Association Analysis of Dyslipidemia-Related Genes in Diabetic Nephropathy

Type 1 diabetes (T1D) increases risk of the development of microvascular complications and cardiovascular disease (CVD). Dyslipidemia is a common risk factor in the pathogenesis of both CVD and diabetic nephropathy (DN), with CVD identified as the primary cause of death in patients with DN. In light of this commonality, we assessed single nucleotide polymorphisms (SNPs) in thirty-seven key genetic loci previously associated with dyslipidemia in a T1D cohort using a casecontrol design. SNPs (n = 53) were genotyped using Sequenom in 1467 individuals with T1D (718 cases with proteinuric nephropathy and 749 controls without nephropathy i.e. normal albumin excretion). Cases and controls were white and recruited from the UK and Ireland. Association analyses were performed using PLINK to compare allele frequencies in cases and controls. In a sensitivity analysis, samples from control individuals with reduced renal function (estimated glomerular filtration rate,60 ml/min/1.73 m2) were excluded. Correction for multiple testing was performed by permutation testing. A total of 1394 samples passed quality control filters. Following regression analysis adjusted by collection center, gender, duration of diabetes, and average HbA1c, two SNPs were significantly associated with DN. rs4420638 in the APOC1 region (odds ratio [OR] = 1.51; confidence intervals [CI]: 1.19–1.91; P = 0.001) and rs1532624 in CETP (OR = 0.82; CI: 0.69–0.99; P = 0.034); rs4420638 was also significantly associated in a sensitivity analysis (P = 0.016) together with rs7679 (P = 0.027). However, no association was significant following correction for multiple testing. Subgroup analysis of end-stage renal disease status failed to reveal any association. Our results suggest common variants associated with dyslipidemia are not strongly associated with DN in T1D among white individuals. Our findings, cannot entirely exclude these key genes which are central to the process of dyslipidemia, from involvement in DN pathogenesis as our study had limited power to detect variants of small effect size. Analysis in larger independent cohorts is required.

Haplotype association analysis of genes within the WNT signalling pathways in diabetic nephropathy

Background: Renal interstitial fibrosis and glomerular sclerosis are hallmarks of diabetic nephropathy (DN) and several studies have implicated members of the WNT pathways in these pathological processes. This study comprehensively examined common genetic variation within the WNT pathway for association with DN.

Methods: Genes within the WNT pathways were selected on the basis of nominal significance and consistent direction of effect in the GENIE meta-analysis dataset. Common SNPs and common haplotypes were examined within the selected WNT pathway genes in a white population with type 1 diabetes, discordant for DN (cases: n = 718; controls: n = 749). SNPs were genotyped using Sequenom or Taqman assays. Association analyses were performed using PLINK, to compare allele and haplotype frequencies in cases and controls. Correction for multiple testing was performed by either permutation testing or using false discovery rate.

Results: A logistic regression model including collection centre, duration of diabetes, and average HbA1c as covariates highlighted three SNPs in GSK3B (rs17810235, rs17471, rs334543), two in DAAM1 (rs1253192, rs1252906) and one in NFAT5 (rs17297207) as being significantly (P< 0.05) associated with DN, however these SNPs did not remain significant after correction for multiple testing. Logistic regression of haplotypes, with ESRD as the outcome, and pairwise interaction analyses did not yield any significant results after correction for multiple testing.

Conclusions: These results indicate that both common SNPs and common haplotypes of WNT pathway genes are not strongly associated with DN. However, this does not completely exclude these or the WNT pathways from association with DN, as unidentified rare genetic or copy number variants could still contribute towards the genetic architecture of DN.© 2013 Kavanagh et al.; licensee BioMed Central Ltd.

Comprehensive Investigation of the Caveolin 2 Gene: Resequencing and Association for Kidney Transplant Outcomes

Caveolae are plasma membrane structures formed from a complex of the proteins caveolin-1 and caveolin-2. Caveolae interact with pro-inflammatory cytokines and are dysregulated in fibrotic disease. Although caveolae are present infrequently in healthy kidneys, they are abundant during kidney injury. An association has been identified between a CAV1 gene variant and long term kidney transplant survival. Chronic, gradual decline in transplant function is a persistent problem in kidney transplantation. The aetiology of this is diverse but fibrosis within the transplanted organ is the common end point. This study is the first to investigate the association of CAV2 gene variants with kidney transplant outcomes. Genomic DNA from donors and recipients of 575 kidney transplants performed in Belfast was investigated for common variation in CAV2 using a tag SNP approach. The CAV2 SNP rs13221869 was nominally significant for kidney transplant failure. Validation was sought in an independent group of kidney transplant donors and recipients from Dublin, Ireland using a second genotyping technology. Due to the unexpected absence of rs13221869 from this cohort, the CAV2 gene was resequenced. One novel SNP and a novel insertion/deletion in CAV2 were identified; rs13221869 is located in a repetitive region and was not a true variant in resequenced populations. CAV2 is a plausible candidate gene for association with kidney transplant outcomes given its proximity to CAV1 and its role in attenuating fibrosis. This study does not support an association between CAV2 variation and kidney transplant survival. Further analysis of CAV2 should be undertaken with an awareness of the sequence complexities and genetic variants highlighted by this study. 

Lipoxins attenuate renal fibrosis by inducing let-7c and suppressing TGFβR1

Lipoxins, which are endogenously produced lipid mediators, promote the resolution of inflammation, and may inhibit fibrosis, suggesting a possible role in modulating renal disease. Here, lipoxin A4 (LXA4) attenuated TGF-ß1-induced expression of fibronectin, N-cadherin, thrombospondin, and the notch ligand jagged-1 in cultured human proximal tubular epithelial (HK-2) cells through a mechanism involving upregulation of the microRNA let-7c. Conversely, TGF-ß1 suppressed expression of let-7c. In cells pretreated with LXA4, upregulation of let-7c persisted despite subsequent stimulation with TGF-ß1. In the unilateral ureteral obstruction model of renal fibrosis, let-7c upregulation was induced by administering an LXA4 analog. Bioinformatic analysis suggested that targets of let-7c include several members of the TGF-ß1 signaling pathway, including the TGF-ß receptor type 1. Consistent with this, LXA4-induced upregulation of let-7c inhibited both the expression of TGF-ß receptor type 1 and the response to TGF-ß1. Overexpression of let-7c mimicked the antifibrotic effects of LXA4 in renal epithelia; conversely, anti-miR directed against let-7c attenuated the effects of LXA4. Finally, we observed that several let-7c target genes were upregulated in fibrotic human renal biopsies compared with controls. In conclusion, these results suggest that LXA4-mediated upregulation of let-7c suppresses TGF-ß1-induced fibrosis and that expression of let-7c targets is dysregulated in human renal fibrosis.

Genetic and Epigenetic Risk Factors for Diabetic Kidney Disease

Diabetes is increasing at daunting rates worldwide, and approximately 40% of affected individuals will develop kidney complications. Diabetic kidney disease (DKD) is the leading cause of end-stage kidney disease, and there are significant healthcare costs providing appropriate renal replacement therapies to affected individuals. For several decades, investigators have sought to discover inherited risk factors and biomarkers for DKD. In recent years, advances in high-throughput laboratory techniques and computational analyses, coupled with the establishment of multicenter consortia, have helped to identify genetic loci that are replicated across multiple populations. Several genome-wide association studies (GWAS) have been conducted for DKD with further meta-analysis of GWAS and comprehensive ”single gene” meta-analyses now published. Despite these efforts, much of the inherited predisposition to DKD remains unexplained. Meta-analyses and integrated–omics pathway studies are being used to help elucidate underlying genetic risks. Epigenetic phenomena are increasingly recognized as important drivers of disease risk, and several epigenome-wide association studies have now been completed. This review describes key findings and ongoing genetic and epigenetic initiatives for DKD.

Epigenetics of diabetic nephropathy

Diabetes is responsible for a large proportion of chronic kidney disease and end-stage renal disease worldwide. Careful monitoring and balanced control of blood glucose for individuals with type 1 diabetes can delay or prevent the onset of micro- and macro-vascular complications, however hyperglycaemia does not explain all of the risk for these diabetic complications. Genetic risk factors for diabetic nephropathy are being identified through international collaborations. However despite these advances a significant proportion of susceptibility remains unexplained, the so-called ‘missing heritability’.

Genome-wide association analysis identifies 13 new risk loci for schizophrenia

Schizophrenia is an idiopathic mental disorder with a heritable component and a substantial public health impact. We conducted a multi-stage genome-wide association study (GWAS) for schizophrenia beginning with a Swedish national sample (5,001 cases and 6,243 controls) followed by meta-analysis with previous schizophrenia GWAS (8,832 cases and 12,067 controls) and finally by replication of SNPs in 168 genomic regions in independent samples (7,413 cases, 19,762 controls and 581 parent-offspring trios). We identified 22 loci associated at genome-wide significance; 13 of these are new, and 1 was previously implicated in bipolar disorder. Examination of candidate genes at these loci suggests the involvement of neuronal calcium signaling. We estimate that 8,300 independent, mostly common SNPs (95% credible interval of 6,300-10,200 SNPs) contribute to risk for schizophrenia and that these collectively account for at least 32% of the variance in liability. Common genetic variation has an important role in the etiology of schizophrenia, and larger studies will allow more detailed understanding of this disorder.

Prospective investigation of body mass index, colorectal adenoma, and colorectal cancer in the prostate, lung, colorectal, and ovarian cancer screening trial

Obesity has consistently been linked to an increased risk of colorectal cancer, particularly among men. Whether body mass index (BMI) differentially influences the risk across the stages of colorectal cancer development remains unclear. We evaluated the associations of BMI with colorectal adenoma incidence, adenoma recurrence, and cancer in the context of a large screening trial, in which cases and controls had an equal chance for disease detection.

The Next Generation Transit Survey (NGTS)

The Next Generation Transit Survey (NGTS) is a new ground-based sky survey designed to find transiting Neptunes and super-Earths. By covering at least sixteen times the sky area of Kepler we will find small planets around stars that are sufficiently bright for radial velocity confirmation, mass determination and atmospheric characterisation. The NGTS instrument will consist of an array of twelve independently pointed 20cm telescopes fitted with red-sensitive CCD cameras. It will be constructed at the ESO Paranal Observatory, thereby benefiting from the very best photometric conditions as well as follow up synergy with the VLT and E-ELT. Our design has been verified through the operation of two prototype instruments, demonstrating white noise characteristics to sub-mmag photometric precision. Detailed simulations show that about thirty bright super-Earths and up to two hundred Neptunes could be discovered. Our science operations are due to begin in 2014.

CD2AP is associated with end-stage renal disease in patients with type 1 diabetes

CD2-associated protein (CD2AP) is essential for podocyte function. CD2AP mutations have been found in patients with focal segmental glomerulosclerosis, a disease histologically resembling diabetic nephropathy and often progressing to end-stage renal disease (ESRD). We hypothesised that variations in the CD2AP gene may contribute to susceptibility to glomerular injury in diabetes and investigated if single-nucleotide polymorphisms (SNPs) in CD2AP are associated with diabetic nephropathy in patients with type 1 diabetes. The discovery cohort consisted of 2,251 Finnish patients with type 1 diabetes. SNPs were selected from the HapMap database to cover the CD2AP gene. The associations between genotyped SNPs and diabetic nephropathy or ESRD were analysed with the chi-squared test and logistic regression. Three SNPs were selected for replication in cohorts from Denmark, Italy, the United Kingdom and Ireland. None of the 15 successfully genotyped SNPs were associated with diabetic nephropathy when compared to patients with normal albumin excretion rate. However, when genotype frequencies in patients with ESRD were compared with all other patients, two CD2AP SNPs, rs9369717 and rs9349417, were found to be associated with ESRD. The meta-analysis of the original and two additional European cohorts resulted in significant p values

Caveolin-1 single nucleotide polymorphism in antineutrophil cytoplasmic antibody associated vasculitis

Immunosuppression is cornerstone treatment of antineutrophil cytoplasmic antibody associated vasculitis (AAV) but is later complicated by infection, cancer, cardiovascular and chronic kidney disease. Caveolin-1 is an essential structural protein for small cell membrane invaginations known as caveolae. Its functional role has been associated with these complications. For the first time, caveolin-1 (CAV1) gene variation is studied in AAV.

DNA hypermethylation and DNA hypomethylation is present at different loci in chronic kidney disease

Genetic risk factors for chronic kidney disease (CKD) are being identified through international collaborations. By comparison, epigenetic risk factors for CKD have only recently been considered using population-based approaches. DNA methylation is a major epigenetic modification that is associated with complex diseases, so we investigated methylome-wide loci for association with CKD. A total of 485,577 unique features were evaluated in 255 individuals with CKD (cases) and 152 individuals without evidence of renal disease (controls). Following stringent quality control, raw data were quantile normalized and β values calculated to reflect the methylation status at each site. The difference in methylation status was evaluated between cases and controls with resultant P values adjusted for multiple testing. Genes with significantly increased and decreased levels of DNA methylation were considered for biological relevance by functional enrichment analysis using KEGG pathways in Partek Genomics Suite. Twenty-three genes, where more than one CpG per loci was identified with Padjusted < 10−8, demonstrated significant methylation changes associated with CKD and additional support for these associated loci was sought from published literature. Strong biological candidates for CKD that showed statistically significant differential methylation include CUX1, ELMO1, FKBP5, INHBA-AS1, PTPRN2, and PRKAG2 genes; several genes are differentially methylated in kidney tissue and RNA-seq supports a functional role for differential methylation in ELMO1 and PRKAG2 genes. This study reports the largest, most comprehensive, genome-wide quantitative evaluation of DNA methylation for association with CKD. Evidence confirming methylation sites influence development of CKD would stimulate research to identify epigenetic therapies that might be clinically useful for CKD.