MiR-27a modulates the MDR1/P-glycoprotein expression by inhibiting FZD7/β-catenin pathway in hepatocellular carcinoma cells.

Chemotherapy has been widely used to treat cancer, however, the appearance of multiple drug resistance (MDR) in cancer patients is regarded as a major clinical obstacle to successful chemotherapy. MicroRNAs (miRNAs) are evolutionary conserved small RNAs that regulate gene expression at the post-transcriptional level and have been shown to regulate cell differentiation, development, proliferation and apoptosis. Nevertheless, the involvement of miRNAs and their roles in the development of MDR in liver cancer are not fully understood. Our study found that the expression of miR-27a was down-regulated in the multidrug-resistant hepatocellular carcinoma cell line BEL-7402/5-fluorouracil (BEL/5-FU) compared with its parental BEL-7402 cell line, while the MDR1/P-glycoprotein expression was elevated. Overexpression of miR-27a by transfecting with miR-27a mimics in the BEL/5-FU cells could reduce the MDR1/P-glycoprotein and β-catenin expressions, enhance the sensitivity of these cells to 5-fluorouracil and 5-fluorouracil-induced apoptosis. Moreover, up-regulation of miR-27a did not decrease the FZD7 mRNA level, but significantly reduce its protein expression in BEL/5-FU cells. It was also confirmed that reduction of FZD7 by RNA interference induced inhibitory effects on the expression of MDR1/P-glycoprotein and β-catenin, similar to miR-27a. Taken together, our findings suggest that miR-27a could function as a novel regulator to reverse MDR in hepatocellular carcinoma cells by inhibiting the FZD7/β-catenin pathway.

Chen Z ，Ma T ，Huang C ，Zhang L ，Lv X ，Xu T ，Hu T ，Li J ... -  《-》

Reversal effect of quercetin on multidrug resistance via FZD7/β-catenin pathway in hepatocellular carcinoma cells.

Chemotherapy has been widely used to treat cancer, but the appearance of multidrug resistance (MDR) is the biggest obstacle to successful chemotherapy. One of the conventional mechanisms of MDR is overexpression of ATP-binding cassette (ABC) transporters such as P-glycoprotein (P-gp/ABCB1) and multidrug resistance-associated proteins (MRPs/ABCCs) that limits the prolonged and efficient use of chemotherapeutic drugs. To enhance the chemosensitivity of tumor cells, attentions have been focused on effective MDR modulators. This study aimed to investigate the reversal effect of quercetin on MDR, and explored its mechanism of action in vitro. The effect and mechanism of quercetin on MDR was examined by using MTT assay, flow cytometry, real-time PCR and western blot analysis in human hepatocellular carcinoma cells. Our data found that the intracellular accumulation of rhodamine-123 (Rh123) and doxorubicin (ADR) were increased, the sensitivity of BEL/5-FU cells to chemotherapeutic drugs were increased, and the expressions of ABCB1, ABCC1 and ABCC2 were all down-regulated, which indicated that the functions and expressions of ABCB1, ABCC1 and ABCC2 efflux pump were inhibited by quercetin treatment. Moreover, the suppression of ABCB1, ABCC1 and ABCC2 by quercetin was dependent on the FZD7 through the Wnt/β-catenin pathway. Further research revealed that reduction of FZD7 by RNA interference (siFZD7) enhanced the sensitivity to chemotherapeutic drugs, increased the cellular accumulation of Rh123 and ADR, and induced inhibitory effects on the expression of FZD7, ABCB1, ABCC1, ABCC2 and β-catenin, similar to quercetin. In the meanwhile, overexpression of FZD7 showed the inversely effect on the expressions. Interesting, it was confirmed that quercetin could inhibit the expression levels of FZD7, ABCB1, ABCC1, ABCC2 and β-catenin in BEL-7402 cells; furthermore, treatment by quercetin combined with siFZD7 in BEL/5-FU cells, the expressions of these genes were effectively decreased in comparison to quercetin combined with siRNA negative control (sncRNA). Overall, these data suggested the effectiveness of using quercetin, at least in part, via inhibiting FZD7 to combat chemoresistance and showed that quercetin could be developed into an efficient natural sensitizer for resistant human hepatocellular carcinoma.

Chen Z ，Huang C ，Ma T ，Jiang L ，Tang L ，Shi T ，Zhang S ，Zhang L ，Zhu P ，Li J ，Shen A ... -  《-》

MicroRNA-504 functions as a tumor suppressor in hepatocellular carcinoma through inhibiting Frizzled-7-mediated-Wnt/β-catenin signaling.

Accumulating evidence suggests that microRNAs (miRNAs) are critical regulators in the development and progression of various malignant tumors, including hepatocellular carcinoma (HCC). Multiple findings have indicated that miRNA-504 (miR-504) is dysregulated in several types of cancers, functioning as an oncogenic miRNA or a tumor suppressive miRNA. However, the role of miR-504 in HCC remains unknown. In this study, we aimed to detect the expression pattern of miR-504 in HCC tissues and cell lines and investigate the precise biological function in HCC cells. Our results showed that miR-504 expression levels were frequently downregulated in both HCC tissues and cell lines. Gain-of-function experiments demonstrated that miR-504 overexpression inhibited the proliferation and invasion in HCC cell lines. By contrast, miR-504 inhibition had the opposite effect. Interestingly, bioinformatics analysis predicted that Frizzled-7 (FZD7) was a potential target gene of miR-504. Dual-luciferase reporter assays confirmed that miR-504 directly targeted the 3'-untranslated region of FZD7 mRNA. In addition, our results showed that miR-504 negatively regulated the mRNA and protein expression of FZD7 in HCC cell lines. Moreover, miR-540 overexpression inhibited the cellular expression of β-catenin and blocked the activation of Wnt signaling in HCC cells. Notably, restoration of FZD7 expression significantly reversed the inhibitory effect of miR-504 on proliferation, invasion, and Wnt/β-catenin signaling in HCC cells. In conclusion, our results demonstrate that miR-504 functions as a tumor suppressive miRNA that inhibits the proliferation and invasion of HCC cells by targeting FZD7 and inhibiting Wnt/β-catenin signaling. Our study provides evidence that miR-504-meidated FZD7/Wnt/β-catenin signaling pathway plays an important role in HCC development and progression and suggests miR-504 as a novel future therapeutic target for treatment of HCC.

Quan H ，Li B ，Yang J 《-》

TFPI-2 downregulates multidrug resistance protein in 5-FU-resistant human hepatocellular carcinoma BEL-7402/5-FU cells.

Tissue factor pathway inhibitor-2 (TFPI-2) is known to induce apoptosis and to suppress tumor metastasis in several types of cancer cells. However, there is little known about its reversal effect on chemoresistant tumor cells. This study investigated the effect of TFPI-2 in 5-fluorouracil (5-FU)-resistant human hepatocellular cancer BEL-7402/5-FU cells in vitro. We constructed TFPI-2 overexpression BEL-7402/5-FU cell lines and explored resistance index (RI) of 5-FU, function of the P-glycoprotein (P-gp) efflux pump, and the mRNA and protein expression of drug resistance gene, including multidrug resistance gene (MDR1), lung-resistance protein (LRP), multidrug resistance-associated protein (MRP1), glutathione-S-transferase-π (GST-π), excision repair cross-complementing gene 1 (ERCC1), and p38 phosphorylation. We found that TFPI-2 improved the RI of 5-FU and inhibited P-gp function. Western blotting and real-time PCR revealed that TFPI-2 also decreased mRNA and protein expression of MDR1, LRP, MRP1, GST-π, and ERCC1, whereas p38 phosphorylation was increased. We considered that TFPI-2 reduces 5-FU resistance in BEL-7402/5-FU cells, and the mechanism appears to involve p38-mediated downregulation of drug resistance gene expression such as MDR1, LRP, MRP1, GST-π, and ERCC1.

Lu F ，Hou YQ ，Song Y ，Yuan ZJ ... -  《-》

miRNA-195 sensitizes human hepatocellular carcinoma cells to 5-FU by targeting BCL-w.

Yang X ，Yin J ，Yu J ，Xiang Q ，Liu Y ，Tang S ，Liao D ，Zhu B ，Zu X ，Tang H ，Lei X ... -  《-》