Yue-Sun Cheung

SIRT2 overexpression in hepatocellular carcinoma mediates epithelial to mesenchymal transition by protein kinase B/glycogen synthase kinase-3β/β-catenin signaling†‡

Sirtuin 1 (SIRT1) has been implicated in telomere maintenance and the growth of hepatocellular carcinoma (HCC). Nevertheless, the role of other sirtuins in the pathogenesis of HCC remains elusive. We found that sirtuin 2 (SIRT2), another member of the sirtuin family, also contributes to cell motility and invasiveness of HCC. SIRT2 is up‐regulated in HCC cell lines and in a subset of human HCC tissues (23/45). Up‐regulations of SIRT2 in primary HCC tumors were significantly correlated with the presence of microscopic vascular invasion (P = 0.001), a more advanced tumor stage (P = 0.004), and shorter overall survival (P = 0.0499). Functional studies by short hairpin RNA–mediated suppression of SIRT2 expression in HCC cell lines revealed significant inhibition of motility and invasiveness. Depletion of SIRT2 also led to the regression of epithelial‐mesenchymal transition (EMT) phenotypes, whereas the ectopic expression of SIRT2 in the immortalized hepatocyte cell line L02 promoted cell motility and invasiveness. Mechanistic studies revealed that SIRT2 regulates the deacetylation and activation of protein kinase B, which subsequently impinges on the glycogen synthase kinase‐3β/β‐catenin signaling pathway to regulate EMT. Conclusions: Our findings have uncovered a novel role for SIRT2 in HCC metastasis, and provide a rationale to explore the use of sirtuin inhibitors in HCC therapy. (HEPATOLOGY 2013;)

Liver stiffness measurement by transient elastography as a predictor on posthepatectomy outcomes.

Background:Liver fibrosis and cirrhosis are well-known risk factors for morbidity after hepatectomy. Liver stiffness measurement (LSM) using transient elastography is a new method for detection of hepatic fibrosis and cirrhosis with high accuracy. Whether LSM can predict posthepatectomy outcomes has not been studied. Methods:This was a prospective cohort study in which consecutive patients underwent hepatectomy for various indications from February 2010 to July 2011. All patients received detailed preoperative assessments including LSM and indocyanine green (ICG) clearance test. The primary outcome was major postoperative complication. Results:One hundred five patients with a mean age of 59 years were included; 75 (71.4%) had chronic viral hepatitis and 76 (72.4%) had hepatocellular carcinoma. Thirty-four patients (32.4%) received major hepatectomy. The median ICG retention rate at 15 minutes was 4.2 (0.1%–32%) and the median LSM was 9.4 (3.3–75 kPa). For posthepatectomy outcomes, only LSM but not ICG showed significant correlation with major postoperative complications on receiver operating characteristic curves, with area under the curve of 0.79 (P < 0.001). Using the calculated cutoff at 12.0 kPa, LSM had sensitivity of 85.7% and specificity of 71.8% in the prediction of major postoperative complications. It was also an independent prognostic factor for major postoperative complications by multivariate analysis. The operative blood loss and transfusion rate were also significantly higher in patients with LSM >12.0 kPa. Conclusions:High LSM (>12.0 kPa) predicted worse posthepatectomy outcomes. Preoperative LSM was better than ICG test in the prediction of major postoperative complications. It was a useful preoperative investigation for risk stratification before hepatectomy.

Antiviral therapy improves post‐hepatectomy survival in patients with hepatitis B virus‐related hepatocellular carcinoma: a prospective‐retrospective study

The effect of antiviral therapy on the post‐hepatectomy long‐term survival in patients with hepatitis B virus (HBV)‐related hepatocellular carcinoma (HCC) remains uncertain.

Feasibility of liver resection without the use of the routine Pringle manoeuver: an analysis of 248 consecutive cases

BACKGROUND New instruments and techniques for hepatectomy have been shown to reduce blood loss during liver resection. The present study aims to evaluate the feasibility and result of our techniques of liver resection without routine inflow occlusion (the Pringle manoeuver). METHODS The cavitron ultrasonic surgical aspirator (CUSA) and saline-linked radio-frequency dissecting sealer (TissueLink) were used together for open hepatectomy, whereas a bipolar vessel sealing device (Ligasure) and TissueLink were used for laparoscopic hepatectomy. Between June 2003 and May 2007, 248 consecutive cases of liver resection were carried out using the above techniques without the routine Pringle manoeuver. The operative and clinical outcome data were prospectively collected and analysed. RESULTS During the study period, a total of 220 cases of open hepatectomy and 28 cases of laparoscopic hepatectomy were performed. The Pringle manoeuver was eventually applied in six patients (2.4%): two for portal vein tumour thrombus extraction and four as a result of heavy bleeding. Median blood loss was 300 ml (20-2700 ml) and the blood transfusion rate was 7.7%. In most of the cases, the liver function tests showed improvement on post-operative day 1 or 2, and the median post-operative hospital stay was 7 days. There were two post-operative deaths (0.8%). Complications occurred in 63 patients (25.4%) and most complications were minor. CONCLUSIONS Refined techniques and instruments for liver resection allow hepatectomy to be done safely without using the routine Pringle manoeuver. Most patients had a quick recovery of liver function and were discharged early.

Elevated Perioperative Transaminase Level Predicts Intrahepatic Recurrence in Hepatitis B-related Hepatocellular Carcinoma After Curative Hepatectomy

OBJECTIVE We aimed to evaluate the role of elevated perioperative alanine aminotransferase (ALT) as a surrogate marker of hepatitis activity in determining the risk of recurrence and survival in hepatitis B-related hepatocellular carcinoma (HCC) after curative hepatectomy. METHODS A retrospective review of the hepatectomy database was performed and 142 patients were found who had hepatitis B-related HCC from January 2001 to March 2006. Their ALT levels preoperatively and 1 month, 3 months, and 6 months postoperatively were recorded. The risk factors for recurrence and prognostic factors of survival were analysed. RESULTS An elevated perioperative ALT level (p = 0.021), multiple tumour nodules in the resected specimen (p < 0.001), and a tumour size greater than 5 cm (p = 0.001) were significant independent risk factors for tumour recurrence. The latter two factors were also independent prognostic factors for overall survival and disease-free survival. An elevated ALT level was an independent prognostic factor for disease-free survival (p = 0.025). CONCLUSION An elevated perioperative ALT level, which reflects increased hepatitis activity, is an independent risk factor for intrahepatic recurrence of hepatitis B-related HCC. It is also associated with a poorer disease-free survival rate.

Cell cycle-related kinase mediates viral-host signalling to promote hepatitis B virus-associated hepatocarcinogenesis

Background Androgen receptor (AR) signalling contributes to male predominance in hepatocellular carcinoma (HCC), which is more pronounced in HBV-endemic areas. Cell cycle-related kinase (CCRK) is essential for AR-induced hepatocarcinogenesis but its molecular function in HBV-associated HCC remains obscure. Objective To determine the molecular function of CCRK in HBV-associated HCC. Design Transcriptional regulation was assessed by chromatin immunoprecipitation, promoter mutation and luciferase reporter assays. Hepatocellular proliferation and tumourigenesis were examined by colony formation, soft agar assays and using HBV X protein (HBx) transgenic mice with low-dose exposure to diethylnitrosamine. Protein expressions were examined in clinical samples and correlated with patient survival by log-rank Mantel–Cox test. Results Overexpression of CCRK, but not its kinase-defective mutant, activated β-catenin/T cell factor signalling through phosphorylation of glycogen synthase kinase-3β (GSK-3β) at Ser9, led to upregulation of AR transcriptional activity and, subsequently, expression of HBx. The viral transactivator in turn induced CCRK expression through enhanced AR signalling, thus forming a positive regulatory loop. RNA interference silencing of CCRK, which suppressed the CCRK/GSK-3β/β-catenin/AR regulatory loop, significantly suppressed HBx-induced hepatocellular proliferation (p=0.001) and transformation (p<0.001) and remarkably reduced >80% diethylnitrosamine-mediated hepatocarcinogenesis in HBx transgenic mice. Finally, patients with HBV-associated HCC with concordant overexpression of CCRK, GSK-3β phosphorylation at Ser9, active dephosphorylated β-catenin and AR phosphorylation at Ser81 had poorer overall (HR=31.26, p<0.0001) and disease-free (HR=3.60, p<0.01) survival rates. Conclusions Our findings highlight the critical role of CCRK in a self-reinforcing circuitry that regulates HBV-associated hepatocarcinogenesis. Further characterisation of this intricate viral-host signalling may provide new prognostic biomarkers and therapeutic targets for HCC treatment.

Outcome of surgical treatment for recurrent pyogenic cholangitis: a single-centre study

BACKGROUND Recurrent pyogenic cholangitis (RPC) is still a common disease in East Asia. The present study reviews the operative results for this disease in a single centre. METHODS The records of 85 patients who underwent surgical treatment for RPC from August 1995 to March 2008 were retrospectively reviewed. RESULTS Patients included 35 men and 50 women with a median age of 61 years. Types of surgery included: hepatectomy (65.9%); hepatectomy plus drainage (9.4%); drainage alone (14.1%), and percutaneous choledochoscopy (10.6%). There was no operative mortality. Complications occurred in 40% of patients and half the complications involved wound infections. The overall incidences of residual stone, stone recurrence and biliary sepsis recurrence were 21.2%, 16.5% and 21.2%, respectively, over a median follow-up of 45.4 months. The drainage-alone group and percutaneous choledochoscopy group had higher incidences of residual stone, stone recurrence and biliary sepsis recurrence. In hepatectomy patients, regardless of whether or not a drainage procedure had been performed, rates of residual stone, stone recurrence and biliary sepsis recurrence were 15.6%, 7.8% and 9.4%, respectively, over a median follow-up of 42.7 months. CONCLUSIONS Hepatectomy is safe and yields the best treatment outcome for RPC. It should be considered as the treatment of choice for suitable patients with RPC.

A CCRK-EZH2 epigenetic circuitry drives hepatocarcinogenesis and associates with tumor recurrence and poor survival of patients

BACKGROUND & AIMS Aberrant chromatin modification is a key feature of hepatocellular carcinoma (HCC), which is characterized by strong sexual dimorphism. Both enhancer of zeste homolog 2 (EZH2) and cell cycle-related kinase (CCRK) contribute to hepatocarcinogenesis, yet whether the two oncogenic factors have functional crosstalk is unknown. METHODS Cellular proliferation and tumorigenicity upon transgenic expression and RNA interference were determined by colony formation and soft agar assays, xenograft, orthotopic and diethylnitrosamine-induced HCC models. Gene regulation was assessed by chromatin immunoprecipitation, site-directed mutagenesis, luciferase reporter, co-immunoprecipitation and expression analyses. Protein levels in clinical specimens were correlated with clinicopathological parameters and patient survival rates. RESULTS Ectopic CCRK expression in immortalized human liver cells increased EZH2 and histone H3 lysine 27 trimethylation (H3K27me3) to stimulate proliferation and tumor formation. Conversely, knockdown of CCRK reduced EZH2/H3K27me3 levels and decreased HCC cell growth, which could be rescued by EZH2 over-expression. Mechanistically, GSK-3β phosphorylation by CCRK activated a β-catenin/TCF/E2F1/EZH2 transcriptional feedback loop to epigenetically enhance androgen receptor (AR) signaling. Simultaneously, the phosphorylation of AKT/EZH2 by CCRK facilitated the co-occupancy of CCRK promoter by EZH2-AR and its subsequent transcriptional activation, thus forming a self-reinforcing circuitry. Lentiviral-mediated knockdown of CCRK, which abrogated the phosphorylation-transcriptional network, prevented diethylnitrosamine-induced tumorigenicity. More importantly, the hyperactivation of the CCRK-EZH2 circuitry in human HCCs correlated with tumor recurrence and poor survival. CONCLUSIONS These findings uncover an epigenetic vicious cycle in hepatocarcinogenesis that operates through reciprocal regulation of CCRK and EZH2, providing novel therapeutic strategy for HCC.

Comparison of postoperative pain between single-incision laparoscopic cholecystectomy and conventional laparoscopic cholecystectomy: prospective case-control study.

Background: Single-incision laparoscopic cholecystectomy (SILC) is an emerging technique used in cholecystectomy. Nevertheless, studies on the difference in postoperative pain between SILC and conventional laparoscopic cholecystectomy are lacking in the literature. Patient and Methods: From August 2009 to July 2010, 20 consecutive patients who underwent SILC (SILC group) were compared with a prospective cohort of 20 patients who underwent conventional 4-port laparoscopic cholecystectomy (LC group) during the same period. Standard postoperative analgesic protocol was applied to all patients. Pain scores were registered before operation and one day after operation using visual analog scale. Satisfactory score was self-assessed by all patients one month after surgery using visual analog scale. Results: There was no significant difference in the baseline characteristics, preoperative pain score, and intraoperative opioid usage between the 2 groups. The postoperative pain score was significantly lower in the SILC group (2.9±1.6 in SILC group vs. 4.8±1.5 in LC group, P<0.01). There was also a lower analgesic consumption and a higher satisfactory score in the SILC group although the results did not reach statistical significance. Conclusions: SILC resulted in significantly less postoperative pain than conventional laparoscopic cholecystectomy. It is a safe alternative to conventional laparoscopic cholecystectomy.

Randomized clinical trial of open hepatectomy with or without intermittent Pringle manoeuvre

The intermittent Pringle manoeuvre (IPM) is commonly applied during liver resection. Few randomized trials have addressed its effectiveness in reducing blood loss and the results have been conflicting. The present study investigated the hypothesis that IPM could reduce blood loss during liver resection by 50 per cent.