The role of microRNAs in in vitro cellular models of colorectal and pancreatic cancer

Based on the previous findings, this study aims to examine the function of microRNA-486-5p in the context of colorectal cancer and microRNA-216a-5p in pancreatic adenocarcinoma. In particular, the role of miRNAs was examined in two in vitro cell culture models: a two-dimensional and a three-dimensional model CSC-enriched. The HT-29 line, representative of the primary tumor, and the T84 line, representative of the metastatic tumor, were utilized as adherent models of colorectal cancer (CRC) and to obtain two models of colorectal cancer stem cells (CCRCs). The BxPC-3 line, representative of the primary tumor, and the AsPC-1 line, representative of the metastatic tumor, were utilized as adherent models of PDAC and to obtain two models of pancreatic cancer stem cells (PaCSCs). To study the effect of miRNAs, the overexpression and inhibition were simulated in all culture models by transfecting synthetic miRNA mimics and miRNA inhibitors. Subsequently, differences in the expression of stemness and EMT markers and clonogenic activity were evaluated following the transfection to elucidate the effect of these miRNAs at the phenotypic and functional levels on the obtained CCSCs and PaCSCs. Results demonstrated that miR-486-5p in CRC models and miR-126a-5p in PDAC models effect on stemness- and EMT reated genes changed depending on the cell lines and culture condition. The expression of miR-486-5p was found to be significantly reduced in CCSCs in comparison to adherent culture models. Furthermore, the overexpression of this microRNA resulted in a notable reduction in colony size. These findings indicate that miR-486-5p functions as an onco-suppressor in CRC, thus enhancing our comprehension of the role of this microRNA in the CCSC phenotype. The results obtained in the study conducted in PDAC models confirmed the upregulation of miR126a-5p in PaCSCs culture models in comparison to adherent culture models. Moreover, it demonstrated a stimulatory effect on the number and size of colonies formed by both cell lines, as compared to colonies where the miRNA was inhibited. The functional study results align with our previous hypothesis regarding the potential dual behavior of miR-216a-5p in PDAC.