Aberrant (pro)renin receptor expression induces genomic instability in pancreatic ductal adenocarcinoma through upregulation of SMARCA5/SNF2H.

(Pro)renin receptor [(P)RR] has a role in various diseases, such as cardiovascular and renal disorders and cancer. Aberrant (P)RR expression is prevalent in pancreatic ductal adenocarcinoma (PDAC) which is the most common pancreatic cancer. Here we show whether aberrant expression of (P)RR directly leads to genomic instability in human pancreatic ductal epithelial (HPDE) cells. (P)RR-expressing HPDE cells show obvious cellular atypia. Whole genome sequencing reveals that aberrant (P)RR expression induces large numbers of point mutations and structural variations at the genome level. A (P)RR-expressing cell population exhibits tumour-forming ability, showing both atypical nuclei characterised by distinctive nuclear bodies and chromosomal abnormalities. (P)RR overexpression upregulates SWItch/Sucrose Non-Fermentable (SWI/SNF)-related, matrix-associated, actin-dependent regulator of chromatin, subfamily a, member 5 (SMARCA5) through a direct molecular interaction, which results in the failure of several genomic stability pathways. These data reveal that aberrant (P)RR expression contributes to the early carcinogenesis of PDAC.

Long-Term Management of Recurrent Cholecystitis after Initial Conservative Treatment: Endoscopic Transpapillary Gallbladder Stenting.

Background: Endoscopic transpapillary gallbladder stenting (ETGBS) is an effective procedure for treating high-risk patients with acute cholecystitis and severe comorbidities. However, the efficacy of ETGBS for recurrent cholecystitis (RC) remains unclear. This study aimed to explore its efficacy in patients with RC for whom cholecystectomy is contraindicated because of its high surgical risk. Methods: Data on 19 high-risk patients who had undergone ETGBS for RC after initial conservative therapy in our institution between June 2006 and May 2012 were retrospectively examined. The primary outcome was the clinical success rate, which was defined as no recurrences of acute cholecystitis after ETGBS until death or the end of the follow-up period. Secondary outcomes were technical success rate and adverse events (AEs). Results: The clinical success rate of ETGBS was 100%, the technical success rate 94.7%, and AE rate 5%: one patient developed procedure-related mild acute pancreatitis. The clinical courses of all patients were as follows: four died of nonbiliary disease, and the remaining 15 were subsequently treated conservatively. The median duration of follow-up was 14.95 months (range 3-42 months). Conclusions: ETGBS is an effective alternative for managing RC in high-risk patients with severe comorbidities.

MicroRNA profiles during galectin-9-induced apoptosis of pancreatic cancer cells.

Pancreatic cancer is the eighth-leading cause of cancer-associated mortality in males and the ninth-leading cause in females worldwide. Even when diagnosed early enough to be potentially resectable, the prognosis of invasive pancreatic cancer is poor. Galectin-9 (Gal-9) is a tandem-repeat type galectin that has recently been demonstrated to possess an anti-proliferative effect on cancer cells. Therefore, the present study evaluated the effects of Gal-9 on the proliferation of human pancreatic cancer cells and examined the microRNAs that are associated with the antitumor effects of Gal-9. Gal-9 suppressed the proliferation of multiple pancreatic cancer cell lines. In addition, Gal-9 treatment increased the levels of caspase-cleaved keratin 18 and the expression of cytochrome c in pancreatic cancer cell lines. This data suggests that Gal-9 induces intrinsic apoptosis in pancreatic cancer cell lines through the caspase-dependent and caspase-independent pathways. In addition, Gal-9 reduced the expression levels of phosphorylated epidermal growth factor receptor and numerous receptor tyrosine kinases (RTKs). In conclusion, Gal-9 may suppress the growth of human pancreatic cancer cells in vitro. These findings suggest that Gal-9 may be a new therapeutic agent for the treatment of pancreatic cancer.

The anti-diabetic drug metformin inhibits pancreatic cancer cell proliferation in vitro and in vivo: Study of the microRNAs associated with the antitumor effect of metformin.

Recent studies suggest that metformin, which is a commonly used oral anti-hyperglycemic agent of the biguanide family, may reduce cancer risk and improve prognosis, yet the detailed mechanisms by which metformin affects various types of cancers, including pancreatic cancer, remain unknown. The aim of the present study was to evaluate the effects of metformin on human pancreatic cancer cell proliferation in vitro and in vivo, and to study microRNAs (miRNAs) associated with the antitumor effect of metformin. We used the human pancreatic cancer cell lines Panc1, PK1 and PK9 to study the effects of metformin on human pancreatic cancer cells. Athymic nude mice bearing xenograft tumors were treated with or without metformin. Tumor growth was recorded after 5 weeks, and the expression of cell cycle-related proteins was determined. In addition, we used miRNA microarray tips to explore the differences in the levels of miRNAs in Panc1 cells and xenograft tumors treated with metformin or without. Metformin inhibited the proliferation of Panc1, PK1 and PK9 cells in vitro. This inhibition was accompanied by a strong decrease in G1 cyclins (particularly in cyclin D1) and retinoblastoma protein (Rb) phosphorylation. In addition, metformin reduced the phosphorylation of epidermal growth factor receptor (EGFR), particularly the phosphorylation of EGFR at Tyr845, and insulin-like growth factor 1 receptor (IGF-1R) in vitro and in vivo. miRNA expression was markedly altered by the treatment with metformin in vitro and in vivo. Our results revealed that metformin inhibits human pancreatic cancer cell proliferation and tumor growth, possibly by suppressing the cell cycle-related molecules via alteration of miRNAs.

Antitumor effect of metformin on cholangiocarcinoma: In vitro and in vivo studies.

Cholangiocarcinoma (CCA) is the most common biliary malignancy and the second most common hepatic malignancy after hepatocellular carcinoma (HCC). Treatment with the anti-diabetic drug metformin has been associated with reduced cancer incidence in patients with type 2 diabetes. Thus, the present study evaluated the effects of metformin on human CCA cell proliferation in vitro and in vivo and identified the microRNAs associated with its antitumor effects. Metformin inhibited the proliferation of the CCA cell lines HuCCT-1 and TFK-1 and blocked the G0 to G1 cell cycle transition, accompanied by AMP kinase pathway activation. Metformin treatment also led to marked decreases in cyclin D1 and cyclin-dependent kinase (Cdk) 4 protein levels and retinoblastoma protein phosphorylation. However, this drug did not affect p27kip protein expression. In addition, it reduced the phosphorylation of Axl, EphA10, ALK and PYK, as well as tumor proliferation in athymic nude mice with xenograft tumors. Furthermore, it markedly altered microRNA expression. These findings suggest that metformin may have clinical use in the treatment of CCA.

Galectin-9 suppresses cholangiocarcinoma cell proliferation by inducing apoptosis but not cell cycle arrest.

Cholangiocarcinoma is the most common biliary malignancy and the second most common hepatic malignancy after hepatocellular carcinoma (HCC). Galectin-9 (Gal-9) is a tandem-repeat-type galectin that has recently been shown to exert antiproliferative effects on cancer cells. Therefore, the present study evaluated the effects of Gal-9 on the proliferation of human cholangiocarcinoma cells in vitro as well as the microRNAs (miRNAs) associated with the antitumor effects of Gal-9. Gal-9 suppressed the proliferation of cholangiocarcinoma cell lines in vitro and the growth of human cholangiocarcinoma cell xenografts in nude mice. Our data further revealed that Gal-9 increased caspase­cleaved keratin 18 (CCK18) levels, and the expression of cytochrome c increased in Gal-9-treated cholangiocarcinoma cell lines. These data suggested that Gal-9 induced cholangiocarcinoma cell apoptosis via the intrinsic apoptosis pathway mediated by caspase-dependent or -independent pathways. In addition, Gal-9 reduced the phosphorylation of the epidermal growth factor receptor (EGFR), insulin-like growth factor and insulin-like growth factor-1 receptor (IGF-1R), hepatocyte growth factor receptor and fibroblast growth factor receptor 3 (FGFR3). These findings suggest that Gal-9 can be a candidate of therapeutic target in the treatment of cholangiocarcinoma.

(Pro)renin receptor is crucial for Wnt/ß-catenin-dependent genesis of pancreatic ductal adenocarcinoma.

Although Wnt/ß-catenin signaling is known to be aberrantly activated in PDAC, mutations of CTNNB1, APC or other pathway components are rare in this tumor type, suggesting alternative mechanisms for Wnt/ß-catenin activation. Recent studies have implicated the (pro)renin receptor ((P)RR) is related to the Wnt/ß-catenin signaling pathway. We therefore investigated the possible role of (P)RR in pancreatic carcinogenesis. Plasma s(P)RR levels were significantly (P < 0.0001) higher in patients with PDAC than in healthy matched controls. We also identified aberrant expression of (P)RR in premalignant PanIN and PDAC lesions and all the PDAC cell lines examined. Inhibiting (P)RR with an siRNA attenuated activation of Wnt/ß-catenin signaling pathway and reduced the proliferative ability of PDAC cells in vitro and the growth of engrafted tumors in vivo. Loss of (P)RR induced apoptosis of human PDAC cells. This is the first demonstration that (P)RR may be profoundly involved in ductal tumorigenesis in the pancreas.

Mechanism of gemcitabine-induced suppression of human cholangiocellular carcinoma cell growth.

Although gemcitabine (2',2'-difluorocytidine monohydrochloride) is a common anticancer agent of cholangiocellular carcinoma (CCC), its growth inhibitory effects and gemcitabine resistance in CCC cells are poorly understood. Our aims were to uncover the mechanism underlying the antitumor effect of gemcitabine and to analyze the mechanism regulating in vitro CCC cell gemcitabine resistance. In addition, we sought to identify miRNAs associated with the antitumor effects of gemcitabine in CCCs. Using a cell proliferation assay and flow cytometry, we examined the ability of gemcitabine to inhibit cell proliferation in three types of human CCC cell lines (HuCCT-1, Huh28, TKKK). We also employed western blotting to investigate the effects of gemcitabine on cell cycle-related molecules in CCC cells. In addition, we used array chips to assess gemcitabine-mediated changes in angiogenic molecules and activated tyrosine kinase receptors in CCC cells. We used miRNA array chips to comprehensively analyze gemcitabine-induced miRNAs and examined clusters of differentially expressed miRNAs in cells with and without gemcitabine treatment. Gemcitabine inhibited cell proliferation in a dose- and time-dependent manner in HuCCT-1 cells, whereas cell proliferation was unchanged in Huh28 and TKKK cells. Gemcitabine inhibited cell cycle progression in HuCCT-1 cells from G0/G1 to S phase, resulting in G1 cell cycle arrest due to the reduction of cyclin D1 expression. In addition, gemcitabine upregulated the angiogenic molecules IL-6, IL-8, ENA-78 and MCP-1. In TKKK cells, by contrast, gemcitabine did not arrest the cell cycle or modify angiogenic molecules. Furthermore, in gemcitabine-sensitive HuCCT-1 cells, gemcitabine markedly altered miRNA expression. The miRNAs and angiogenic molecules altered by gemcitabine contribute to the inhibition of tumor growth in vitro.

Molecular Biologic Approach to the Diagnosis of Pancreatic Carcinoma Using Specimens Obtained by EUS-Guided Fine Needle Aspiration.

We review the utility of endoscopic ultrasound-guided fine needle aspiration (EUS-FNA), a rapid, safe, cost-effective, and accurate diagnostic modality for evaluating pancreatic tumors. EUS-FNA is currently used for the diagnosis and staging of pancreatic tumors. The sensitivity of EUS-FNA for pancreatic malignancy ranges from 75% to 94%, and its specificity approaches 100% in most studies. However, EUS-FNA has some limitations in the diagnosis of well-differentiated or early-stage cancers. Recent evidence suggests that molecular biological analysis using specimens obtained by EUS-FNA improves diagnostic sensitivity and specificity, especially in borderline cytological cases. It was also reported that additional information regarding patient response to chemotherapy, surgical resectability, time to metastasis, and overall survival was acquired from the genetic analysis of specimens obtained by EUS-FNA. Other studies have revealed that the analysis of KRAS, MUC, p53, p16, S100P, SMAD4, and microRNAs is helpful in making the diagnosis of pancreatic carcinoma. In this paper, we describe the present state of genetic diagnostic techniques for use with EUS-FNA samples in pancreatic diseases. We also discuss the role of molecular biological analyses for the diagnosis of pancreatic carcinoma.