and effects of miRNA-19b/20a/92a on gastric cancer stem cells and the related mechanism.

We aimed to analyze the and effects of miRNA-19b/20a/92a on gastric cancer stem cells (GCSCs) and the related mechanism. GCSCs were cultured until adherence and differentiation, and subjected to miRNA microarray analysis to find and to verify miRNA deletion. Cells stably expressing lentivirus carrying miRNA-19b/20a/92a were constructed by transfection. The relationship between miRNA-19b/20a/92a and renewal of GCSCs was studied by the tumor sphere assay, and that between miRNA-19b/20a/92a and their proliferation was explored with MTT and colony formation assays. Target genes of miRNA for promoting the proliferation and self-renewal of GCSCs were found by using bioinformatics database, and verified by the reporter gene assay and Western blot. The expressions of miRNA-19b/20a/92a gradually decreased during the adherence and differentiation of GCSCs. The expressions of lentivirus carrying miRNA-17-19 gene in MKN28 and CD44/EpCAM cells were increased significantly. Transient transfection with pre-miRNA-19b/20a/92a elevated miRNA expressions in CD44/EpCAM and MKN28 cells, whereas transfection with pre-miRNA-19b/20a/92a antagonists reduced the expressions in SGC7901 and CD44/EpCAM cells. Overexpression of lenti-miRNA-19b/20a/92a significantly enhanced the capability of GCSCs to form tumor spheres. In the presence of chemotherapeutic agent, the survival of lenti-miRNA-19b/20a/92a-infected cells was prolonged. Transient transfection with pre-miRNA-19b/20a/92a significantly increased the number of CD44/EpCAM cells, but transfection with antagonists had the opposite outcomes. The stable miRNA-19b/20a/92a expression groups proliferated faster than the control group did. The proliferation of cells transfected with pre-miRNA-19b/20a/92a was accelerated, whereas that of cells transfected with the antagonists was decelerated. Compared with the control group, the number of colonies in the former group was higher, but that in the latter group was lower. miRNA-19b and miRNA-92a could bind the 3' untranslated region of HIPK1, while miRNA-20a was able to bind that of E2F1. Expressions of miRNA-20a and miRNA-92a in gastric cancer samples were negatively correlated with the prognosis of patients. miRNA-19b/20a/92a facilitated the self-renewal of GCSCs by targeting E2F1 and HIPK1 on the post-transcriptional level and activating the β-catenin signal transduction pathway. miRNA-92a was an independent factor and index predicting the prognosis of gastric cancer.