| Talinl is a macromolecular adaptor protein that conjugates cell adhesion molecules(such as integrins.FAK and vinculin)to the actin cytoskeleton and regulates integrins and focal adhesion signaling.Several studies have found that Talinl is over-expressed in several tumor types and promotes tumor progression,including prostate tumor,oral squamous cell carcinoma,glioblastoma multiforme and ovarian serous carcinoma.However,the explicit role of Talin1 in initation and progression of HCC is still unclear and its functional mechanism remains largely unknown.In this study,we detected the expression levels of Talinl on a tissue microarray of HCC progression by immunohistochemical analysis.We found that the high expression rate of Talinl was 100%(5/5)in normal liver,83.3%(5/6)in hepatocirrhosis,100%(2/2)in liver hyperplasia,83.3%(15/18)in the corresponding adjacent non-tumor,42.1%(8/19)in primary HCC and 14.3%(1/7)in metastatic foci.These results suggested that the expression of Talinl was decreased gradually through the initiation and progression of HCC.Meanwhile,Talinl was significantly down-regulated in tumor tissues(P<0.05)and metastatic foci(P<0.01)compared with non-tumor tissues.Furthermore.we detected the expression of Talin1 in 196 adjacent non-tumor tissues and 200 HCC tissues by immunohistochemical analysis.We found that 67.5%of HCC tissues showed low Talin1 expression,whereas only 7.2%of adjacent non-tumor tissues showed low Talin1 expression(P<0.001).Pearson' s chi-square test showed that Talinl down-regulation significantly correlated with younger age(P = 0.032),larger tumor size(P = 0.006),higher alpha-fetoprotein(AFP)levels(P = 0.003)and tumor recurrence(P<0.001).Kaplan-Meier and log-rank test analysis revealed that patients with low Talin1 expression exhibited worse overall survival(P<0.001)and disease-free survival rates(P = 0.001)than those with high Talinl expression.The univariate and multivariate analysis demonstrated that Talin1 expression(P = 0.010)was independent prognostic factor for overall survival in HCC patients.We found by western blot analysis that higher Talin1 levels were found in the HCC cell lines with low-malignancy(SK-Hep-1/HepG2)rather than in the more aggressive HCC cell lines(SMCC-7721/MHCC97H/HCCLM3).Hence,SK-Hep-1 and HepG2 cells were selected for Talin1 knockdown by using small interfering RNAs(siRNAs).We found that a dramatic morphological change emerged in Talin1-knockdown SK-Hep-1 and HepG2 cells,with polygonal or oval cells becoming spindle-like fibroblastic cells,which is regarded as a main characteristic of EMT.At the molecular level,a decrease of the epithelial marker E-cadherin and an increase of mesenchymal markers N-cadherin and vimentin,was also observed in Talin1-knockdown cells.Meanwhile,silencing of Talinl in SK-Hep-1 cells promoted the growth of lamellipodia.Transwell migration and invasion assays showed that Talin1-knockdown HCC cells exhibited a significant increase of migration and invasion capacities compared with the corresponding control cells(All P<0.05).Mechanistically,we found by screening several pathways that phosphorylated ERK1/2 was significantly elevated in Talin1-knockdown cells,while phosphorylated smad2/3,phosphorylated AKT,RhoA and Rac1 exhibited no obvious changes.The U0126 is a specific ERK1/2 inhibitor.We found that the promoting effects of Talinl knockdown on EMT,migration,and invasion were reversed by the U0126.Above all,these results demonstrated that inhibition of Talinl can act as a positive regulator of EMT,migration and invasion in HCC cells via activation of the ERK1/2 pathway.Taken together,our results suggested that Talinl might serve as a tumor suppressor in HCC and a potential prognostic biomarker for HCC patients. |
PDF Full Text Request