AXIN2 gene silencing reduces apoptosis through regulating mitochondria-associated apoptosis signaling pathway and enhances proliferation of ESCs by modulating Wnt/β-catenin signaling pathway.

OBJECTIVE:Embryonic stem cells (ESCs) mainly originate from totipotent cells in early-stage of mammalian embryo and could proliferate in a manner of un-limitation. This study aimed to investigate roles of Axin2 in proliferation of ESCs and explore the associated mechanisms. MATERIALS AND METHODS:Axis inhibition protein 2 (AXIN2) over-expression (LV5-AXIN2) and AXIN2 RNA interfere vectors were structured and transfected into H9 cells. 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) was used to evaluate cell proliferative activity. Flow cytometry analysis was employed to measure apoptosis of H9 cells. AXIN2, β-catenin, transcription factor 4 (TCF4), c-myc, c-jun and Cyclin D mRNA levels and protein expressions were determined using quantitative real-time PCR (qRT-PCR) and Western blotting assay. RESULTS:LV5-AXIN2 and were successfully structured with higher transfecting efficacy. AXIN2 gene silencing remarkably increased proliferative activity and AXIN2 treatment significantly induced apoptosis of H9 cells, comparing with blank vector group (p<0.05). AXIN2 gene silencing significantly enhanced B-cell lymphoma-2 (Bcl-2) expression and remarkably inhibited cleaved caspase-3 expression comparing to that in blank vector group (p<0.05). treatment significantly enhanced and AXIN2 over-expression significantly reduced β-catenin and TCF4 expression, comparing to that in blank vector group (p<0.05). AXIN2 gene silence activated down-stream molecules of Wnt/β-catenin signaling pathway, including c-jun, c-myc, and Cyclin D1 (p<0.05). CONCLUSIONS:AXIN2 gene silencing reduced apoptosis by regulating mitochondria-associated apoptosis signaling pathway and enhanced proliferation by modulating molecules in Wnt/β-catenin signaling pathway. Therefore, targeting of aberrant apoptosis and AXIN2 might be a novel clinical strategy to inhibit aging and enhance self-renewal of ESCs.

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