MiR-501-5p regulates CYLD expression and promotes cell proliferation in human hepatocellular carcinoma.

OBJECTIVE: Previous studies have shown that the micro-ribonucleic acid miR-501-5p is upregulated in hepatocellular carcinoma cells and tissues with high hepatitis B virus replication, and that miR-501 overexpression significantly promotes hepatitis B virus replication. We further analysed a published microarray-based high-throughput dataset (NCBI/GEO/GSE36915) and found that miR-501-5p was upregulated in hepatocellular carcinoma tumour tissues. We therefore investigated the possible function of miR-501-5p during the development of hepatocellular carcinoma. METHODS: Expression of miR-501-5p in human hepatocellular carcinoma specimens and cell lines was assessed, using real-time polymerase chain reaction. Luciferase reporter assays were used to confirm CYLD as a target of miR-501-5p. The effect of miR-501-5p on cell proliferation was confirmed, using tetrazolium and colony formation assays. Gene and protein expression were examined, using real-time polymerase chain reaction and western blotting, respectively. RESULTS: MiR-501-5p was upregulated in hepatocellular carcinoma specimens and cell lines, and directly targeted the 3' untranslated region of CYLD. MiR-501-5p upregulation corresponded with a downregulation of CYLD in the same tissues and cell lines, and overexpression of MiR-501-5p decreased CYLD expression, increased expression of cyclin D1 and c-myc and promoted the proliferation of hepatocellular carcinoma cells in vitro. CONCLUSIONS: This study suggests that miR-501-5p may play an important role in the development of hepatocellular carcinoma by promoting cell proliferation, and indicates that miR-501-5p may represent a novel therapeutic target for hepatocellular carcinoma.

[Effect of pectins of different degree of esterification on in-vitro sophoridine release of hydrophilic matrix tablets containing total alkaloids of Sophora alopecuroides].

OBJECTIVE: To prepare colon-targetting tablets of total alkaloids of Sophora alopecuroides and evaluate the effect of pectins of different degree of esterification (DE) on sophoridine release profiles in-vitro. METHOD: Wet granulation technique was employed to prepare petin-based matrix tablets, then tablets were coated the optimal formulation with Kollicoat MAE 30 DP based on the optimal formulation and analysed their release. RESULT: Coated formulation E could target total alkaloids of S. alopecuroides to colon and various DE of pectin exerted different effects on sophoridine release. The release of low DE pectin-based matrix tablets coating with Kollicoat MAE 30 DP approximatedly fitted zere-order eqution, which was erosion depended. CONCLUSION: Low DE pectin-based matrix tablet coating with Kollicoat MAE 30 DP can deliver sophoridine to colon, hence improve the effectiveness of sophoridine.

RNA interfering targeting human leukocyte antigen-G enhanced immune surveillance mediated by the natural killer cells on hepatocellular carcinoma.

BACKGROUND: Up-regulation of Human Leukocyte Antigen-G (HLA-G) expression is thought to contribute to the escape in immune surveillance by suppressing natural killer (NK) cell function. However, little is known about the expression of HLA-G in hepatocellular carcinoma (HCC) and its relationship to NK cell-mediated cytotoxicity. In this study, we aimed to investigate the expression of HLA-G in human HCC cell lines and determine whether its expression was related to inhibition of NK cell cytolysis. MATERIALS AND METHODS: The levels of HLA-G gene expression in human HCC cell lines were assessed by indirect immunofluorescence analysis (IFA), Real time RT-PCR and Western Blot. Vectors containing small interfering RNA (siRNA) specifically targeting the HLA-G gene were constructed and applied to diminish HLA-G expression. The cells were examined by flow cytometry and fluorescent microscope 24 h after transfection as well as 2-3 weeks after G418 selection. The steady-state levels of HLA-G mRNA and protein were then checked by real time RT-PCR and Western Blot analysis, respectively. A nonradioactive cytotoxicity assay was used to evaluate the effect of HLA-G on NK-mediated cytotoxicity against the siRNA treated cells. RESULTS: Both HLA-G mRNA and protein can be detected in human HCC cell lines. Levels of HLA-G mRNA and protein were diminished 88.73% and 75.91% respectively by targeting siRNA. In the stable HLA-G gene knock-down cell lines, a significant increase in NK cell-mediated lysis occurred. CONCLUSIONS: Abnormal expression of HLA-G in HCC cell lines plays an important role in protecting against NK cell attack. The significant correlation between HLA-G expression and NK cell lysis implies that the abnormal expression of HLA-G might contribute to the mechanism of escape from host immune surveillance in HCC.