Cholangiocarcinoma (CCA) is a destructive malignancy with limited responsiveness to conventional chemotherapy. Although angiotensin receptor blockers (ARBs) have gained attention for their potential ...anticancer activity, little is known about their effects on CCA. The transcriptional co-activator, Yes-associated protein (YAP) is a critical oncogene in several cancers, including CCA. Following recent evidence showing that YAP is regulated by angiotensin II (AT-II), we investigated the effects of an ARB, losartan, on two human CCA cell lines (KKU-M213 and HuCCT-1) with regards to YAP oncogenic regulation. Losartan suppressed AT-II-induced CCA cell proliferation in a dose-dependent manner, induced apoptosis, decreased YAP (Ser127), and downregulated the YAP target genes CTGF, CYR61, ANKRD1, and MFAP5. However, losartan did not affect epithelial–mesenchymal transition, differentiation, or stemness in the CCA cells. Xenograft tumor growth assay showed that oral administration of a low clinical dose of losartan considerably reduced subcutaneous tumor burden and attenuated intratumor vascularization in CCA cell-derived xenograft tumors in BALB/c nude mice. These results indicate that ARB therapy could serve as a potential novel strategy for CCA treatment.
•Angiotensin receptor blocker suppresses human cholangiocarcinoma cell growth.•Angiotensin receptor blocker inhibits YAP oncogenic activity.•Angiotensin receptor blocker shows anti-angiogenic effect in cholangiocarcinoma.
Aim
Intestinal endotoxin is important for the progression of non‐alcoholic steatohepatitis (NASH). Circulating endotoxin levels are elevated in most animal models of diet‐induced non‐alcoholic fatty ...liver disease (NAFLD) and NASH. Furthermore, plasma endotoxin levels are significantly higher in NAFLD patients, which is associated with small intestinal bacterial overgrowth and increased intestinal permeability. By improving the gut microbiota environment and restoring gut‐barrier functions, probiotics are effective for NASH treatment in animal models. It is also widely known that hepatic fibrosis and suppression of activated hepatic stellate cells (Ac‐HSCs) can be attenuated using an angiotensin‐II type 1 receptor blocker (ARB). We thus evaluated the effect of combination probiotics and ARB treatment on liver fibrosis using a rat model of NASH.
Methods
Fisher 344 rats were fed a choline‐deficient/L‐amino acid‐defined (CDAA) diet for 8 weeks to generate the NASH model. Animals were divided into ARB, probiotics, and ARB plus probiotics groups. Therapeutic efficacy was assessed by evaluating liver fibrosis, the lipopolysaccharide Toll‐like receptor (TLR)4 regulatory cascade, and intestinal barrier function.
Results
Both probiotics and ARB inhibited liver fibrosis, with concomitant HSC activation and suppression of liver‐specific transforming growth factor‐β and TLR4 expression. Probiotics reduced intestinal permeability by rescuing zonula occludens‐1 disruption induced by the CDAA diet. Angiotensin‐II type 1 receptor blocker was found to directly suppress Ac‐HSCs.
Conclusions
Probiotics and ARB are effective in suppressing liver fibrosis through different mechanisms. Currently both drugs are in clinical use; therefore, the combination of probiotics and ARB is a promising new therapy for NASH.
DNA methyltransferase 1 (DNMT1) is an essential regulator maintaining both epigenetic reprogramming during DNA replication and genome stability. We investigated the role of DNMT1 in the regulation of ...postnatal liver histogenesis under homeostasis and stress conditions. We generated Dnmt1 conditional knockout mice (Dnmt1Δalb) by crossing Dnmt1fl/fl with albumin‐cyclization recombination transgenic mice. Serum, liver tissues, and primary hepatocytes were collected from 1‐week‐old to 20‐week old mice. The Dnmt1Δalb phenotype was assessed by histology, confocal and electron microscopy, biochemistry, as well as transcriptome and methylation profiling. Regenerative growth was induced by partial hepatectomy and exposure to carbon tetrachloride. The impact of Dnmt1 knockdown was also analyzed in hepatic progenitor cell lines; proliferation, apoptosis, DNA damage, and sphere formation were assessed. Dnmt1 loss in postnatal hepatocytes caused global hypomethylation, enhanced DNA damage response, and initiated a senescence state causing a progressive inability to maintain tissue homeostasis and proliferate in response to injury. The liver regenerated through activation and repopulation from progenitors due to lineage‐dependent differences in albumin‐cyclization recombination expression, providing a basis for selection of less mature and therefore less damaged hepatic progenitor cell progeny. Consistently, efficient knockdown of Dnmt1 in cultured hepatic progenitor cells caused severe DNA damage, cell cycle arrest, senescence, and cell death. Mx1‐cyclization recombination‐driven deletion of Dnmt1 in adult quiescent hepatocytes did not affect liver homeostasis. Conclusion: These results establish the indispensable role of DNMT1‐mediated epigenetic regulation in postnatal liver growth and regeneration; Dnmt1Δalb mice provide a unique experimental model to study the role of senescence and the contribution of progenitor cells to physiological and regenerative liver growth. (Hepatology 2016;64:582‐598)
Objecive Patients with autoimmune hepatitis (AIH) reportedly have an impaired quality of life (QOL), mainly due to depression, even during remission. In addition, hypozincaemia has been demonstrated ...in patients with chronic liver disease, including AIH, and is known to be related to depression. Corticosteroids are known to cause mental instability. We therefore investigated the longitudinal association between zinc supplementation and changes in the mental status among AIH patients treated with corticosteroids. Materials This study enrolled 26 patients with serological remission of AIH routinely treated at our facility after excluding 15 patients who either discontinued polaprezinc (150 mg/day) within 24 months or interrupted treatment. Two questionnaires, the Chronic Liver Disease Questionnaire (CLDQ) and SF-36, were adopted to evaluate the QOL before and after zinc supplementation. Results Serum zinc levels were significantly elevated after zinc supplementation (p<0.0001). The CLDQ worry subscale significantly improved after zinc supplementation (p=0.017), but none of the SF-36 subscales was affected. Multivariate analyses demonstrated that daily prednisolone dosing was inversely related to both the CLDQ worry domain score (p=0.036) and the SF-36 mental health component (p=0.031). There was a significant negative correlation between the changes in the daily steroid dose and the CLDQ worry domain scores before and after zinc supplementation (p=0.006). No serious adverse events occurred during the observation period. Conclusion Zinc supplementation safely and efficiently improved mental impairment, possibly caused by continuous treatment with corticosteroids, in patients with AIH.
The relative contribution of hepatocyte growth factor (HGF)/MET and epidermal growth factor (EGF)/EGF receptor (EGFR), two key signal transduction systems in the normal and diseased liver, to fate ...decisions of adult hepatic progenitor cells (HPCs) has not been resolved. Here, we developed a robust culture system that permitted expansion and genetic manipulation of cells capable of multilineage differentiation in vitro and in vivo to examine the individual roles of HGF/MET and EGF/EGFR in HPC self-renewal and binary cell fate decision. By employing loss-of-function and rescue experiments in vitro, we showed that both receptors collaborate to increase the self-renewal of HPCs through activation of the extracellular signal-regulated kinase (ERK) pathway. MET was a strong inducer of hepatocyte differentiation by activating AKT and signal transducer and activator of transcription (STAT3). Conversely, EGFR selectively induced NOTCH1 to promote cholangiocyte specification and branching morphogenesis while concomitantly suppressing hepatocyte commitment. Furthermore, unlike the deleterious effects of MET deletion, the liver-specific conditional loss of Egfr facilitated rather than suppressed progenitor-mediated liver regeneration by switching progenitor cell differentiation toward hepatocyte lineage. These data provide new insight into the mechanisms regulating the stemness properties of adult HPCs and reveal a previously unrecognized link between EGFR and NOTCH1 in directing cholangiocyte differentiation.
Scope
This study investigated the combined effect of the angiotensin II (AT‐II) receptor blocker losartan and branched‐chain amino acids (BCAAs) on skeletal muscle atrophy in rats with cirrhosis and ...steatohepatitis.
Method and Results
Fischer 344 rats are fed a choline‐deficient l‐amino acid‐defined (CDAA) diet for 12 weeks and treated with oral losartan (30 mg kg−1 day−1) and/or BCAAs (Aminoleban EN, 2500 mg kg−1 day−1). Treatment with losartan and BCAAs attenuated hepatic inflammation and fibrosis and improved skeletal muscle atrophy and strength in CDAA‐fed rats. Both agents reduced intramuscular myostatin and pro‐inflammatory cytokine levels, resulting in inhibition of the ubiquitin–proteasome system (UPS) through interference with the SMAD and nuclear factor‐kappa B pathways, respectively. Losartan also augmented the BCAA‐mediated increase of skeletal muscle mass by promoting insulin growth factor‐I production and mitochondrial biogenesis. Moreover, losartan decreased the intramuscular expression of transcription factor EB (TFEB), a transcriptional inducer of E3 ubiquitin ligase regulated by AT‐II. In vitro assays illustrated that losartan promoted mitochondrial biogenesis and reduced TFEB expression in AT‐II‐stimulated rat myocytes, thereby potentiating the inhibitory effects of BCAAs on the UPS and caspase‐3 cleavage.
Conclusion
These results indicate that this regimen could serve as a novel treatment for patients with sarcopenia and liver cirrhosis.
This study demonstrates that angiotensin‐II receptor blockers efficiently augment the inhibitory effects of branched‐chain amino acids on skeletal muscle atrophy in a rat liver cirrhosis model, which is based on the prevention of liver inflammation and fibrosis resulting in the myokine signaling inhibition and insulin growth factor‐1 restoration as well as promoted mitochondrial biogenesis in myotubes.
Aim
Acute‐on‐chronic liver failure (ACLF) is associated with a high risk of short‐term mortality after progression to multiple organ failure. A disintegrin‐like and metalloproteinase with ...thrombospondin type‐1 motifs 13 (ADAMTS13) is a metalloproteinase that specifically cleaves multimeric von Willebrand factor (VWF). An imbalance between ADAMTS13 enzyme and VWF substrate is associated with liver cirrhosis progression that induces ACLF. This study examined the relationship between ADAMTS13 and VWF and ACLF development to determine whether ADAMTS13 and VWF are useful predictive biomarkers for ACLF development and prognosis of patients with liver cirrhosis.
Methods
The study enrolled 67 patients with Child–Pugh class A and B liver cirrhosis. ADAMTS13 activity (ADAMTS13:AC) and VWF antigen (VWF:Ag) were measured using enzyme‐linked immunosorbent assays. The ratio of VWF:Ag to ADAMTS13:AC (VWF:Ag/ADAMTS13:AC) was used to divide patients into two groups according to the classification and regression tree based on Gray model survival analysis.
Results
Compared with patients with Child–Pugh class A liver cirrhosis, class B patients had a higher VWF:Ag/ADAMTS13:AC and a higher risk of ACLF development. Cumulative incidence of ACLF was significantly higher in patients with high (>7.9) versus low (≤7.9) VWF:Ag/ADAMTS13:AC (hazard ratio HR, 6.50; 95% CI, 2.31–18.29; p < 0.001). Cumulative survival was significantly lower in cirrhotic patients with high versus low VWF:Ag/ADAMTS13:AC (HR 5.11; 95% CI, 1.85–14.14; p = 0.002).
Conclusions
For patients with liver cirrhosis, VWF:Ag/ADAMTS13:AC is associated with functional liver reserve and predicts the development of ACLF and the prognosis.
ABSTRACT
To investigate the role of enhancer of zeste homolog (EZH) 1 and EZH2 in liver homeostasis, mice were generated that carried Ezh1‐/‐ and EZH2fl/fl alleles and an Alb‐Cre transgene. Only the ...combined loss of EZH1 and EZH2 in mouse hepatocytes caused a depletion of global trimethylation on Lys 27 of histone H3 (H3K27me3) marks and the specific loss of over ~1900 genes at 3 mo of age. Ezh1‐/‐,Ezh2fl/flAlb‐Cre mice exhibited progressive liver abnormalities manifested by the development of regenerative nodules and concomitant periportal fibrosis, inflammatory infiltration, and activation of A6‐positive hepatic progenitor cells at 8 mo of age. In response to chronic treatment with carbon tetrachloride, all experimental mice, but none of the controls (n = 27 each), showed increased hepatic degeneration associated with liver dysfunction and reduced ability to proliferate. After two‐thirds partial hepatectomy, mutant mice (n = 5) displayed increased liver injury and a blunted regenerative response. Genomewide analyses at 3 mo of age identified 51 genes that had lost H3K27me3 marks, and their expression was significantly increased. These genes were involved in regulation of cell survival, fibrosis, and proliferation. H3K27me3 levels and liver physiology were unaffected in mice lacking either EZH1 globally or EZH2 specifically in hepatocytes. This work demonstrates a critical redundancy of EZH1 and EZH2 in maintaining hepatic homeostasis and regeneration.—Bae, W. K., Kang, K., Yu, J. H., Yoo, K. H., Factor, V. M., Kaji, K., Matter, M., Thorgeirsson, S., Hennighausen, L. The methyltransferases enhancer of zeste homolog (EZH) 1 and EZH2 control hepatocyte homeostasis and regeneration. FASEB J. 29, 1653‐1662 (2015). www.fasebj.org
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