Insulin-like growth factor II induces apoptosis in osteoblasts.
The ability of insulin-like growth factor II (IGF-II) to modulate apoptosis was studied in murine osteoblasts. At 72 h of culture, 0.01, 0.1, and 1.0 nM IGF-II produced a dose-dependent increase in apoptosis assayed by TdT-mediated dUTP-biotin nick end labeling (TUNEL) and confirmed with acridine orange-ethidium bromide staining. A maximal increase of 5.0-fold above control was found with 1 nM IGF-II. A time course of treatment with 0.1 nM IGF-II demonstrated a significant increase in apoptosis compared to vehicle-treated cells by 48 h. IGF-II-induced apoptosis could not be inhibited by a blocking antibody to the IGF-I receptor. Human osteoblast cultures demonstrated a similar dose-dependent increase in apoptosis with IGF-II. No significant effect of IGF-II was found on proliferation in murine osteoblast cultures. Western blot analysis demonstrated that IGF-II decreased Bcl-2 protein levels, but not Bax, resulting in a significant reduction in the Bcl-2/Bax ratio. To determine if overexpression of Bcl-2 could block IGF-II-induced apoptosis, osteoblasts were isolated from a transgenic mouse that overexpresses human Bcl-2 in bone through a construct utilizing the 2.3 kb promoter region of the Type I collagen gene linked to a 1.8 kb region of human Bcl-2 (Col2.3Bcl-2). At 72 h, IGF-II significantly increased apoptosis in a dose-dependent manner in osteoblast cultures from the control littermates. In osteoblasts from Col2.3Bcl-2 mice, no significant effect on apoptosis was found with 0.01, 0.1, or 1.0 nM IGF-II. Western blot analysis of Bcl-2 and Bax levels demonstrated a transient decrease in the Bcl-2/Bax ratio at 24 h with no decrease in the ratio at 48 or 72 h. Thus, IGF-II appears to promote osteoblast apoptosis, and overexpression of Bcl-2 is able to block IGF-II-induced apoptosis.
Gronowicz
Gloria A
GA
Department of Orthopaedic Surgery, University of Connecticut Health Center, Farmington, CT 06030, USA. gronowicz@nso1.uchc.edu
McCarthy
Mary-Beth
MB
Zhang
Hai
H
Zhang
Wenjian
W
D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D002452:Cell Count / Q000379:methods; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D006801:Humans; D007335:Insulin-Like Growth Factor II / Q000494:pharmacology*; D051379:Mice; D008822:Mice, Transgenic; D010006:Osteoblasts / Q000166:cytology / Q000187:drug effects* / Q000502:physiology
D007335:Insulin-Like Growth Factor II