In vitro investigation of possible biomarkers of chemoresistant hepatocellular carcinoma cells

Hepatocellular carcinoma (HCC) is the second-leading cause of cancer-related deaths worldwide. Complete EMT has long been considered as a crucial step for initiating metastasis. However, it is now known that many carcinoma cells can metastasize with incomplete loss of epithelial traits or incomplete gain of mesenchymal phenotype, which is named as partial EMT. Here, in order to determine the similarities and differences between complete and partial EMT phenotype in HCC, the EMT associated transcription factor Slug was overexpressed in different HCC cell lines and the cells were compared for their EMT phenotype, chemoresistance and stemness. Conditioned medium was also collected from a HCC cell line overexpressing Slug and was analyzed for markers of chemoresistance and partial EMT. We observed that Slug overexpression resulted in the induction of both complete and partial EMT in different HSK cell lines. Complete EMT was characterized by the down regulation of E-cadherin and up-regulation of ZEB2 mRNA. Partial EMT was characterized by the up-regulation of E-cadherin and down-regulation of vimentin and ZEB2 mRNA. Interestingly, Slug mediated chemoresistance through the down-regulation of ABCB1 and up-regulation of ABCG2 mRNAs as well as expression of CD133, a stemness marker was similar in both complete and partial EMT phenotype. On the other hand, partial EMT was associated with enhanced secretion of post translationally modified fibronectin1, collagen type II alpha 1 and native fibrinogen gamma chain upregulations in the conditioned medium of HSK cells. These proteins can therefore be considered as potential biomarkers of chemoresistance and partial EMT in HSK. Similarly, in the conditioned medium, hsa-miR-296-3p, hsa-miR-101-3p, hsa-miR-885-5p, miRNAs were found to be up regulated respectively. It has been suggested that these microRNAs can also be used as chemoresistance and partial EMT biomarkers in HCC.