of these alleles, (ii) the strong secondary structure derived from the high GC-rich content of the template and (iii) the high frequency of class I/class III heterozygous in which the amplification of the short class I alleles is strongly favored over the long class III alleles. In fact, long alleles may only be detected by traditional Southern blot hybridization with an internal radioactive-specific probe, and this is a laborious, time-consuming technique unsuitable for large population studies requiring large amounts of purified, high molecular weight DNA, which is not always available. The analysis of diallelic markers in the 5' region of the insulin gene, especially the -23/HphI RFLP, has proven useful as an alternative to Southern blot typing of the INS-VNTR, at least in populations of European descent, in which the VNTR locus may be considered diallelic (class I/class III) and tight linkage disequilibrium between -23/ HphI and VNTR alleles has been demonstrated (4). However, it might be necessary to investigate both the frequency of class II alleles and the degree of linkage disequilibrium between the proposed marker and the INS-VNTR locus when a non-Caucasian population is investigated for the first time. We report a set of PCR conditions that allow efficient amplification of INSVNTR alleles of all classes, offering a nonradioactive and rapid alternative to the conventional, tedious Southern blot technique for reliable VNTR typing in large samples from any population. Genomic DNA was isolated from blood samples using NucleoSpin® Blood kit (Macherey-Nagel, Duren, Germany). DNA (150 ng) was subjected to PCR amplification with flanking primers (5) (5FP1: 5' -CACCTTGGCCCATCCATGGCGGCATC-3' and 5FP2: 5'-CTCCAGGAGAGCAAAGCCCTCACCTG-3', which anneal 21 and 64 bp beyond the 5' and 3' ends of the VNTR, respectively). The downstream primer is labeled with fluorochrome Cy5 on its 5' end, so the pair of primers is routinely used for INSVNTR class I subtyping on an ALFexpressTM automatic DNA sequencing apparatus (Amersham Pharmacia Biotech, Uppsala, Sweden). PCR amplifications were performed in 100-mL reactions containing NH4-based PCR buffer (16 mM ammonium sulfate, 67 mM TrisHCl, pH 8.8, and 0.01% Tween® 20), 2 mM MgCl2, 10% glycerol, 0.4 mM Benchmarks