Role of HSA-MIR-141-3P and HSA-MIR-200B-3P in gastrointestinal stromal tumor pathogenesis

Abstract

Introduction. Gastrointestinal stromal tumor (GIST) is the most common mesenchymal tumor of the gastrointestinal tract. It is now known that the main event in GIST tumorigenesis is mutations in KIT or PDGFRA, which lead to uncontrolled cell growth and tumor formation. However, it has been determined that changes in gene expression are responsible for the gradual malignant transformation in GIST, but, little is known about the underlying mechanisms regulating these expression signatures. MicroRNAs (miRNAs) are a class of small non-coding RNAs involved in posttranscriptional regulation of gene expression. Various studies have shown that expression deregulation of miRNAs has a crucial role in cancer and is associated with tumorigenesis, tumor progression, metastasis and may have influence in tumor drug resistance. Biological functional determination of specific miRNAs and their target genes could contribute to understanding of cancer pathogenesis and may improve its diagnostics or even treatment. AIM The aim of the study was to assess the functional role of microRNAs (hsa-miR-141-3p and hsa-miR- 200b-3p) in gastrointestinal stromal tumor pathogenesis. Methods. GIST-T1 cell line was used as a model system for this study. miRNAs of interest were chosen from previous non-coding RNA profiling study (2). Cells have been transfected using two miRNAs mimics (at concentrations 80-100 nM): hsa-miR-141-3p and hsa-miR-200b-3p. MTT assay was used for assessment of cell viability 72 h after transfection. „Wound Healing“ assay was used to determine the migration of cells after transfection. Further, target genes of miRNAs were chosen with TargetScan 7: hsa-miR-141-3p: STAT1, STAT5A, STAT5B, EGFR, KIT and PDGFRA, for hsa-miR-200b-3p: STAT1, EGFR and ETV1. Their expression was validated using qRT-PCR (internal control - GAPDH) and analysed using 2-ΔΔCt method. Finally, protein expression of targets was analyzed using Wester [...].