Jingyuan Xie

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.

Interleukin-10 deficiency aggravates kidney inflammation and fibrosis in the unilateral ureteral obstruction mouse model

Interleukin-10 functions as a general immunosuppressive cytokine, which also negatively regulates inflammatory responses through complex mechanisms. Recent studies suggested that IL-10 may also inhibit fibrosis in various diseased models. However, the role of IL-10 in renal fibrosis has not been demonstrated. Here, we investigated the effects of IL-10 in the development of renal tubulointerstitial fibrosis by creating the unilateral ureteral obstruction (UUO) model in IL-10 knockout (−/−) mice. We performed sham or unilateral ureteral obstruction surgery in 8-week-old IL-10−/− male mice and age and sex-matched wild type littermates. Mice were killed at 7 days or 14 days post surgery and renal tissues were obtained for RNA, protein, and immunohistochemical analysis. Our results found IL-10 deficiency resulted in enhanced renal fibrosis demonstrated by more severe tubular injury and collagen deposition and higher expression of pro-fibrotic genes (including α-SMA, MMP-2, fibronectin, FSP-1 and vimentin). Our results also found IL-10−/− UUO mice developed more severe renal inflammation with a significant increase in inflammatory cells infiltration, and upregulation of inflammatory chemokines (MCP-1 and RANTES), and cytokines (TNF-α, IL-6, IL-8, and M-CSF). Further study revealed that enhanced renal inflammation and fibrosis was associated with significantly increased activation of both TGF-β/Smad3 and NF-κB signaling pathways. In summary, our study provides the direct evidence that IL-10 is an endogenous cytokine that has a key role in protecting against development of renal inflammation and fibrosis. Enhancement of IL-10 expression could be a potential anti-fibrosis therapy for patients with chronic kidney diseases.

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.

Diuretics associated acute kidney injury: clinical and pathological analysis

Abstract Objective: In order to evaluate the clinical and pathological characteristics of diuretics associated acute kidney injury (AKI) and its management. Methods: We performed a retrospective study including 131 cases that diagnosed as diuretics associated AKI from 1 January 1999 to 1 January 2010 in Ruijin Hospital affiliated to Shanghai Jiao Tong University. Drug applications and its related clinical, laboratory and histological data were collected. Results: The male to female ratio was 2:1. The proportion of ages <20 years, 20–40 years, 40–60 years and ≥60 years were 6.9%, 17.6%, 27.5% and 48.1% respectively. Most patients (96.2%) had at least one complication of which chronic kidney disease (CKD) occurred most frequently (72 in 131, 55.0%). We divided all the patients to diuretic group (N = 131) and non-diuretic group (N = 185) based on diuretics history. We found patients in diuretic group had higher rates of CVD (38.9% vs. 18.4%), hypertension (42.0% vs. 29.2%), CKD (55.0% vs. 27.0%) and DM (17.6% vs. 4.3%) than non-diuretic group. Of 131 diuretics associated AKI, 36 cases (27.5%) were caused by diuretics only, 39 cases (29.8%) were caused by the combination of diuretics and other drugs like antibiotics, contrast media, ACEI or NSAIDs, and 56 cases (42.7%) had other AKI risk factors such as operation, infection, acute heart failure or hepatorenal syndrome. In addition, our data suggested the severity of RIFLE classification and pathological injury of glomerular basement membrane was higher in large-dosage furosemide group (> = 120 mg/d) than in low-dosage group (<120 mg/d). The most common lesion induced by diuretics was vacuolar degeneration of tubular epithelial cell. Logistic regression analysis showed predictors of all-cause mortality were age, gender, RIFLE classification when AKI onset. Age and RIFLE classification were predictive factor of non-complete recovery. Conclusion: This article firstly focuses on diuretics associated AKI, whose onset was related to aging, primary diseases and diuretic dosage. The combination of diuretics with other drugs such as antibiotics, contrast media, ACEI, NSAIDs, etc. would synergistically induced AKI. The pathological lesion of diuretics associated AKI may be mostly manifested vacuolar degeneration of tubular epithelial cell. Aging, gender, severity of RIFLE classification may be predictive factors of all-cause mortality of diuretics associated AKI.

Distinct metabolic profile of primary focal segmental glomerulosclerosis revealed by NMR-based metabolomics.

Background Primary focal segmental glomerulosclerosis (FSGS) is pathological entity which is characterized by idiopathic steroid-resistant nephrotic syndrome (SRNS) and progression to end-stage renal disease (ESRD) in the majority of affected individuals. Currently, there is no practical noninvasive technique to predict different pathological types of glomerulopathies. In this study, the role of urinary metabolomics in the diagnosis and pathogenesis of FSGS was investigated. Methods NMR-based metabolomics was applied for the urinary metabolic profile in the patients with FSGS (n = 25), membranous nephropathy (MN, n = 24), minimal change disease (MCD, n = 14) and IgA nephropathy (IgAN, n = 26), and healthy controls (CON, n = 35). The acquired data were analyzed using principal component analysis (PCA) followed by orthogonal projections to latent structure discriminant analysis (OPLS-DA). Model validity was verified using permutation tests. Results FSGS patients were clearly distinguished from healthy controls and other three types of glomerulopathies with good sensitivity and specificity based on their global urinary metabolic profiles. In FSGS patients, urinary levels of glucose, dimethylamine and trimethylamine increased compared with healthy controls, while pyruvate, valine, hippurate, isoleucine, phenylacetylglycine, citrate, tyrosine, 3-methylhistidine and β-hydroxyisovalerate decreased. Additionally, FSGS patients had lower urine N-methylnicotinamide levels compared with other glomerulopathies. Conclusions NMR-based metabonomic approach is amenable for the noninvasive diagnosis and differential diagnosis of FSGS as well as other glomerulopathies, and it could indicate the possible mechanisms of primary FSGS.

GWAS for serum galactose-deficient IgA1 implicates critical genes of the O-glycosylation pathway

Aberrant O-glycosylation of serum immunoglobulin A1 (IgA1) represents a heritable pathogenic defect in IgA nephropathy, the most common form of glomerulonephritis worldwide, but specific genetic factors involved in its determination are not known. We performed a quantitative GWAS for serum levels of galactose-deficient IgA1 (Gd-IgA1) in 2,633 subjects of European and East Asian ancestry and discovered two genome-wide significant loci, in C1GALT1 (rs13226913, P = 3.2 x 10−11) and C1GALT1C1 (rs5910940, P = 2.7 x 10−8). These genes encode molecular partners essential for enzymatic O-glycosylation of IgA1. We demonstrated that these two loci explain approximately 7% of variability in circulating Gd-IgA1 in Europeans, but only 2% in East Asians. Notably, the Gd-IgA1-increasing allele of rs13226913 is common in Europeans, but rare in East Asians. Moreover, rs13226913 represents a strong cis-eQTL for C1GALT1 that encodes the key enzyme responsible for the transfer of galactose to O-linked glycans on IgA1. By in vitro siRNA knock-down studies, we confirmed that mRNA levels of both C1GALT1 and C1GALT1C1 determine the rate of secretion of Gd-IgA1 in IgA1-producing cells. Our findings provide novel insights into the genetic regulation of O-glycosylation and are relevant not only to IgA nephropathy, but also to other complex traits associated with O-glycosylation defects, including inflammatory bowel disease, hematologic disease, and cancer.

Novel mutations in the inverted formin 2 gene of Chinese families contribute to focal segmental glomerulosclerosis

Here, we report a genetic study of an extended family of Chinese ancestry with focal segmental glomerulosclerosis (FSGS), with one of the affected members also concurrently diagnosed with IgA nephropathy (IgAN). By genome-wide linkage analysis and subsequent sequencing, we identified an S85W mutation in the inverted formin 2 (INF2) gene that perfectly cosegregated with the kidney disease phenotype. The entire INF2 coding region was sequenced in 200 healthy controls, 55 families with FSGS, and 34 families with IgAN. This analysis identified a novel insertion, S129_Q130insVRQLS, in another FSGS pedigree. In vitro studies found that α-actinin 4 expression was decreased and INF2 showed perinuclear localization in S85W-transfected podocytes. Phosphorylation of serum response factor, and that its nuclear translation was decreased in S85W podocytes, indicated decreased activation in mutants. Abnormal actin organization was also found in S85W podocytes, while no change of microtubule structure was observed. Co-immunoprecipitation and immunofluorescence found decreased interaction between INF2 and Cdc42 in S85W podocytes. However, all these changes were not found in S129_Q130insVRQLS podocytes. The overall frequency of INF2 mutations was ~3.6% among Chinese familial FSGS, which was considerably lower than that from studies of European FSGS families. Thus, S85W but not the S129_Q130insVRQLS variant leads to podocyte cytoskeletal abnormalities, probably by impaired serum response factor phosphorylation.

Underweight Is an Independent Risk Factor for Renal Function Deterioration in Patients with IgA Nephropathy.

Studies on the relationship between body mass index (BMI) and renal progression in IgA Nephropathy (IgAN) were limited, especially for underweight patients with IgAN. To elucidate the clinical features and effect of underweight on renal function deterioration in this disease, we recruited IgAN patients with diagnostic age ≥18 years old and a baseline estimated glomerular filtration rate (eGFR) ≥15 ml/min/1.73m2 from our center between 1985 and 2014. Patients secondary to systemic diseases or follow-up less than 6 months were excluded. All patients’ clinical data at renal biopsy and during follow-up were recorded. Renal outcome was defined as end-stage kidney disease (ESRD). Baseline body mass index (BMI) was calculated by weight (kg) over squared height (m2). According to WHO Asian guideline, BMI was categorized as follows: <18.5kg/m2 (underweight), 18.5–22.99kg/m2 (normal weight), 23–27.49kg/m2 (overweight) and obese (≥27.5 kg/m2). Of 930 primary IgAN patients enrolled in this study, mean age at renal biopsy was 37.6 years and 49.2% were men. Totally, 114 (12.3%) ESRD occurred after a mean follow-up of 47.1 months. More ESRD happened in underweight patients (17.3%) compared to patients with normal weight (13.2%), overweight (11.0%) or obesity (9.5%). By multivariate Cox regression analysis, underweight was independently associated with a higher risk of ESRD after adjustment for demographic characteristics and clinical variables (HR: 3.5, 95% CI: 1.3–9.5, P = 0.01) comparing to normal weight. Underweight patients had lower hemoglobin, serum uric acid, triglycerides, cholesterol and lymphocyte counts than patients with normal weight. Furthermore, BMI was positively correlated with serum C3 (r = 0.25, p <0.001). Our research finds that underweight is an independent risk factor for kidney disease progression in IgAN, which might be associated with malnutrition status and decreased C3 levels.

Angiotensin II-mediated activation of fibrotic pathways through ERK1/2 in rat peritoneal mesothelial cells

Introduction: Peritoneal fibrosis is a common complication of peritoneal dialysis (PD) although the pathway involved is unclear. Of this article, angiotensin II (Ang II)-mediated upregulation of mitogen-activated protein kinase (MAPK) pathway as well as their downstream profibrotic genes including transforming growth factor (TGF)-β1 and fibronectin (FN) was investigated. Methods: Rat peritoneal mesothelial cells (RPMCs) were obtained by enzymatic digestion from the colic omentum. After incubated with Ang II, real-time PCR, ELISA, and Western blot analysis were used to determine RPMCs cellular and secretory (supernatants) levels of TGF-β1, FN, tissue inhibitor of metalloproteinase-1 (TIMP-1), and plasminogen activator inhibitor-1 (PAI-1) as well as the phosphorylation of extracellular signal-regulated kinase (ERK1/2), p38 MAPK, and stress-activated protein kinase/c-Jun NH2-terminal kinase (JNK). We also determined the downstream pathways using the specific inhibitors including PD98059 (ERK1/2), SB230580 (p38 MAPK), SP600125 (JNK), and losartan [Ang II type-1 (AT1] receptor blocker). Results: Ang II increased mRNA and protein levels of TGF-β1, FN, TIMP-1, and PAI-1 in a time- and dose-dependent manner in RPMCs. Ang II induced a 1.5–2-fold increase in both mRNA and protein levels of the above molecules at 10 nmol/L. Ang II also upregulated the phosphorylation of ERK1/2 and p38 but not of JNK. Finally, inhibition of either AT1 or ERK1/2 was able to suppress Ang II-induced expression of FN. Conclusion: In cultured RPMCs, Ang II upregulated profibrotic signaling pathways through AT1-mediated ERK1/2 phosphorylation.

Genetic studies of IgA nephropathy: what have we learned from genome-wide association studies.

Primary IgA nephropathy (IgAN) is the most common glomerulonephritis in the world. It is most common in Asian populations, followed by Caucasians, yet relatively infrequent amongst African populations. The striking difference in the prevalence of IgAN between world populations, together with the known familial aggregation of disease, suggests an inherited mechanism. Recently three genome-wide association studies (GWAS) of IgAN have identified seven susceptibility loci, providing initial insight into the genetic architecture of this trait. While genetic studies of complex traits are challenging, applying new techniques and methods of analysis, especially Next-Generation Sequencing, will push the genetic studies of IgAN forward.