Xiangyu Zhong

MicroRNA-421 functions as an oncogenic miRNA in biliary tract cancer through down-regulating farnesoid X receptor expression

MicroRNAs (miRNAs) are involved in the development of most cancers. However, few studies have been conducted to determine their relationship to biliary tract cancer (BTC). Farnesoid X receptor (FXR) has been reported to be a tumor suppressor for hepatocellular carcinoma and breast cancer; but few studies have focused on its correlation with BTC. In this study, we identified miR-421 as a potential regulator of FXR expression. We found that their expression amount was inversely correlated as FXR was aberrantly down-regulated in both primary tumor specimens and cell lines; while miR-421 was significantly up-regulated. Ectopic expression of miR-421 significantly decreased FXR protein concentration in BTC cells and promoted cell proliferation, colony formation and migration in vitro. Furthermore, a decrease in miR-421 expression induced G(0)/G(1) cell cycle arrest. In conclusion, our study identified microRNA-421 functions as an oncomiR in BTC by targeting FXR. This finding may provide a novel therapeutic strategy for treatment of biliary tract cancer.

IL-6/STAT3/TFF3 signaling regulates human biliary epithelial cell migration and wound healing in vitro

Interleukin-6 (IL-6), through activation of the signal transducer and activator of transcription 3 (STAT3) and trefoil factor family 3 (TFF3), has been implicated in the promotion of mouse biliary epithelial cell (BEC) proliferation and migration. However, it is still unclear whether the IL-6/STAT3/TFF3 signaling had similar effects on human BECs. Here, we showed that exposure of human BECs to recombinant IL-6 resulted in STAT3 phosphorylation and increased the expression of TFF3 at both mRNA and protein levels. Moreover, inhibition of STAT3 using RNA interference significantly abrogated IL-6-induced TFF3 expression. In an in-vitro wound healing model, IL-6 facilitated human BEC migration. This promotion of cell migration by IL-6 was blocked when STAT3 was knocked down. Interestingly, the addition of exogenous TFF3 could rescue the cell migration defects caused by STAT3 silencing. In conclusion, our data indicate that STAT3 plays a critical role in IL-6-induced TFF3 expression in human BECs and the IL-6/STAT3/TFF3 signaling is involved in human BEC migration and wound healing.

Impact of Pancreatic Margin Status and Lymph Node Metastases on Recurrence after Resection for Invasive and Noninvasive Intraductal Papillary Mucinous Neoplasms of the Pancreas: A Meta-Analysis

Background: Accurate information is currently lacking regarding the values of positive margins (M+) and lymph node (LN) metastases as independent predictors of postoperative recurrence in invasive and noninvasive intraductal papillary mucinous neoplasms (IPMNs) of the pancreas. Methods: A comprehensive online literature search identified all types of primary studies that included M+ and LN metastases as risk factors and defined recurrence as an outcome in patients with IPMNs. Suitable articles were also identified by manually researching references in qualifying articles. A meta-analysis of the result was performed using a random effects model. Results: The recurrence rate in noninvasive IPMNs was 3.72% in patients with negative margin (M–) versus 9.56% in those with M+ (odds ratio, OR = 0.37, 95% confidence interval, 95% CI: 0.17–0.78, p = 0.010). The recurrence rate in invasive M– IPMNs in was 33.85% compared to 53.66% in M+ IPMNs (OR = 0.47, 95% CI: 0.25–0.88, p = 0.020). The recurrence rate in invasive IPMNs with positive LN was 76.92% compared to 30.86% with negative LN; OR = 0.15, 95% CI: 0.06–0.37, p < 0.0001). Conclusions: M+ were associated with disease recurrence in all patients with IPMN, and nodal metastases were significantly associated with recurrence in invasive IPMN.

GPC1 regulated by miR-96-5p, rather than miR-182-5p, in inhibition of pancreatic carcinoma cell proliferation.

To determine the relationships between miR-96-5p/-182-5p and GPC1 in pancreatic cancer (PC), we conducted the population and in vitro studies. We followed 38 pancreatic cancer patients, measured and compared the expression of miR-96-5p/-182-5p, GPC1, characteristics and patients’ survival time of different miR-96-5p/-182-5p expression levels in PC tissues. In an in vitro study, we investigated the proliferation, cycle and apotosis in cells transfected with mimics/inhibitors of the two miRNAs, and determine their effects on GPC1 by dual-luciferase assay. In the follow-up study, we found that the expressions of miR-96-5p/-182-5p were lower/higher in PC tissues; patients with lower/higher levels of miR-96-5p/-182-5p suffered poorer characteristics and decreased survival time. In the in vitro study, the expressions of miR-96-5p/-182-5p were different in cells. Proliferation of cells transfected with miR-96-5p mimics/inhibitors was lower/higher in Panc-1/BxPC-3; when transfected with miR-182-5p mimics/inhibitors, proliferation of cells were higher/lower in AsPC-1/Panc-1. In a cell cycle study, panc-1 cells transfected with miR-96-5p mimics was arrested at G0/G1; BxPC-3 cells transfected with miR-96-5p inhibitors showed a significantly decrease at G0/G1; AsPC-1 cells transfected with miR-182-5p mimics was arrested at S; Panc-1 cells transfected with miR-182-5p inhibitors showed a decrease at S. MiR-96-5p mimics increased the apoptosis rate in Panc-1 cells, and its inhibitors decreased the apoptosis rate in BxPC-3. Dual luciferase assay revealed that GPC1 was regulated by miR-96-5p, not -182-5p. We found that miR-96-5p/-182-5p as good markers for PC; miR-96-5p, rather than -182-5p, inhibits GPC1 to suppress proliferation of PC cells.

The Wnt antagonist sFRP1 as a favorable prognosticator in human biliary tract carcinoma.

Inactivation of Secreted Frizzled-Related Protein-1 (SFRP1) and overexpression of β-catenin play important roles in the development and progression of a wide range of malignancies. We sought to determine whether the expression of SFRP1 and β-catenin correlates with clinicopathologic parameters in human biliary tract cancer (BTC) and to evaluate the potential roles of these proteins as prognostic indicators. The expression of SFRP1 and β-catenin in 78 patients with BTC and 36 control patients as investigated by immunohistochemistry. A wide variety of statistical parameters were assessed to determine the association between these proteins and the occurrence, clinical features, and overall survival rate in BTC.SFRP1 and β-catenin had an inverse correlation (r = −0.636, P<0.0001) as assessed by Spearman rank analysis, with 52 (66.7%) of the BTC samples negative for SFRP1 expression and 53 (68.0%) positive for β-catenin expression. Expression of each protein was associated with the histological type and lymph node invasion of BTC. A significantly poorer overall survival rate was observed for patients with low SFRP1 expression (P<0.0001) or high β-catenin expression (P = 0.007). SFRP1 expression (P<0.0001), β-catenin expression (P<0.01) and histological type (P<0.01) were correlated with overall survival rate as assessed by univariate analysis; while multivariate analysis suggested that SFRP1 (hazard ratio, 10.514; 95% confidence intervals, 2.381–39.048; P<0.0001) may serve as an independent prognostic factor for BTC. Collectively, these results demonstrate that SFRP1 is a favorable prognostic factor for human BTC and that its expression inversely correlates with that of β-catenin.

Clinical and biological significance of Cdk10 in hepatocellular carcinoma

Cyclin-dependent kinase 10 (Cdk10) is a Cdc2-related kinase and plays an essential role in the progression from the G2 to M phase of the cell cycle. However, relative little is known about its expression pattern, clinical relevance, and biological function in hepatocellular carcinoma (HCC). In the present study, we investigated the mRNA and protein expression levels of Cdk10 in 127 pairs of HCC samples and adjacent nontumorous liver tissues and evaluated its clinical significance. Additionally, we assessed the effects of restoration of Cdk10 on cell proliferation and drug sensitivity in HCC cells. We showed that the Cdk10 mRNA and protein expression was markedly decreased in HCC samples compared to adjacent nontumorous liver tissues. Quantitative real-time polymerase chain reaction and immunohistochemical studies revealed that reduced Cdk10 expression was significantly associated with alpha-fetoprotein levels, tumor size, and tumor stage. Ectopic expression of Cdk10 reduced HCC cell proliferation, blocked the cell cycle at the G0-G1 phase, as well as inhibited cell migration and anchorage-independent growth. Additionally, Cdk10 overexpression enhanced the chemosensitivity of HCC cells to cisplatin and epidoxorubicin, two chemotherapeutic agents commonly used in HCC. These data collectively demonstrate that reduced Cdk10 expression is closely linked to HCC development and progression. Restoration of its expression may have therapeutic benefits in treating this malignancy.

CDK10 functions as a tumor suppressor gene and regulates survivability of biliary tract cancer cells

Cyclin-dependent kinase 10 (CDK10) is a member of the Cdc2 family of kinases, and has been demonstrated to be an important determinant of resistance to endocrine therapy for breast cancer. To investigate the expression and possible function of CDK10 in biliary tract cancer (BTC), we systematically examined CDK10 in tissues and cell lines. We found that expression of CDK10 was downregulated in both biliary tract tumors and cell lines. Remarkably, the expression of CDK10 correlated with clinical characteristics. Overexpression or knockdown of CDK10, respectively, inhibited or promoted cell proliferation, colony formation and migration. This suggests that CDK10 functions as a tumor suppressor gene in BTC. Overexpression of CDK10 caused malignant cells to become sensitive to chemotherapy and other hostile environments, suggesting that CDK10 functions to regulate survivability of BTC cells. We investigated the expression of six genes to resolve the mechanism. c-RAF was negatively regulated by CDK10 in both cells and specimens. Our results indicate that CDK10 plays a crucial role in the growth and survivability of biliary tract cancer, and offers a potential therapeutic target for this fatal disease.

SP1-induced upregulation of lncRNA SPRY4-IT1 exerts oncogenic properties by scaffolding EZH2/LSD1/DNMT1 and sponging miR-101-3p in cholangiocarcinoma

BackgroundAccumulating evidence has indicated that long non-coding RNAs (lncRNAs) behave as a novel class of transcription products during multiple cancer processes. However, the mechanisms responsible for their alteration in cholangiocarcinoma (CCA) are not fully understood.MethodsThe expression of SPRY4-IT1 in CCA tissues and cell lines was determined by RT-qPCR, and the association between SPRY4-IT1 transcription and clinicopathologic features was analyzed. Luciferase reporter and chromatin immunoprecipitation (ChIP) assays were performed to explore whether SP1 could bind to the promoter region of SPRY4-IT1 and activate its transcription. The biological function of SPRY4-IT1 in CCA cells was evaluated both in vitro and in vivo. ChIP, RNA binding protein immunoprecipitation (RIP) and luciferase reporter assays were performed to determine the molecular mechanism of SPRY4-IT1 in cell proliferation, apoptosis and invasion.ResultsSPRY4-IT1 was abnormally upregulated in CCA tissues and cells, and this upregulation was correlated with tumor stage and tumor node metastasis (TNM) stage in CCA patients. SPRY4-IT1 overexpression was also an unfavorable prognostic factor for patients with CCA. Additionally, SP1 could bind directly to the SPRY4-IT1 promoter region and activate its transcription. Furthermore, SPRY4-IT1 silencing caused tumor suppressive effects via reducing cell proliferation, migration and invasion; inducing cell apoptosis and reversing the epithelial-to-mesenchymal transition (EMT) process in CCA cells. Mechanistically, enhancer of zeste homolog 2 (EZH2) along with the lysine specific demethylase 1 (LSD1) or DNA methyltransferase 1 (DNMT1) were recruited by SPRY4-IT1, which functioned as a scaffold. Importantly, SPRY4-IT1 positively regulated the expression of EZH2 through sponging miR-101-3p.ConclusionsOur data illustrate how SPRY4-IT1 plays an oncogenic role in CCA and may offer a potential therapeutic target for treating CCA.

Use of a cocktail regimen consisting of soluble galectin-1, rapamycin and histone deacetylase inhibitor may effectively prevent type 1 diabetes.

Type 1 diabetes (T1D) is an autoimmune disorder that results in destruction of insulin-releasing beta-cells of the pancreas. During the pathogenesis of T1D, at least two phases of beta-cell death occur: an initiation event wherein macrophage-derived inflammatory cytokines induce beta-cell necrosis and release of beta-cell-specific antigens, and a second, antigen-driven event in which T-cell-mediated immune response is directed against beta-cells. In contrast to macrophages and autoreactive T cells, regulatory T cells play a key role in inducing and maintaining immunological tolerance to self antigens. Therefore, modulation of the immune system may prevent the development of T1D. Herein, we proposed a cocktail regimen consisting of soluble galectin-1, rapamycin and histone deacetylase inhibitor (HDACi) for the treatment of T1D because (a) HDACi has been reported to protect against IL-1beta-mediated loss in beta-cell viability, (b) HDACi and rapamycin have the ability to promote the generation and function of regulatory T cells and thus suppress the cytotoxic T-cell function, and (c) administration of soluble galectin-1 can trigger apoptosis of the beta-cell-reactive T cells. This cocktail regimen may not only block T-cell- and cytokine-mediated autoimmunity but also restore self-tolerance to beta-cell antigens, therefore representing a novel alternative for treatment of T1D.

Long non-coding RNA UCA1 indicates an unfavorable prognosis and promotes tumorigenesis via regulating AKT/GSK-3β signaling pathway in cholangiocarcinoma

Long non-coding RNAs (lncRNAs) have been documented to play key roles in a wide range of pathophysiological processes, including cancer initiation and progression. Recently, the aberrant expression of urothelial carcinoma associated 1 (UCA1) was observed in many types of cancers. However, its clinical relevance and exact effects in cholangiocarcinoma (CCA) remains unknown. In the present study, we aimed to investigate the clinical significance of UCA1 and evaluate its prognostic value in patients with CCA. Besides, the functional roles of UCA1 were detected both in vitro and in vivo. Moreover, potential signaling pathways were explored to clarify the molecular mechanisms underlying CCA cell proliferation. The results indicated that UCA1 transcription is enhanced in both CCA tissue samples and cell lines, and this overexpression is associated with tumor stage (P = 0.007), lymph node invasion (P = 0.027), TNM stage (P = 0.004) and postoperative recurrence (P = 0.033) of CCA patients. Besides, UCA1 could function as an independent prognostic predictor for overall survival in patients with CCA (P = 0.014). For the part of functional assays, knockdown of UCA1 could attenuate CCA cell growth both in vitro and in vivo. Besides, UCA1 facilitates apoptosis via Bcl-2/caspase-3 pathway. In addition, UCA1 regulates migration and invasion potential of CCA cells by affecting EMT. Furthermore, AKT/GSK-3β axis was activated to upregulate CCND1 expression due to overexpression of UCA1 in CCA. To summary, UCA1 might be a potentially useful prognostic biomarker and therapeutic target for CCA.