Genotyping and investigation of mirna levels in HCV infected patients

Hepatitis C virus (HCV) is a significant public health issue due to its high genetic variability, which makes vaccination impossible, its high rate of chronicity, and complications such as cirrhosis and hepatocellular carcinoma. Determining the genotype of the causative agent is important for predicting the prognosis and selecting the appropriate treatment. MicroRNAs (miRNAs), a type of non-coding ribonucleic acid (RNA), play a role in the post-transcriptional regulation of protein synthesis. The observation that miRNAs remain stable in the blood has led to the idea that they could be used in the diagnosis of diseases. The aim of this study is to identify the genotypes among patients infected with HCV and compare laboratory finding between the genotypes; compare the plasma miRNA levels of healthhy individuals with those of HCV infected patients; and investigate the relationship between miRNA levels by comparing them across different genotypes. Fifty HCV RNA-positive patients who visited the Central Laboratory of Pamukkale University Hospital were selected as the patient group, and 50 voluntary individuals were selected as the control group. HCV RNA and HCV genotype determination were performed using a real-time polymerase chain reaction (RT-PCR) device. RT-PCR was conducted on the plasma of participants from both the patient and control groups to determine the expression levels of miRNA-122, miRNA-155, miRNA-224, and miRNA-21 genes. Gene expression was analyzed using the 2-ΔΔCT method in the presence of the U6 reference gene (housekeeping gene). Among the 50 patients who tested positive for HCV RNA, 13 (26%) were infected with genotype 1a, 14 (28%) with genotype 1b, 1 (2%) with genotype 2, and 22 (44%) with genotype 3. Among the patients infected with genotype 1a, 2 (15.4%) were women and 11 (84.6%) were men; among those infected with genotype 1b, 10 (71.4%) were women and 4 (28.6%) were men; and among those infected with genotype 3, 4 (18.2%) were women and 18 (81.8%) were men (p=0.001). It was observed that there were significantly more men among the patients infected with genotypes 1a and 3. It was found that patients infected with genotype 1b were significantly older than those infected with genotypes 1a and 3 (p=0.004). No significant difference was observed between the three genotypes in terms of AST, ALT, ALP, and HCV RNA levels. In the control group, 28 (56%) were women and 22 (44%) were men, while in the patient group, 16 (32%) were women and 34 (68%) were men (p=0.016). It was observed that the number of males in the patient group was significantly higher. The average age of the control group was 36.42 ± 9.22, while the average age of the patient group was 32.18 ± 11.71. According to these findings, the patient group was significantly younger than the control group (p=0.005). No significant difference was found between the two groups in terms of miRNA-122 and miRNA-224 expression. However, the expression of miRNA-155 (p=0.014) and miRNA-21 (p<0.0001) was significantly reduced in the patient group. Furthermore, there was no significant relationship between the expression levels of the four miRNAs and age, ALT, AST, ALP, or HCV RNA levels. It was also found that the expression levels of the four miRNAs were positively correlated with each other. In the ROC analysis, miRNA-155 (p=0.014) and miRNA-21 (p<0.001) were found to be statistically significant in distinguishing between patients and healthy individuals. miRNA-155 could differentiate between patients and healthy individuals with an accuracy of 64.3%, while miRNA-21 could distinguish between patients and healthy individuals with an accuracy of 95.1%. These results suggest that miRNA-21 and miRNA-155 could be used as biomarkers in the diagnosis of HCV infection. No significant difference was found between the three genotypes in terms of the expression levels of miRNA-122, miRNA-155, miRNA-224, and miRNA-21. In the future, further studies are needed as miRNAs require standardization before they can be used as biomarkers. Our study suggests that the distribution of HCV genotypes in our country may have changed due to factors such as changes in transmission routes and migration. It highlights the need for further research in this area and efforts to reduce the incidence of HCV. Key words: HCV, HCV genotype, miRNA, microRNA