Zhaohui Wang

Genome-wide association study identifies susceptibility loci for IgA nephropathy

We carried out a genome-wide association study of IgA nephropathy, a major cause of kidney failure worldwide. We studied 1,194 cases and 902 controls of Chinese Han ancestry, with targeted follow up in Chinese and European cohorts comprising 1,950 cases and 1,920 controls. We identified three independent loci in the major histocompatibility complex, as well as a common deletion of CFHR1 and CFHR3 at chromosome 1q32 and a locus at chromosome 22q12 that each surpassed genome-wide significance (P values for association between 1.59 × 10−26 and 4.84 × 10−9 and minor allele odds ratios of 0.63–0.80). These five loci explain 4–7% of the disease variance and up to a tenfold variation in interindividual risk. Many of the alleles that protect against IgA nephropathy impart increased risk for other autoimmune or infectious diseases, and IgA nephropathy risk allele frequencies closely parallel the variation in disease prevalence among Asian, European and African populations, suggesting complex selective pressures.

Knockdown of Stat3 activity in vivo prevents diabetic glomerulopathy

Recent studies suggest that Stat3, a transcription factor that mediates cytokine signaling, plays a critical role in the pathogenesis of diabetic nephropathy. Complete Stat3 gene knockout is embryonic lethal; therefore, we crossed Stat3+/- mice with Stat3 mutant mice (SA/SA) that lack full Stat3 activity. This strategy generated Stat3SA/- mice (25% activity) and Stat3SA/+ mice (75% activity), which were made diabetic using streptozotocin in order to define the role of Stat3 in diabetic kidney disease. While the glomerular number was not different between these two groups of mice, the diabetic SA/- mice had significantly less proteinuria, mesangial expansion, glomerular cell proliferation, and macrophage infiltration than the diabetic SA/+ mice. The reduction in Stat3 activity did not affect glomerular hyperfiltration seen after the induction of diabetes, as it was increased to the same degree in both groups of mice. Phosphorylation of Stat3 was markedly increased in the glomeruli of diabetic SA/+ mice compared to diabetic SA/- mice. The expression of inflammatory markers, IL-6, MCP-1, and activated NF-kappaB; type IV collagen, TGF-beta, and ICAM-1 mRNA; or type IV collagen and TGF-beta protein, were all found to be significantly less in glomeruli isolated from diabetic SA/- mice, as compared with diabetic SA/+ mice. Our study shows that Stat3 plays a critical role in the regulation of inflammation and abnormal matrix synthesis at an early stage of DN.

Identification and functional analysis of a novel TRPC6 mutation associated with late onset familial focal segmental glomerulosclerosis in Chinese patients

OBJECTIVEnMutations in the TRPC6 gene are responsible for a late onset form of familial focal segmental glomerulosclerosis (FSGS). However, the role of TRPC6 variation in Chinese patients with late onset familial FSGS remains unclear. Here, we screened 31 Chinese pedigrees with late onset familial FSGS for changes in TRPC6 by DNA sequence analysis.nnnMETHODSnGenomic DNA was extracted from peripheral leukocytes. We PCR-amplified each of 13 exons of TRPC6 for sequence analysis. When a novel nucleotide change seemed likely to cause FSGS, we carried out an in vitro research to determine the effects of the mutation on TRPC6 function. HEK 293 cells were transfected stably with vectors containing mutant or wild type TRPC6 cDNA. We then compared the expression of mutant TRPC6 to wild type TRPC6 using Western blot. For the observation of the function of mutant TRPC6 channel compared with wild type TRPC6 channel, Intracellular Ca(2+) concentration was detected using fluorescent indicator Fluo-3 among different groups of cells upon stimulation with 1-oleoyl-2-acetyl sn-glycerol (OAG).nnnRESULTSnAll the 31 pedigrees with late onset familial FSGS were collected in our department from September 1997 to October 2007. A novel TRPC6 mutation (cytosine 2664 adenine resulting in Glutamine 889 Lysine substitution, Q889K) was identified in one of these pedigrees. Mutant TRPC6 (TRPC6(Q889K)) or wild type TRPC6 was stably expressed in HEK293 cells by Western blot. The mutant TRPC6 expression was a little increased without significant difference compared with wild type TRPC6 expression, whereas the intracellular Ca(2+) level in cells expressing mutant TRPC6 was significantly increased compared with that in the cells expressing wild TRPC6 upon stimulation.nnnCONCLUSIONnWe identified a novel TRPC6 mutation Q889K associated with late onset FFSGS in Chinese pedigrees and this mutation was demonstrated to be gain of function by an in vitro functional research.

COL4A3 mutations cause focal segmental glomerulosclerosis

Focal segmental glomerulosclerosis (FSGS) is a histologically identifiable glomerular injury often leading to proteinuria and renal failure. To identify its causal genes, whole-exome sequencing and Sanger sequencing were performed on a large Chinese cohort that comprised 40 FSGS families, 50 sporadic FSGS patients, 9 independent autosomal recessive Alports syndrome (ARAS) patients, and 190 ethnically matched healthy controls. Patients with extrarenal manifestations, indicating systemic diseases or other known hereditary renal diseases, were excluded. Heterozygous COL4A3 mutations were identified in five (12.5%) FSGS families and one (2%) sporadic FSGS patient. All identified mutations disrupted highly conserved protein sequences and none of them was found in either public databases or the 190 healthy controls. Of the FSGS patients with heterozygous COL4A3 mutations, segmental thinning of the glomerular base membrane (GBM) was only detected in the patient with electronic microscopy examination results available. Five ARAS patients (55.6%) had homozygous or compound-heterozygous mutations in COL4A3 or COL4A4. Serious changes in the GBM, hearing loss, and ocular abnormalities were found in 100%, 80%, and 40% of the ARAS patients, respectively. Overall, a new subgroup of FSGS patients resulting from heterozygous COL4A3 mutations was identified. The mutations are relatively frequent in families diagnosed with inherited forms of FSGS. Thus, we suggest screening for COL4A3 mutations in familial FSGS patients.

The risk predictive values of UK-PBC and GLOBE scoring system in Chinese patients with primary biliary cholangitis: the additional effect of anti-gp210

Adequate risk stratification is critical for the management of the patients with primary biliary cholangitis (PBC). The UK‐PBC and GLOBE scoring systems for prognosis of PBC have been proposed recently, but have not been validated in Asian population.

ACTN4 gene mutations and single nucleotide polymorphisms in idiopathic focal segmental glomerulosclerosis.

Aim: To investigate the association between mutations or single nucleotide polymorphisms (SNPs) of the gene ACTN4 in Chinese patients with idiopathic focal segmental glomerulosclerosis (FSGS). Materials and Methods: Genomic DNA of 82 Chinese idiopathic FSGS patients and 70 healthy people were used to analyze ACTN4 gene mutations by polymerase chain reaction, direct sequencing and GenBank matching. Hair follicle DNA of novel mutated patients’ parents were sequenced and α-actinin-4 expression in patients’ kidney was examined by immunofluorescence. For SNPs, after the Hardy-Weinberg equilibrium test, allele association and the frequencies of genotypes were analyzed, followed by association analysis between genotypes and clinical diagnosis. Results: We found a heterozygous candidate mutation 184T>A (S62T) in 1 patient and a 5′ UTR candidate mutation 1-34C>T in another patient. Both patients had non-nephrotic syndrome FSGS with reduced kidney α-actinin-4 expression. Promoter activity analysis suggests that the 1-34C>T candidate mutation may affect the transcriptional regulation of ACTN4 gene. Additionally, 6 novel silent variants and 2 novel SNPs were also found in this study. Novel SNP 484 + 87C>G had a significant association with the level of urine protein excretion in these idiopathic FSGS patients. Conclusions: Our data suggest that mutations and SNP of ACTN4 gene may contribute to be associated with Chinese idiopathic FSGS.

Identification of five novel variants in the thiazide-sensitive NaCl co-transporter gene in Chinese patients with Gitelman syndrome.

Aim:u2003 Gitelman syndrome (GS) is an autosomal recessive renal tubulopathy characterized by hypokalaemic metabolic alkalosis, significant hypomagnesemia, low urinary calcium, secondary aldosteronism and normal blood pressure. GS is caused by inactivating variants in the SLC12A3 gene, which encodes the thiazide‐sensitive NaCl co‐transporter. So far, more than 100 variants have been described in the SLC12A3 gene in Gitelman syndrome.

A successful screening for Fabry disease in a Chinese dialysis patient population.

Fig. 1. Family pedigree of index patient 1 (a) and index patient 2 (b). a, Left ventricular hypertrophy; b, hypertension; c, proteinuria; d, end stage of renal disease. impairment, acroparesthesiae, cardiac hypertrophy, corneal dystrophy, hypohidrosis, and impaired temperature regulatio (2, 3). FD is often underdiagnosed in China and only few Chinese cases have been reported (4–8). Several screening studies performed in patients on dialysis showed a high incidence of FD, and therefore, we initiated a