Western Blot Analysis of Protein Expression

The western blot analyses were performed as previously described (Liao et al., 2013 (link)). The proteins were loaded on 10% sodium dodecyl sulphate polyacrylamide gels, transferred to polyvinylidene fluoride membranes (Millipore, Billerica, MA, USA) and blocked with 5% non-fat milk powder in PBST (containing 0.1% Tween). The membranes were probed overnight with the following primary antibodies: β-actin (Cell Signalling, Beverly, MA, USA), homeodomain-containing factor A10 (HOXA10) (Santa Cruz, Dallas, TX, USA) and Forkhead box O1 (FoxO1) (Cell Signalling). The membranes were incubated with peroxidase-conjugated secondary antibody (Boster, Wuhan, China). The blots were visualized through an enhanced chemiluminescence kit (Amersham Biosciences, Piscataway, NJ, USA) according to the recommended instructions.