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ABSTRACT: Glypican-3 (GPC3) is a membrane-associated heparan sulfate proteoglycan involved in regulation of cell proliferation, cell survival, cell migration and differentiation process. MicroRNAs (miRNAs) are single-stranded, noncoding functional RNAs that are important in many biological processes. GPC3 and miRNAs have been found to play essential roles in the development and progression of hepatocellular carcinoma (HCC). However, little information about the relationship between GPC3 and miRNAs is available nowadays. In the present study, we demonstrated the relationship between GPC3 and miRNAs. The results of dual-luciferase reporter assay showed that miR-520c-3p could target GPC3. The results of fluorescence quantitative PCR and Western blot analysis indicated that GPC3 protein levels decreased with unchanged transcription efficiency after miRNA transfection, and there was negative correlation of miR-520c-3p expression in HCC in relate to GPC3 protein levels. Moreover, Annexin V-FITC, Edu and transwell assays showed that miR-520c-3p not only could induce HCC cell apoptosis but also could inhibit the growth and invasion of the HCC cells. Interestingly, overexpression of GPC3 could effectively reverse apoptosis induced by miR-520c-3p transfection in HCC. Taken together, these results supported that miR-520c-3p may decrease GPC3 protein levels to inhibit proliferation of HCC cells. Therefore, GPC3 could be a new target for genetic dianosis and treatment of HCC.
Hepatology Research 04/2013; · 2.20 Impact Factor
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ABSTRACT: Glypican-3 (GPC3), a membrane-associated heparan sulfate proteoglycan, is frequently upregulated in hepatocellular carcinoma (HCC). Yes-associated protein (YAP) is also found over-expressed in HCC and has been identified as a key effector molecule in Hippo pathway, which could control the organ size in animals through the regulation of cell proliferation and apoptosis and plays an important role in the development of malignant tumors. Studies have reported that GPC3 and YAP might collaborate to regulate the development of HCC. To elucidate the role of GPC3 in the development of HCC and its relationship with YAP, siRNA technique was employed to knock down GPC3 in Huh7 HCC cells. Moreover, recombinant human YAP-1 was used to examine the effects of GPC3 on Huh7 cells. The results of flow cytometric analysis and Annexin-V-FLUOS apoptosis assay showed that knockdown of GPC3 induced apoptosis in Huh7 cells, resulting in inhibition of cell proliferation as examined by EdU incorporation assay, migration and invasion. GPC3 knockdown also suppressed the expression of YAP in mRNA and protein levels, as examined by fluorescence quantitative PCR and Western blot analysis. Moreover, addition of recombinant human YAP-1 effectively rescued the cells from apoptosis triggered by GPC3 knockdown. Taken together, our findings suggest that GPC3 regulates HCC cell proliferation with the involvement of Hippo pathway. J. Cell. Biochem. © 2012 Wiley Periodicals, Inc.
Journal of Cellular Biochemistry 10/2012; · 2.87 Impact Factor
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ABSTRACT: To investigate the expression of myeloid differentiation protein-2 (MD-2), MD-2B (a splicing isoform of MD-2 that can block Toll-like receptor 4 (TLR4)/MD-2 LPS-mediated signal transduction) and TLR4 in the liver of acute cholangitis rats.
Male Sprague-Dawley rats (SPF level) were randomly divided into four groups: (A) sham-operated group; (B) simple common bile duct ligation group; (C) acute cholangitis group; and (D) acute cholangitis anti-TLR4 intervention group (n = 25 per group). Rat liver tissue samples were used to detect TLR4, MD-2 and MD-2B mRNA expression by fluorescence quantitative PCR in parallel with pathological changes.
In acute cholangitis, liver TLR4 and MD-2 mRNA expression levels at 6, 12, 24, 48 and 72 h were gradually up-regulated but MD-2B mRNA expression gradually down-regulated (P < 0.05). After TLR4 antibody treatment, TLR4 and MD-2 mRNA expression were lower compared with the acute cholangitis group (P < 0.05). However, MD-2B mRNA expression was higher than in the acute cholangitis group (P < 0.05). MD-2 and TLR4 mRNA expressions were positively correlated (r = 0.94981, P < 0.05) and MD-2B mRNA expression was negatively correlated with MD-2 and TLR4 mRNA (r = -0.89031, -0.88997, P < 0.05).
In acute cholangitis, MD-2 plays an important role in the process of TLR4- mediated inflammatory response to liver injury while MD-2B plays a negative regulatory role.
World journal of hepatology. 06/2010; 2(6):233-8.
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ABSTRACT: To investigate the effect of arsenic trioxide (As2O3) on expression of anti-oncogene RAS association domain family gene 1A(RASSF1A) in nasopharyngeal carcinoma cell line CNE-2Z.
CNE-2Z cells were treated with various concentrations of As2O3 for different times. The IC(50) values were detected by trypan blue stain assay. Cell cycle redistribution was analyzed by flow cytometry. The final concentration 2 micromol/L, 1 micromol/L, 0.5 micromol/L of As2O3 was added to CNE-2Z cell for succedent experiments. The controls and no drugs of CNE-2Z cells were cultivated for 48 h. Methylation specific PCR was used to detect the change of methylation status of RASSF1A gene. The expression of RASSF1A gene was detected by reverse transcription PCR and Western blot at mRNA and protein level.
The suppression of cell proliferation by As2O3 was time and dose-dependent. After being treated with As2O3, the IC(50) values of As2O3 were (1.50 +/- 0.05), (1.09 +/- 0.13), (0.65 +/- 0.04) micromol/L at 24, 48, and 72 h, respectively. As2O3 also arrested CNE-2Z cells in G2/M phase of cell cycle. After the effect of As2O3, the methylation of RASSF1A gene became weaker by increasing the concentration of As2O3; and the expression of RASSF1A gene became stronger at mRNA and protein level. Between different concentration of As2O3 group and no drugs group, the differences had statistical significance (P < 0.05). Along with increasing the concentration of As2O3, the expression of RASSF1A gene became stronger at mRNA and protein level, the methylation of RASSF1A gene became weaker and weaker.
As2O3 can activate the expression of RASSF1A gene to inhibit the cell cycle progress of nasopharyngeal carcinoma cell line.
Zhonghua er bi yan hou tou jing wai ke za zhi = Chinese journal of otorhinolaryngology head and neck surgery 10/2009; 44(10):866-70.
