Point-of-care HCV RNA testing improves hepatitis C testing rates and allows rapid treatment initiation among people who inject drugs attending a medically supervised injecting facility.

BACKGROUND: To achieve hepatitis C virus (HCV) elimination targets, simplified care engaging people who inject drugs is required. We evaluated whether fingerstick HCV RNA point-of-care testing (PoCT) increased the proportion of clients attending a supervised injecting facility who were tested for hepatitis C. METHODS: Prospective single-arm study with recruitment between 9 November 2020 and 28 January 2021 and follow-up to 31 July 2021. Clients attending the supervised injecting facility were offered HCV RNA testing using the Xpert® HCV Viral Load Fingerstick (Cepheid, Sunnyvale, CA) PoCT. Participants with a positive HCV RNA test were prescribed direct acting antiviral (DAA) therapy. The primary endpoint was the proportion of clients who engaged in HCV RNA PoCT, compared to a historical comparator group when venepuncture-based hepatitis C testing was standard of care. RESULTS: Among 1618 clients who attended the supervised injecting facility during the study period, 228 (14%) engaged in PoCT. This was significantly higher than that observed in the historical comparator group (61/1,775, 3%; p < 0.001). Sixty-five (28%) participants were HCV RNA positive, with 40/65 (62%) receiving their result on the same day as testing. Sixty-one (94%) HCV RNA positive participants were commenced on DAA therapy; 14/61 (23%) started treatment on the same day as diagnosis. There was no difference in the proportion of HCV RNA positive participants commenced on treatment with DAA therapy when compared to the historical comparator group (61/65, 94% vs 22/26, 85%; p = 0.153). However, the median time to treatment initiation was significantly shorter in the PoCT cohort (2 days (IQR 1-20) vs 41 days (IQR 22-76), p < 0.001). Among participants who commenced treatment and had complete follow-up data available, 27/36 (75%) achieved hepatitis C cure. CONCLUSIONS: HCV RNA PoCT led to a significantly higher proportion of clients attending a supervised injecting facility engaging in hepatitis C testing, whilst also reducing the time to treatment initiation.

Single-cell atlas of the liver myeloid compartment before and after cure of chronic viral hepatitis.

BACKGROUND & AIMS: Chronic viral infections present serious public health challenges; however, direct-acting antivirals (DAAs) are now able to cure nearly all patients infected with hepatitis C virus (HCV), representing the only cure of a human chronic viral infection to date. DAAs provide a valuable opportunity to study immune pathways in the reversal of chronic immune failures in an in vivo human system. METHODS: To leverage this opportunity, we used plate-based single-cell RNA-seq to deeply profile myeloid cells from liver fine needle aspirates in patients with HCV before and after DAA treatment. We comprehensively characterised liver neutrophils, eosinophils, mast cells, conventional dendritic cells, plasmacytoid dendritic cells, classical monocytes, non-classical monocytes, and macrophages, and defined fine-grained subpopulations of several cell types. RESULTS: We discovered cell type-specific changes post-cure, including an increase in MCM7+STMN1+ proliferating CD1C+ conventional dendritic cells, which may support restoration from chronic exhaustion. We observed an expected downregulation of interferon-stimulated genes (ISGs) post-cure as well as an unexpected inverse relationship between pre-treatment viral load and post-cure ISG expression in each cell type, revealing a link between viral loads and sustained modifications of the host's immune system. We found an upregulation of PD-L1/L2 gene expression in ISG-high neutrophils and IDO1 expression in eosinophils, pinpointing cell subpopulations crucial for immune regulation. We identified three recurring gene programmes shared by multiple cell types, distilling core functions of the myeloid compartment. CONCLUSIONS: This comprehensive single-cell RNA-seq atlas of human liver myeloid cells in response to cure of chronic viral infections reveals principles of liver immunity and provides immunotherapeutic insights. CLINICAL TRIAL REGISTRATION: This study is registered at ClinicalTrials.gov (NCT02476617). IMPACT AND IMPLICATIONS: Chronic viral liver infections continue to be a major public health problem. Single-cell characterisation of liver immune cells during hepatitis C and post-cure provides unique insights into the architecture of liver immunity contributing to the resolution of the first curable chronic viral infection of humans. Multiple layers of innate immune regulation during chronic infections and persistent immune modifications after cure are revealed. Researchers and clinicians may leverage these findings to develop methods to optimise the post-cure environment for HCV and develop novel therapeutic approaches for other chronic viral infections.

LOXL-2 and TNC-C are markers of liver fibrogenesis in HCV/HIV-, HIV- and HCV-infected patients.

Background: Lysil oxidase like enzyme-2 (LOXL-2) and TNC-C play important roles in organ fibrosis. We assessed circulating LOXL-2 and TNC-C levels and their relationship to fibrosis severity in HIV- and/or HCV-infected individuals. Methods: Healthy controls (n = 22), HIV mono- (n = 15), HCV mono- (n = 52) and HCV/HIV-co-infected (n = 92) subjects were included. Results: LOXL-2 and TNC-C levels were significantly higher in HCV mono- and HCV/HIV-co-infected individuals with F0 compared to healthy controls. In addition, in HCV/HIV-co-infected individuals, LOXL-2 levels were higher in intermediate fibrosis compared to no/mild fibrosis. Conclusion: In HCV/HIV-co-infected study participants, both LOXL-2 and TNC-C were significantly higher in intermediate fibrosis compared to no/mild fibrosis, but did not further increase with advanced fibrosis. Furthermore, both markers were elevated among HCV/HIV-positive individuals with mild/no fibrosis.

The Importance of Prisons in Achieving Hepatitis C Elimination: Insights from the Australian Experience.

Following the availability of highly effective direct-acting antivirals (DAAs) to treat hepatitis C infection, the uptake of treatment by people living with hepatitis C rose dramatically in high- and middle-income countries but has since declined. To achieve the World Health Organization's (WHO) 2030 target to eliminate hepatitis C as a public health threat among people who inject drugs, an increase in testing and treatment is required, together with improved coverage of harm reduction interventions. The population that remains to be treated in high- and middle-income countries with high hepatitis C prevalence are among the most socially disadvantaged, including people who inject drugs and are involved in the criminal justice system, a group with disproportionate hepatitis C prevalence, compared with people in the wider community. Imprisonment provides an unrivalled opportunity for screening and treating large numbers of people for hepatitis C, who may not access mainstream health services in the community. Despite some implementation challenges, evidence of the efficacy, acceptability, and cost-effectiveness of in-prison hepatitis treatment programs is increasing worldwide, and evaluations of these programs have demonstrated the capacity for treating people in high numbers. In this Perspective we argue that the scale-up of hepatitis C prevention, testing, and treatment programs in prisons, along with the investigation of new and adapted approaches, is critical to achieving WHO elimination goals in many regions; the Australian experience is highlighted as a case example. We conclude by discussing opportunities to improve access to prevention, testing, and treatment for people in prison and other justice-involved populations, including harnessing the changed practices brought about by the COVID-19 pandemic.

Hepatitis B and Hepatitis C Virus Infection Promote Liver Fibrogenesis through a TGF-ß1-Induced OCT4/Nanog Pathway.

Hepatitis B virus (HBV)/hepatitis C virus (HCV) coinfection accelerates liver fibrosis progression compared with HBV or HCV monoinfection. Octamer binding transcription factor 4 (OCT4) and Nanog are direct targets of the profibrogenic TGF-ß1 signaling cascade. We leveraged a coculture model to monitor the effects of HBV and HCV coinfection on fibrogenesis in both sodium taurocholate cotransporting polypeptide-transfected Huh7.5.1 hepatoma cells and LX2 hepatic stellate cells (HSCs). We used CRISPR-Cas9 to knock out OCT4 and Nanog to evaluate their effects on HBV-, HCV-, or TGF-ß1-induced liver fibrogenesis. HBV/HCV coinfection and HBx, HBV preS2, HCV Core, and HCV NS2/3 overexpression increased TGF-ß1 mRNA levels in sodium taurocholate cotransporting polypeptide-Huh7.5.1 cells compared with controls. HBV/HCV coinfection further enhanced profibrogenic gene expression relative to HBV or HCV monoinfection. Coculture of HBV and HCV monoinfected or HBV/HCV coinfected hepatocytes with LX2 cells significantly increased profibrotic gene expression and LX2 cell invasion and migration. OCT4 and Nanog guide RNA independently suppressed HBV-, HCV-, HBV/HCV-, and TGF-ß1-induced α-SMA, TIMP-1, and Col1A1 expression and reduced Huh7.5.1, LX2, primary hepatocyte, and primary human HSC migratory capacity. OCT4/Nanog protein expression also correlated positively with fibrosis stage in liver biopsies from patients with chronic HBV or HCV infection. In conclusion, HBV and HCV independently and cooperatively promote liver fibrogenesis through a TGF-ß1-induced OCT4/Nanog-dependent pathway.

Fatty Acids Activate the Transcriptional Coactivator YAP1 to Promote Liver Fibrosis via p38 Mitogen-Activated Protein Kinase.

BACKGROUND & AIMS: Patients with simple steatosis (SS) and nonalcoholic steatohepatitis can develop progressive liver fibrosis, which is associated with liver-related mortality. The mechanisms contributing to liver fibrosis development in SS, however, are poorly understood. SS is characterized by hepatocellular free fatty acid (FFA) accumulation without lobular inflammation seen in nonalcoholic steatohepatitis. Because the Hippo signaling transcriptional coactivator YAP1 (YAP) has previously been linked with nonalcoholic fatty liver disease (NAFLD)-related fibrosis, we sought to explore how hepatocyte FFAs activate a YAP-mediated profibrogenic program. METHODS: We analyzed RNA sequencing data from a GEO DataSet (accession: GSE162694) consisting of 143 patients with NAFLD. We also performed immunohistochemical, immunofluorescence, immunoblot, and quantitative reverse-transcription polymerase chain reaction analyses (qRT-PCR) in liver specimens from NAFLD subjects, from a murine dietary NAFLD model, and in FFA-treated hepatic spheroids and hepatocytes. RESULTS: YAP-target gene expression correlated with increasing fibrosis stage in NAFLD patients and was associated with fibrosis in mice fed a NAFLD-inducing diet. Hepatocyte-specific YAP deletion in the murine NAFLD model attenuated diet-induced fibrosis, suggesting a causative role of YAP in NAFLD-related fibrosis. Likewise, in hepatic spheroids composed of Huh7 hepatoma cells and primary human hepatic stellate cells, Huh7 YAP silencing reduced FFA-induced fibrogenic gene expression. Notably, inhibition of p38 mitogen-activated protein kinase could block YAP activation in FFA-treated Huh7 cells. CONCLUSIONS: These studies provide further evidence for the pathological role of YAP in NAFLD-associated fibrosis and that YAP activation in NAFLD may be driven by FFA-induced p38 MAPK activation.

The risk of hepatitis C virus recurrence in hepatitis C virus-infected patients treated with direct-acting antivirals after achieving a sustained virological response: A comprehensive analysis.

BACKGROUND & AIMS: The risk for hepatitis C virus (HCV) recurrence persists after HCV eradication with direct-acting antivirals (DAAs), particularly in patients with ongoing high-risk behaviours. Our aim was to assess the risk of HCV recurrence (late relapse and/or reinfection) post-sustained virological response (SVR). METHODS: We searched the literature for studies reporting HCV recurrence rates post-SVR in PubMed, Web of Science and the Cochrane Library. Identified publications were divided into groups based on patient risk for HCV reinfection: low-risk HCV mono-infection, high-risk HCV mono-infection and a human immunodeficiency virus (HIV)/HCV coinfection. The HCV recurrence rate for each study was calculated by using events divided by the person-years of follow-up (PYFU). HCV recurrence was defined as confirmed, detectable HCV RNA post-SVR. RESULTS: In the 16 studies of low-risk patients, the pooled recurrence rate was 0.89/1000 PYFU (95% confidence interval [CI], 0.16-2.03). For the 19 studies of high-risk patients, the pooled recurrence rate was 29.37/1000 PYFU (95% CI, 15.54-46.91). For the eight studies of HIV/HCV-coinfected patients, the pooled recurrence rate was 23.25/1000 PYFU (95% CI, 4.24-53.39). The higher pooled estimates of recurrence in the high-risk and HIV/HCV-coinfected populations were predominantly driven by an increase in reinfection rather than late relapse. CONCLUSIONS: The HCV recurrence risk after achieving SVR with all-oral DAAs therapy is low, and the risk of HCV recurrence in high-risk and HIV/HCV-coinfected populations was driven by an increase in reinfection rather than late relapse.

Helicobacter pylori antimicrobial resistance in Melbourne, Australia. Time to review therapeutic guidelines?

BACKGROUND: Antimicrobial resistance (AMR) in Helicobacter pylori is a global concern. The AMR data to inform the Australian Therapeutic Guidelines are based on data over 20 years old. AIMS: To evaluate the frequency of AMR in H. pylori isolates from gastric biopsy specimens received in our laboratory in Melbourne, Australia. To review the literature on resistance rates in Australia and compare historic data. METHODS: A retrospective, observational study summarising AMR rates in all H. pylori isolates from our laboratory from 2015 to June 2020. Microbiology laboratory in metropolitan Melbourne, Australia, receiving referrals from private hospitals, gastroenterology clinics and endoscopy suites. Population minimum inhibitory concentration distributions and frequency of resistance to clarithromycin, amoxicillin, metronidazole and tetracycline in H. pylori isolates. RESULTS: Three hundred and eighty-six H. pylori isolates with susceptibility testing data were identified. The frequency of resistance in this cohort was: clarithromycin 89.9%, amoxicillin 23.5%, metronidazole 66.1% and tetracycline 4.4%. Comparison with historical data may suggest increasing AMR rates in Australia. The main limitation is the lack of treatment history to correlate AMR results. CONCLUSIONS: Definitive conclusions from this cohort cannot be made, but trends suggest rising levels of primary H. pylori AMR rates in Australia. This has important implications for empirical treatment decision making and treatment outcomes. Primary H. pylori AMR requires dedicated studies and current Australian therapeutic guideline recommendations may require re-evaluation. We propose considerations for improving the management of H. pylori in Australia. A centralised public health approach to H. pylori AMR surveillance should be established.

A qualitative exploration of enablers for hepatitis B clinical management among ethnic Chinese in Australia.

An estimated 18% of people living with chronic hepatitis B (CHB) in Australia were born in China. While guideline-based care, including regular clinical monitoring and timely treatment, prevent CHB-related cirrhosis, cancer and deaths, over three-quarters of people with CHB do not receive guideline-based care in Australia. This qualitative study aimed to identify enablers to engagement in CHB clinical management among ethnic Chinese people attending specialist care. Participants self-identified as of Chinese ethnicity and who attended specialist care for CHB clinical management were interviewed in Melbourne in 2019 (n = 30). Semi-structured interviews covered experiences of diagnosis and engagement in clinical management services, and advice for people living with CHB. Interviews were recorded with consent; data were transcribed verbatim and thematically analysed. Receiving clear information about the availability of treatment and/or the necessity of long-term clinical management were the main enablers for participants to engage in CHB clinical management. Additional enablers identified to maintain regular clinical monitoring included understanding CHB increases risks of cirrhosis and liver cancer, using viral load indicators to visualize disease status in patient-doctor communication; expectations from family, peer group and medical professionals; use of a patient recall system; availability of interpreters or multilingual doctors; and largely subsidized healthcare services. In conclusion, to support people attending clinical management for CHB, a holistic response from community, healthcare providers and the public health sector is required. There are needs for public health programmes directed to communicate (i) CHB-related complications; (ii) availability of effective and cheap treatment; and that (iii) long-term engagement with clinical management and its benefits.

Non-invasive fibrosis algorithms are clinically useful for excluding cirrhosis in prisoners living with hepatitis C.

BACKGROUND AND AIMS: Prison-based HCV treatment rates remain low due to multiple barriers, including accessing transient elastography for cirrhosis determination. The AST-to-platelet ratio index (APRI) and FIB-4 scores have excellent negative predictive value (NPV) in hospital cohorts to exclude cirrhosis. We investigated their performance in a large cohort of prisoners with HCV infection. METHODS: This was a retrospective cohort study of participants assessed by a prison-based hepatitis program. The sensitivity, specificity, NPV and positive predictive value (PPV) of APRI and FIB-4 for cirrhosis were then analysed, with transient elastography as the reference standard. The utility of age thresholds as a trigger for transient elastography was also explored. RESULTS: Data from 1007 prisoners were included. The median age was 41, 89% were male, and 12% had cirrhosis. An APRI cut-off of 1.0 and FIB-4 cut-off of 1.45 had NPVs for cirrhosis of 96.1% and 96.6%, respectively, and if used to triage prisoners for transient elastography, could reduce the need for this investigation by 71%. The PPVs of APRI and FIB-4 for cirrhosis at these cut-offs were low. Age ≤35 years alone had a NPV for cirrhosis of 96.5%. In those >35 years, the APRI cut-off of 1.0 alone had a high NPV >95%. CONCLUSION: APRI and FIB-4 scores can reliably exclude cirrhosis in prisoners and reduce requirement for transient elastography. This finding will simplify the cascade of care for prisoners living with hepatitis C.

COVID-19 induced liver function abnormality associates with age.

BACKGROUND: Coronavirus disease 2019 (COVID-19) is a novel infectious disease that may cause fever, dry cough, fatigue and shortness of breath. The impact of COVID-19 on liver function is not well described. RESULTS: We found that the overall frequency of LFT abnormality was 17.6%. Frequency of LFT abnormality was significantly greater in patients with severe/critical (SC) COVID-19 compared to those with mild/moderate (MM) COVID-19 (32.4% vs 11.6%, p=0.011). Among patients with LFT abnormality, the median age was significantly higher in the SC group compared to the MM group (52 vs 39 years, p=0.021). CONCLUSION: COVID-19 is frequently associated with mild liver function abnormality, particularly in individuals with severe/critical COVID-19 who were older. Liver function should be monitored carefully during infection, with judicious use of hepatotoxic agents where possible and avoidance of prolonged hypotension to minimize liver injury in older patients. METHODS: The No. 2 People's Hospital of Fuyang City in China has admitted a total of 159 patients with confirmed COVID-19 since the outbreak from January 2020 to March 2020. We analyzed the incidence of liver function test (LFT) abnormality in these patients with confirmed COVID-19 infection.

Hepatitis B-related outcomes following direct-acting antiviral therapy in Taiwanese patients with chronic HBV/HCV co-infection.

BACKGROUND & AIMS: The outcome of HBV infection, including the dynamics of HBsAg and HBV virological reactivation, among patients coinfected with HCV receiving direct-acting antivirals (DAAs) remains unclear. Thus, we aimed to analyze HBV-related outcomes in these patients. METHODS: Serial HBsAg and HBV DNA levels were measured in 79 HBV/HCV-coinfected patients receiving DAAs (13 receiving anti-HBV nucleot(s)ide analog [NUC] therapy simultaneously). The endpoints included HBsAg dynamics and seroclearance, HBV reactivation (HBV DNA >1 log increase or >100 IU/ml if undetectable at baseline) and HBV-related clinical reactivation. RESULTS: HBsAg levels declined from a median of 73.3 IU/ml at baseline to 16.2 IU/ml at the end-of-DAA treatment and increased to 94.1 IU/ml at 12 months post-treatment. During a mean 11.1-months of follow-up, 8 (10.1%) patients experienced HBsAg seroclearance and 30 (38.0%) HBV reactivation (12-month cumulative incidence, 10.3% and 40.4%, respectively). Patients with pre-treatment HBsAg ≤10 IU/ml had a significantly higher rate of HBsAg seroclearance (hazard ratio [HR] 8.52; 95% CI 1.048-69.312) and lower risk of HBV reactivation than those with pre-treatment HBsAg >10 IU/ml (HR 2.88; 95% CI 1.057-7.844) in multivariate analyses. Six patients (4 cirrhotics) not receiving NUC therapy experienced HBV-related clinical reactivation; 3 of the 4 cirrhotics developed liver failure and 2 died despite immediate NUC therapy. Compared to untreated HBV-monoinfected patients, HBV/HCV-coinfected patients without NUC prophylaxis had a similar rate of HBsAg seroclearance, but a significantly higher risk of HBV reactivation following DAA therapy (HR 6.59; 95% CI 2.488-17.432). CONCLUSIONS: DAA-treated HBV/HCV-coinfected patients had significantly higher rates of HBV seroclearance, particularly among those with low pre-treatment HBsAg titer, but were at higher risk of HBV reactivation, particularly among those with higher pre-treatment HBsAg titer. Prophylactic anti-HBV therapy is essential for cirrhotic patients, irrespective of baseline HBV DNA levels. LAY SUMMARY: We studied outcomes relating to hepatitis B virus (HBV) in patients coinfected with both hepatitis B and C. Patients receiving direct-acting antiviral treatment for hepatitis C were more likely to experience seroclearance (or functional cure of HBV), but were also more likely to experience HBV reactivation, which can lead to hepatitis, liver failure and death. In coinfected cirrhotic patients being treated for HCV, prophylactic treatment for HBV is mandatory.

Hepatitis C Virus Infection in Pregnancy: An Update.

Parenteral transmission is the major route of hepatitis C virus transmission in adults; however, vertical transmission is most common in children. There are several factors that have been shown to be associated with vertical transmission of hepatitis C virus, including hepatitis C virus RNA, human immunodeficiency virus coinfection, and peripheral blood mononuclear cell infection. As there is no effective vaccine to prevent hepatitis C virus infection, and there are no human data describing the safety of the new direct acting antiviral agents in pregnancy, the only preventive strategy for vertical transmission is to treat the hepatitis C virus infection before becoming pregnant. Direct acting antiviral agents are interferon-free, and many are also ribavirin-free. Based on animal studies, sofosbuvir plus ledipasvir may be the best safety profile during pregnancy for now; however, it is too early to recommend treating hepatitis C virus-infected pregnant women with these direct acting antiviral agents currently.

Microrna-130a Downregulates HCV Replication through an atg5-Dependent Autophagy Pathway.

We previously identified that miR-130a downregulates HCV replication through two independent pathways: restoration of host immune responses and regulation of pyruvate metabolism. In this study, we further sought to explore host antiviral target genes regulated by miR-130a. We performed a RT² Profiler™ PCR array to identify the host antiviral genes regulated by miR-130a. The putative binding sites between miR-130a and its downregulated genes were predicted by miRanda. miR-130a and predicted target genes were over-expressed or knocked down by siRNA or CRISPR/Cas9 gRNA. Selected gene mRNAs and their proteins, together with HCV replication in JFH1 HCV-infected Huh7.5.1 cells were monitored by qRT-PCR and Western blot. We identified 32 genes that were significantly differentially expressed more than 1.5-fold following miR-130a overexpression, 28 of which were upregulated and 4 downregulated. We found that ATG5, a target gene for miR-130a, significantly upregulated HCV replication and downregulated interferon stimulated gene expression. miR-130a downregulated ATG5 expression and its conjugation complex with ATG12. ATG5 and ATG5-ATG12 complex affected interferon stimulated gene (ISG) such as MX1 and OAS3 expression and subsequently HCV replication. We concluded that miR-130a regulates host antiviral response and HCV replication through targeting ATG5 via the ATG5-dependent autophagy pathway.