Fig. 5

Effect of silencing of AKT isoforms, PP1α and PP1γ on GSK3 isoforms in insulin sensitive and insulin resistant condition: Three days post-proliferation, N2A cells were transfected with A–F AKT1, AKT2, and AKT3) specific siRNA and then differentiated in 2% DMSO for 3 days. Cells were stimulated with or without 100 nM insulin for 30 min prior to cell lysis. G–J Proliferated N2a cells were transfected with non-specific (scrambled) and PP1α and PP1γ specific siRNA. Post transfection cells were differentiated in the absence (MF; insulin sensitive) or chronic presence of 100 nM insulin (MFI; insulin resistant) for 3 days. Transfected N2a cells were treated with or without 100 nM insulin for 30 min. Lysates was subjected to western blotting, followed by probing with relevant primary antibodies. Bar represents relative change in A–C, G and J pGSK3α (Ser 21) when probed with anti-GSK3α antibody. D–F, H and I Bar represents relative change in pGSK3β (Ser9) when probed with anti-GSK3β antibody. Experiments were executed three times and a representative result is shown. Data expressed are mean ± SE. ***p < 0.001 as compared to Lane 1; ^###p < 0.001, ^##p < 0.01 as compared to Lane 2, ^θθp < 0.01 as compared to Lane 4. IB Immunoblot. Open bars: MF, filled bars: MFI, IP Immunoprecipitation IB Immunoblot, SC Scrambled