Hepatic Hepatitis C Virus RNA Research Articles

HCV RNA in serum and liver samples of patients undergoing living donor liver transplantation.

ObjectiveTo compare hepatitis C virus (HCV) RNA levels from serum and explanted native liver samples from patients undergoing living donor liver transplantation (LDLT).MethodsThis was a prospective observational study. Serum and liver samples were collected from consecutive serum anti-HCV-positive transplant recipients between February 2016 to August 2019. HCV RNA was extracted from liver samples and subjected to one-step reverse-transcription qPCR. using the TopScript One Step qRT-PCR Probe Kit with HCV qPCR probe assay and human GAPDH qPCR probe assay on a ViiA7 Real-Time PCR System.ResultsAmong the 80 patients, 36% (29/80) were HCV RNA positive in serum and 85% (68/80) had positive hepatic HCV RNA. Post-liver transplantation, 4% (3/80) patients were serum positive.ConclusionsOur study suggests that pre-transplant serum HCV RNA levels may give an underestimate of the number of positive HCV RNA cases and that hepatic HCV RNA data may be more accurate.

Kinetics of miR-122 Expression in the Liver during Acute HCV Infection

The relationships among micro RNA-122 (miR-122) expression in the liver, hepatitis C virus (HCV) replication and hepatic damage were analyzed in three chimpanzees observed for 180 days after inoculation with HCV genotype 1a. Levels of miR-122 in the liver and serum were measured by real-time RT PCR in serial liver biopsies and serum samples. Hepatic miR-122 levels were normalized separately for each of three chimpanzees with small RNAs and microRNAs that are endogenous to the liver and are stably expressed. Two- to 4-fold rise in hepatic miR-122 levels was observed at the onset of HCV infection (the first 4 weeks) when HCV titers in the liver and serum increased rapidly in all three chimpanzees in concordance with in vitro data indicating the miR-122 significance for HCV replication. Between 10 to 14 weeks after inoculation, when hepatic and serum HCV RNA titers exceeded 3 logs and alanine aminotransferase (ALT) activity was elevated, hepatic miR-122 levels were in decline. Cumulative data derived from all three chimpanzees from 180 days of observation documented an inverse (negative) correlation between hepatic miR-122 and HCV RNA in the liver and serum and positive correlation between level of serum miR-122 and HCV replication. Subsequent rise of miR-122 level during HCV clearance and ALT normalization suggested a tri-phasic occurrence of the relationship among hepatic miR-122 expression, HCV replication and hepatic destruction, which was the most apparent in one chimpanzee but less evident in two other animals. In vivo kinetics of hepatic and serum miR-122, HCV replication and hepatic destruction reflects complexities of the virus-host interaction during the acute phase of HCV infection.

Alteration of hepatic nuclear receptor-mediated signaling pathways in hepatitis C virus patients with and without a history of alcohol drinking

The current study tests a hypothesis that nuclear receptor signaling is altered in chronic hepatitis C patients and that the altered pattern is specific to alcohol drinking history. The expression of a panel of more than 100 genes encoding nuclear receptors, coregulators, and their direct/indirect targets was studied in human livers. Gene expression pattern was compared between 15 normal donor livers and 23 hepatitis C virus (HCV) genotype 1-positive livers from patients without a drinking history (matched for age, sex, and body mass index). HCV infection increased the expression of nuclear receptors small heterodimer partner and constitutive androstane receptor (CAR) as well as genes involved in fatty acid trafficking, bile acid synthesis and uptake, and inflammatory response. However, the expression of retinoid X receptor (RXR) α, peroxisomal proliferator-activated receptor (PPAR) α and β as well as steroid regulatory element-binding protein (SREBP)-1c was decreased in HCV-infected livers. Gene expression pattern was compared in chronic hepatitis C patients with and without a drinking history. Alcohol drinking increased the expression of genes involved in fatty acid uptake, trafficking, and oxidation, but decreased the expression of genes responsible for gluconeogenesis. These changes were consistent with reduced fasting plasma glucose levels and altered expression of upstream regulators that include RXRα, PPARα, and CAR. The messenger RNA levels of fibroblast growth factor 21, interleukin-10, and fatty acid synthase, which are all regulated by nuclear receptors, showed independent correlation with hepatic HCV RNA levels. Our findings suggest that those genes and pathways that showed altered expression could potentially be therapeutic targets for HCV infection and/or alcohol drinking-induced liver injury.

Durability of a sustained virologic response in patients with chronic hepatitis C treated with peginterferon and ribavirin

A Sustained virologic response is durable in patients with chronic hepatitis C with peginterferon alfa-2a and ribavirin. Swain MG, Lai MY, Shiffman ML, Cooksley WG, Zeuzem S, Dieterich DT, Abergel A, Pessoa MG, Lin A, Tietz A, Connell EV, Diago M. Gastroenterology 2010;139(5):1593-1601. The current standard therapy for chronic hepatitis C virus (HCV) infection is the combination of pegylated interferon and ribavirin. The ultimate goal of treatment in patients with chronic HCV infection is to completely eradicate HCV in infected hosts, and prevent liver-related morbidity and mortality. However, these goals are difficult to measure and it takes long time to assess the achievement of these goals. A sustained virologic response (SVR) has been used as a surrogate marker of these ultimate goals in treatment of chronic hepatitis C. An SVR is defined as undetectable serum HCV RNA at 24 weeks after the end of treatment. To be a useful surrogate marker, achievement of SVR should result in the reduction of liver-related morbidity and mortality, complete eradication of HCV in infected hosts, and no late virologic relapse. Achievement of SVR has been associated with improvement in liver histology and health-related quality of life, as well as a reduced risk of hepatocellular carcinoma and liver-related mortality.1-3 The survival of patients who achieved SVR was reported to be comparable to that of general population, matched for age and sex.4 However, the development of hepatocellular carcinoma was reported despite achievement of SVR in patients with chronic hepatitis C and advance fibrosis.5 There is some debate as to whether achievement of SVR results in complete eradication of HCV. In a study of 2089 patients with chronic hepatitis C from 3 clinical trials, hepatic HCV RNA was found to correlate with serum HCV RNA in all cases, and 98% of sustained virologic responders had undetectable hepatic HCV RNA.6 Several small studies have described patients who achieved SVR but have residual HCV RNA detected in peripheral blood mononuclear cells with highly sensitive reverse transcription and nested polymerase chain reaction assays.7,8 Whether these findings represent a replication-competent virus or have any clinical significance is unclear. Several small studies reported SVR achieved with conventional interferon alone or in combination with ribavirin was durable. In these reports, serum HCV RNA remained undetectable in 92-100% of patients who achieved SVR with 2 to 13 years of follow-up.4,5,9-12 However, there are limited data on the durability of SVR achieved with peginterferon in patients with chronic hepatitis C. Swain et al. studied the durability of SVR achieved with peginterferon in 1343 patients.13 They analyzed the durability of SVR in large cohort of patients who were enrolled in one of 9 randomized controlled clinical trials,14-22 which evaluated the efficacy of peginteferon alfa-2a alone or in combination with ribavirin. Total 1343 treatment-naive patients who achieved SVR and had at least one post-baseline HCV RNA evaluation were assessed. One thousand three hundred thirty one of 1343 patients (99.1%) remained HCV RNA undetectable in serum at mean of 3.9 years after completion of treatment. The durability of SVR in this study was independent of patient characteristics such as HCV mono-infection, HCV-HIV co-infection, or ALT level. The durability of SVR also appeared to be unaffected by the type of treatment. HCV RNA was detected in the serum of 12 patients who obtained SVR in their original studies at mean of 666 days after completing treatment. Among the 12 patients with reappearance of HCV RNA, there were no baseline or treatment factors associated with reappearance of HCV RNA including HCV genotype, sex, age, race, risk factor for infection, or baseline viral load. It was unknown whether these cases reflect relapse or re-infection because of the paucity of paired samples. The findings of this study were comparable with those reported from smaller studies with conventional interferon. There was one domestic report on the durability of interferon-based therapy in patients with chronic hepatitis C.23 In this study, 73 patients who achieved SVR with conventional interferon or peginterferon in combination with ribavirin were included. HCV RNA reappeared in 8 of 73 (11%), but persistent viremia remained only in one patient (1.4%). Transient positive tests probably represent false-positive test, and the durability of SVR in Korean patients with chronic hepatitis C appears to be similar with those reported in other studies. In conclusion, this study showed SVR achieved with peginterferon alfa-2a alone or in combination with ribavirin is durable in 99% of patients for at least 4 years after completion of therapy and is independent of patient or treatment characteristics.

Virus‐specific T‐cell responses associated with hepatitis C virus (HCV) persistence in the liver after apparent recovery from HCV infection

Hepatitis C virus (HCV) RNA persistence in the liver has been described even after apparent resolution of HCV infection. Because T-cell reactivity plays a role in recovery from HCV infection, virus-specific T-cell responses were investigated in apparently recovered individuals in whom hepatic HCV RNA persistence was documented: 15 sustained virological responders to interferon (IFN)-treatment and 9 asymptomatic aviremic anti-HCV carriers. HCV-specific CD4(+) T-cell proliferative responses were detected significantly more often in apparently recovered individuals (sustained virological responders: 60%; asymptomatic anti-HCV carriers: 66%) compared with 50 chronic hepatitis C patients (28%; P < 0.05). However, T-cell frequencies and numbers tended to decline over time and the number of HCV proteins targeted by CD4(+) T-cell proliferative responses was limited. Interestingly, liver viral load correlated inversely with virus-specific immune responses. Thus, CD4(+) T-cell responders showed significantly lower hepatic HCV RNA levels (P < 0.05). HCV-specific IFN-gamma-secreting CD4(+) T-cells were not detected in all the apparently recovered patients although they were found significantly more often compared with chronic hepatitis C patients (P < 0.05). Also, HCV NS3-specific CD8(+) T-cells were detected in 11 HLA-A2-positive apparently recovered individuals (8 sustained virological responders and 3 asymptomatic anti-HCV carriers); T-cell frequencies tended to be greater in those patients who had lower hepatic viral levels. In conclusion, HCV-specific T-cells are detectable in apparently recovered individuals in whom HCV RNA can persist in the liver indicating that HCV replication may be prolonged in the face of an insufficient or inadequate virus-specific CD4(+) and CD8(+) T-cell response.

Successful Treatment of Fulminant Hepatitis C by Therapy with Alpha Interferon and Ribavirin

Acute hepatitis C virus (HCV) infection has been identified as an important cause of fulminant hepatic failure (FHF), characterized by rapid deterioration of liver function from massive hepatic necrosis leading to encephalopathy and multiorgan failure with a high mortality rate of 65 to 92% (2, 3). Alpha interferon-ribavirin is effective in treating acute and chronic HCV infections (4, 8). We report the details of successful antiviral therapy for a patient with HCV-associated FHF, accompanied by HCV eradication and prevention of chronicity. The present study was approved by the ethics committee of Kaohsiung Municipal HsiaoKang Hospital. Informed consent was obtained from the patient's next of kin. A 49-year-old Taiwanese woman presented in March 2002 with a 7-day history of anorexia and tea-colored urine. She had received an illegal percutaneous administration of folk remedies 2 months ago and had no experience with interferon-ribavirin previously. Rapid deterioration of liver function with coagulopathy preceding an irritable mood and mental disturbance developed at day 4. Seroconversion of antibodies to HCV with HCV genotype 1b infection was detected. She had serum levels of bilirubin of 14.95, 20.12, and 27.57 mg/dl at days 0, 3, and 7, respectively; alanine aminotransferase levels of 1,198, 1,896, and 2,100 U/liter, respectively; and HCV RNA levels of 7.22, 7.85, and 8.37 logs (IU/ml), respectively (Fig. ​(Fig.1).1). Based on the assessment of clinical and virologic measurements, HCV-associated FHF was diagnosed (2). Other conditions predisposing to fulminant hepatitis, including hepatitis A, B, and E viruses, cytomegalovirus, Epstein-Barr virus, and autoimmune or alcohol- or drug-related causes were excluded. Serologic markers of human immunodeficiency virus were negative. FIG. 1. Clinical course and biochemical and molecular profiles of a patient with fulminant hepatitis C treated with alpha interferon and ribavirin combination therapy. Serum HCV RNA was determined by using a quantitative branched DNA assay (VERSANT HCV RNA 3.0 ... Because of the rapid hepatic decompensation and the low probability of an available liver donor, we conducted a treatment protocol at day 7 with recombinant alpha interferon 2b (3 × 106 U given intramuscularly daily) plus 800 mg of oral ribavirin daily for 2 weeks, followed by alpha interferon 2b (6 × 106 U thrice weekly) plus ribavirin (1,000 mg/day) for 22 weeks. Her psychoneurological and gastrointestinal manifestations were reversed, her liver biochemistry normalized, and seronegativity of HCV RNA developed 1 month later and was sustained throughout the 2-year follow-up period. Only mild and self-limited side effects, such as hemolytic anemia, were noted. Liver histopathology revealed acute submassive necrosis at 10 days after antiviral therapy, minimal necroinflammatory activity at the end of treatment, and complete remission to normal liver histology at 6 months after the end of treatment. Hepatic HCV RNA was undetectable in the latter two specimens. Orthotopic liver transplantation, an established treatment option for FHF, is associated with a number of problems, including donor shortages, the risk of death while waiting for a transplant, postoperative mortality, universal HCV recurrence, and rapid progression of HCV-related disease in liver transplant recipients (1, 2). Our patient was characterized by continuous viral replication and high HCV RNA levels, and the extent of liver damage correlated with the magnitude of viral replication (3), implying that viral factors are important in the pathogenesis of HCV-related FHF. Effective antiviral therapy, which quickly lowers the viral loads (6), may rescue patients with HCV-associated FHF from rapid, extensive destruction of hepatocytes and provide an opportunity for adequate regeneration of native liver. However, alpha interferon is not recommended for patients with hepatitis B-related hepatic decompensation because of the risk of developing overt liver failure (5, 7). Physicians should be careful when treating FHF patients with interferon.

Intrahepatic HCV RNA loads in 37 HIV-HCV co-infected patients with controlled HIV infection.

Serum and intrahepatic hepatitis C virus (HCV) RNA were measured in 37 HIV-HCV co-infected patients with controlled human immunodeficiency virus (HIV) infection and correlated with clinical, biological, and histological parameters. Thirty-seven interferon-naive patients underwent liver biopsy. HCV-induced activity (A) and fibrosis (F) were evaluated with METAVIR score. The 37 patients included had HIV plasma loads < 10,000 copies/ml, CD4(+) count > 250/microl. All the patients but two were receiving antiretroviral treatment. Liver tissue and sera were used for measurement of HCV RNA by the Cobas Amplicor HCV Monitor. All patients had serum and liver HCV RNA, and both levels were correlated (r = 0.47; P = 0.003). Intrahepatic HCV load did not depend on age, sex, duration of HCV infection, CD4(+), HCV genotype, or fibrosis. AST levels correlated with intrahepatic HCV load (r = 0.52; P = 0.001). Patients with METAVIR A1/A2 had significantly lower levels of liver HCV-RNA than were found in patients with METAVIR A3 (P = 0.026). Highly active antiretroviral therapy (HAART) including protease inhibitors(PI)-treated patients had significantly lower intrahepatic HCV load (P = 0.04). A weak but significant correlation between serum and liver HCV RNA was found. The amount of hepatic HCV RNA was correlated with AST levels, histological activity, but not with HCV genotype or fibrosis. The immune improvement associated with PI regimens could help reduce HCV load, supporting a protective effect of PI-induced immune restoration.

Hepatic HCV RNA before and after treatment with interferon alone or combined with ribavirin

The clinical use of measuring hepatic hepatitis C virus (HCV) RNA before and after therapy in patients with chronic hepatitis C has been assessed in a number of small clinical trials. Viral clearance from the liver may be a better marker of long-term response than eradication of serum HCV RNA. The aim of this study was to evaluate quantitative hepatic HCV-RNA measurements before and after antiviral therapy. Two thousand eighty-nine chronic hepatitis C patients were enrolled in 3 published clinical trials evaluating interferon alfa-2b alone or with ribavirin either as initial therapy or for interferon relapse. Hepatic HCV-RNA quantitation was performed with a modified reverse-transcription polymerase chain reaction (RT-PCR) before and 24 weeks after therapy in 951 and 1,316 patients, respectively. Pretherapy hepatic HCV-RNA concentrations correlated best with serum HCV-RNA concentrations (R = .236, P = .0001) and negatively correlated with alanine transaminase (ALT) values (−0.178, P = .0001), duration of infection (−0.09, P = .02), parenchymal injury (−0.135, P = .0001), histologic activity index (HAI) inflammatory score (−0.085, P = .01), Knodell fibrosis score (−0.072, P = .03), and body weight (−0.078, P = .02). In paired liver biopsy specimens (n = 534), change in hepatic HCV RNA correlated with the change in the HAI (R = .346, P = .0001). Of 400 sustained virologic responders (SVR), 393 (98%) had undetectable hepatic HCV RNA, whereas 7 (2%) had detectable hepatic HCV RNA; 5 have been followed and 2 have had reappearance of serum HCV RNA 12 months after therapy. In conclusion, measurement of hepatic HCV RNA before or after therapy reflects changes observed in serum HCV RNA, and correlates inversely with hepatic inflammation and fibrosis, but otherwise has minimal clinical use. (HEPATOLOGY 2002;35:688-693.)

Plasma hepatitis C virus (HCV) RNA is comparable to hepatic HCV RNA when obtained while on therapy at the end of treatment as a predictor of sustained response in patients with chronic hepatitis C infection

Plasma hepatitis C virus (HCV) RNA is comparable to hepatic HCV RNA when obtained while on therapy at the end of treatment as a predictor of sustained response in patients with chronic hepatitis C infection

Plasma hepatitis C virus (HCV) RNA is comparable to hepatic HCV RNA when obtained while on therapy at the end of treatment as a predictor of sustained response in patients with chronic hepatitis C infection

Plasma Hepatitis C Virus (HCV) RNA is Comparable to Hepatic HCV RNA When Obtained While on Therapy at the End of Treatment as a Predictor of Sustained Response in Patients with Chronic Hepatitis C Infection Line J. Deguzman, Jose M. Niato, Arrowhead Regional Medical Ctr, Colton, CA; Shelley A. Deguzman, Inland Empire Digest Disease & Liver Ctr, Rediands, CA; Andrew Conrad, National Genetics institute, Cuiver City, CA; Bradley Collins, Arrowhead Regidna/Medical Ctr, Colton, CA

Patients co-infected with human immunodeficiency virus and hepatitis C virus demonstrate higher levels of hepatic HCV RNA.

Serum and liver hepatitis C virus (HCV) RNA levels in patients with hepatitis C have previously been quantified using different techniques. In this work, we used an automated, multicycle, polymerase chain reaction (PCR)-based technique to quantify HCV RNA in 1-2 mm of frozen liver tissue, and in serum, from 70 patients with antibodies to HCV (anti-HCV), with and without human immunodeficiency virus (HIV) co-infection. Stored liver tissue and sera collected at the time of liver biopsy were used for measurement of HCV RNA. Forty-eight HCV patients and 22 HIV/HCV co-infected patients were studied. Co-infected patients had significantly higher median serum and liver HCV RNA (6.7 log copies ml-1 serum and 2.90 log copies microg-1 liver nucleic acids) than patients with HCV alone (6.2 log copies ml-1 serum and 2.19 log copies microg-1 liver nucleic acids). There was only a weak correlation between serum and liver HCV RNA (r = 0.43). There was no correlation between liver and serum HCV RNA and host factors such as duration of disease, CD4 counts, alanine aminotransferase levels or histological score. There was no correlation with HCV genotype. Co-infected patients were more likely to harbour HCV genotype 1 (85%) when compared to patients with HCV alone (58%). An identical genotype was found in liver and serum in 89% of those tested; in 11%, a mixed genotype was present in serum. Patients with HCV genotypes 1 and non-1 had similar histological scores. Hence, an automated PCR-based technique is useful for measuring both liver and serum HCV RNA. Serum HCV genotypes closely paralleled those found in liver tissue. HIV co-infection was associated with higher serum, as well as intrahepatic, HCV RNA levels, by mechanisms not directly related to CD4 counts. The lack of correlation between liver HCV RNA and histology suggests that HCV is not directly cytopathic.

Clinical significance of hepatic HCV RNA in patients with chronic hepatitis C demonstrating long-term sustained response to interferon-alpha therapy

Whether sustained biochemical response and absence of serum HCV RNA in the 6-12 months following suspension of interferon-alpha (IFN-alpha) therapy reflect definitive viral clearance in patients with chronic hepatitis C virus (HCV) infection is controversial. To obtain more information on this topic, HCV RNA was sought in both liver and serum samples of 25 long-term responders who were followed for a median period of 39 months (range 21-79) after discontinuation of IFN-alpha. Liver biopsy was undertaken before and 6 to 12 months after IFN-alpha withdrawal. Liver and serum HCV RNA were tested by a nested polymerase chain reaction. Twenty-two patients (88%) tested negative for both liver and serum HCV RNA, two patients had detectable HCV RNA in both liver and serum, and one patient showed persistent HCV RNA only in the liver. Post-treatment liver histology improved markedly in all patients, including those with viral persistence. During further follow-up, biochemical remission was maintained in all patients except one in whom both serum and liver specimens remained HCV RNA positive. The data indicate that the large majority of long-term responders test negative for HCV RNA in the liver, which suggests definitive eradication of HCV RNA infection.

Hepatitis C virus: Quantitation and distribution in liver

The optimal method for viral quantitation and the most appropriate site for determining viral load in patients with chronic hepatitis C virus (HCV) infection are unknown. We developed a method for measuring HCV RNA in the liver with the following features: 1) efficient extraction of RNA from tissue (89% of RNA recovered); 2) accurate amplification using branched DNA with strong concordance between a single sample tested on multiple occasions either in the same or in different runs; 3) good sensitivity (95%) and specificity (100%). HCV RNA was detected in as little as 2 mg of tissue, and viral load determined in a needle biopsy was representative of viral load in other parts of the liver. Within individual livers, 68% of the samples quantitated were within 1.5-fold of the geometric mean, and 95% were within 2.2-fold of the geometric mean. The mean ratio of virus in the liver and serum was 103, range 17.4-286. A delay of 30 minutes before freezing the liver tissue resulted in a reduction in the measured viral load in some, but not all instances. A sensitive, specific and reproducible method for quantitating HCV RNA in the liver has been developed. Measurement of viral load at one site was representative of viral load at other sites. While hepatic HCV RNA levels are consistently greater than serum levels, the ratio of liver of serum viral load varies widely. The clinical use of measurement of viral load in the liver remains to be defined.

Hepatic hepatitis C virus RNA as a predictor of a long-term response to interferon-alpha therapy.

To identify predictors of a long-term response to interferon-alpha therapy in chronic hepatitis C and to determine whether hepatitis C virus (HCV) was eradicated in patients with chronic hepatitis C who had a long-term response to therapy. A retrospective analysis. In- and outpatient liver clinic of a municipal hospital in Japan. 47 patients with chronic hepatitis C who responded to interferon-alpha were divided into two groups: 22 patients with a long-term response (serum aminotransferase levels remained normal for > 1 year after therapy) and 25 patients with a short-term response (serum aminotransferase levels increased again after therapy). Genotyping of HCV, titers of HCV RNA in liver and serum samples (using the reverse transcriptase-polymerase chain reaction), histologic activity index, and liver histologic tests during and 1 year after therapy. Among the 22 long-term responders, HCV RNA was no longer detectable in liver and serum samples of 21 (95%) at the end of therapy and remained undetectable in the serum of 20 (91%) and in the liver of 19 (86%) 1 year after therapy. Liver histologic tests improved substantially immediately after therapy and 1 year after therapy in the long-term responders; however, 18 (82%) of these patients still had mild, chronic hepatitis. Among the 25 short-term responders, HCV RNA was still detected in the liver of 19 (76%) and in the serum of 9 (36%) at the end of therapy. Multivariate logistic regression analysis showed that the persistent presence of hepatic HCV RNA at the end of therapy was the strongest predictor of relapse. These findings suggest that HCV infection was eradicated in most of the long-term responders to interferon-alpha therapy because HCV RNA could no longer be detected in their serum and liver samples and because a significant improvement gradually occurred in their liver histologic results. The persistent presence of hepatic HCV RNA at the end of therapy was the most important predictor of relapse.