Swayam Prakash

Association of HLA‐G promoter and 14‐bp insertion–deletion variants with acute allograft rejection and end‐stage renal disease

The aim of this study was to investigate the HLA-G 14-bp insertion/deletion (I/D) polymorphism among end-stage renal disease (ESRD) patients. Cytomegalovirus (CMV) infection, acute allograft rejection (AR) and overall survival after renal transplantation was investigated in 300 ESRD patients and 302 age, sex and ethnicity-matched controls. Sequencing was performed to evaluate the impact of HLA-G promoter region single-nucleotide polymorphisms (SNPs) whereas semi-quantitative PCR method was used to determine the probable HLA-G expression pattern among ESRD and AR cases. Further, soluble human leukocyte antigen (HLA)-G (sHLA-G) expression levels were compared in AR vs non-AR cases in the light of HLA-G 14-bp I/D polymorphism. Increased risk was found for 14-bp D/D (deletion-DD) genotype and 14-bp D allele [DD: odds ratio (OR) = 1.46, 95% confidence interval (CI) = 1.03-2.06, P value = 0.0358; D: OR = 1.29, 95% CI = 1.03-1.62, P value = 0.0277], respectively for ESRD and CMV infection (DD: OR = 2.70, 95% CI = 1.45-5.05, P value = 0.0021; D: OR = 1.94, 95% CI = 1.22-3.08, P value = 0.0052). Nearly fourfold (OR = 3.62, 95%CI = 1.61-8.14, p = 0.0039) risk was observed for 14-bp I/I (insertion-II) genotype for AR. Survival analysis showed increased overall survival (OS) (AR or death) for 14-bp D/D genotype. HLA-G promoter region sequencing was carried out among 60 ESRD patients and 100 normal controls which showed increased risk for -964 G>A, -725 C>G/T and -486 A>C SNPs. -964 G>A and -725 C>G/T SNPs showed risk association for AR patients. High level of HLA-G transcripts was observed among non-AR patients. Further soluble HLA-G (sHLA-G) showed increased levels in ESRD patients (mean ± SEM; 62.16 ± 2.43 U/ml) as compared to controls (mean ± SEM; 21.06 ± 3.89 U/ml) (P = <0.0001). The 14-bp I/I, 14-bp I/D and 14-bp D/D genotypes showed significantly higher levels of sHLA-G among non-AR as compared to AR patients.

Vascular endothelial growth factor gene polymorphisms in North Indian patients with end stage renal disease

CONTEXT Vascular endothelial growth factor (VEGF) is involved in the development and differentiation of the vascular system. VEGF is expressed constitutively by epithelial cells from embryonic to adult kidneys and may play a key role in progression of kidney diseases. It is required for the growth and proliferation of glomerular and peritubular endothelial cells. In the kidney VEGF expression is prominently found in glomerular podocytes and in tubular epithelial cells, while VEGF receptors are mainly seen on preglomerular, glomerular, and peritubular endothelial cells. OBJECTIVES We have investigated the role of VEGF gene polymorphisms (-2578C/A,-2549 18 bp I/D, -1154 G/A and +936 C/T) as a susceptibility marker for end stage renal disease (ESRD). PARTICIPANTS AND METHODS We genotyped VEGF gene polymorphism in three hundred patients and three hundred and fifty ethnically matched unrelated healthy controls free from any renal disease. These markers were studied using ARMS-PCR and PCR-RFLP methods. Patients were categorized on the basis of the histo-pathological subtypes into chronic glomerulonephritis (CGN=109), hypertensive nephrosclerosis (HTN=106) and chronic interstitial nephritis (CIN=60). RESULTS VEGF -2578C and -2549D alleles were found to be ESRD causative alleles. It was observed that there was significant differences in the frequencies of the T allele of +936C/T polymorphism among CGN, HTN and CIN respectively. VEGF -1154AA genotype and A allele were associated significantly with CGN. T-G-A-D, T-A-C-I,C-G-A-D,C-A-C-D,C-G-C-I,C-A-A-D and T-G-C-D were seven haplotypes concurred in all the ESRD patients irrespective of underlying disease. While C-G-C-D & C-G-A-I haplotypes showed risk association in CGN & CIN, C-A-C-I was observed to play predisposing role in HTN. CONCLUSION The results highlight the role of studied VEGF polymorphisms in end stage renal disease at large and subsequently in the three primary kidney diseases among the North Indian population.

HLA-G gene expression influenced at allelic level in association with end stage renal disease and acute allograft rejection.

BACKGROUND Human leukocyte antigen (HLA)-G is a non-classical major-histocompatibility complex class-I molecule associated with immunosuppressive function. We have evaluated the impact of HLA-G allele associated with untranslated-region (UTR)-haplotype in end stage renal disease (ESRD) and acute allograft rejection (AR) cases. The mRNA levels of different HLA-G isoforms were evaluated in ESRD and AR cases. Subsequently, the total HLA-G mRNA levels and protein concentration were evaluated against its UTR-haplotype among ESRD and AR cases. METHODOLOGY Sequence based typing of the promoter region was carried-out to evaluate the impact of HLA-G haplotype in 350 ESRD cases and 300 controls. HLA-G gene expression was evaluated at the transcriptional level using semi-quantitative and quantitative PCR, whereas protein concentration was determined by ELISA among both cases and control. RESULTS Increased risk was observed for G*01:01:01:03, G*01:01:02, G*01:06 and G*01:05:N haplotypes while G*01:01:01:01 and G*01:04:01 haplotypes showed a protective effect in ESRD and AR cases. Higher level of soluble HLA-G isoforms (G5 and G6) was observed among ESRD cases. Reduced levels of soluble isoform (G5) and increased levels of membrane bound (G1 and G3) isoforms were found among AR cases, revealing risk association. Decreased HLA-G expression was observed at both mRNA and protein level for G*01:01:01:03 and G*01:05:N haplotypes in ESRD and AR cases. CONCLUSIONS These results suggest that the variation in the expression profile of membrane bound and soluble isoforms may modulate the risk for ESRD and AR. UTR-haplotypes appear to be involved in different HLA-G expression patterns at transcriptional and translational levels.

Interleukin-1 gene cluster variants and recurrent pregnancy loss among North Indian women: retrospective study and meta-analysis

The aim of this study was to determine whether or not interleukin-1 alpha (IL-1a), interleukin-1 beta (IL-1b) and IL-1 receptor antagonist (IL-1RA) polymorphisms are associated with risk of unexplained recurrent pregnancy loss (RPL) among North Indian women. This retrospective case–control study examine 200 well-characterized RPL cases for IL-1 gene cluster variants, determined by restriction fragment length polymorphism-PCR. The observed allele, genotype and haplotype distributions were compared with those obtained from 300 ethnically matched negative controls. Invariant distribution of IL-1 gene cluster single-nucleotide polymorphisms was observed among RPL cases and controls. Meta-analysis of IL-1b _511, +3953 and IL-1RN 86-bp variable number tandem repeat from the reported literature and this study did not reveal any significant association with the risk of RPL. In conclusion, no significant difference between RPL and control groups was observed at the allele, genotype or haplotype levels when tested for association using the dominant, recessive and additive models of inheritance for IL-1 gene cluster variants. As far as is known, this is the first report from India pertaining to IL-1 gene cluster variants’ association with the risk of RPL from North India.

Distribution of Killer cell immunoglobulin like receptor genes in end stage renal disease among North Indian population

INTRODUCTION NK cell function is regulated by cell surface inhibitory and activating receptors including the C-type lectin receptors and Killer Immunoglobulin-like receptors (KIR). The effect of immune modulating cytokines produced by NK cells in the pathogenesis of end stage renal disease (ESRD) remained intriguing. In this regard the present study assesses the combinatorial association of KIR gene content and KIR receptor-HLA ligand in the North Indian ESRD patients. MATERIAL AND METHODS KIR gene polymorphism as a susceptible marker in ESRD among 512 patients and 512 ethnically matched controls was analyzed. PCR-SSP based genotyping for KIR gene content and HLA-A, B, C typing was carried out. RESULTS Significant difference in frequencies of KIR2DS1-HLA-C2 (p≤0.0001, OR=1.98, CI=1.50-2.61), KIR2DS2-HLAC1 (p≤0.0001, OR=1.87, CI=1.42-2.46), KIR3DS1-HLA-Bw4 (p=0.0038, OR=1.46, CI=1.13-1.88) combinations for ESRD was found. In the combinatorial analysis Bw4(+)/3DL1(-)/3DS1(+) (p≤0.0001, OR=4.90, CI=2.75-8.71) and C1(+)/2DL2(-)/2DL3(-)/2DS2(+)/2DS3(+) (p=0.0037, OR=2.50, CI=1.35-4.63) showed risk association. KIR3DS1 was observed to be susceptible for all four primary kidney disease groups. CONCLUSION NK cell de-regulation due to HLA ligand binding KIR receptors may be involved in the patho-physiology of ESRD. Upon analyzing the data in this context it was found that C2/C2 donor might improve the clinical outcome of patients having C2 ligands.

Vascular endothelial growth factor single-nucleotide polymorphism in gall bladder cancer

Angiogenesis plays a key role in growth, progression, and metastasis of various cancers. Vascular endothelial growth factor (VEGF) polymorphism has been associated with several cancers. Role of VEGF has not been reported in gallbladder cancer (GBC). Present study was designed to investigate the role of VEGF polymorphism in GBC and in other (benign) gallbladder diseases, that is chronic cholecystitis (CC) and xanthogranulomatous cholecystitis (XGC).

Genetic association of adipokine and UCP2 polymorphism with recurrent miscarriage among non-obese women

The adipokines produced from adipose tissues influence energy homeostasis, resulting in alterations of the adipokine concentrations. This process may be associated with fertility impairment, resulting in recurrent miscarriage. The present study investigated whether there was any association between the UCP2 45-bp indel polymorphism and the adipokine gene polymorphisms, namely leptin 2549 (C/A), adeponectin 276 (G/T) and 45 (T/G) and resistin 420 (C/G) in 200 non-obese recurrent miscarriage patients and 300 ethnically matched negative controls. These markers were studied using gene-specific PCR single specific primer and restriction fragment length polymorphism. For leptin 2549 and adeponectin 276, the A allele and G allele showed 3.42-fold (P=0.0001) and 1.36-fold (P=0.036) increased risk of recurrent miscarriage, respectively. Combined analysis of UCP2 45-bp indel and leptin 2549 showed U0-L0 and U1-L0 variants to be at 2- and 3-fold increased associative risk, respectively. Combined analysis of leptin 2549 and adeponectin 276 showed L0-D0 and L0-D1 variants to be at 2- and 4-fold increased associative risk, respectively. The combination U1-L0-D1-A1-R1 was 4.39-fold higher (P=0.0007) among recurrent miscarriage patients. In conclusion, the results highlight the role of the studied adipokine and UCP2 polymorphisms in recurrent miscarriage among the North Indian non-obese population. Pregnancy invokes a large shift in maternal metabolism. The normal concentrations of adipokines, which maintain the integrity of the hypothalamus-pituitary-gonadal axis, regular ovulatory processes and successful embryo implantation, are altered because of the influence of energy homeostasis, which in turn leads to fertility impairment and recurrent miscarriage of unknown aetiology. Recurrent miscarriage is reported in higher frequency among obese women. The UCP2 45-bp indel polymorphism and the adipokine gene polymorphisms namely leptin 2549 (C/A), adeponectin 276 (G/T), adeponectin 45 (T/G) and resistin 420 (C/G) have been shown to be associated with obesity. Most of the adipokine-related studies done previously have taken into consideration the metabolic function and obesity. However, there exist very few studies to evaluate the role of adipokines in non-obese recurrent miscarriage with no cause of repeated pregnancy losses. The present study focused at evaluating the independent effect of these single-nucleotide polymorphisms in non-obese women undergoing recurrent miscarriage.

Genetic associations of killer immunoglobulin like receptors and class I human leukocyte antigens on childhood acute lymphoblastic leukemia among north Indians.

BACKGROUND Molecular interactions between KIRs and their cognate HLA class-I ligands, play a central role in the regulation of natural killer (NK) cell responses in malignancies. We aimed to determine the role of KIR genes and their HLA ligands in genetic predisposition of childhood acute lymphoblastic leukemia (ALL). METHODS Genotyping of 16 KIR genes, along with HLA class-I groups C1/C2 and Bw4 super-type ligands, was carried-out in 137 childhood ALL cases and 274 healthy controls. RESULTS We observed an increased incidence of activating KIRs namely; 2DS2 (OR=2.23, p=<0.001), 2DS3 (OR=1.74, p=0.011), 3DS1 (OR=2.22, p=<0.001), 2DS5 (OR=2.10, p=0.001), 2DS1 (OR=4.42, p=<0.001) and 2DS4 (OR=2.88, p=<0.001) genes in childhood ALL cases compared to controls. Frequency of BB genotype that possess 2-6 activating KIR genes was predominant in cases compared to controls (OR=2.55, p=<0.001). KIR-receptor/HLA-ligand combinations analysis revealed a moderate risk of almost 2-fold for activating KIR-ligand combinations namely; KIR2DS1-HLAC2, KIR2DS2-HLAC1 and KIR3DS1-HLABw4 in childhood ALL cases. CONCLUSION Our data suggests the role for KIR genes and their HLA ligands in aetiology of childhood ALL.

MicroRNAs involvement in renal pathophysiology: A bird's eye view

MicroRNAs (miRNAs) are known to suppress gene expression by binding to messenger RNAs and in turn regulate different pathophysiological processes. Transforming growth factor-β, mitogen-activated protein kinase signaling, and Wnt signaling-like major pathways associated with miRNAs are involved with kidney diseases. The discovery of miRNAs has provided new insights into kidney pathologies and may provide effective therapeutic strategies. Research has demonstrated the role of miRNAs in a variety of kidney diseases including diabetic nephropathy, lupus nephritis, hypertension, nephritic syndrome, acute kidney injury, renal cell carcinoma, and renal fibrosis. miRNAs are implicated as playing a role in these diseases due to their role in apoptosis, cell proliferation, differentiation, and development. As miRNAs have been detected in a stable condition in different biological fluids, they have the potential to be tools to study the pathogenesis of human diseases with a great potential to be used in disease diagnosis and prognosis. The purpose of this review is to examine the role of miRNA in kidney disease.

Vascular endothelial growth factor gene polymorphisms and association with age related macular degeneration in Indian patients

Background Age-related macular degeneration (AMD) is an important cause of visual impairment in elderly people. AMD is a multifactorial disease in which both environmental and genetic factors have been implicated. Various single nucleotide polymorphisms (SNPs) have been found to be associated with AMD. Aim This study was aimed to investigate the association of polymorphisms in VEGF genes with age related macular degeneration (AMD) in Indian patients. Method Genotyping for the VEGF − 1154 (G > A), − 2578 (C > A), + 405 (G > C) and − 460 (C > T) SNPs was performed in 100 AMD patients and 100 controls by polymerase chain reaction (PCR), restriction fragment length polymorphism (PCR-RFLP) and sequencing method. Results Out of the four SNPs, heterozygous genotypes of VEGF − 1154 G > A (OR = 2.58, p = 0.0035), + 460 C > T (OR = 2.90, p = 0.0046), and + 405 G > C (OR = 2.02, p = 0.02) have shown susceptible association with AMD. However, VEGF − 2578 C > A did not show any statistical significance. Further A-A-G-T haplotype comprising of three mutant alleles revealed risk association (OR = 12.7, p = 0.0030) with AMD. Conclusion The present study suggests significant genetic associations for VEGF − 1154 G > A, + 460 C > T, and + 405 G > C polymorphisms with AMD. Early detection of individuals with risk to these SNPs could lead to strategies for prevention, early diagnosis, and management of AMD.