Circulating microRNA-196a as a candidate diagnostic biomarker for chronic hepatitis C

Abstract

Previous studies have demonstrated the inhibitory effect of microRNA (miR)-196a on hepatitis C virus (HCV) expression in human hepatocytes. However, the clinical implications of aberrant miR-196a expression and the application of circulating miR-196a in the diagnosis and management of chronic hepatitis C (CHC) require further investigation. The present study aimed to examine the possibility of using serum miR-196a as a biomarker for CHC. The Affymetrix miRNA array platform was used for miRNA expression profiling in adenovirus (Ad)-HCV core-infected (HepG2-HCV) and Ad-enhanced green fluorescence protein (EGFP)-infected HepG2 cells (HepG2-control). miR-196a downregulation and levels were analyzed using stem-loop reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis of the sera of 43 patients with CHC and 22 healthy controls. A total of six miRNAs were identified as significantly different (≥ 1.5 fold; P ≤ 0.05) between the two groups. Of note, significant miR-196a downregulation was observed in HepG2-HCV as compared with HepG2‑EGFP. Furthermore, as compared with that of the healthy control group, serum miR-196a was demonstrated to be significantly lower in patients with CHC. In addition, analysis of the receiver operating characteristic (ROC) curve for serum miR-196a revealed an area under the ROC curve of 0.849 (95% confidence interval, 0.756-0.941; P<0.001) with 81.8% sensitivity and 76.7% specificity in discriminating chronic HCV infection from healthy controls at a cut-off value of 6.115 x 10(-5), demonstrating significant diagnostic value for CHC. However, no correlation was identified between serum miR-196a and alanine aminotransferase, aspartate aminotransferase or HCV-RNA. In conclusion, the present study identified circulating miR-196a as a specific and noninvasive candidate biomarker for the diagnosis of CHC.

Figure 1

Expression profile of microRNAs in HepG2-hepatitis C virus and HepG2-control cells.

Figure 2

miR-196a was significantly downregulated by Ad-HCV infection. (A) Confirmation of HCV core protein expression using western blot analysis following Ad-HCV infection for 48 h. (B) Relative expression levels of miR-196a in HepG2-HCV and HepG2-control cells. Values are presented as mean ± standard deviation; each experiment was conducted in triplicate. U6 was used as an internal control for miRNA reverse transcription quantitative polymerase chain reaction experiments; ^*P<0.05. miR, microRNA; HCV, hepatitis C virus.

Figure 3

Reduced serum miR-196a levels in patients with CHC. (A) Comparison of miR-196a concentrations between CHC and healthy controls. Serum miR-196a concentrations in correlation with (B) ALT, (C) AST and (D) HCV-RNA. ^**P<0.001. miR, microRNA; CHC, chronic hepatitis C; ALT, alanine aminotransferase; AST, aspartate aminotransferase; HCV, hepatitis C virus.

Figure 4

ROC curve analysis of circulating miR-196a levels for discriminating chronic hepatitis C from healthy controls. miR-196a, microRNA-196a; ROC, receiver operating characteristic; AUC, area under the ROC curve.