Figure 7.

Partial rescue of GH-induced STAT5 signaling by an IGF-IR lacking the intracellular domain of its β-chain. A, Upper panel, Floxed-IGF-IR primary osteoblasts were infected with either Ad-GFP only (1600 MOI), Ad-GFP plus Ad-Cre (800 MOI each simultaneously), or Ad-IGF-IR_Δ950 plus Ad-Cre (800 MOI each simultaneously). Serum-starved cells were treated with vehicle or GH (250 ng/ml) for 10 min, after which detergent extracts were resolved by SDS-PAGE and sequentially immunoblotted with anti-pSTAT5, anti-STAT5, and anti-IGF-IRα. Lower panel, Densitometric quantitation of pSTAT5 signals from three independent experiments. In each experiment, the maximum signal was considered 100%. Data are plotted as mean ± se. P values are indicated. B, The experiment in A was repeated, except that Ad-IGF-IR_Δ950 was used at 8, 80, or 400 MOI. Note that rescue of GH-induced STAT5 activity was afforded by Ad-IGF-IR_Δ950, despite the reduced level of expression of the truncation mutant achieved with 8–400 MOI compared with 800 MOI used in A. C, Floxed IGF-IR primary osteoblasts were infected as in A. Serum-starved cells were treated with vehicle or GH (50 ng/ml) for 6 h. Total RNA was extracted, and quantitative real-time PCR for IGF-I (normalized for β-actin) was performed. Pooled data from two separate experiments (one in triplicate and one in duplicate) are represented as mean ± se. In each case, the value of the vehicle-treated samples is considered 100%. P values are indicated.