In Situ Enzymatic Oligonucleotide Amplification of Hepatitis C Virus-Rna in Liver Biopsy Specimens (Reverse Transcriptase in Situ Polymerase Chain Reaction) After Orthotopic Liver Transplantation for Hepatitis C-Related Liver Disease

Fragulidis, Georgios P.; Cirocco, Robert; Weppler, Deborah; Berho, Mariana; Gillian, G.; Markou, M.; Viciana, Ana L.; Esquenazi, Violet; Nery, Jose; Miller, Joshua; Reddy, K. Rajender; Tzakis, Andreas G.

doi:10.1097/00007890-199812150-00010

Cited by 12 publications

(9 citation statements)

References 19 publications

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“…The presence of viral RNA in liver samples has been shown by in situ hybridization 19 or reverse-transcriptase in situ polymerase chain reaction. 20 However, these methods have technical difficulties and have not been widely used.…”

mentioning

confidence: 99%

Timing of reinfection and mechanisms of hepatocellular damage in transplanted hepatitis C virus[ndash ]reinfected liver

Ballardini¹

2002

Liver Transplantation

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Pathogenic mechanisms and dynamics of hepatitis C virus (HCV) reinfection in orthotopic liver transplantation (OLT) are poorly defined. This study focuses on these aspects by studying 55 frozen biopsy specimens from transplant recipients with various histological diagnoses obtained from 4 days to 4 years post-OLT and 10 patients with HCV-related chronic hepatitis. The percentage of HCV-infected hepatocytes, number and distribution of CD8 and natural killer cells, and rates of hepatocellular apoptosis and proliferation were quantified by immunohistochemistry. HCV antigens were detected in 37% of biopsy specimens obtained within 20 days and 90% of biopsy specimens obtained from 21 days to 6 months after OLT. The number of HCV-infected hepatocytes was never less than 40% in acute hepatitis specimens and never greater than 30% in the other cases. Hepatocellular apoptosis was high in biopsy specimens of acute hepatitis and moderate in those from transplant recipients with normal histological characteristics, but still greater than in specimens of chronic active hepatitis. Proliferation correlated significantly with apoptosis. Lymphocyte infiltration was high and similar among cases of acute hepatitis, chronic hepatitis, and rejection. These data: (1) show that the detection of liver HCV antigens is sensitive enough to be used in clinical practice as a diagnostic tool to detect infection of the transplanted liver and might be useful, combined with conventional histological evaluation to detect hepatitic damage, for therapeutic decision making;

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confidence: 99%

Timing of reinfection and mechanisms of hepatocellular damage in transplanted hepatitis C virus[ndash ]reinfected liver

Ballardini¹

2002

Liver Transplantation

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“…The specificity of ISH is high but its sensitivity low, and it is difficult to detect low copy numbers of the HBV or HCV genome in tissue. PCR technology has been adapted to in situ amplification of viral genomes or their replicative intermediates in liver tissue sections, but sensitivity and specificity remain major challenges to the application of this approach (13,18,23,25,26,30,31). Here, we describe the use of a novel, highly specific and sensitive PCR-ISH method to determine the distribution and localization of HBV DNA, HBV RNA, and HCV RNA in both normal and cancerous liver tissues.…”

Section: Discussionmentioning

confidence: 99%

Detection of Hepatitis B and C Viruses in Almost All Hepatocytes by Modified PCR-Based In Situ Hybridization

et al. 2010

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Although PCR-based in situ hybridization (PCR-ISH) can be used to determine the distribution and localization of pathogens in tissues, this approach is hampered by its low specificity. Therefore, we used a highly specific and sensitive PCR-ISH method to reveal the lobular distribution and intracellular localization of hepatitis B virus (HBV) and HCV in chronic liver disease and to clarify the state of persistent HBV and HCV infection in the liver. HBV genomic DNA was detected in almost all hepatocytes, whereas HBV RNA or protein was differentially distributed only in a subset of the HBV DNA-positive region. Further, HCV genomic RNA was detected in almost all hepatocytes and was localized to the cytoplasm. HCV RNA was also detected in the epithelium of the large bile duct but not in endothelial cells, portal tracts, or sinusoidal lymphocytes. In patients with HBV and HCV coinfection, HCV RNA was localized to the noncancerous tissue, whereas HBV DNA was found only in the cancerous tissue. Using this novel PCR-ISH method, we could visualize the staining pattern of HBV and HCV in liver sections, and we obtained results consistent with those of real-time detection (RTD)-PCR analysis. In conclusion, almost all hepatocytes are infected with HBV or HCV in chronic liver disease; this finding implies that the viruses spreads throughout the liver in the chronic stage.Hepatitis B virus (HBV) and hepatitis C virus (HCV) are the primary causative agents of chronic liver disease (2, 9, 17). HBV infection remains a global health problem; it is estimated that 350 million individuals are persistently infected with the virus and that approximately 15% to 25% of these individuals will die due to the sequelae of the infection (23, 29). Further, more than 170 million people are infected with HCV worldwide (21). HCV has a single-stranded RNA genome (8, 19), does not have canonical oncogenes, and can easily establish chronic infection without integration into the host genome (3,20), resulting in hepatic steatosis and hepatocellular carcinoma (HCC) (28). The viruses share a similar route of transmission, such as via the transfusion of infected blood or body fluids or use of contaminated needles.Several studies have shown that 10% to 35% of the individuals infected with HBV also have HCV infection, although the prevalence varies depending on the population studied (4, 32, 34). The relationship between coinfection and acceleration of malignant transformation remains unclear, but HBV and HCV coinfection seems to alter the natural history of both HBVrelated and HCV-related liver disease (2, 12). HCV has been shown to inhibit HBV gene expression (7, 15). The high prevalence of occult HBV infection may indicate that HCV also inhibits HBV replication (34). Most epidemiological studies of HBV have been performed by using diagnostic serological assays (16). We recently used a novel, highly sensitive diagnostic PCR method to demonstrate that the HBV genome is detectable in the sera of a substantial proportion of patients with chronic HCV infectio...

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“…Although it is easy to show systemic HCV infection through serum HCV-RNA levels, it is more difficult to evaluate the actual state of HCV infection in the grafted liver. The presence of HCV-RNA in liver biopsy specimens has been shown by in situ hybridization (Agnello et al, 1998) or by reverse-transcriptase in situ polymerase chain reaction (Fragulidis et al, 1998). However, these methods have technical difficulties and have not been widely used.…”

Section: Immunohistochemical Detection Of Hcv Antigens For Recurrent mentioning

confidence: 99%

Immunohistochemical Staining of Liver Grafts for Recurrent Hepatitis C After Liver Transplantation

Shinmoto¹,

Ogino²,

Yagi³

et al. 2011

Liver Biopsy in Modern Medicine

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In Situ Enzymatic Oligonucleotide Amplification of Hepatitis C Virus-Rna in Liver Biopsy Specimens (Reverse Transcriptase in Situ Polymerase Chain Reaction) After Orthotopic Liver Transplantation for Hepatitis C-Related Liver Disease

Abstract: Our findings indicate that the HCV in situ RT-PCR assay may be helpful in the differentiation of recurrent hepatitis C disease from rejection. This may further help in the adjustment of immunosuppression.

Cited by 12 publications

References 19 publications

Timing of reinfection and mechanisms of hepatocellular damage in transplanted hepatitis C virus[ndash ]reinfected liver

Timing of reinfection and mechanisms of hepatocellular damage in transplanted hepatitis C virus[ndash ]reinfected liver

Detection of Hepatitis B and C Viruses in Almost All Hepatocytes by Modified PCR-Based In Situ Hybridization

Immunohistochemical Staining of Liver Grafts for Recurrent Hepatitis C After Liver Transplantation

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