Ski sirna

Ski specific siRNA (Ski-siRNA) and scrambled sequence siRNA (negative control siRNA, NC-siRNA) were purchased from Thermo Fisher Scientific. The target sequence of Ski-siRNA was 5′-CGGACCTTGGCTGGTTCCTCCAATA-3′, and the sequence of 5′-TTCTCCGAACGTGTCACGT-3′ was considered to be NC-siRNA. The MG-63 and U2OS cells were transfected with Ski-siRNA or NC-siRNA using Lipofectamine^® RNAiMAX Reagent (Thermo Fisher Scientific) in accordance with the manufacture's protocol. Following 48 h of transfection, the cells were collected for the used in the following experiments. Western blot analyses were used to assess the silencing effect of Ski-siRNA. The cells were pre-treated with basic culture medium DMEM/F12 with or without 50 µM LY294002 (a specific PI3K inhibitor, S1737, Beyotime) for 6 h prior to transfection.