Virus Genotype-Dependent Transcriptional Alterations in Lipid Metabolism and Inflammation Pathways in the Hepatitis C Virus-infected Liver

Despite advances in antiviral therapy, molecular drivers of Hepatitis C Virus (HCV)-related liver disease remain poorly characterised. Chronic infection with HCV genotypes (1 and 3) differ in presentation of liver steatosis and virological response to therapies, both to interferon and direct acting antivirals. To understand what drives these clinically important differences, liver expression profiles of patients with HCV Genotype 1 or 3 infection (n = 26 and 33), alcoholic liver disease (n = 8), and no liver disease (n = 10) were analysed using transcriptome-wide microarrays. In progressive liver disease, HCV genotype was the major contributor to altered liver gene expression with 2151 genes differentially expressed >1.5-fold between HCV Genotype 1 and 3. In contrast, only 6 genes were altered between the HCV genotypes in advanced liver disease. Induction of lipogenic, lipolytic, and interferon stimulated gene pathways were enriched in Genotype 1 injury whilst a broad range of immune-associated pathways were associated with Genotype 3 injury. The results are consistent with greater lipid turnover in HCV Genotype 1 patients. Moreover, the lower activity in inflammatory pathways associated with HCV genotype 1 is consistent with relative resistance to interferon-based therapy. This data provides a molecular framework to explain the clinical manifestations of HCV-associated liver disease

Widespread GLI expression but limited canonical hedgehog signaling restricted to the ductular reaction in human chronic liver disease

Canonical Hedgehog (Hh) signaling in vertebrate cells occurs following Smoothened activation/translocation into the primary cilia (Pc), followed by a GLI transcriptional response. Nonetheless, GLI activation can occur independently of the canonical Hh pathway. Using a murine model of liver injury, we previously identified the importance of canonical Hh signaling within the Pc+ liver progenitor cell (LPC) population and noted that SMO-independent, GLI-mediated signals were important in multiple Pc-ve GLI2+ intrahepatic populations. This study extends these observations to human liver tissue, and analyses the effect of GLI inhibition on LPC viability/gene expression. Human donor and cirrhotic liver tissue specimens were evaluated for SHH, GLI2 and Pc expression using immunofluorescence and qRT-PCR. Changes to viability and gene expression in LPCs in vitro were assessed following GLI inhibition. Identification of Pc (as a marker of canonical Hh signaling) in human cirrhosis was predominantly confined to the ductular reaction and LPCs. In contrast, GLI2 was expressed in multiple cell populations including Pc-ve endothelium, hepatocytes, and leukocytes. HSCs/myofibroblasts (gt;99%) expressed GLI2, with only 1.92% displaying Pc. In vitro GLI signals maintained proliferation/viability within LPCs and GLI inhibition affected the expression of genes related to stemness, hepatocyte/biliary differentiation and Hh/Wnt signaling. At least two mechanisms of GLI signaling (Pc/SMOdependent and Pc/SMO-independent) mediate chronic liver disease pathogenesis. This may have significant ramifications for the choice of Hh inhibitor (anti-SMO or anti-GLI) suitable for clinical trials. We also postulate GLI delivers a pro-survival signal to LPCs whilst maintaining stemness

Impact of Intensive Care Unit Discharge Delay on Liver Transplantation Outcomes

Background: For general intensive care unit (ICU) patients, ICU discharge delay (ICUDD) has been associated with an increased hospital length of stay (LOS) and the acquisition of multi-resistant organism (MRO) infections. The impact of ICUDD on liver transplant (LT) recipients is unknown. Methods: We retrospectively studied consecutive adult LT between 2011 and 2019. ICUDD was defined as >8 h between a patient being cleared for discharge to ward and the patient leaving the ICU. Results: 550 patients received LT and the majority (68.5%) experienced ICUDD. The median time between clearance for ward and the patient leaving the ICU was 25.6 h. No donor or recipient variables were associated with ICUDD. Patients cleared for discharge early in the week (Sunday-Tuesday) and those discharged outside routine work hours were more likely to experience ICUDD (p = 0.001 and p < 0.001, respectively). The median hospital LOS was identical (18 days, p = 0.96) and there were no differences in other patient outcomes. Patients who became colonized with MRO in the ICU spent a longer time there compared to those who remained MRO-free (9 vs. 6 days, p < 0.001); however, this was not due to ICUDD. Conclusion: ICUDD post-LT is common and does not prolong hospital LOS. ICUDD is not associated with adverse patient outcomes or MRO colonization

Accumulation of Deleterious Passenger Mutations Is Associated with the Progression of Hepatocellular Carcinoma

<div><p>In hepatocellular carcinoma (HCC), somatic genome-wide DNA mutations are numerous, universal and heterogeneous. Some of these somatic mutations are drivers of the malignant process but the vast majority are passenger mutations. These passenger mutations can be deleterious to individual protein function but are tolerated by the cell or are offset by a survival advantage conferred by driver mutations. It is unknown if these somatic deleterious passenger mutations (DPMs) develop in the precancerous state of cirrhosis or if it is confined to HCC. Therefore, we studied four whole-exome sequencing datasets, including patients with non-cirrhotic liver (n = 12), cirrhosis without HCC (n = 6) and paired HCC with surrounding non-HCC liver (n = 74 paired samples), to identify DPMs. After filtering out putative germline mutations, we identified 187±22 DPMs per non-diseased tissue. DPMs number was associated with liver disease progressing to HCC, independent of the number of exonic mutations. Tumours contained significantly more DPMs compared to paired non-tumour tissue (258–293 per HCC exome). Cirrhosis- and HCC-associated DPMs do not occur predominantly in specific genes, chromosomes or biological pathways and the effect on tumour biology is presently unknown. Importantly, for the first time we have shown a significant increase in DPMs with HCC.</p></div

Driver mutations in non-tumour tissue.

<p>Patient samples were separated based on the number of mutations in putative driver genes (x-axis, defined as the 20 top recurrently mutated genes in HCC according to COSMIC database, listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0162586#pone.0162586.s011" target="_blank">S3 Table</a>) and analysed the number of benign missense SNVs (A and C) and DPMs (B and D). Significant correlation between DPMs and putative driver mutations (p<0.0001, Spearman rank correlation test) was observed in non-tumour tissue of WES 2. No significant correlation was seen in HCC tissues (p>0.05, Spearman rank correlation test).</p

Clinical characteristics of patients analysed in WES 1: non-HCC liver injury samples.

<p>Clinical characteristics of patients analysed in WES 1: non-HCC liver injury samples.</p

Summary data of publicly-available WES datasets used in this study.

<p>Summary data of publicly-available WES datasets used in this study.</p

DPMs in HCC and surrounding non-tumour tissue.

<p>Variants were classified based on the predicted effect on the amino acid sequence (A). Total benign missense variants (B and D) and DPMs (C and E) in the datasets 1000G and WES 1–4 are shown as a percentage of all somatic exonic mutations. Significantly more DPMs (but not benign missense SNVs) were detected in tumour compared to paired non-tumour tissue (* p<0.05, ** p<0.01, *** p<0.001 and **** p<0.0001, Wilcoxon matched-pairs signed-rank test). Lines link matched non-tumour and tumour tissues samples. NC-non-cirrhosis; C-cirrhosis; NT-non-tumour; T-tumour.</p

Summary statistics for normalised DPMs between datasets.

<p>Summary statistics for normalised DPMs between datasets.</p