Figure 6. Activation of IR signalling in 3T3-L1 adipocytes.

(A) Top panel: 3T3-L1 pre-adipocytes before and after differentiation into adipocytes were incubated with vehicle or 100 nM insulin at 37 °C for 15 min as described in the Experimental section. Adipocyte differentiation was determined by Western blotting of equal amounts of soluble protein with Abs directed against IRβ or GLUT4. Bottom panel: equal amounts of soluble protein from adipocytes were used for Western blotting with Abs specific for the phosphorylated forms of IRβ, IRS-1, Akt, GSK3β and ERK. The blots are representative of three independent experiments. Anti-pY, anti-pTyr. (B) Differentiated 3T3-L1 adipocytes were incubated in the presence or absence of 1 μg/ml CG7 for 15 min at 37 °C. Cells were also incubated simultaneously with 1 nM insulin in the presence or absence of CG7. Cells incubated with 100 nM insulin served as controls. Equal amounts of soluble proteins from each preparation were Western blotted with Abs specific for the phosphorylated forms of IRβ, Akt and GSK3β. Blots were stripped and re-probed with Ab directed against IRβ. The data are one representative for three separate experiments. IB, immunoblot.