Urinary Exosomes Derived CircRNAs As Biomarkers For Chronic Renal Fibrosis

Objective: Renal fibrosis is the major pathological basis for the progression of chronic kidney disease to end-stage renal failure,which can reflect the severity of renal destruction.At present,the gold standard for the diagnosis of renal fibrosis is renal biopsy pathology,and there is still a lack of accurate and non-invasive detection methods to reflect the progression of renal fibrosis.In recent years,relevant studies have shown that urinary exosomes carry biological information derived from renal intrinsic cells,and circular RNA is highly enriched and stably expressed in exosomes.Therefore,urinary exosome-derived circ RNA have the potential to become biomarker related to kidney disease.The aim of this study is to investigate the expression of urinary exosome-derived circ RNA and to explore the possibility that urinary exosome-derived circ RNA could be a novel biomarker reflecting renal fibrosis.Methods: Morning urine samples from 3 chronic kidney disease patients without renal fibrosis and 3 renal fibrosis patients confirmed by renal puncture biopsy were collected,and urine exosomes were extracted by ultracentrifuge.Human circ RNAs microarray were performed to detect the circ RNAs expression profiles of patients with renal fibrosis.Screening of candidate circ RNA with significant differences and statistical significance,expanding the sample size to validate the screening results.urinary exosomes were collected from 110 patients with CKD confirmed by renal biopsy(CKD group)and54 healthy controls.The urine exosomal RNA was extracted,and the relative expression level of hsa＿circ＿0036649 was detected by real-time fluorescence quantitative PCR.The differences in the expression of hsa＿circ＿0036649 between CKD patients and healthy controls were analyzed,and its correlation with renal function parameters and pathological indicators was further explored.The receiver operating characteristic(ROC)curve indicate urinary exosomes hsa＿circ＿0036649 in the diagnosis of renal fibrosis.Results: Human circ RNA microarray demonstrated that 365 circ RNAs up regulated and195 down regulated in urinary exosomes from renal fibrosis patients compared to without renal fibrosis CKD patients.Among the circ RNAs with down regulated expression,hsa＿circ＿0036649 was the largest decrease(P<0.05).The expression of hsa＿circ＿0036649in renal fibrosis patients was lower than that in healthy control group.Spearman correlation showed that the expression of hsa＿circ＿0036649 in urinary exosomes were negatively correlated with serum creatinine(Scr,rs=-0.366,P<0.001),urea nitrogen(BUN,rs=-0.215,P=0.006),cystatin C(rs=-0.424,P<0.001),24-hour proteinuria(rs=-0.214,P=0.024),renal tubulointerstitial fibrosis score(TIF,rs=-0.360,P<0.001)and glomerulosclerosis score(rs=-0.273,P=0.004),and positively correlated with estimated glomerular filtration rate(e GFR,rs=0.374,P<0.001).Stepwise multivariate logistic regression showed that the expression of hsa＿circ＿0036649 is not only a risk indicator for renal fibrosis,but also can effectively distinction between degrees of severity of renal fibrosis(OR=0.125;95%CI:0.026-0.599;P=0.009).The ROC curve suggested that urinary exosomes derived hsa＿circ＿0036649(AUC=0.706,95%CI:0.606-0.807,P=0.001)could be used as a biomarker for the diagnose renal fibrosis at a cutoff value of 0.597 with a sensitivity of 45.5% and specificity of 87.9%.Conclusion:1.Human circ RNAs microarray reveals the expression profile of circ RNAs in patients with renal fibrosis.2.The expression level of hsa＿circ＿0036649 derived from urinary exosomes was correlated with renal function parameters and pathological indicators.3.ROC curves indicate that the expression of hsa＿circ＿0036649 derived from urinary exosomes may be used as a biomarker for the diagnosis of renal fibrosis.