الفهرس | Only 14 pages are availabe for public view |
Abstract Hepatitis C is a major global public health problem and is one of the main causes of cirrhosis and Hepatocellular carcinoma. About 200 million people; 3% of the world’s population are infected with hepatitis C virus and 3 to 4 million persons are newly infected each year with a global 170 million chronic carriers at risk of developing liver cirrhosis and/or liver cancer. At least 85% of infected persons become chronically infected. Blood bank and community-based surveys conducted in Egypt have reported sero-prevalence rates of HCV to be as high as 40% in some parts of the country. Blood transfusion is thought to be the main route of transmission. While hepatocytes are the major site of viral replication, the virus can also replicate at the extrahepatic sites including PBMC. This is reflected by a higher frequency of extrahepatic complications and diseases associated with chronic HCV infection, including mixed cryoglobulinaemia, cutaneous vasculitis, glomerulo-nephritis, neuropathy and lymphoproliferafive disorders in HCV-infected persons. There is strong support from recent studies that hepatocytes are the major site of viral replication, the virus can also replicate at the extrahepatic sites including PBMCs. Patients with detectable negative-strand HCV RNA in PBMCs had lower IFN sustained response rates compared to those without detectable negative-strand HCV RNA in PBMCs. Low-level replication of HCV in PBMCs may lead to Summary - 121 - reactivation of HCV after termination of therapy. Because of the frequency and the severity of the disease, many tests have been developed for diagnosis of hepatitis C virus. They include non specific tests e.g. liver transaminases and auto antibodies detection; specific tests which include the antibody detection (serological) methods as ELISA and RIBA tests, detection of the HCV RNA itself through the use of NATs and finally, genotype testing. While traditional HCV ”viral load” assays quantify HCV RNA in the serum, they are unable to distinguish positive- from negative-strand HCV RNA and thus have limited utility in assessing extrahepatic replication. The use of ”strand-specific” RT-PCR to detect the replicative intermediate negative-strand HCV RNA is often cited as evidence for viral replication and the use of nested RTPCR (in which the products of a first round of PCR were reamplified using a second, nested set of primers) gave extreme- sensitivity and increased specificity. This study was done on 30 HCV infected patients who had sero-positive HCV results (using 3rd generation ELISA technique) including 21 patients positive for HCV RNA in the serum and 9 patients negative for HCV RNA in the serum by Real time PCR. Blood samples were collected from the Outpatient of Internal Medicine Hospital of Ain Shams University. All cases were subjected to full history taking and complete general and local examination.Twenty healthy controls matched age and sex also included in this study. Summary - 122 - All cases were also subjected to HCV positive- and negative-stranded RNA detection in PBMCs samples by using thermal cycler (Gene Amp PCR System 9700) for conventional RT-PCR qualitative assay, also nested PCR was performed. A statistical study was done among the results and this study revealed that: the HCV-RNA positive-strand was detected in PBMCs from 20 out of 30 sero-positive patients (66.7%) and also from 20 out of 21 serum HCV-RNA positive patients (95.2%). There is highly statistically significant association (p<0.01) between HCV-Ab positive cases and the presence of HCV-RNA positive-strand and also between serum HCV-RNA positive cases and the presence of HCV-RNA positive strand in PBMCs. The HCV-RNA negative-strand was detected in PBMCs from 7 out of 30 sero-positive patients (23.3%) and this association was statistically significant (p<0.05). Also HCV-RNA negative-strand was detected in PBMCs from 7 out of 21 serum HCV-RNA positive patients (33.3%), so there is highly statistically significant association (p<0.01) between HCV-RNA positive serum and HCV-RNA negative-strand in PBMCs. There was a high statistically significant association between positive- and negativestrand HCV-RNA (p<0.01). No positive results for both positiveand negative-strands HCV- RNA were detected in PBMCs of the 9 patients with negative serum HCV-RNA nor in the controls. Summary - 123 - from this study and other studies we concluded that PBMCs act as possible site for extrahepatic replication and may represent a reservoir for HCV which cause reinfection of hepatocytes |