Glycolate oxidase-1 gene variants influence the risk of hyperoxaluria and renal stone development.

PURPOSE: Oxalate is an excellent calcium ion attractor with great abundance in the human body, and the liver is the major source of oxalate. The Glycolate oxidase-1 (GOX1) gene is solely responsible for the glycolate and glyoxylate metabolism and produces oxalate. This study has been designed to comprehend the association of genetic variants of the GOX1 gene with the risk of hyperoxaluria and renal stone disease in the Indian population. METHOD: The present study is a candidate gene approach prospective case-control study carried out on 300 participants (150 cases and 150 controls) at Muljibhai Patel Urological Hospital, Gujarat, India. Biochemical parameters, including serum levels of calcium, creatinine, parathyroid hormone, and 24-h urine metabolites, were performed. The genotyping of GOX1 gene variants rs6086287, rs2235250, rs2255183, and rs2294303 was performed using a customized TaqMan assay probe by RT-PCR. RESULT: Parathyroid hormone, serum creatinine, and urine metabolites were significantly elevated in nephrolithiasis compared to healthy individuals. All mutated homozygous genotypes GG (rs6086287), TT (rs2235250), GG (rs2255183), and CC (rs2294303) were significantly associated with a high risk of renal stone disease. Individuals diagnosed with hyperoxaluria and carrying TG (rs6086287), AG (rs2255183), and TT (rs2294303) genotypes have a significantly high risk of renal stone disease. Moreover, haplotype analysis and correlation analysis also confirmed the strong association between genetic variants and nephrolithiasis. CONCLUSION: Genetic variants of the GOX1 genes were associated with renal stone disease. In the presence of risk genotype and hyperoxaluria, the susceptibility to develop renal stone disease risk gets modulated.

INF2 and ROBO2 gene mutation in an Indian family with end stage renal failure and follow-up of renal transplantation.

BACKGROUND: Accurate genetic diagnosis of end-stage renal disease patients with a family history of renal dysfunction is very essential. It not only helps in proper prognosis, but becomes crucial in designating donor for live related renal transplant. We here present a case of family with deleterious mutations in INF2 and ROBO2 and its importance of genetic testing before preparing for kidney transplantation. CASE PRESENTATION: We report the case of a 29-year-female with end-stage renal disease and rapidly progressive renal failure. Mutational analysis revealed an Autosomal Dominant inheritance pattern and mutation in exon 4 of the INF2 gene (p. Thr215Ser) and exon 26 of the ROBO2 gene (p. Arg1371Cys). Her mother was diagnosed for CKD stage 4 with creatinine level of 4.3 mg/dL. Genetic variants (INF2 and ROBO2) identified in proband were tested in her sisters and mother. Her elder sister was positive for both heterozygous variants (INF2 and ROBO2). Her mother was positive for mutation in INF2 gene, and her donor elder sister did not showed mutation in INF2 gene and had mutation in ROBO2 gene without any clinical symptoms. CONCLUSION: This case report emphasize that familial genetic screening has allowed us in allocating the donor selection in family where family member had history of genetic defect of Chronic Kidney Disease. Information of the causative renal disorder is extremely valuable for risk-assessment and planning of kidney transplantation.

A systematic review and meta-analysis recite the efficacy of Tacrolimus treatment in renal transplant patients in association with genetic variants of CYP3A5 gene.

Tacrolimus is an immunosuppressant with a narrow therapeutic index and pharmacokinetic variability. This variability may be attributed to genetic variants in gene CYP3A5 associated with Tacrolimus metabolism. Studies focusing on genetic variants in the CYP3A5 gene associated with Tacrolimus metabolism have been published, a meta-analysis of these published articles may provide a direction that can change the future research and clinical management of renal transplant patients. In this systematic review and meta-analysis, we have reviewed and analyzed the studies and clinical trials conducted to determine the association between genetic variants of CYP3A5 and Tacrolimus metabolism from the PubMed database and clinical trials (www.clinicaltrials.gov). This meta-analysis also assessed the correlation of CYP3A5 genotype (rs776746) with concentration/dose (Co/D) of Tacrolimus in renal transplant patients. The 59 published articles on genetic association of the CYP3A5 on Tacrolimus doses were reviewed for this systematic review. Meta-analysis showed that the Tacrolimus Co/D ratio is significantly lower in the CYP3A5 expressor group as compared with non-expressor in Asian, European as well as in mixed populations at any post-transplant period (P<0.0001). Our study further confirmed that the CYP3A5 variant (rs776746) is clinically relevant for the dose determination of Tacrolimus. Variations in Tacrolimus Co/D have been found to be significantly linked to the patient's CYP3A5 genetic variant (rs776746). The addition of other genetic variants involved in the pharmacokinetic of Tacrolimus may determine efficient regimen for drug dose. Our meta-analysis confirmed that the CYP3A5 genetic variant (rs776746) analysis is relevant in personalizing the Tacrolimus dose determination in renal transplant patients.

Suppression of TLR signaling by IRAK-1 and -4 dual inhibitor decreases TPF-resistance-induced pro-oncogenic effects in HNSCC.

Combination of docetaxel, cisplatin and 5-FU, known as TPF, is an FDA-approved treatment for head and neck squamous cell carcinoma (HNSCC). Acquired chemo-resistance to TPF, a primary reason for non-responsiveness to the treatment and relapse of tumor is a major concern for treatment failure, especially in elder patients. In this study, we investigated the role of Interleukin-1 receptor-associated kinases (IRAK) mediated Toll-like receptor (TLR)-signaling in chemo-resistance using a cell line-based in-vitro TPF-resistant HNSCC model of laryngeal origin. TPF chemo-resistant state showed over-expression and phosphorylation of the active downstream kinases IRAK-1 and IRAK-4 along with enhanced proliferative potential, survival, stemness and metastatic capability as compared to the parent cell line. Pharmacological inhibition of IRAK-1 and -4 had a cytostatic effect on chemo-resistant cells and re-sensitized them to chemotherapy. The treatment also decreased the pro-oncogenic effects of the chemo-resistant cells. Our study provides insights into the pro-oncogenic role of amplified IRAK-1 and-4 mediated TLR signaling in TPF-resistant HNSCC. Pharmacological inhibition of IRAK-1 and-4 signaling is a promising therapeutic strategy for TPF-resistant HNSCC. It can also be used as a combination therapy or a chemo-drug sparing regimen in HNSCC.

Haplotype of CaSR gene is associated with risk of renal stone disease in West Indian population.

Calcium is the most abundant metabolite involved in the stone matrix. The CaSR gene controls calcium homeostasis, and genetic variation in the CaSR gene could lead to the development of renal stone disease. Therefore, the current study has been designed to assess the association of genetic variants of CaSR gene polymorphisms with renal stone disease. A single-centric prospective study has been carried out on a total of 300 participants (150 cases and 150 controls). Serum levels of calcium, creatinine, parathyroid hormone, and 24 h urine metabolites were measured. Two polymorphisms, rs1801725 and rs1042636, of the CaSR gene, have been genotyped for each participant. T test, binary logistic regression, and Chi-square analysis were used for statistical analysis. Renal stone patients had significantly higher levels of serum parathyroid hormone, creatinine, and 24-h urine metabolites in comparison to the controls. CaSR gene variants, rs1801725 (GG) and rs1042636 (AA), both have shown significant association with renal stone disease. In addition, individuals having specific genotypes along with metabolic abnormalities such as hypercalcemia and hyperparathyroidism are found to be at a higher significant risk of developing the renal stone disease. In the present study, the haplotype of the CaSR gene has shown an association with renal stone disease. Individuals with hyperparathyroidism and hypercalcemia and risk genotype have a higher susceptibility to developing renal stone disease.

Multi-Omics Identifies Circulating miRNA and Protein Biomarkers for Facioscapulohumeral Dystrophy.

The development of therapeutics for muscle diseases such as facioscapulohumeral dystrophy (FSHD) is impeded by a lack of objective, minimally invasive biomarkers. Here we identify circulating miRNAs and proteins that are dysregulated in early-onset FSHD patients to develop blood-based molecular biomarkers. Plasma samples from clinically characterized individuals with early-onset FSHD provide a discovery group and are compared to healthy control volunteers. Low-density quantitative polymerase chain reaction (PCR)-based arrays identify 19 candidate miRNAs, while mass spectrometry proteomic analysis identifies 13 candidate proteins. Bioinformatic analysis of chromatin immunoprecipitation (ChIP)-seq data shows that the FSHD-dysregulated DUX4 transcription factor binds to regulatory regions of several candidate miRNAs. This panel of miRNAs also shows ChIP signatures consistent with regulation by additional transcription factors which are up-regulated in FSHD (FOS, EGR1, MYC, and YY1). Validation studies in a separate group of patients with FSHD show consistent up-regulation of miR-100, miR-103, miR-146b, miR-29b, miR-34a, miR-454, miR-505, and miR-576. An increase in the expression of S100A8 protein, an inflammatory regulatory factor and subunit of calprotectin, is validated by Enzyme-Linked Immunosorbent Assay (ELISA). Bioinformatic analyses of proteomics and miRNA data further support a model of calprotectin and toll-like receptor 4 (TLR4) pathway dysregulation in FSHD. Moving forward, this panel of miRNAs, along with S100A8 and calprotectin, merit further investigation as monitoring and pharmacodynamic biomarkers for FSHD.

PARP1 Differentially Interacts with Promoter region of DUX4 Gene in FSHD Myoblasts.

OBJECTIVE: The goal of the study is to identity proteins, which interact with the promoter region of double homeobox protein 4 (DUX4) gene known to be causative for the autosomal dominant disorder Facioscapulohumeral Muscular Dystrophy (FSHD). METHODS: We performed a DNA pull down assay coupled with mass spectrometry analysis to identify proteins that interact with a DUX4 promoter probe in Rhabdomyosarcomca (RD) cells. We selected the top ranked protein poly (ADP-ribose) polymerase 1 (PARP1) from our mass spectrometry data for further ChIP-qPCR validation using patients' myoblasts. We then treated FSHD myoblasts with PARP1 inhibitors to investigate the role of PARP1 in the FSHD myoblasts. RESULTS: In our mass spectrometry analysis, PARP1 was found to be the top ranked protein interacting preferentially with the DUX4 promoter probe in RD cells. We further validated this interaction by immunoblotting in RD cells (2-fold enrichment compared to proteins pulled down by a control probe, p<0.05) and ChIP-qPCR in patients' myoblasts (65-fold enrichment, p<0.01). Interestingly, the interaction was only observed in FSHD myoblasts but not in the control myoblasts. Upon further treatment of FSHD myoblasts with PARP1 inhibitors, we showed that treatment with a PARP1 inhibitor, 3-aminobenzamide (0.5 mM), for 24 h had a suppression of DUX4 (2.6 fold, p<0.05) and ZSCAN4, a gene previously shown to be upregulated by DUX4, (1.6 fold, p<0.01) in FSHD myoblasts. Treatment with fisetin (0.5 mM), a polyphenol compound with PARP1 inhibitory property, for 24 h also suppressed the expression of DUX4 (44.8 fold, p<0.01) and ZSCAN4 (2.2 fold, p<0.05) in the FSHD myoblasts. We further showed that DNA methyltransferase 1 (DNMT1), a gene regulated by PARP1 was also enriched at the DUX4 promoter in RD cells through immunoblotting (2-fold, p<0.01) and immortalized FSHD myoblasts (42-fold, p<0.01) but not control myoblasts through ChIP qPCR. CONCLUSION: Our results showed that PARP1 and DNMT1 interacted with DUX4 promoter and may be involved in modulating DUX4 expression in FSHD.

Transcriptional Pathways Associated with Skeletal Muscle Changes after Spinal Cord Injury and Treadmill Locomotor Training.

The genetic and molecular events associated with changes in muscle mass and function after SCI and after the implementation of candidate therapeutic approaches are still not completely known. The overall objective of this study was to identify key molecular pathways activated with muscle remodeling after SCI and locomotor training. We implemented treadmill training in a well-characterized rat model of moderate SCI and performed genome wide expression profiling on soleus muscles at multiple time points: 3, 8, and 14 days after SCI. We found that the activity of the protein ubiquitination and mitochondrial function related pathways was altered with SCI and corrected with treadmill training. The BMP pathway was differentially activated with early treadmill training as shown by Ingenuity Pathway Analysis. The expression of several muscle mass regulators was modulated by treadmill training, including Fst, Jun, Bmpr2, Actr2b, and Smad3. In addition, key players in fatty acids metabolism (Lpl and Fabp3) responded to both SCI induced inactivity and reloading with training. The decrease in Smad3 and Fst early after the initiation of treadmill training was confirmed by RT-PCR. Our data suggest that TGFß/Smad3 signaling may be mainly involved in the decrease in muscle mass observed with SCI, while the BMP pathway was activated with treadmill training.

Culture Conditions Affect Expression of DUX4 in FSHD Myoblasts.

Facioscapulohumeral muscular dystrophy (FSHD) is believed to be caused by aberrant expression of double homeobox 4 (DUX4) due to epigenetic changes of the D4Z4 region at chromosome 4q35. Detecting DUX4 is challenging due to its stochastic expression pattern and low transcription level. In this study, we examined different cDNA synthesis strategies and the sensitivity for DUX4 detection. In addition, we investigated the effects of dexamethasone and knockout serum replacement (KOSR) on DUX4 expression in culture. Our data showed that DUX4 was consistently detected in cDNA samples synthesized using Superscript III. The sensitivity of DUX4 detection was higher in the samples synthesized using oligo(dT) primers compared to random hexamers. Adding dexamethasone to the culture media significantly suppressed DUX4 expression in immortalized (1.3 fold, p < 0.01) and primary (4.7 fold, p < 0.01) FSHD myoblasts, respectively. Culture medium with KOSR increased DUX4 expression and the response is concentration dependent. The findings suggest that detection strategies and culture conditions should be carefully considered when studying DUX4 in cultured cells.

Morpholino treatment improves muscle function and pathology of Pitx1 transgenic mice.

Paired-like homeodomain transcription factor 1 (PITX1) was proposed to be part of the disease mechanisms of facioscapulohumeral muscular dystrophy (FSHD). We generated a tet-repressible muscle-specific Pitx1 transgenic mouse model which develops phenotypes of muscular dystrophy after the PITX1 expression is induced. In this study, we attempted to block the translation of PITX1 protein using morpholinos. Three groups of the transgenic mice received intravenous injections of phosphorodiamidate morpholino oligomers (PMO) (100 mg/kg), octaguanidinium dendrimer-conjugated morpholino (vivo-morpholino) (10 mg/kg), or phosphate-buffered saline (PBS) after the PITX1 expression was induced. Immunoblotting data showed that PITX1 expression in the triceps and quadriceps was significantly reduced 70% and 63% by the vivo-morpholino treatment, respectively. Muscle pathology of the mice treated with the vivo-morpholino was improved by showing 44% fewer angular-shaped atrophic myofibers. Muscle function determined by grip strength was significantly improved by the vivo-morpholino treatment. The study showed that systemic delivery of the vivo-morpholino reduced the PITX1 expression and improved the muscle phenotypes. Aberrant expression of DUX4 from the last unit of the D4Z4 array has been proposed to be the cause of FSHD. The findings of this study suggest that the same principle may be applied to suppress the aberrantly expressed DUX4 in FSHD.

Conditional expression of TGF-ß1 in skeletal muscles causes endomysial fibrosis and myofibers atrophy.

To study the effects of transforming growth factor beta 1 (TGF-ß1) on fibrosis and failure of regeneration of skeletal muscles, we generated a tet-repressible muscle-specific TGF-ß1 transgenic mouse in which expression of TGF-ß1 is controlled by oral doxycycline. The mice developed muscle weakness and atrophy after TGF-ß1 over-expression. We defined the group of mice that showed phenotype within 2 weeks as early onset (EO) and the rest as late onset (LO), which allowed us to further examine phenotypic differences between the groups. While only mice in the EO group showed significant muscle weakness, pathological changes including endomysial fibrosis and smaller myofibers were observed in both groups at two weeks after the TGF-ß1 was over-expressed. In addition, the size of the myofibers and collagen accumulation were significantly different between the two groups. The amount of latent and active TGF-ß1 in the muscle and circulation were significantly higher in the EO group compared to the LO or control groups. The up-regulation of the latent TGF-ß1 indicated that endogenous TGF-ß1 was induced by the expression of the TGF-ß1 transgene. Our studies showed that the primary effects of TGF-ß1 over-expression in skeletal muscles are muscle wasting and endomysial fibrosis. In addition, the severity of the pathology is associated with the total amount of TGF-ß1 and the expression of endogenous TGF-ß1. The findings suggest that an auto-feedback loop of TGF-ß1 may contribute to the severity of phenotypes.

Gene expression profiling of gastrocnemius of "minimuscle" mice.

Few studies have investigated heterogeneity of selection response in replicate lines subjected to equivalent selection. We developed four replicate lines of mice based on high levels of voluntary wheel running (high runner or HR lines) while also maintaining four nonselected control lines. This led to the unexpected discovery of the HR minimuscle (HRmini) phenotype, recognized by a 50% reduction in hindlimb muscle mass, which became fixed in 1 of the four HR selected lines. Here, we report genome-wide expression profiling describing transcriptome differences between HRnormal and HRmini medial gastrocnemius. Consistent with the known reduction of type IIB fibers in HRmini, Myh4 gene expression was -8.82-fold less (P = 0.0001) in HRmini, which was closely associated with differences in the "calcium signaling" canonical pathway, including structural genes (e.g., Mef2c, twofold greater in HRmini, P = 0.0003) and myogenic factors (e.g., Myog, 3.8-fold greater in HRmini, P = 0.0026) associated with slow-type myofibers. The gene that determines the HRmini phenotype is known to reside in a 2.6335-Mb interval on mouse chromosome 11 and 7 genes (Myh10, Chrnb1, Acadvl, Senp3, Gabarap, Eif5a, and Clec10a) from this region were differentially expressed. Verification by real-time PCR confirmed 1.5-fold greater (P < 0.05) expression of very long chain acyl-CoA dehydrogenase (Acadvl) in HRmini. Ten other genes associated with fatty acid metabolism were also upregulated in HRmini, suggesting differences in the ability to metabolize fatty acids in HRnormal and HRmini muscles. This work provides a resource for understanding differences in muscle phenotypes in populations exhibiting high running capacity.

Functional polymorphisms of the cyclooxygenase (PTGS2) gene and risk for gallbladder cancer in a North Indian population.

BACKGROUND: Cyclooxygenase-2 (PTGS2) overexpression has been implicated in various cancers. We aimed to evaluate the role of PTGS2 -1195G>A [reference sequence (rs) 689466], -765G>C (rs20417) and +8473T>C (rs5275) polymorphisms in conferring interindividual susceptibility to gallbladder cancer. MATERIALS AND METHODS: The study included 167 gallbladder cancer cases and 184 controls. Genotyping was done by polymerase chain reaction-restriction fragment length polymorphism. Risk was estimated using unconditional logistic regression. RESULTS: Significant risk was observed in the presence of PTGS2 -1195GA (P=0.006; odds ratio=2.00; 95% confidence interval=1.2-3.3) and AA genotypes (P=0.050; odds ratio=2.98; 95% confidence interval=1.0-8.9). Combined risk due to GA+AA genotypes was 2.12 (P=0.002; 95% confidence interval=1.3-3.3; P-trend=0.001). Sub-grouping showed a risk due to the PTGS2 -1195(GA+AA) genotype in males (P=0.007; odds ratio=2.97; 95% confidence interval=1.3-6.5), patients without gallstones (P=0.001; odds ratio=2.53; 95% confidence interval=1.4-4.7) and with late-onset gallbladder cancer (P=0.012; odds ratio=1.99; 95% confidence interval=1.1-3.4). Gallbladder cancer patients who used tobacco were at increased risk in the presence of the PTGS2 -765GC genotype (P=0.018; odds ratio=2.96; 95% confidence interval=1.2-7.2). DISCUSSION: An association of PTGS2 -1195G>A polymorphism with gallbladder cancer, particularly in patients without gallstones, suggests a direct role of PTGS2 in cancer development.

Single-nucleotide polymorphisms of DNA repair genes OGG1 and XRCC1: association with gallbladder cancer in North Indian population.

BACKGROUND: DNA damage by endogenous or exogenous source of reactive oxygen species (ROS) plays an important role in induction and progression of various cancers. Physiologically, gallbladder is likely to be exposed to various ROS which leads to extensive DNA damage. Cells overcome the DNA damage by repair mechanisms. Genetic variants of OGG1 and XRCC1, important enzymes participating in base excision repair pathway, may confer interindividual variations in susceptibility to gallbladder cancer (GBC). This study was aimed to examine the role of OGG1 Ser326Cys (rs1052133) and XRCC1 Arg194Trp (C > T) (rs25487) and Arg399Gln (G > A) (rs1799782) polymorphisms in GBC susceptibility. METHODS: The study included 173 GBC patients and 204 controls. Genotyping was done by polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) method. Differences in the frequencies were estimated by chi-square test and risk was estimated by using unconditional logistic regression after adjusting for age and gender. RESULTS: OGG1 Cys/Cys genotype frequency was significantly higher in GBC patients [odds ratio (OR) = 2.93; 95% confidence interval (CI) = 1.14-7.51]. The increased risk was more pronounced in female GBC patients (OR = 5.92; 95%CI = 1.20-29.13), patients with gallstone (OR = 5.50; 95%CI = 1.99-15.16), female gender, and late onset of disease (OR = 4.72, 95%CI = 1.43-15.53). In XRCC1 Arg399Gln polymorphism, significant differences in frequencies of Gln/Gln and Arg/Gln genotypes conferred significantly low risk for GBC (OR = 0.62; 95%CI = 0.39-0.97 and OR = 0.37; 95%CI = 0.19-0.71 respectively). However, XRCC1 Arg194Trp polymorphism was not associated with GBC. The carriers of Arg-Gln haplotype consisting of 194Arg and 399Gln alleles of XRCC1 were also at significant low risk for GBC (OR = 0.59, 95%CI = 0.42-0.82). Interaction of genotypes and tobacco usage did not modulate the risk. CONCLUSION: Results suggest that Cys/Cys genotype of OGG1 Ser326Cys polymorphism is associated with increased risk of GBC. However, Arg399Gln polymorphism and Arg-Gln haplotype comprising XRCC1 Arg194Trp and Arg399Gln polymorphisms conferred low risk for GBC susceptibility.

Single nucleotide polymorphism in the ABCG8 transporter gene is associated with gallbladder cancer susceptibility.

BACKGROUND: Gallbladder cancer (GBC) usually arises against the background of gallstone disease, which may be causatively related to supersaturation of cholesterol in bile. An imbalance in cholesterol homeostasis because of oversecretion of cholesterol in the gallbladder promotes gallstone formation. The excretion of cholesterol from the liver is regulated by adenosine triphosphate-binding cassette transporter ABCG8. A common genetic polymorphism D19H of ABCG8 associated with gallstone disease may be causatively related to the genetic predisposition of GBC. AIM: We aimed to examine the role of ABCG8 D19H (rs11887534) polymorphism in susceptibility to GBC. METHODOLOGY: This study included 171 confirmed GBC patients and 221 controls. Genotyping for the ABCG8 D19H polymorphism was performed by the polymerase chain reaction-restriction fragment length polymorphism method. RESULTS: We observed that in our population the ABCG8 DH genotype frequency was significantly higher in GBC patients [P=0.011; odds ratio (OR)=1.79; 95% confidence interval (CI)=1.1-2.8]. Also, at the allele level, ABCG8H conferred an increased risk for GBC (P=0.023; OR=1.60; 95% CI=1.0-2.4). The risk was more pronounced in GBC patients with gallstones (P=0.027; OR=1.85; 95% CI=1.0-3.1), and in patients with an early onset of the disease (P=0.013; OR=2.55, 95% CI=1.2-5.3). However, there was no modulation of GBC risk because of the ABCG8 polymorphism in a gender-specific manner. CONCLUSION: The results suggest that the DH genotype and the H allele of the ABCG8 D19H polymorphism are associated with GBC susceptibility. The GBC patients with gallstone disease harbouring the ABCG8 variant allele are at a higher risk, while the effect of this polymorphism on GBC patients without gallstones appears to be small.

IL-1 gene polymorphisms and genetic susceptibility of gallbladder cancer in a north Indian population.

Long-standing gallstones are generally present in 65-80% patients of gallbladder cancer (GBC). It has also been suggested that inflammation caused by gallstones may be involved in the development of GBC. Interleukin-1 receptor antagonist (IL-1RN) and interleukin-1 beta (IL-1B) are proinflammatory cytokine genes at the interleukin-1 locus, and polymorphisms of these genes have been associated with various inflammatory diseases. The aim of this study was to investigate whether polymorphism in the IL-1RN and IL-1B genes are associated with GBC patients with and without gallstones. Polymorphisms within the IL-1RN 86-base pair VNTR (variable number tandem repeat) and IL-1B (-511C --> T) were genotyped using polymerase chain reaction (PCR) and PCR restriction fragment length polymorphism in 166 healthy subjects and 124 GBC patients. The frequency of the IL-1RN, VNTR 2/2 genotype was significantly higher in GBC patients [P = 0.017; odds ratio (OR) = 3.25; 95% confidence interval (CI) = 1.23-8.58]. CC genotype and 'C' allele of the -511IL-1B C --> T polymorphism also showed high risk for GBC (P = 0.033; OR = 3.36; 95%CI = 1.52-7.43, P = 0.047, OR = 1.41; 95%CI = 1.00-1.98, respectively). The higher cancer risk due to the IL-1RN, 2/2 genotype was observed in GBC patients with or without stones (P = 0.038; OR = 3.58; 95%CI = 1.08-11.65, P = 0.035; OR = 3.33; 95%CI = 1.08-10.61). Risk due to the CC genotype of IL-1B, however, was confined to GBC patients harboring gallstones (P = 0.0003; OR = 6.92; 95%CI = 2.65-18.03). The haplotype 1/C of IL-1RN and IL-1B was found to confer a significantly enhanced risk of GBC in cancer patients with gallstones (P = 0.022; OR = 2.19; 95%CI = 1.12-4.27), while higher risk resulting from 2/C haplotype was of borderline significance (P = 0.061; OR = 3.04; 95%CI = 0.95-9.70). Individuals with 1/C and 2/C haplotypes of IL-1RN VNTR and -511IL-1B C --> T polymorphisms were more susceptible to develop GBC with gallstones compared to healthy controls in north India.

Genetic susceptibility of epidermal growth factor +61A>G and transforming growth factor beta1 -509C>T gene polymorphisms with gallbladder cancer.

Epidermal growth factor (EGF) and transforming growth factor beta1 (TGFbeta1) play important roles in tumor biology. Single nucleotide polymorphisms in EGF and TGFB1 genes alter the expression of these growth factors and influence the tumorigenesis process. The aim of our present study was to determine the association of EGF+61A>G (rs4444903) and TGFB1-509C>T (rs1800469) gene polymorphism with susceptibility to gallbladder cancer (GBC). The present case-control association study was carried out in 126 confirmed GBC patients and 190 healthy subjects. Genotyping was performed by polymerase chain reaction-restriction fragment length polymorphism methods. The GG genotype of EGF+61A>G was significantly associated with GBC [p=0.012, odds ratio (OR)=2.22, 95% confidence interval (CI)=1.19-4.15] in comparison to healthy subjects. Analysis based on gender indicated risk due to GG genotype was limited to female GBC patients (p=0.003, OR=3.45, 95% CI=1.52-7.82). Upon stratification of GBC patients on the basis of the presence or absence of gallstones, the risk due to EGF polymorphism was not modulated by the status of gallstones. The TGFB1-509C>T polymorphism was not associated with GBC. Also, we did not find any association of this polymorphism when GBC patients were subdivided on the basis of gender. However, after stratification of GBC patients on the status of gallstones, we determined that the CT genotype of TGFB1 was associated with increased risk of GBC without gallstones (p value=0.030, OR=2.90, 95% CI=1.26-6.69). Furthermore, the combination of the GG genotype of EGF and the CT genotype of TGFB1 demonstrated synergistic increase in risk of GBC. In conclusion, the higher producing +61G allele of EGF and -509 CT genotype of TGFB1 synergistically increase the susceptibility of gallbladder cancer (p value=0.003). Further study in large samples size is required to confirm our findings.

Association of CYP1A1 Msp1 polymorphism with tobacco-related risk of gallbladder cancer in a north Indian population.

Polymorphisms in the gene encoding the metabolic enzyme cytochrome P450 1A1 (CYP1A1) might contribute to the variability in individual susceptibility to gallbladder cancer (GBC). This study comprised 142 consecutive cases of proven GBC and 171 healthy participants of similar ethnicity. CYP1A1 6235 T/C transition was analyzed using polymerase chain reaction-restriction fragment length polymorphism. The CC genotype of CYP1A1 was significantly associated with a risk of GBC [odds ratio (OR) 2.3, 95% confidence interval (CI)=1.1-4.5, P=0.026], and the age- and sex-adjusted OR was 2.0. The CYP1A1 C allele frequency was also higher in GBC patients (OR 1.4, 95% CI=1.0-2.0, P=0.039). After sex stratification, the TC genotype and C allele were significantly higher in men and conferred a risk for GBC (OR 4.2, 95% CI=1.8-9.7, P=0.001; OR 2.2, CI=1.3-3.7, P=0.005). In addition, the TC genotype and C allele frequencies were significantly higher in GBC patients without associated gallstones (OR 2.4, 95% CI=1.2-4.7, P=0.011; OR 1.6, CI=1.0-2.4, P=0.023). The usage of tobacco (smoking or nonsmoking) by GBC patients showed a significant increase in cancer risk with the TC genotype (OR 4.1, 95% CI=1.3-11.9, P=0.012). It seems that the higher risk of the TC genotype in men may be related to tobacco usage.