Prognostic significance of c-Met, ß-catenin and FAK in patients with hepatocellular carcinoma following surgery.

The present study aimed to investigate the prognostic value of specific molecular markers in patients with hepatocellular carcinoma (HCC) who had received surgery. Immunohistochemical analysis was used to measure the expression of hepatocyte growth factor receptor (c-Met), ß-catenin and focal adhesion kinase (FAK) in patients with HCC. c-Met expression was identified to be high in patients with larger tumors, higher α-fetoprotein (AFP) levels, higher Edmondson grades, portal vein invasion and higher tumor-node-metastasis (TNM) stages. FAK expression was high in patients with portal vein invasion, higher Edmondson grades and higher TNM stages. ß-catenin expression was high in patients with larger tumors, hepatitis B virus (HBV) infection, portal vein invasion, higher Edmondson grades and higher TNM stages. Following multivariate analysis, FAK (P=0.002) and ß-catenin (P=0.006) expression levels were demonstrated to be significantly associated with Edmondson grade. Additionally, the tumor size (P=0.009) and HBV infection status (P=0.002) were revealed to be associated with ß-catenin expression. Kaplan-Meier survival curve analysis demonstrated that patients with HCC with higher FAK expression, higher ß-catenin expression, portal vein invasion, higher Edmondson grades, higher TNM stages, younger ages and higher AFP levels had significantly poorer prognoses. Cox's regression analysis revealed that the survival period was correlated with the Edmondson grade, age, AFP level, and FAK and ß-catenin expression. Univariate analysis of c-Met, ß-catenin and FAK identified a significant correlation between FAK and ß-catenin (P=0.015). Correlation analysis revealed no significant correlation between the three molecular markers, but ß-catenin and c-Met were markedly correlated (P=0.052). No significant correlation between FAK, c-Met or ß-catenin expression was identified. FAK and ß-catenin expression demonstrated a correlation with a range of clinicopathological factors, and high FAK and ß-catenin expression levels were identified to be correlated with a poor survival rate of patients with HCC. Thus, patients with higher FAK and ß-catenin expression may require more aggressive therapy. The results of the present study suggest that FAK and ß-catenin expression possess more prognostic value than c-Met expression in patients with HCC.

miR-338-3p inhibits epithelial-mesenchymal transition and metastasis in hepatocellular carcinoma cells.

Down-regulation of the miRNA miR-338-3p correlates with the invasive ability of hepatocellular carcinoma (HCC) cells. However, it is currently unclear whether down-regulation of miR-338-3p induces epithelial-mesenchymal transition (EMT), which may be the underlying mechanism governing HCC invasion. Here, we demonstrate that restoration of miR-338-3p expression via transfection of a miR-338-3p mimic reversed EMT and inhibited the motility and invasiveness of HCC cells. Conversely, silencing of endogenous miR-338-3p expression with a miR-338-3p-specific inhibitor induced EMT and enhanced HCC cell motility. Additionally, Snail1 (an upstream regulatory protein of EMT) and Gli1 (a key transcription factor in the sonic hedgehog (SHH) signaling pathway) expression was up-regulated in cells treated with the miR-338-3p inhibitor and down-regulated by the miR-338-3p mimic. Further analyses demonstrated that miR-338-3p inhibitor-induced EMT in HCC cells was blocked by treatment with a small interfering RNA (siRNA) targeting Snail1, that the SHH signaling pathway was required for both miR-338-3p inhibitor-induced EMT and up-regulation of Snail1, and that miR-338-3p targeted a sequence within the 3'-untranslated region of N-cadherin mRNA. Notably, miR-338-3p expression was significantly down-regulated in HCC samples from patients with metastases and was associated with poor metastasis-free survival rates. Lastly, correlations between the expression levels of miR-338-3p and E-cadherin, Smoothened (SMO), Gli1, Snail1, N-cadherin, and vimentin were confirmed in HCC xenograft tumors and HCC patient specimens. Our findings suggest that miR-338-3p suppresses EMT and metastasis via both inhibition of the SHH/Gli1 pathway and direct binding of N-cadherin. miR-338-3p is a potential therapeutic target for metastatic HCC.

MicroRNA-24 increases hepatocellular carcinoma cell metastasis and invasion by targeting p53: miR-24 targeted p53.

MicroRNA-24 (miR-24), a member of the miRNA family, functions as an oncogene in various types of human cancer. However, the underlying mechanisms of miR-24 involvement in the development and progression of hepatocellular carcinoma (HCC) remain poorly understood. The present study revealed that miRNA-24 down-regulates p53 through binding to the 3'-UTR of p53 mRNA based on a luciferase reporter assay, and that the expression level of miR-24 could affect the invasion of HCC lines via p53. Down-regulation of p53 significantly attenuated the inhibitory effects of miR-24 knockdown on the invasion of HCC cells, suggesting that miR-24 could be a potential target for HCC treatment. Moreover, our results revealed that miR-24 expression was significantly increased in HCC metastatic tumor tissues compared with matched non-metastatic tumor tissues, and that the up-regulation of miR-24 was significantly associated with down-regulation of p53 in the HCC tissues. In conclusion, this study demonstrates that miR-24 functions as an oncogene in HCC, at least partly by promoting cell invasion through down-regulation of p53. Therefore, miR-24 may be a potential therapeutic target for treatment of HCC.

Suppression of RNA interference pathway in vitro by Grass carp reovirus.

The means of survival of genomic dsRNA of reoviruses from dsRNA-triggered and Dicer-initiated RNAi pathway remains to be defined. The present study aimed to investigate the effect of Grass carp reovirus (GCRV) replication on the RNAi pathway of grass carp kidney cells (CIK). The dsRNA-triggered RNAi pathway was demonstrated unimpaired in CIK cells through RNAi assay. GCRV-specific siRNA was generated in CIK cells transfected with purified GCRV genomic dsRNA in Northern blot analysis; while in GCRV-infected CIK cells, no GCRV-specific siRNA could be detected. Infection and transfection experiments further indicated that replication of GCRV correlated with the increased transcription level of the Dicer gene and functional inhibition of in vitro synthesized egfp-siRNA in silencing the EGFP reporter gene. These data demonstrated that although only the genomic dsRNA of GCRV was sensitive to the cellular RNAi pathway, unidentified RNAi suppressor protein(s) might contribute to the survival of the viral genome and efficient viral replication.

[Application of nucleic acid sequence-based amplification in detection of cytomegalovirus mRNA of recipients after allogeneic peripheral blood stem cell transplantation].

The study was purposed to investigate diagnostic value of late-mRNA detection by nucleic acid sequence-based amplification (NASBA) technique for human cytomegalovirus (HCMV) infection of the recipients after allogeneic peripheral blood stem cell transplantation (allo-PBSCT) and to evaluate the clinical significance for guiding antiviral therapy. 352 samples were collected from 128 transplant patients after allo-PBSCT. A molecular biological diagnostic technique--NASBA was used to detect human cytomegalovirus (HCMV) late mRNA encoding the viral structural protein PP67 (UL65) expression in peripheral blood of recipients after allo-PBSCT, and the detected results were compared with HCMV DNA detection by PCR. The sensitivity, specificity and early diagnostic value of HCMV mRNA detection for HCMV disease were evaluated. The results showed that out of 352 detected blood specimens from 84 patients 183 specimens (51.99%) were positive of HCMV DNA by PCR, 105 specimens (29.83%) were positive of HCMV mRNA by NASBA. 45 patients were infected by HCMV. The sensitivity and specificity of HCMV DNA and HCMV mRNA for detecting HCMV disease were 95.56% (43/45), 93.33% (42/45) and 60.24% (50/83), 97.59% (81/83). The results of specificity showed significant difference between two groups of HCMV mRNA and HCMV DNA (P < 0.05). It is concluded that the detection of late-mRNA of HCMV by NASBA technique is rapid, sensitive and specific detection for HCMV active infection. The detected result correlates with clinical symptoms. It can monitor HCMV infection of allo-PBSCT transplanted recipients and provide indication to antiviral therapy.