67 laminin receptor promotes the malignant potential of tumour cells up-regulating lysyl oxidase-like 2 expression in cholangiocarcinoma.

BACKGROUND: 67 laminin receptor (67LR) plays an important role in the invasion and metastasis of cholangiocarcinoma, but its mechanism remains unclear. AIMS: We investigated the clinical significance of 67LR and its relation to lysyl oxidase-like 2 (LOXL2) in 67LR-mediated invasion and metastasis in cholangiocarcinoma. METHODS: The clinical significance of 67LR and LOXL2 expression and the prognosis of patients were investigated in 73 cancerous and 32 paracancerous tissues by immunohistochemistry. The impact of LOXL2 on invasion, metastasis and 67LR expression was evaluated in cholangiocarcinoma cells by shRNA or expressed-plasmid transfection. RESULTS: Expression of 67LR was recognized in 35.62% cholangiocarcinoma tissue, and none in paracancerous tissues. LOXL2 was positively correlated with expression of 67LR. Expression of 67LR or LOXL2 in cholangiocarcinomas was significantly associated with lymph node metastasis, differentiation and poor overall survival. Cox analysis showed that 67LR can act as an independent prognostic biomarker of prognosis in cholangiocarcinoma patients. Expression of LOXL2 decreased by knockdown of 67LR and increased by overexpression of 67LR in cholangiocarcinoma cells. Knockdown of LOXL2 reduced invasion and metastasis in vitro and in vivo. CONCLUSION: 67LR may regulate the expression of LOXL2 to promote invasion and metastasis in cholangiocarcinoma cells. It could be used as an independent prognostic marker in cholangiocarcinoma patients.

LPS increases MUC5AC by TACE/TGF-α/EGFR pathway in human intrahepatic biliary epithelial cell.

BACKGROUND: Mucin 5AC (MUC5AC) overproduction plays important roles in stone formation and recurrence of hepatolithiasis. We aim to investigate the involved mechanism and the potential target to block this process. METHODS: 42 bile duct samples from hepatolithiasis and 15 normal bile duct samples from hemangioma patients were collected for detecting MUC5AC expression by immunohistochemistry. MUC5AC and phosphoepidermal growth factor receptor (pEGFR) expressions in human intrahepatic biliary epithelial cells (HIBECs) cultured with or without lipopolysaccharide (LPS) were detected by real-time PCR and western blot analysis. Transforming growth factor-α (TGF-α) secretion in HIBECs was detected by ELISA. RESULTS: MUC5AC was overexpressed in bile ducts of hepatolithiasis samples compared with bile ducts from hemangioma samples. LPS upregulated MUC5AC expression in HIBECs. LPS promoted EGFR activation, and inhibiting EGFR activation by AG1478 significantly decreased LPS-induced MUC5AC overexpression in HIBECs. Moreover, LPS increased TGF-α secretion, and inhibiting tumor necrosis factor-α converting enzyme (TACE), which has been implicated in ectodomain cleavage of TGF-α, significantly inhibited LPS-induced EGFR activation and subsequent MUC5AC overexpression in HIBECs. CONCLUSION: Our results suggested that LPS increases MUC5AC expression through the TACE/TGF-α/EGFR pathway in HIBECs. This new finding might give light to the prevention of stone formation and recurrence of hepatolithiasis.

Aberrant expression of GATA binding protein 6 correlates with poor prognosis and promotes metastasis in cholangiocarcinoma.

AIM: GATA6, a zinc-finger transcription factor, functions as a tumour promoter or suppresser according to different tumour origins. We investigated the clinical significance of GATA6 and its role in invasion and metastasis in cholangiocarcinoma (CCA). METHODS: Expression of GATA6 in 87 cancerous, 24 paracancerous, 32 lymph-node metastatic and 8 liver metastatic samples from 87 CCA patients undergoing surgical resection was detected by immunohistochemistry. Impact of GATA6 on invasion, metastasis and 67kDa laminin receptor expression (67LR) was evaluated in CCA cells by shRNA lentivirus or expressed-plasmid transfection. RESULTS: Aberrant expression of GATA6 in CCAs was significantly associated with lymph-node metastasis. GATA6 expression was higher in lymph-node and liver metastatic tissues compared with primary cancerous tissues. Kaplan-Meier analysis showed GATA6 expression correlated with poor overall survival and early recurrence in CCAs. Cox analysis suggested GATA6 was an independent prognostic marker for overall survival and recurrence-free survival. CCA cell invasion and migration were decreased by GATA6 knockdown and enhanced by GATA6 overexpression in vitro. Knockdown of GATA6 reduced CCA cell metastasis by xenotransplantation into nude mice. 67LR, which is overexpressed in CCAs and promotes invasion and metastasis through several pathways, positively correlated with GATA6 expression in 87 CCAs. Both mRNA and protein levels of 67LR were regulated by GATA6 in CCA cells. Moreover, ChIP analysis showed GATA6 bound to 67LR gene promoter in CCA cells. CONCLUSION: Aberrant expression of GATA6 correlates with poor prognosis and promotes invasion and metastasis in CCA.

Surgical treatment of congenital biliary duct cyst.

BACKGROUND: It is acknowledged that total cyst excision is a safe and ideal surgical treatment for congenital biliary duct cyst, compared to simple internal drainage. The aim of this study was to determine the optimal operation occasion and the effect of laparoscopy on congenital biliary duct cyst based upon total cyst excision. METHODS: From January 2002 to January 2011, 217 patients were admitted to Southwest Hospital for congenital biliary duct cyst. To determine the optimal surgery occasion, we divided these subjects into three groups, the infant group (age ≤ 3 years), the immaturity group (3 < age ≤ 18 years), and the maturity group (age > 18 years), and then evaluated the feasibility, risk and long-term outcome after surgery in the three groups. To analyze the effect of laparoscopic technique on congenital biliary duct cyst, we divided the patients into the laparoscopy and the open surgery groups. RESULTS: Among the three groups, the morbidity from cholangiolithiasis before surgical treatment had obvious discrepancy (p < 0.05) (lowest in the infant group), and intraoperative blood loss also had apparent diversity (p < 0.05). Furthermore, long-term outcomes (secondary cholangiolithiasis, stoma stenosis and cholangiocarcinoma) showed no significant difference between different groups (p > 0.05).Similarly, no significant discrepancy was observed in the morbidity from postoperative complications or long-term postoperative complications (p > 0.05) between the laparoscopic and the open surgery groups. CONCLUSIONS: We conclude that total cyst excision should be performed as early as possible. The optimal treatment occasion is the infant period, and laparoscopic resection may be a new safe and feasible minimally invasive surgery for this disease.

Downregulation of mucins in graft bile ducts after liver transplantation in rats.

BACKGROUND: Nonanastomotic biliary strictures represent a serious complication after orthotopic liver transplantation (OLT). This study investigates the potential role of mucins in bile duct injury after OLT. METHODS: Sprague-Dawley rats were divided into four groups: normal group (Normal, n=5), sham-operated group (Sham, n=20), OLT group with 1 hr donor cold ischemic time (n=20), and OLT group with 12 hr donor cold ischemic time (OLTn=20). Expression of mucins and GATA factors in bile ducts was examined by real-time polymerase chain reaction, immunohistochemistry, and immunoblotting. Bile was collected for biochemical analysis, and the histological changes associated with bile duct injury were evaluated. RESULTS: In normal bile ducts, Muc1, Muc2, Muc3A, Muc4, and Muc6 mRNA were expressed, whereas Muc5AC mRNA was undetectable. The expression of Muc1, Muc3A, and Muc4 but not Muc2 and Muc6 at mRNA level in graft bile ducts decreased remarkably early after OLT. The decreased expression of Muc1 and Muc4 was further confirmed at protein level by immunohistochemistry and immunoblotting. Downregulation of Muc1 and Muc3A expression by prolonged cold ischemic time was significantly associated with the injury severity scores of large but not small bile ducts. Among six GATA factors, GATA3, GATA4, and GATA6 mRNA were expressed in normal bile ducts. GATA4 and GATA6 mRNA levels decreased significantly after OLT. CONCLUSION: Downregulation of Muc1 and Muc3A expression by prolonged cold ischemic time may play a potential role in large bile duct injury early after OLT.

Role of cholangiocyte bile Acid transporters in large bile duct injury after rat liver transplantation.

BACKGROUND: The pathogenesis of nonanastomotic strictures with a patent hepatic artery remains to be investigated. This study focuses on the role of cholangiocyte bile acid transporters in bile duct injury after liver transplantation. METHODS: Sprague-Dawley rats were divided into three groups (n=20 for each): the sham-operated group (Sham), the transplant group with 1-hr donor liver cold preservation (CP-1h), and the transplant group with 12-hr donor liver cold preservation (CP-12h). Bile was collected for biochemical analysis. The histopathologic evaluation of bile duct injury was performed and the cholangiocyte bile acid transporters apical sodium-dependent bile acid transporter (ASBT), ileal lipid binding protein (ILBP), and Ostalpha/Ostbeta were investigated. RESULTS.: The immunohistochemical assay suggested that ASBT and ILBP were expressed exclusively on large bile duct epithelial cells, whereas Ostalpha and Ostbeta were expressed on both small and large bile ducts. Western blot and quantitative polymerase chain reaction analysis showed that the expression levels of these transporters dramatically decreased after transplantation. It took seven to 14 days for ILBP, Ostalpha, and Ostbeta to recover, whereas ASBT recovered within 3 days and even reached a peak above the normal level seven days after operation. In the CP-12h group, the ratios of the ASBT/ILBP, ASBT/Ostalpha and ASBT/Ostbeta expression levels were correlated with the injury severity scores of large but not small bile ducts. CONCLUSIONS: The results suggest that the unparallel alteration of cholangiocyte bile acid transporters may play a potential role in large bile duct injury after liver transplantation with prolonged donor liver preservation.

Effects of simulated weightlessness on liver Hsp70 and Hsp70mRNA expression in rats.

Space flight is known to induce a number of hepatic physiological alterations. In this study, we investigated Hsp70 expressing features of rat liver under simulated weightlessness. Tail-suspension was used to simulate the weightlessness animal model. Forty-eight male Wistar rats were randomly assigned to 6 experimental groups and Hsp70 protein and mRNA expressions in the liver were detected by Western blot and RT-PCR respectively. The tail-suspension significantly increased Hsp70mRNA expression levels in rat liver (P<0.05). The semi-quantitative PCR showed that Hsp70mRNA was upregulated as early as 6 hours of suspension. Western blot analysis indicated that Hsp70 protein was significantly upregulated in the early stage of suspension as compared with controls (P<0.05). The results suggest that simulated weightlessness acts as a kind of stress to elevate liver Hsp70 expression both at protein and mRNA levels. This may be meaningful in astronaut's trainings by preadaptation to non-damaging stress exposures or other environmental factors to foster the astronaut's ability of weightless tolerance.

Epithelial-mesenchymal transition induced by hepatitis C virus core protein in cholangiocarcinoma.

BACKGROUND: Cholangiocarcinoma (CC) is associated with chronic hepatitis C virus (HCV) infection. We investigated the effect of hepatitis C virus core protein (HCVc) on epithelial-mesenchymal transition (EMT) in CC and tried to identify its target trigger. METHODS: First, we examined expression of HCVc and epithelial and mesenchymal markers in CC tissues. Then we transient-transfected HCVc gene into a CC cell line and examined expression of lysyl oxidase-like 2 (LOXL2) and epithelial and mesenchymal markers by quantitative real-time polymerase chain reaction (PCR) and Western blotting. Finally, LOXL2 gene silencing was shown in QBC939/HCVc cells by RNA interference (RNAi), and we further examined expression of epithelial and mesenchymal markers by quantitative real-time PCR and Western blotting. RESULTS: Through immunohistochemical staining, the present study showed that HCVc is significantly associated with CC invasion and metastasis. In vitro study showed that HCVc expression induces EMT in CC cell line QBC939, and a mechanism through LOXL2 pathway is suggested. Expression of HCVc was significantly correlated with greater migratory and invasive potential of CC cells. CONCLUSIONS: These observations indicate that HCVc plays a critical role in promoting invasion and metastasis of CC cells.

Ultrasonography guided percutaneous radiofrequency ablation for hepatic cavernous hemangioma.

AIM: Hepatic cavernous hemangioma (HCH) is the most common benign tumor of the liver and its management is still controversial. Recent success in situ radiofrequency ablation of hepatic malignancies has led us to consider using this technique in patients with HCH. This study was to assess the efficacy, safety, and complications of percutaneous radiofrequency ablation (PRFA) under ultrasonography guidance in patients with HCH. METHODS: Twelve patients (four men and eight women, age ranged 33-56 years, mean age was 41.7 years) with 15 hepatic cavernous hemangiomas (2.5 cm to 9.5 cm) were treated using the RF-2000 generator and 10-needle LeVeen electrode percutaneously guided by B-ultrasound. Lesions larger than 3 cm were treated by multiple overlapping ablations that encompass the entire lesion as well as a rim of normal liver tissue (approximately 0.5 cm). RESULTS: All the patients who received PRFA therapy had no severe pain, bleeding or bile leakage during and after the procedures. Nine to 34 months' follow-up (mean, 21 months) by ultrasound and/or spiral CT scan demonstrated that the ablated lesions in this group were shrunk remarkably, and the shrunken range was 38-79 % (mean, 67 % per 21 months). The contrast enhancement was disappeared within the tumor or at its periphery in all cases on spiral CT scans obtained 3 to 6 months after treatment. CONCLUSION: The results of this study suggest that PRFA therapy is a mini-invasive, simple, safe, and effective method for the treatment of selected patients with HCH.