Specific fate decisions in adult hepatic progenitor cells driven by MET and EGFR signaling.

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.

Gut dysbiosis associated with clinical prognosis of patients with primary biliary cholangitis.

AIM: Although some relationships between gut microbiota and liver diseases have been reported, it remains uncertain whether changes in gut microbiota owing to differences in race, food and living environment have similar effects. Response to ursodeoxycholic acid (UDCA) may predict the long-term prognosis of patients with primary biliary cholangitis (PBC); however, little is known about the significance of the gut microbiome in patients with PBC. We elucidated the relationships among clinical profiles, biochemical response to UDCA and gut microbiome composition in patients with PBC. METHODS: Fecal samples from 76 patients with PBC treated at our hospital were collected; patients whose UDCA intake period was <1 year were excluded. The microbiome structures of patients were determined using 16S ribosomal RNA gene sequencing and were statistically compared with those of healthy subjects. The structures of patients in the UDCA responder (n = 43) and non-responder (n = 30) groups were compared according to the Nara criteria (reduction rate of gamma-glutamyl transpeptidase, ≥69%, after 1 year). RESULTS: Compared with healthy subjects, bacterial diversity was lower in patients with PBC, with a decreased abundance of the order Clostridiales and increased abundance of Lactobacillales. The UDCA non-responder group had a significantly lower population of the genus Faecalibacterium, known as butyrate-producing beneficial bacteria (P < 0.05), although no significant differences in gender, body mass index, medicated drugs or other serological data were indicated between these two groups. CONCLUSIONS: Gut dysbiosis with loss of beneficial Clostridiales commensals was observed in patients with PBC. Decrease in Faecalibacterium abundance might predict the long-term prognosis of patients with PBC.

Combined Treatment with Sodium-Glucose Cotransporter-2 Inhibitor (Canagliflozin) and Dipeptidyl Peptidase-4 Inhibitor (Teneligliptin) Alleviates NASH Progression in A Non-Diabetic Rat Model of Steatohepatitis.

Hepatocellular carcinoma (HCC) is the strongest independent predictor of mortality in non-alcoholic steatohepatitis (NASH)-related cirrhosis. The effects and mechanisms of combination of sodium-dependent glucose cotransporter inhibitor and canagliflozin (CA) and dipeptidyl peptidase-4 inhibitor and teneligliptin (TE) on non-diabetic NASH progression were examined. CA and TE suppressed choline-deficient, L-amino acid-defined diet-induced hepatic fibrogenesis and carcinogenesis. CA alone or with TE significantly decreased proinflammatory cytokine expression. CA and TE significantly attenuated hepatic lipid peroxidation. In vitro studies showed that TE alone or with CA inhibited cell proliferation and TGF-ß1 and α1 (I)-procollagen mRNA expression in Ac-HSCs. CA+TE inhibited liver fibrogenesis by attenuating hepatic lipid peroxidation and inflammation and by inhibiting Ac-HSC proliferation with concomitant attenuation of hepatic lipid peroxidation. Moreover, CA+TE suppressed in vivo angiogenesis and oxidative DNA damage. CA or CA+TE inhibited HCC cells and human umbilical vein endothelial cell (HUVEC) proliferation. CA+TE suppressed vascular endothelial growth factor expression and promoted increased E-cadherin expression in HUVECs. CA+TE potentially exerts synergistic effects on hepatocarcinogenesis prevention by suppressing HCC cell proliferation and angiogenesis and concomitantly reducing oxidative stress and by inhibiting angiogenesis with attenuation of oxidative stress. CA+TE showed chemopreventive effects on NASH progression compared with single agent in non-diabetic rat model of NASH, concurrent with Ac-HSC and HCC cell proliferation, angiogenesis oxidative stress, and inflammation. Both agents are widely, safely used in clinical practice; combined treatment may represent a potential strategy against NASH.

VWF/ADAMTS13 ratio as a potential biomarker for early detection of hepatocellular carcinoma.

BACKGROUND: To investigate the von Willebrand factor to ADAMTS13 ratio as a potential biomarker for early detection of hepatocellular carcinoma (HCC) in cirrhosis. METHODS: Serum levels of alpha-fetoprotein, des-γ-carboxy prothrombin, Lens culinaris agglutinin-reactive fraction of alpha-fetoprotein (alpha-fetoprotein-L3%), vascular endothelial growth factor, and vascular endothelial growth factor receptor-2, as well as the plasma levels of von Willebrand factor antigen (von Willebrand factor: Ag) and ADAMTS13 activity (ADAMTS13:AC), were evaluated in 41 cirrhotic patients with HCC undergoing radiofrequency ablation and in 20 cirrhotic patients without HCC. The diagnostic accuracy of each biomarker was evaluated using the receiver operating characteristic curve analysis. RESULTS: The von Willebrand factor: Ag and von Willebrand factor: Ag/ADAMTS13:AC ratios were significantly higher in cirrhotic patients with HCC than in those without HCC (p < 0.05 and p < 0.01, respectively), whereas ADAMTS13:AC was significantly lower in those with HCC than those without HCC (p < 0.05). However, no relationship was observed between the von Willebrand factor: Ag/ADAMTS13:AC ratio and serum tumor markers such as alpha-fetoprotein, des-γ-carboxy prothrombin, and alpha-fetoprotein-L3%. Multivariate regression analysis identified von Willebrand factor: Ag/ADAMTS13:AC ratio and alpha-fetoprotein-L3% as significant factors of HCC development. Receiver operating characteristic analysis showed that the von Willebrand factor: Ag/ADAMTS13:AC ratio and alpha-fetoprotein-L3% had a better performance than alpha-fetoprotein, des-γ-carboxy prothrombin, alpha-fetoprotein-L3%, vascular endothelial growth factor, and vascular endothelial growth factor receptor-2, von Willebrand factor: Ag, and ADAMTS13:AC. The von Willebrand factor: Ag/ADAMTS13:AC ratio was exclusively correlated with tumor volume and stage as well as serum vascular endothelial growth factor levels. CONCLUSIONS: The von Willebrand factor: Ag/ADAMTS13:AC ratio can potentially serve as a novel biomarker for early diagnosis of HCC in cirrhotic patients.

Blocking development of liver fibrosis augments hepatic progenitor cell-derived liver regeneration in a mouse chronic liver injury model.

AIM: The roles of hepatic progenitor cells (HPCs) in regeneration of a diseased liver are unclear. Hepatic stellate cells (HSCs) contribute to liver fibrosis but are also a component of the HPC niche. Hepatic progenitor cells expand along with HSC activation and liver fibrosis. However, little is known about the interplay of liver fibrosis and HPC-mediated liver regeneration. This study aimed to investigate HSCs and HPCs in liver regeneration. METHODS: Liver injury in mice was induced with 3,5-diethoxycarbonyl-1,4-dihydrocollidine, and HPC expansion and fibrosis were assessed. An angiotensin II type 1 receptor blocker (ARB) was administered to assess its effect on fibrosis and regeneration. RESULTS: Treatment with ARB attenuated fibrosis and expansion of α-smooth muscle actin-positive activated HSCs as indicated by increased liver weight and Ki-67-positive hepatocytes. Immunohistochemical staining suggested that HPC differentiation was shifted toward hepatocytes (HCs) when ARB treatment decreased HPC encapsulation by HSCs and extracellular matrix. Conditioned medium produced by culturing the human HSC LX-2 line strongly augmented differentiation to biliary epithelial cells (BECs) but inhibited that to HCs. Activated HSCs expressed Jagged1, a NOTCH ligand, which plays a central role in differentiation of HPCs toward BECs. CONCLUSIONS: Hepatic stellate cells, the HPC niche cells, control differentiation of HPCs, directing them toward BECs rather than HCs in a diseased liver model. Antifibrosis treatment with an ARB preferentially redirects HPC differentiation toward HCs by blocking the NOTCH pathway in the HPC niche, resulting in more efficient HPC-mediated liver regeneration.

Combining probiotics and an angiotensin-II type 1 receptor blocker has beneficial effects on hepatic fibrogenesis in a rat model of non-alcoholic steatohepatitis.

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-ß 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.

Proton pump inhibitor therapy does not increase serum endotoxin activity in patients with cirrhosis.

AIM: Proton pump inhibitors (PPIs) are frequently prescribed in patients with cirrhosis, but this therapy entails potential complications. We aimed to investigate the influence of PPI use on intestinal permeability in patients with cirrhosis. METHODS: We recruited 228 patients with cirrhosis and divided them into four groups. Group (Gp)1 comprised patients receiving a PPI with concurrent neomycin (NEO) (PPI-NEO group, n = 14 [6.1%]), Gp2 and Gp3 comprised those receiving either PPI or NEO (PPI group, n = 91 [39.9%]; and NEO group, n = 11 [4.4%]), and Gp4 comprised those receiving neither of these medications (control group; n = 112 [49.1%]). We assessed the intestinal permeability by measuring endotoxin activity (EA) using a luminol chemiluminescence method. RESULTS: Endotoxin activity levels were significantly higher in patients with Child B cirrhosis than in those with Child A cirrhosis, but we found no significant differences in EA levels between patients with Child C cirrhosis and those with either Child A or B cirrhosis. We observed no significant differences in EA levels among groups 1-4. Patients without antibiotic exposure (n = 203), comprising 91 patients on PPI therapy (Gp2) and 112 no-PPI-therapy controls (Gp4), were subdivided according to Child-Pugh (CP) classification. We found no significant differences in EA levels between Gp2 and Gp4 in either CP class. CONCLUSION: Our results suggest that PPI usage does not have a significant impact on serum levels of gut-derived endotoxins, which are already elevated because of the increased intestinal permeability in patients with cirrhosis.

Identification of clinical risk factors for histological progression of primary biliary cholangitis.

AIM: To identify laboratory predictors of histological progression (HP) of primary biliary cholangitis (PBC). METHODS: Sequential biopsies were carried out on 35 (11.4%) of 308 patients with PBC treated with ursodeoxycholic acid (UDCA). Patients were divided into high γ-glutamyl transpeptidase (GGT) (n = 18) and low GGT (n = 17) groups, based on the median value of GGT at baseline. Patients were then categorized as showing HP (progressive group, PG) or lacking HP (non-progressive group, NPG) according to the Scheuer and Nakanuma classifications, with the latter grading liver fibrosis (fibrosis score) and bile duct loss (BDL score). RESULTS: According to the Scheuer definition, 12 patients had HP and 23 did not. According to the Nakanuma definition, 8 and 27 patients were in the PG and NPG groups, respectively. The fibrosis and BDL scores progressed in 13 and 8 patients, respectively, whereas 22 and 25 patients did not show HP, respectively. Fisher's exact probability test analysis revealed that the rate of HP using the Nakanuma fibrosis score was significantly higher in the high GGT group compared to the low GGT group (P < 0.05). However, no significant correlation was found between the HP of PBC and the biochemical response to UDCA therapy. Both univariate and multivariate logistic regression analyses indicated that the serum GGT level at baseline is an independent risk factor for an increased Nakanuma fibrosis score. CONCLUSIONS: The level of serum GGT at baseline is significantly associated with liver fibrosis progression in PBC, and therefore could help to predict the HP of PBC.

Combined effect of a farnesoid X receptor agonist and dipeptidyl peptidase-4 inhibitor on hepatic fibrosis.

AIM: Non-alcoholic steatohepatitis (NASH) has a broad clinicopathological spectrum (inflammation to severe fibrosis). The farnesoid X receptor agonist obeticholic acid (OCA) ameliorates the histological features of NASH; satisfactory antifibrotic effects have not yet been reported. Here, we investigated the combined effects of OCA + a dipeptidyl peptidase-4 inhibitor (sitagliptin) on hepatic fibrogenesis in a rat model of NASH. METHODS: Fifty Fischer 344 rats were fed a choline-deficient L-amino-acid-defined (CDAA) diet for 12 weeks. The in vitro and in vivo effects of OCA + sitagliptin were assessed along with hepatic fibrogenesis, lipopolysaccharide-Toll-like receptor 4 (TLR4) regulatory cascade and intestinal barrier function. Direct inhibitory effects of OCA + sitagliptin on activated hepatic stellate cells (Ac-HSCs) were assessed in vitro. RESULTS: Treatment with OCA + sitagliptin potentially inhibited hepatic fibrogenesis along with Ac-HSC proliferation and hepatic transforming growth factor (TGF)-ß1, α1(I)-procollagen, and tissue inhibitor of metalloproteinase-1 (TIMP-1) mRNA expression and hydroxyproline levels. Obeticholic acid inhibited hepatic TLR4 expression and increased hepatic matrix metalloproteinase-2 expression. Obeticholic acid decreased intestinal permeability by ameliorating CDAA diet-induced zonula occludens-1 disruption, whereas sitagliptin directly inhibited Ac-HSC proliferation. The in vitro suppressive effects of OCA + sitagliptin on TGF-ß1 and α1(I)-procollagen mRNA expression and p38 phosphorylation in Ac-HSCs were almost consistent. Sitagliptin directly inhibited the regulation of Ac-HSC. CONCLUSIONS: Treatment with OCA + sitagliptin synergistically affected hepatic fibrogenesis by counteracting endotoxemia induced by intestinal barrier dysfunction and suppressing Ac-HSC proliferation. Thus, OCA + sitagliptin could be a promising therapeutic strategy for NASH.

Exogenous Administration of Low-Dose Lipopolysaccharide Potentiates Liver Fibrosis in a Choline-Deficient l-Amino-Acid-Defined Diet-Induced Murine Steatohepatitis Model.

Various rodent models have been proposed for basic research; however, the pathogenesis of human nonalcoholic steatohepatitis (NASH) is difficult to closely mimic. Lipopolysaccharide (LPS) has been reported to play a pivotal role in fibrosis development during NASH progression via activation of toll-like receptor 4 (TLR4) signaling. This study aimed to clarify the impact of low-dose LPS challenge on NASH pathological progression and to establish a novel murine NASH model. C57BL/6J mice were fed a choline-deficient l-amino-acid-defined (CDAA) diet to induce NASH, and low-dose LPS (0.5 mg/kg) was intraperitoneally injected thrice a week. CDAA-fed mice showed hepatic CD14 overexpression, and low-dose LPS challenge enhanced TLR4/NF-κB signaling activation in the liver of CDAA-fed mice. LPS challenge potentiated CDAA-diet-mediated insulin resistance, hepatic steatosis with upregulated lipogenic genes, and F4/80-positive macrophage infiltration with increased proinflammatory cytokines. It is noteworthy that LPS administration extensively boosted pericellular fibrosis with the activation of hepatic stellate cells in CDAA-fed mice. Exogenous LPS administration exacerbated pericellular fibrosis in CDAA-mediated steatohepatitis in mice. These findings suggest a key role for LPS/TLR4 signaling in NASH progression, and the authors therefore propose this as a suitable model to mimic human NASH.

Sodium glucose cotransporter 2 inhibitor canagliflozin attenuates liver cancer cell growth and angiogenic activity by inhibiting glucose uptake.

Sodium-glucose cotransporter 2 inhibitors (SGLT2-Is) comprise a new class of antidiabetic agents that inhibit glucose reabsorption in the renal proximal tubules. Although a recent report demonstrated the potential ability of SGLT2-Is to attenuate cancer growth of SGLT2-expressing cancer cells, little is known about the effects of SGLT2-Is on hepatocellular carcinoma (HCC). Here, we investigate the anti-cancer properties of a SGLT2-I, canagliflozin, against human liver cancer cells. SGTL2 mRNA and protein expression were detected in Huh7 and HepG2 cells, although not in HLE as well as primary human hepatocytes and hepatic stellate cells. Canagliflozin exerted antiproliferative effects on SGLT2-expressing Huh7 and HepG2 cells in a dose-dependent manner by inhibiting glycolytic metabolism including glucose uptake, lactate and intracellular ATP production. This agent also induced G2/M arrest and apoptosis with inhibited phosphorylation of ERK, p38 and AKT and cleavage of caspase3. Xenograft tumor growth assay showed that oral administration of canagliflozin (10 mg/kg/day) significantly reduced subcutaneous tumor burdens in a glycemic status-independent manner, and attenuated intratumor vascularization in Huh7- and HepG2-derived xenograft tumors in BALB/c nude mice. In vitro, canagliflozin suppressed the increased human umbilical vein endothelial cell (HUVEC) proliferation and tubular formation which were observed in Huh7 or HepG2 co-cultures. By contrast, canagliflozin had no effect on tumor growth and intratumor angiogenesis in SGLT2-null HLE-derived xenograft models. These results indicate that SGLT2-I therapy is a potential new strategy for the treatment of HCC.

Acyclic retinoid and angiotensin-II receptor blocker exert a combined protective effect against diethylnitrosamine-induced hepatocarcinogenesis in diabetic OLETF rats.

BACKGROUND: Insulin resistance (IR) is closely associated with the progression of hepatocellular carcinoma (HCC). Acyclic retinoid (ACR) targets retinoid X receptor α and reportedly prevents HCC recurrence in clinical practice. Angiotensin-II receptor blocker (ARB) can also inhibit experimental hepatocarcinogenesis and HCC development. These are reported to suppress IR-based hepatocarcinogenesis; however, limited data are available regarding the combined effects of both these agents. This study aimed to investigate the combined chemopreventive effect of ACR and ARB on liver tumorigenesis on rats with congenital diabetes. METHODS: Male diabetic Otsuka Long-Evans Tokushima Fatty (OLETF) and non-diabetic Long-Evans Tokushima Otsuka (LETO) rats underwent 70% partial hepatectomy following a single intraperitoneal injection of diethylnitrosamine to induce hepatocarcinogenesis and the administration of ACR (peretinoin, 40 mg/kg/day), ARB (losartan, 30 mg/kg/day), and a combination of ACR and ARB. Six weeks thereafter, we assessed the size and number of the pre-neoplastic lesions (PNL) as well as the altered angiogenesis, oxidative stress, and chronic inflammation in the liver. Moreover, we assessed the effects exerted by ACR and ARB on in vitro cell growth in human HCC cell lines and human umbilical vascular endothelial cells (HUVECs). RESULTS: OLETF rats showed increase in the size and number of PNLs compared to LETO rats. ACR suppressed the augmentation in size and number of PNLs in the OLETF rats with suppression of cell growth, intrahepatic angiogenesis, lipid peroxidation, oxidative DNA damage, and proinflammatory cytokine production. Combining ACR with ARB enhanced the tumor-suppressive effect and ameliorated intrahepatic angiogenesis, lipid peroxidation, and proinflammatory status; however, cell growth and oxidative DNA damage remained unchanged. IR-mimetic condition accelerated in vitro proliferative activity in human HCC cells, while ACR inhibited this proliferation with G0/G1 arrest and apoptosis. Furthermore, ACR and ARB significantly attenuated the HUVECs proliferation and tubular formation under the IR-mimetic condition, and a combination of both agents demonstrated greater inhibitory effects on HUVEC growth than each single treatment. CONCLUSIONS: ACR and ARB exert a combined inhibitory effect against IR-based hepatocarcinogenesis by the inhibition of cell growth, intrahepatic angiogenesis, and oxidative stress. Thus, this combination therapy appears to hold potential as a chemopreventive treatment therapy against HCC.

Treatment of long-segment Barrett's adenocarcinoma by complete circular endoscopic submucosal dissection: a case report.

BACKGROUND: We present the first description of en bloc endoscopic submucosal dissection (ESD) for total circumferential Barrett's adenocarcinoma, predominantly of the long-segment Barrett's esophagus (LSBE), with a 2-year follow-up and management strategies for esophageal stricture prevention. CASE PRESENTATION: A 59-year-old man was diagnosed with LSBE and Barrett's adenocarcinoma by esophagogastroduodenoscopy (EGD). A 55-mm-long circumferential tumor was completely resected by ESD. Histopathology revealed a well-differentiated adenocarcinoma within the LSBE superficial muscularis mucosa. For post-ESD stricture prevention, the patient underwent an endoscopic triamcinolone injection administration, oral prednisolone administration, and preemptive endoscopic balloon dilatation. Two years later, there is no evidence of esophageal stricture or recurrence. CONCLUSIONS: ESD appears to be a safe, effective option for total circumferential Barrett's adenocarcinoma in LSBE.

Platelet hyperaggregability is associated with decreased ADAMTS13 activity and enhanced endotoxemia in patients with acute cholangitis.

AIM: Insufficient ADAMTS13 activity (ADAMTS13:AC) leads to increased levels of unusually large von Willebrand factor (VWF) multimers and causes microcirculatory disturbance and multiple organ failure (MOF). Endotoxin (Et) triggers the activation of coagulation and cytokine cascades, leading to MOF in severe inflammatory response syndrome. Here, we investigated the potential role of endotoxemia-related ADAMTS13 in acute cholangitis. METHODS: Twenty-four patients with acute cholangitis, including 7 with severe acute cholangitis, were recruited in this study. The levels of ADAMTS13:AC, VWF antigen (VWF:Ag), interleukin (IL)-6, IL-8, and tumor necrosis factor (TNF)-α in each patient were determined by enzyme-linked immunosorbent assay, whereas Et levels were determined by Et activity assay (EAA) analysis. RESULTS: The ADAMTS13:AC and VWF:Ag levels were significantly lower and higher, respectively, in patients with acute cholangitis than in controls. The EAA levels were higher in patients with acute cholangitis than in controls, and were inversely correlated with that of ADAMTS13:AC. Patients with severe acute cholangitis had significantly lower ADAMTS13:AC and higher VWF:Ag levels than those with mild to moderate cholangitis. Notably, ADMTS13:AC was directly correlated with platelet counts and inversely correlated with IL-6 levels, and the VWF:Ag/ADAMTS13:AC ratio was directly correlated with IL-8 and TNF-α levels. CONCLUSIONS: Imbalance of ADAMTS13:AC and VWF:Ag levels might be associated with severe acute cholangitis, reflecting platelet hyperaggregability. Severe acute cholangitis has severe pathophysiological features and is complicated by endotoxemia and MOF. Notably, this is the first report indicating an association between the levels of ADAMTS13:AC and VWF:Ag and those of EAA and cytokines in acute cholangitis.

[Short-term sustained virological response in a patient with liver cirrhosis, hepatitis C, and declining hepatic spare ability during treatment with direct-acting antivirals].

A 75-year-old female patient with liver cirrhosis and hepatitis C was treated with direct-acting antivirals (DAA) (Sofosbuvir+Ledipasvir). The hepatitis C virus (HCV) -RNA level decreased to negative 4 weeks after the start of the treatment. Six weeks later, she developed ascites and showed declining hepatic spare ability. Accordingly, DAA treatment was stopped. She was started on furosemide 20mg/day and spironolactone 50mg/day. After 7 days, she started taking tolvaptan 7.5mg/day because furosemide and spironolactone proved to be ineffective. This new regimen resolved the ascites. The HCV-RNA level remained negative, although DAA was not restarted. Finally, she achieved a sustained virological response (SVR). The hepatic spare ability at the time of SVR recovered than that at the time of DAA treatment.

Predictive parameter of tolvaptan effectiveness in cirrhotic ascites.

AIMS: The efficacy of the vasopressin V2 receptor antagonist tolvaptan for difficult-to-treat cirrhotic ascites has recently been reported. However, its effect is variable among patients. This study aimed to clarify the predictive factors for obtaining a good response to tolvaptan in patients with difficult-to-treat ascites. METHODS: Data were collected from 50 patients with liver cirrhosis having ascites (hepatitis B, n = 1; hepatitis C, n = 22; alcoholism, n = 11; and others, n = 16) after treatment with tolvaptan (3.75-7.5 mg/day) in addition to conventional diuretics. A follow-up assessment was carried out after 7-day tolvaptan treatment for all patients. RESULTS: After an uneventful 7-day tolvaptan treatment, 18 patients (36.0%) lost more than 2 kg of their body weight (responders). Twenty-six patients (52.0%) showed an increase in urine volume (>300 mL) on day 2. Tolvaptan was also effective for patients with pleural effusion, portal vein thrombosis, and hepatocellular carcinoma. Basal blood urea nitrogen (BUN) levels, plasma renin activity, and aldosterone levels were significantly higher in the poor responders (<2 kg weight loss), who were considered to be in the relative vascular underfilling state, than in the responders. Basal BUN was extracted as a predictive factor of responsiveness by multivariate logistic regression analysis. CONCLUSIONS: Tolvaptan is useful and safe for the treatment of cirrhotic ascites. This report showed that BUN will predict the response of tolvaptan even when measured before tolvaptan treatment.

Combined treatment with dipeptidyl peptidase-4 inhibitor (sitagliptin) and angiotensin-II type 1 receptor blocker (losartan) suppresses progression in a non-diabetic rat model of steatohepatitis.

AIM: Dipeptidyl peptidase-4 (DPP4) inhibitors (DPP4-I) are oral glucose-lowering drugs for type 2 diabetes mellitus. Previously, we reported that DPP4-I (sitagliptin) exerted suppressive effects on experimental liver fibrosis in rats. Blockade of the renin-angiotensin system by angiotensin-II type 1 receptor blocker (losartan), commonly used in the management of hypertension, has been shown to significantly alleviate hepatic fibrogenesis and carcinogenesis. We aimed to elucidate the effects and possible mechanisms of a sitagliptin + losartan combination on the progression of non-diabetic non-alcoholic steatohepatitis (NASH) in a rat model. METHODS: To induce NASH, Fischer 344 rats were fed a choline-deficient L-amino acid-defined diet for 12 weeks. We elucidated the chemopreventive effects of sitagliptin + losartan, especially in conjunction with hepatic stellate cell (HSC) activation, angiogenesis, and oxidative stress, all known to play important roles in the progression of NASH. RESULTS: Sitagliptin + losartan suppressed choline-deficient L-amino acid-defined diet-induced hepatic fibrogenesis and carcinogenesis. The combination treatment exerted a greater inhibitory effect than monotherapy. These inhibitory effects occurred almost concurrently with the suppression of HSC activation, neovascularization, and oxidative stress. In vitro studies showed that sitagliptin + losartan inhibited angiotensin II-induced proliferation and expression of transforming growth factor-ß1 and α1 (I)-procollagen mRNA of activated HSC and in vitro angiogenesis, in parallel with the suppression observed in in vivo studies. CONCLUSIONS: The widely and safely used sitagliptin + losartan combination treatment in clinical practice could be an effective strategy against NASH.

Relationship between hepatic progenitor cell-mediated liver regeneration and non-parenchymal cells.

Hepatic progenitor cells (HPCs) are thought to reside in the canals of Hering and can be activated and contribute to liver regeneration in response to liver injury by proliferating and differentiating towards both hepatocytes and biliary epithelial cells. In this setting, several cytokines, chemokines, and growth factors related to liver inflammation and other liver cells comprising the HPC niche, namely hepatic stellate cells (HSCs), play crucial roles in HPC activation and differentiation. In response to several types of liver injury, tumor necrosis factor-like weak inducer of apoptosis (TWEAK) is secreted by several inflammatory cells, including monocytes, T lymphocytes, and macrophages, and acts as an initiator of the HPC niche and HSC activation. Following TWEAK-induced activation of the HPC niche, fibroblast growth factor 7 and hepatocyte growth factor released from activated HSC play central roles in maintaining HPC proliferation. In contrast, HGF-MET and Wnt3a-ß-catenin signals are the predominant mediators of the hepatocyte differentiation of HPC, whereas epidermal growth factor receptor-NOTCH signaling controls HPC differentiation towards biliary epithelial cells. These signals are maintained exclusively by activated HSC and inflammatory cells surrounding HPC. Together, HSC and inflammatory cells surrounding HPC are responsible for the precise control of HPC proliferation and differentiation fate. In this review, we discuss recent progress in understanding of interactions between HPC and other liver cells in HPC-mediated liver regeneration in the setting of liver inflammation.

Curcumin effectively inhibits oncogenic NF-κB signaling and restrains stemness features in liver cancer.

BACKGROUND & AIMS: The cancer stem cells (CSCs) have important therapeutic implications for multi-resistant cancers including hepatocellular carcinoma (HCC). Among the key pathways frequently activated in liver CSCs is NF-κB signaling. METHODS: We evaluated the CSCs-depleting potential of NF-κB inhibition in liver cancer achieved by the IKK inhibitor curcumin, RNAi and specific peptide SN50. The effects on CSCs were assessed by analysis of side population (SP), sphere formation and tumorigenicity. Molecular changes were determined by RT-qPCR, global gene expression microarray, EMSA, and Western blotting. RESULTS: HCC cell lines exposed to curcumin exhibited differential responses to curcumin and were classified as sensitive and resistant. In sensitive lines, curcumin-mediated induction of cell death was directly related to the extent of NF-κB inhibition. The treatment also led to a selective CSC-depletion as evidenced by a reduced SP size, decreased sphere formation, down-regulation of CSC markers and suppressed tumorigenicity. Similarly, NF-κB inhibition by SN50 and siRNA against p65 suppressed tumor cell growth. In contrast, curcumin-resistant cells displayed a paradoxical increase in proliferation and expression of CSC markers. Mechanistically, an important component of the CSC-depleting activity of curcumin could be attributed to a NF-κB-mediated HDAC inhibition. Co-administration of the class I/II HDAC inhibitor trichostatine sensitized resistant cells to curcumin. Further, integration of a predictive signature of curcumin sensitivity with human HCC database indicated that HCCs with poor prognosis and progenitor features are most likely to benefit from NF-κB inhibition. CONCLUSIONS: These results demonstrate that blocking NF-κB can specifically target CSC populations and suggest a potential for combined inhibition of NF-κB and HDAC signaling for treatment of liver cancer patients with poor prognosis.

Modeling pathogenesis of primary liver cancer in lineage-specific mouse cell types.

BACKGROUND & AIMS: Human primary liver cancer is classified into biologically distinct subgroups based on cellular origin. Liver cancer stem cells (CSCs) have been recently described. We investigated the ability of distinct lineages of hepatic cells to become liver CSCs and the phenotypic and genetic heterogeneity of primary liver cancer. METHODS: We transduced mouse primary hepatic progenitor cells, lineage-committed hepatoblasts, and differentiated adult hepatocytes with transgenes encoding oncogenic H-Ras and SV40LT. The CSC properties of transduced cells and their ability to form tumors were tested by standard in vitro and in vivo assays and transcriptome profiling. RESULTS: Irrespective of origin, all transduced cells acquired markers of CSC/progenitor cells, side populations, and self-renewal capacity in vitro. They also formed a broad spectrum of liver tumors, ranging from cholangiocarcinoma to hepatocellular carcinoma, which resembled human liver tumors, based on genomic and histologic analyses. The tumor cells coexpressed hepatocyte (hepatocyte nuclear factor 4α), progenitor/biliary (keratin 19, epithelial cell adhesion molecule, A6), and mesenchymal (vimentin) markers and showed dysregulation of genes that control the epithelial-mesenchymal transition. Gene expression analyses could distinguish tumors of different cellular origin, indicating the contribution of lineage stage-dependent genetic changes to malignant transformation. Activation of c-Myc and its target genes was required to reprogram adult hepatocytes into CSCs and for tumors to develop. Stable knockdown of c-Myc in transformed adult hepatocytes reduced their CSC properties in vitro and suppressed growth of tumors in immunodeficient mice. CONCLUSIONS: Any cell type in the mouse hepatic lineage can undergo oncogenic reprogramming into a CSC by activating different cell type-specific pathways. Identification of common and cell of origin-specific phenotypic and genetic changes could provide new therapeutic targets for liver cancer.