The TRPC1 gene on chromosome 3q22–24 resides within the linkage region for diabetic nephropa-thy (DN) in type 1 (T1D) and type 2 diabetes mellitus (T2D). A recent study has demonstrated that TRPC1 expression is reduced in the kidney of diabetic ZDF- and STZ-treated rats. The present study aimed to evaluate the genetic and functional role of TRPC1 in the development of DN.
Genetic association study was performed with two independent cohorts, including 1,177 T1D European Americans with or without DN from GoKinD population and 850 African-American subjects with T2D-associated end-stage renal disease (ESRD), or with hypertensive (non-diabetic) ESRD, and nondiabetic controls. Seven tag SNP markers derived from HapMap data (phase II) were genotyped. TRPC1 gene expression was examined using real time RT-PCR.
No significant association of TRPC1 DNA polymorphisms with DN or ERSD was found in GoKinD and African-American populations. TRPC1 gene mRNA expression in kidney was found to be trendily reduced in 12-week and significantly in 26-week-old db/db mice.
TRPC1 genetic polymorphism may not fundamentally contribute to the development of DN, while reduction of the gene expression in kidney may be a late phenomenon of DN as seen in diabetic animal models.
TRPC1 gene; Single-nucleotide polymorphism; Diabetic nephropathy; End-stage renal disease; Diabetes types 1 and 2
Variants in the engulfment and cell motility 1 (ELMO1) gene are associated with nephropathy due to type 2 diabetes mellitus (T2DM) in a Japanese cohort. We comprehensively evaluated this gene in African American (AA) T2DM patients with end-stage renal disease (ESRD). Three hundred nine HapMap tagging SNPs and 9 reportedly associated SNPs were genotyped in 577 AA T2DM-ESRD patients and 596 AA non-diabetic controls, plus 43 non-diabetic European American controls and 45 Yoruba Nigerian samples for admixture adjustment. Replication analyses were conducted in 558 AAs with T2DM-ESRD and 564 controls without diabetes. Extension analyses included 328 AA with T2DM lacking nephropathy and 326 with non-diabetic ESRD. The original and replication analyses confirmed association with four SNPs in intron 13 (permutation p-values for combined analyses = 0.001-0.003), one in intron 1 (P=0.004) and one in intron 5 (P=0.002) with T2DM-associated ESRD. In a subsequent combined analysis of all 1,135 T2DM-ESRD cases and 1,160 controls, an additional 7 intron 13 SNPs produced evidence of association (P = 3.5×10-5 – P=0.05). No associations were seen with these SNPs in those with T2DM lacking nephropathy or with ESRD due to non-diabetic causes. Variants in intron 13 of the ELMO1 gene appear to confer risk for diabetic nephropathy in AA.
Association; Nephropathy; African American; Diabetes
Genetic studies in Turkish, Native American, European American, and African American (AA) families have linked chromosome 18q21.1-23 to susceptibility for diabetes associated nephropathy. In this study we have carried out fine linkage mapping in the 18q region previously linked to diabetic nephropathy in AAs by genotyping both microsatellite and single nucleotide polymorphisms (SNPs) for linkage analysis in an expanded set of 223 AA families multiplexed for type 2 diabetes associated ESRD (T2DM-ESRD). Several approaches were used to evaluate evidence of linkage with the strongest evidence for linkage in ordered subset analysis with an earlier age of T2DM diagnosis compared to the remaining pedigrees (LOD 3.9 at 90.1cM, ΔP=0.0161, NPL P value = 0.00002). Overall, the maximum LODs and LOD-1 intervals vary in magnitude and location depending upon analysis. The linkage mapping was followed up by performing a dense SNP map, genotyping 2,814 SNPs in the refined LOD-1 region in 1,029 AA T2DM-ESRD cases and 1,027 AA controls. Of the top 25 most associated SNPs, 10 resided within genic regions. Two candidate genes stood out: NEDD4L and SERPINB7. SNP rs512099, located in intron 1 of NEDD4L, was associated under a dominant model of inheritance (P value = 0.0006; Odds ratio (95% Confidence Interval) (OR (95%CI)) = 0.70 (0.57-0.86)). SNP rs1720843, located in intron 2 of SERPINB7, was associated under a recessive model of inheritance (P value = 0.0017; OR (95% CI) = 0.65 (0.50-0.85)). Collectively, these results suggest that multiple genes in this region may influence diabetic nephropathy susceptibility in AAs.
African American; diabetes type 2; nephropathy; linkage analysis; SNP; association analysis
The purpose of this study was to examine whether known genetic risk factors for type 1 diabetes (HLA-DRB1, -DQA1, and -DQB1 and insulin locus) play a role in the etiology of diabetic nephropathy.
RESEARCH DESIGN AND METHODS
Genetic analysis of HLA-DRB1, -DQA1, -DQB1 and the insulin gene (INS) was performed in the Genetics of Kidneys in Diabetes (GoKinD) collection of DNA (European ancestry subset), which includes case patients with type 1 diabetes and nephropathy (n = 829) and control patients with type 1 diabetes but not nephropathy (n = 904). The availability of phenotypic and genotypic data on GoKinD participants allowed a detailed analysis of the association of these genes with diabetic nephropathy.
Diabetic probands who were homozygous for HLA-DRB1*04 were 50% less likely to have nephropathy than probands without any DRB1*04 alleles. In heterozygous carriers, a protective effect of this allele was not as clearly evident; the mode of inheritance therefore remains unclear. This association was seen in probands with both short (<28 years, P = 0.02) and long (≥28 years, P = 0.0001) duration of diabetes. A1C, a marker of sustained hyperglycemia, was increased in control probands with normoalbuminuira, despite long-duration diabetes, from 7.2 to 7.3 to 7.7% with 0, 1, and 2 copies of the DRB1*04 allele, respectively. This result is consistent with a protective effect of DRB1*04 that may allow individuals to tolerate higher levels of hyperglycemia, as measured by A1C, without developing nephropathy.
These data suggest that carriers of DRB1*04 are protected from some of the injurious hyperglycemic effects related to nephropathy. Interestingly, DRB1*04 appears to be both a risk allele for type 1 diabetes and a protective allele for nephropathy.
MCF2L2, ADIPOQ and SOX2 genes are located in chromosome 3q26-27, which is linked to diabetic nephropathy (DN). ADIPOQ and SOX2 genetic polymorphisms are found to be associated with DN. In the present study, we first investigated the association between MCF2L2 and DN, and then evaluated effects of these three genes on the development of DN.
A total of 1177 type 1 diabetes patients with and without DN from the GoKinD study were genotyped with TaqMan allelic discrimination. All subjects were of European descent.
Leu359Ile T/G variant in the MCF2L2 gene was found to be associated with DN in female subjects (P = 0.017, OR = 0.701, 95%CI 0.524-0.938) but not in males. The GG genotype carriers among female patients with DN had tendency decreased creatinine and cystatin levels compared to the carriers with either TT or TG genotypes. This polymorphism MCF2L2-rs7639705 together with SNPs of ADIPOQ-rs266729 and SOX2-rs11915160 had combined effects on decreased risk of DN in females (P = 0.001).
The present study provides evidence that MCF2L2, ADIPOQ and SOX2 genetic polymorphisms have effects on the resistance of DN in female T1D patients, and suggests that the linkage with DN in chromosome 3q may be explained by the cumulated genetic effects.
Conflicting reports exist as to whether sickle cell trait is a risk factor for the progression of nephropathy. In order to determine whether African Americans with sickle cell trait are at increased risk for kidney disease, we assessed the genetic association between sickle cell trait and end-stage renal disease (ESRD). Hemoglobin S, non-muscle myosin heavy chain 9 (MYH9), and apolipoprotein L1 (APOL1) risk variants were genotyped in 3258 unrelated African Americans: 1085 with non-diabetic ESRD, 996 with type 2 diabetes-associated ESRD, and 1177 controls. Since APOL1 is strongly associated with ESRD in African Americans, interactions between APOL1 and MYH9 risk variants and hemoglobin S were assessed using case-only and case-control centered two-way logistic regression interaction analyses. The sickle cell trait genotype frequencies were 8.7% in non-diabetic ESRD, 7.1% in type 2 diabetes-ESRD, and 7.2% in controls. There was no age-, gender-, and admixture-adjusted significance for sickle cell trait association with non-diabetic ESRD (odds ratio 1.16); type 2 diabetes-ESRD (odds ratio 1.01); or all-cause ESRD (combined non-diabetic and type 2 diabetic-ESRD patients compared to the controls; odds ratio 1.05) in dominant models. In addition, no evidence of APOL1 or MYH9 interactions with sickle cell trait was detected. Hence, sickle cell trait is not associated with diabetic or non-diabetic ESRD in a large sample of African Americans.
African American; APOL1; diabetes; end-stage kidney disease; hemoglobin S; hypertension
Familial clustering of disparate kidney diseases including clinically diagnosed hypertensive and diabetic nephropathy, idiopathic focal segmental glomerulosclerosis (FSGS) and Human Immunodeficiency Virus-associated nephropathy are often observed in African Americans. Admixture mapping recently identified the non-muscle myosin heavy chain 9 gene (MYH9) as a susceptibility factor strongly associated with several non-diabetic etiologies of end-stage renal disease (ESRD) in African Americans, less strongly with diabetes-associated ESRD. MYH9-associated nephropathies reside in the spectrum of FSGS/focal global glomerulosclerosis. The renal histology in proteinuric African Americans homozygous for MYH9 risk variants with longstanding type 2 diabetes mellitus is unknown. We report a case of coincident idiopathic FSGS, collapsing variant; and diabetic nephropathy in an African American homozygous for the MYH9 E1 risk haplotype. This case demonstrates that diabetic African Americans with overt proteinuria can have mixed renal lesions, including those in the spectrum of MYH9-associated nephropathy. Careful interpretation of kidney biopsies in proteinuric African Americans with diabetes is necessary to exclude coincident non-diabetic forms of nephropathy, precisely define etiologies of kidney disease, and determine the natural history and treatment response in mixed lesions of diabetes-associated and MYH9-associated kidney disease.
We report a case of coincident idiopathic FSGS, collapsing variant; and diabetic nephropathy in an African American homozygous for the MYH9 E1 risk haplotype.
African American; collapsing variant focal segmental glomerulosclerosis; diabetes; diabetic nephropathy; MYH9
Background. Although MYH9 is strongly associated with biopsy-proven idiopathic and HIV-associated focal segmental glomerulosclerosis (FSGS) and clinically diagnosed ‘hypertension-associated’ end-stage renal disease (ESRD) in African Americans, its role in type 2 diabetes mellitus (T2DM)-associated ESRD is unclear.
Methods. To assess whether MYH9 was associated with T2DM-ESRD, 751 African Americans with T2DM-ESRD, 227 with T2DM lacking nephropathy and 925 non-diabetic non-nephropathy controls were genotyped for 14 MYH9 SNPs. Association analyses used SNPGWA and Dandelion.
Results. Comparing T2DM-ESRD cases with non-diabetic controls, single SNP associations were detected with 8 of 14 SNPs, gender- and admixture-adjusted P-values 0.047–0.005 [recessive model, odds ratio (OR) range 1.30–1.55]. The previously associated MYH9 E1 and L1 haplotypes were associated with T2DM-ESRD (E1: OR 1.27, 95% CI 1.04–1.56, P = 0.021 recessive and L1: OR 1.43, 95% CI 1.09–1.87, P = 0.009 dominant). Contrasting the 751 T2DM-ESRD cases with 227 T2DM non-nephropathy controls revealed that E1 haplotype SNPs rs4821480, rs2032487 and rs4821481 were associated with kidney failure (OR 1.38–1.40 recessive, all P < 0.048). Among E1 and L1 risk homozygotes, respectively, mean (SD) diabetes duration prior to renal replacement therapy was 16.6 (9.7) and 16.4 (10.0) years, and 65% had diabetic retinopathy.
Conclusions. Genetic dissection of T2DM-associated ESRD reveals that MYH9 underlies a portion of this clinically diagnosed disorder in African Americans. It is likely that a subset of African Americans with T2DM and coincident nephropathy have primary MYH9-related kidney disease (e.g. FSGS or global glomerulosclerosis), although renal biopsy studies need to be performed.
African American; diabetic nephropathy; kidney; MYH9; type 2 diabetes mellitus
The fat mass and obesity associated (FTO) gene has an important genetic effect on body mass index (BMI) and risk of obesity, and obesity contributes to the progression of renal diseases, including diabetic nephropathy. We thus conducted a genetic association study to evaluate whether the FTO gene confers the risk susceptibility to the development of diabetic nephropathy. Genotyping experiments of the common FTO polymorphism, rs9939609, in 1170 type 1 diabetes patients with (n = 597) or without diabetic nephropathy (n = 573) were performed with TaqMan allelic discrimination. All subjects are of European descent and selected from the Genetics of Kidney Diseases in Diabetes (GoKinD) study. The frequency of T allele of this polymorphism was 0.414 in the studied population. There was no allelic association of this polymorphism with diabetic nephropathy. But, the risk susceptibility of A allele conferring to the increased BMI among type 1 diabetes patients was observed. The subjects carrying with AA genotype had higher BMI compared to the carriers with TA and/or TT genotype(s) (P ≤ 0.019). The present study provides evidence that the common FTO genetic polymorphism, rs9939609, is associated with increased BMI in type 1 diabetes but not with diabetic nephropathy.
diabetic nephropathy; fat mass and obesity associated; genetic association; single nucleotide polymorphism
The Genetics of Kidneys in Diabetes (GoKinD) study was initiated to facilitate research aimed at identifying genes involved in diabetic nephropathy (DN) in type 1 diabetes (T1D). In this review, we present on overview of this study and the various reports that have utilized its collection. At the forefront of these efforts is the recent genome-wide association (GWA) scan implemented on the GoKinD collection. We highlight the results from our analysis of these data and describe compelling evidence from animal models that further support the potential role of associated loci in the susceptibility of DN. To enhance our analysis of genetic associations in GoKinD, using genome-wide imputation (GWI), we expanded our analysis of this collection to include genotype data from more than 2.4 million common SNPs. We illustrate the added utility of this enhanced dataset through the comprehensive fine-mapping of candidate genomic regions previously linked with DN and the targeted investigation of genes involved in candidate pathway implicated in its pathogenesis. Collectively, GWA and GWI data from the GoKinD collection will serve as a springboard for future investigations into the genetic basis of DN in T1D.
genome-wide association; diabetic nephropathy; type 1 diabetes; imputation
Background & Aims
Excessive Ca2+ influx mediates many cytotoxic processes, including those associated with autoimmune inflammatory diseases such as acute pancreatitis and Sjögren's syndrome. TRPC3 is a major Ca2+ influx channel in pancreatic and salivary gland cells. We investigated whether genetic or pharmacological inhibition of TRPC3 protects pancreas and salivary glands from Ca2+-dependent damage.
We developed a Ca2+-dependent model of cell damage for salivary gland acini. Acute pancreatitis was induced by injection of cerulein into wild-type and Trpc3−/− mice. Mice were also given the Trpc3-selective inhibitor pyrazole 3 (Pyr3).
Salivary glands and pancreas of Trpc3−/− mice were protected from Ca2+-mediated cell toxicity. Analysis of Ca2+ signaling in wild-type and Trpc3−/− acini showed that Pyr3 is highly specific inhibitor of Tprc3; it protected salivary glands and pancreas cells from Ca2+-mediated toxicity by inhibiting the Trpc3-mediated component of Ca2+ influx.
TRPC3-mediated Ca2+ influx mediates damage to pancreas and salivary glands. Pharmacological inhibition of TRPC3 with the highly selective TRPC3 inhibitor Pyr3 might be developed for treatment of patients with acute pancreatitis and Sjögren's syndrome.
Ca2+ influx; inflammation; cell death; therapeutics
To examine the association between single nucleotide polymorphisms (SNPs) in the engulfment and cell motility 1 (ELMO1) gene, a locus previously shown to be associated with diabetic nephropathy in two ethnically distinct type 2 diabetic populations, and the risk of nephropathy in type 1 diabetes.
RESEARCH DESIGN AND METHODS
Genotypic data from a genome-wide association scan (GWAS) of the Genetics of Kidneys in Diabetes (GoKinD) study collection were analyzed for associations across the ELMO1 locus. In total, genetic associations were assessed using 118 SNPs and 1,705 individuals of European ancestry with type 1 diabetes (885 normoalbuminuric control subjects and 820 advanced diabetic nephropathy case subjects).
The strongest associations in ELMO1 occurred at rs11769038 (odds ratio [OR] 1.24; P = 1.7 × 10−3) and rs1882080 (OR 1.23; P = 3.2 × 10−3) located in intron 16. Two additional SNPs, located in introns 18 and 20, respectively, were also associated with diabetic nephropathy. No evidence of association for variants previously reported in type 2 diabetes was observed in our collection.
Using GWAS data from the GoKinD collection, we comprehensively examined evidence of association across the ELMO1 locus. Our investigation marks the third report of associations in ELMO1 with diabetic nephropathy, further establishing its role in the susceptibility of this disease. There is evidence of allelic heterogeneity, contributed by the diverse genetic backgrounds of the different ethnic groups examined. Further investigation of SNPs at this locus is necessary to fully understand the commonality of these associations and the mechanism(s) underlying their role in diabetic nephropathy.
Background. Non-diabetic forms of nephropathy commonly lead to end-stage renal disease (non-DM ESRD). Previous studies have demonstrated that African Americans are more susceptible to non-DM ESRD compared to other ethnic groups, and this risk has a strong genetic component. A genome-wide scan for ESRD in African American families enriched for non-DM ESRD showed evidence for linkage in chromosome 13q33.3, and a candidate gene in this region, klotho, was selected for a detailed analysis in a follow-up case-control association study.
Methods. Thirty-four single-nucleotide polymorphisms (SNPs) in the klotho gene were genotyped in 317 unrelated African American non-DM ESRD cases and 354 non-nephropathy controls, including 12 SNPs identified by re-sequencing a region around exon 4.
Results. Two SNPs demonstrated modest admixture-adjusted evidence of association with non-DM ESRD, rs650439 (P = 0.013, recessive model) and rs643780 (P = 0.017, recessive model), while rs17643698 approached significance (P = 0.0953, two degrees of freedom test). Eight of the most significant SNPs were tested for replication in a second case-control collection (557 African American non-DM ESRD cases and 187 controls), and there was no evidence of association in replicate cases and controls; nor when the samples were combined for a total of 874 non-DM cases and 541 controls. Cox proportional hazards models were computed to test for association between polymorphisms in klotho and age at onset of ESRD. A three-SNP haplotype, rs526906, rs525014 and rs571118 (T/T/A), was associated with age of onset of ESRD [P = 0.007, recessive model; hazard ratio (HR) = 0.70]. Subjects homozygous for this haplotype had a mean 4 years later onset of ESRD, suggesting a slower disease progression. HapMap subjects homozygous for this haplotype had increased expression of klotho, further supporting a protective role of this variant in ESRD.
Conclusion. We conclude that three SNPs in intron 1 of the klotho gene are associated with delayed age at onset of non-DM ESRD in African Americans.
genetics; klotho; non-diabetic ESRD
The Genetics of Kidneys in Diabetes (GoKinD) study is an initiative that aims to identify genes that are involved in diabetic nephropathy. A large number of individuals with type 1 diabetes were screened to identify two subsets, one with clear-cut kidney disease and another with normal renal status despite long-term diabetes. Those who met additional entry criteria and consented to participate were enrolled. When possible, both parents also were enrolled to form family trios. As of November 2005, GoKinD included 3075 participants who comprise 671 case singletons, 623 control singletons, 272 case trios, and 323 control trios. Interested investigators may request the DNA collection and corresponding clinical data for GoKinD participants using the instructions and application form that are available at http://www.gokind.org/access. Participating scientists will have access to three data sets, each with distinct advantages. The set of 1294 singletons has adequate power to detect a wide range of genetic effects, even those of modest size. The set of case trios, which has adequate power to detect effects of moderate size, is not susceptible to false-positive results because of population substructure. The set of control trios is critical for excluding certain false-positive results that can occur in case trios and may be particularly useful for testing gene—environment interactions. Integration of the evidence from these three components into a single, unified analysis presents a challenge. This overview of the GoKinD study examines in detail the power of each study component and discusses analytic challenges that investigators will face in using this resource.
This meta-analysis assessed the pooled effect of each genetic variant reproducibly associated with diabetic nephropathy.
PubMed, EMBASE and Web of Science were searched for articles assessing the association between genes and diabetic nephropathy. All genetic variants statistically associated with diabetic nephropathy in an initial study, then independently reproduced in at least one additional study, were selected. Subsequently, all studies assessing these variants were included. The association between these variants and diabetic nephropathy (defined as macroalbuminuria/proteinuria or end-stage renal disease [ESRD]) was calculated at the allele level and the main measure of effect was a pooled odds ratio. Pre-specified subgroup analyses were performed, stratifying for type 1/type 2 diabetes mellitus, proteinuria/ESRD and ethnic group.
The literature search yielded 3,455 citations, of which 671 were genetic association studies investigating diabetic nephropathy. We identified 34 replicated genetic variants. Of these, 21 remained significantly associated with diabetic nephropathy in a random-effects meta-analysis. These variants were in or near the following genes: ACE, AKR1B1 (two variants), APOC1, APOE, EPO, NOS3 (two variants), HSPG2, VEGFA, FRMD3 (two variants), CARS (two variants), UNC13B, CPVL and CHN2, and GREM1, plus four variants not near genes. The odds ratios of associated genetic variants ranged from 0.48 to 1.70. Additional variants were detected in subgroup analyses: ELMO1 (Asians), CCR5 (Asians) and CNDP1 (type 2 diabetes).
This meta-analysis found 24 genetic variants associated with diabetic nephropathy. The relative contribution and relevance of the identified genes in the pathogenesis of diabetic nephropathy should be the focus of future studies.
Electronic supplementary material
The online version of this article (doi:10.1007/s00125-010-1996-1) contains supplementary material, which is available to authorised users.
Diabetes mellitus; Diabetic nephropathy; Genetic association studies; Genetic polymorphism; Meta-analysis
A genome-wide association study was performed using the Affymetrix 6.0 chip to identify genes associated with diabetic nephropathy in African Americans. Association analysis was performed adjusting for admixture in 965 type 2 diabetic African American patients with end-stage renal disease (ESRD) and in 1029 African Americans without type 2 diabetes or kidney disease as controls. The top 724 single nucleotide polymorphisms (SNPs) with evidence of association to diabetic nephropathy were then genotyped in a replication sample of an additional 709 type 2 diabetes-ESRD patients and 690 controls. SNPs with evidence of association in both the original and replication studies were tested in additional African American cohorts consisting of 1246 patients with type 2 diabetes without kidney disease and 1216 with non-diabetic ESRD to differentiate candidate loci for type 2 diabetes-ESRD, type 2 diabetes, and/or all-cause ESRD. Twenty-five SNPs were significantly associated with type 2 diabetes-ESRD in the genome-wide association and initial replication. Although genome-wide significance with type 2 diabetes was not found for any of these 25 SNPs, several genes, including RPS12, LIMK2, and SFI1 are strong candidates for diabetic nephropathy. A combined analysis of all 2890 patients with ESRD showed significant association SNPs in LIMK2 and SFI1 suggesting that they also contribute to all-cause ESRD. Thus, our results suggest that multiple loci underlie susceptibility to kidney disease in African Americans with type 2 diabetes and some may also contribute to all-cause ESRD.
The incidence of end-stage renal disease (ESRD) in the US is rising at an alarming rate, with the largest increase among African-American populations. The key risk factors for kidney disease are hypertension and diabetes, which are both becoming more prevalent in the US, and particularly in African Americans. Although African Americans make up 12.6% of the US population, the incidence of diabetes-related ESRD is four times higher than for whites, and the prevalence of ESRD due to hypertension is twice that of white patients. Approximately 30 to 40% of all patients with diabetes will develop nephropathy and many will progress to ESRD, necessitating dialysis or kidney transplantation. Recent studies in patients with type 2 diabetes indicate a significant delay in progression or development of diabetic nephropathy following blockade of the renin-angiotensin-aldosterone system with the use of angiotensin receptor antagonists. Early intervention in patients with hypertension is necessary to prevent kidney damage, and data from the African American Study of Kidney Disease and Hypertension suggest that angiotensin-converting enzyme inhibitors are effective in this population. Although African-American patients receiving hemodialysis appear to have increased survival compared with whites, racial factors and poor access to medical care contribute to the increased risk of kidney disease in minorities. A concerted effort is necessary to raise awareness in minority populations and provide strategies for prevention and early treatment thereby attenuating the increasing prevalence of kidney failure in these groups.
Recent studies have reported that aberrant expression of transient receptor potential channel C6 (TRPC6) in a variety of human cancers is associated with aggressive behavior. However, the functional significance of TRPC6 in human cervical cancer is not known. This study was planned to detect whether TRPC6 is expressed in cervical cancer tissue and to evaluate the association between TPRC6 expression and clinicopathologic features.
Tissue samples were collected from the West China Second UNIV Hospital of Sichuan University. TRPC6 expression was detected by quantitative real-time reverse transcription polymerase chain reaction and Western blotting. TRPC6 expression was evaluated by immunohistochemistry analysis of 40 cervical cancer specimens, and correlations were sought between elevated expression of TRPC6 and clinicopathologic features.
Increased expression of TRPC6 was detected in 25 of the 40 cervical cancer samples. Positive cells found in cervical carcinomas were significantly increased in numbers compared with specimens without lymphovascular space invasion. Elevated expression of TRPC6 was neither related to International Federation of Gynecology and Obstetrics stage nor pelvic lymph metastases. Indeed, the clinicopathologic analysis indicated that overexpression of TRPC6 was significantly associated with lymphovascular space invasion.
These results indicate that elevated expression of TRPC6 might be associated with an aggressive cervical cancer phenotype.
cervical cancer; transient receptor potential channel C6; overexpression; invasion
African Americans (AAs) have increased susceptibility to non-diabetic nephropathy relative to European Americans.
Follow-up of a pooled genome-wide association study (GWAS) in AA dialysis patients with nondiabetic nephropathy; novel gene-gene interaction analyses.
Setting & Participants
Wake Forest sample: 962 AA nondiabetic nephropathy cases; 931 non-nephropathy controls. Replication sample: 668 Family Investigation of Nephropathy and Diabetes (FIND) AA nondiabetic nephropathy cases; 804 non-nephropathy controls.
Individual genotyping of top 1420 pooled GWAS-associated single nucleotide polymorphisms (SNPs) and 54 SNPs in six nephropathy susceptibility genes.
APOL1 genetic association and additional candidate susceptibility loci interacting with, or independently from, APOL1.
The strongest GWAS associations included two non-coding APOL1 SNPs, rs2239785 (odds ratio [OR], 0.33; dominant; p = 5.9 × 10−24) and rs136148 (OR, 0.54; additive; p = 1.1 × 10−7) with replication in FIND (p = 5.0 × 10−21 and 1.9 × 10−05, respectively). Rs2239785 remained significantly associated after controlling for the APOL1 G1 and G2 coding variants. Additional top hits included a CFH SNP(OR from meta-analysis in above 3367 AA cases and controls, 0.81; additive; p = 6.8 × 10−4). The 1420 SNPs were tested for interaction with APOL1 G1 and G2 variants. Several interactive SNPs were detected, the most significant was rs16854341 in the podocin gene (NPHS2) (p = 0.0001).
Non-pooled GWAS have not been performed in AA nondiabetic nephropathy.
This follow-up of a pooled GWAS provides additional and independent evidence that APOL1 variants contribute to nondiabetic nephropathy in AAs and identified additional associated and interactive non-diabetic nephropathy susceptibility genes.
African American; APOL1; CFH; end-stage renal disease; FIND; FSGS; hypertension
The surface expression and channel activation of transient receptor potential canonical 6 (TRPC6) were regulated by tyrosine phosphorylation and resultant binding with stimulatory PLC-γ1 and inhibitory nephrin. Disease-causing mutations made the TRPC6s insensitive to nephrin suppression, suggesting that the cell-type–specific regulation of TRPC6 might be involved in the pathogenesis.
Transient receptor potential canonicals (TRPCs) play important roles in the regulation of intracellular calcium concentration. Mutations in the TRPC6 gene are found in patients with focal segmental glomerulosclerosis (FSGS), a proteinuric disease characterized by dysregulated function of renal glomerular epithelial cells (podocytes). There is as yet no clear picture for the activation mechanism of TRPC6 at the molecular basis, however, and the association between its channel activity and pathogenesis remains unclear. We demonstrate here that tyrosine phosphorylation of TRPC6 induces a complex formation with phospholipase C (PLC)-γ1, which is prerequisite for TRPC6 surface expression. Furthermore, nephrin, an adhesion protein between the foot processes of podocytes, binds to phosphorylated TRPC6 via its cytoplasmic domain, competitively inhibiting TRPC6–PLC-γ1 complex formation, TRPC6 surface localization, and TRPC6 activation. Importantly, FSGS-associated mutations render the mutated TRPC6s insensitive to nephrin suppression, thereby promoting their surface expression and channel activation. These results delineate the mechanism of TRPC6 activation regulated by tyrosine phosphorylation, and imply the cell type–specific regulation, which correlates the FSGS mutations with deregulated TRPC6 channel activity.
Numerous studies have documented the presence of racial disparities among Americans in health outcomes with respect to cardiovascular disease, infant mortality, cancer, and kidney disease. With regard to kidney diseases, these disparities are more dramatic. African, Hispanic, and Native Americans have the highest risks of end-stage renal disease (ESRD). The incidence of ESRD is four times higher in African Americans than in whites. Diseases causing chronic kidney failure, such as diabetes mellitus, hypertension, systemic lupus erythematosus, and human immunodeficiency virus-associated nephropathy, are particularly prevalent among African-American patients. In addition to the higher prevalence, the morbidity associated with kidney complications of these diseases appears worse in African-American patients. African Americans also have worse outcomes and a relatively reduced access to kidney transplantation--the best therapy for ESRD. It is highly likely that social and environmental factors play a very significant role in the persistence of these disparities. A detailed understanding of these socioeconomic and environmental factors will be critical in formulating rational public health strategies to redress these disparities. This paper reviews the social, economic and environmental factors that impact on the incidence of ESRD in minority groups.
Transient receptor potential channels Trpc2 and Trpc3 are expressed on normal murine erythroid precursors, and erythropoietin stimulates an increase in intracellular calcium ([Ca2+]i) through TRPC2 and TRPC3. Because modulation of [Ca2+]i is an important signaling pathway in erythroid proliferation and differentiation, Trpc2, Trpc3, and Trpc2/Trpc3 double knockout mice were utilized to explore the roles of these channels in erythropoiesis. Trpc2, Trpc3, and Trpc2/Trpc3 double knockout mice were not anemic, and had similar red blood cell counts, hemoglobins, and reticulocyte counts as wild type littermate controls. Although the erythropoietin induced increase in [Ca2+]i was reduced, these knockout mice showed no defects in red cell production. The major phenotypic difference at steady state was that the mean corpuscular volume, mean corpuscular hemoglobin, and hematocrit of red cells were significantly greater in Trpc2 and Trpc2/Trpc3 double knockout mice, and mean corpuscular hemoglobin concentration was significantly reduced. All hematological parameters in Trpc3 knockout mice were similar to controls. When exposed to phenyhydrazine, unlike the Trpc3 knockouts, Trpc2 and Trpc2/Trpc3 double knockout mice showed significant resistance to hemolysis. This was associated with significant reduction in hydrogen peroxide-induced calcium influx in erythroblasts. While erythropoietin induced calcium influx through TRPC2 or TRPC3 is not critical for erythroid production, these data demonstrate that TRPC2 plays an important role in oxidative stress-induced hemolysis which may be related to reduced calcium entry in red cells in the presence of Trpc2 depletion.
TRP Channels; Trpc2; Trpc3; erythropoietin; oxidative stress
Transient receptor potential (TRP) C1 and C3 (TRPC1 and TRPC3) are expressed in vascular smooth muscle cells and are thought to be involved in vascular contractility. In the present study, we determined the effect of systemic hypertension on TRPC1/TRPC3 channel expression and vascular contractility in rat carotid artery (CA). CA were studied from male spontaneously hypertensive rats (SHR), Wistar-Kyoto (WKY), and Long Evans (LE) rats. TRPC1/3 expression was determined by RT-PCR and Western blot. TRP channel function was evaluated by whole-cell patch clamp, using UTP (60 μM) to stimulate TRPC1/3 channels. Contractions of endothelium-denuded CA segments to UTP (1–300 μM) and phenylephrine (Phe; 0.1 nM–10 μM) were measured in an isometric tension bath. TRPC1 and TRPC3 mRNA was present in CA of both WKY and SHR. Western blot demonstrated 3.1 ± 1.2 times greater TRPC3 expression and 0.5 ± 0.2 times TRPC1 in SHR versus WKY CA. Isolated CA showed potentiated contraction to UTP in the SHR versus WKY. Activation of voltage-dependent Ca2+ channels (VDCC) in UTP-mediated constriction only occurred in SHR CA. Contraction to Phe was unaltered between WKY and SHR CA and involved equal significant VDCC activation in both groups. Patch clamp demonstrated that the UTP-stimulated current (Iutp) was greater in SHR compared to the normotensive WKY and LE rats with peak Iutp (at −110 mV) of −63 ± 24 pA compared to −25 ± 4 pA, respectively. We demonstrate that UTP-mediated but not Phe-mediated constrictions are potentiated in the CA during hypertension. Expression of TRPC1 is decreased whereas TRPC3 is increased in SHR CA. Interestingly, VDCC activation only contributes to UTP-mediated contraction of SHR CAs whereas it contributes substantially and equally in Phe-mediated contraction. We speculate that the alteration of TRPC channel expression in hypertension leads to greater smooth muscle depolarization, VDCC activation, and vascular contractility in the UTP (but not Phe) signaling pathway.
calcium; carotid artery; hypertension; transient receptor potential channel; vascular smooth muscle
The vascular ectonucleotidase ENTPD1 protects against renal injury and modulates glucose homeostasis in mouse models. We sought to determine whether human variation in ENTPD1 influences predisposition to diabetes or diabetic nephropathy.
RESEARCH DESIGN AND METHODS
We analyzed ENTPD1 single nucleotide polymorphisms (SNPs) in 363 African American control subjects, 380 subjects with type 2 diabetes and end-stage renal disease (DM-ESRD), and 326 subjects with ESRD unrelated to diabetes (non–DM-ESRD). Using human cell lines, we correlated disease-associated ENTPD1 haplotypes with ENTPD1 gene expression. Finally, we studied consequences of ENTPD1 deletion in a mouse model of type 2 diabetes (db/db).
A common ENTPD1 two-SNP haplotype was associated with increased risk for DM-ESRD (P = 0.0027), and an uncommon four-SNP haplotype was associated with protection against DM-ESRD (P = 0.004). These haplotypes correlated with ENTPD1 gene expression levels in human cell lines in vitro. Subjects with high ENTPD1-expressing haplotypes were enriched in the DM-ESRD group. By crossing ENTPD1-null mice with db mice, we show that ENTPD1 deletion has prominent effects on metabolic syndrome traits. Specifically, deletion of ENTPD1 lowered glucose levels in control (db/−) mice with one functional leptin receptor and dramatically lowered weights in db/db mice with no functional leptin receptors. Similar effects were seen in aged ENTPD1-null mice with normal leptin receptors.
ENTPD1 polymorphisms appear to influence susceptibility to type 2 diabetes and/or diabetic nephropathy in African Americans. Studies in human cell lines and in vivo mouse data support a potential role for ENTPD1 genetic variation in susceptibility to type 2 diabetes.
Diabetic retinopathy (DR) and diabetic nephropathy (DN) are serious microvascular complications of diabetes mellitus. Correlations between severity of DR and DN and computed heritability estimates for DR were determined in a large, multiethnic sample of diabetic families. The hypothesis was that (1) the severity of DR correlates with the presence and severity of nephropathy in individuals with diabetes mellitus, and (2) the severity of DR is under significant familial influence in members of multiplex diabetic families.
The Family Investigation of Nephropathy and Diabetes (FIND) was designed to evaluate the genetic basis of DN in American Indians, European Americans, African Americans, and Mexican Americans. FIND enrolled probands with advanced DN, along with their diabetic siblings who were concordant and discordant for nephropathy. These diabetic family members were invited to participate in the FIND-Eye study to determine whether inherited factors underlie susceptibility to DR and its severity. FIND-Eye participants underwent eye examinations and had fundus photographs taken. The severity of DR was graded by using the Early Treatment Diabetic Retinopathy Study Classification (ETDRS). Sib–sib correlations were calculated with the SAGE 5.0 program FCOR, to estimate heritability of retinopathy severity.
This report summarizes the results for the first 2368 diabetic subjects from 767 families enrolled in FIND-Eye; nearly 50% were Mexican American, the largest single ethnicity within FIND. The overall prevalence of DR was high; 33.4% had proliferative DR; 7.5%, 22.8%, and 9.5% had severe, moderate, and mild nonproliferative DR, respectively; 26.6% had no DR. The severity of DR was significantly associated with severity of DN, both by phenotypic category and by increasing serum creatinine concentration (χ2 = 658.14, df = 20; P < 0.0001). The sib–sib correlation for DR severity was 0.1358 in the total sample and 0.1224 when limited to the Mexican-American sample. Broad sense heritabilities for DR were 27% overall and 24% in Mexican-American families. The polygenic heritability of liability for proliferative DR approximated 25% in this FIND-Eye sample.
These data confirm that the severity of DR parallels the presence and severity of nephropathy in individuals with diabetes mellitus. The severity of DR in members of multiplex diabetic families appears to have a significant familial connection.