MicroRNAs and Liver Fibrosis in Hepatitis C Patients

Abstract

HCV is a major cause of chronic liver disease. Unfortunately, Egypt has by far the highest national-level HCV prevalence in the world, and the highest prevalence of hepatitis C virus genotype 4 (HCV-4), which is responsible for almost 90% of infections and is considered a major cause of chronic hepatitis, liver cirrhosis, hepatocellular carcinoma, and liver transplantation in the country.

The therapeutic decisions and prognostic evaluations of chronic liver diseases depend on the accurate assessment of the degree of liver fibrosis where liver biopsy was considered as a gold standard, it has potential risk due its limitations. So searching for alternative non invasive methods for assessing the severity of liver fibrosis has significantly increased. But these non invasive biomarkers have limitations. A great necessity of specific highly sensitive, minimally invasive bio-diagnostic tool has been elucidated. Circulating microRNA levels have become a rapidly growing area of clinical research based on their altered expression found in different diseases of liver. Several miRNAs play important roles in HCV related inflammation, fibrosis and HCC development. Circulating miR-29 as a potential new hepatic stellate cell (HSC) activation marker and miR-155 as a positive regulator of inflammation. For this purpose: Thirty five individuals who enrolled to the hospitals of AL Azhar University were divided into 2 groups: The first control group contained 6 healthy volunteers without any evidence of liver disease and confirmed by clinical examination. The second Chronic HCV Egyptian patients (CHCV) group which contained 29 untreated patients with a serological, virological and histological diagnosis of chronic HCV infection. Patients who infected with both HCV and HBV and patients with chronic liver disease of etiologic agent other than HCV are ruled out. All persons in the study were subjected to: 1- Laboratory investigations of all enrolled persons, for Aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activities AST/ALT ratio was calculated, platelets count, prothrombin index determination and Aspartate aminotransferase platelet ratio index (APRI) was calculated by the following formula: APRI = (AST/upper limit of normal for AST) × 100 platelet count (× 109/L) 2- Quantification of HCV–RNA for the chronic HCV group: was detected using Polymerase chain reaction (PCR) assay 3- Liver biopsy and histopathology examination for the chronic HCV group, staging and grading of liver histopathology were performed METAVIR scoring system. 4- Total RNA extraction: Total RNA was isolated from serum samples, total RNA concentration and purity were analyzed by using the nanodrop spectrophotometer. 5- MicroRNA-specific quantitative real-time RT-PCR, cDNA was synthesized from total RNA and universal Primer, the expression profiles of the studied microRNAs were measured and normalized to GAPDH as a house keeping gene and the fold changes in gene expression were calculated using the 2−ΔΔCt method.