FIGURE 4.

Effects of IRS-2 or myr-Akt overexpression on glucose uptake. 3T3-L1 adipocytes were infected with adenovirus containing cDNA encoding LacZ or IRS-2. Forty eight hours after infection, cells were serum-starved for 2 h and then pretreated with or without 100 nm GH for 24 h, followed by stimulation with or without 0.1 nm insulin for 20 min. A, cells were solubilized with Tris/Triton lysis buffer. Whole cell lysates were separated by SDS-PAGE and immunoblotted (IB) with anti-phospho-Akt (Ser-478) antibody and anti-Akt antibody. B, cells were assayed for glucose uptake as described under “Experimental Procedures.” The results are presented as the means ± S.E. of five wells. The difference between insulin-stimulated cells with and without GH pretreatment is significant with p < 0.01 (*). C, 3T3-L1 adipocytes were infected with adenovirus containing cDNA encoding LacZ or myristoylated Akt (myr-Akt). Forty eight hours after infection, cells were serum-starved for 2 h, then pretreated with or without 100 nm GH for 24 h followed by stimulation with or without 0.1 nm insulin for 20 min, and assayed for glucose uptake as described under “Experimental Procedures.” The results are presented as the means ± S.E. of five wells. The difference between insulin-stimulated cells with and without GH pretreatment is significant with p < 0.01 (*).