IGF-I inhibits apoptosis induced by serum withdrawal, but potentiates TNF-alpha-induced apoptosis, in 3T3-L1 preadipocytes.
We have previously shown that human preadipocytes in primary culture undergo apoptosis in response to serum deprivation and addition of tumour necrosis factor alpha (TNF-alpha), and have proposed that regulation of preadipocyte apoptosis in vivo may contribute to the overall control of adipose mass. In the present study we have investigated both pro- and anti-apoptotic factors, and the signalling pathways by which they act, in murine 3T3-L1 preadipocytes. Apoptotic indices (fraction of cells undergoing apoptosis) were determined by microscopic examination of acridine orange-stained cells, fluorescence-activated cell sorting of propidium iodide-stained cells, or phase-contrast video microscopy. Murine 3T3-L1 cells were more susceptible to apoptosis than human preadipocytes. In medium containing 10% newborn calf serum, the basal apoptotic index was very low (<2%), but the number of apoptotic cells increased significantly following serum withdrawal (10% after 24 h). Addition of TNF-alpha (6 nM) stimulated apoptosis in both serum-containing and serum-free media (apoptotic indices of 12% and 20% respectively after 24 h). IGF-I inhibited by approximately 50% the apoptosis induced by serum withdrawal, but increased by 25% the apoptosis induced by TNF-alpha in serum-free medium. It was shown by using specific inhibitors of lipid and protein kinases (LY294002, rapamycin, PD98059, SB203580) that both phosphoinositide 3-kinase and MAP kinase pathways contribute to the anti-apoptotic action of IGF-I on serum-starved cells, while phosphoinositide 3-kinase but not MAP kinase activity is required for the paradoxical pro-apoptotic action of IGF-I in the presence of TNF-alpha. We conclude that, in addition to its previously described anti-apoptotic action, IGF-I can also be pro-apoptotic in 3T3-L1 cells in the presence of TNF-alpha, and that both the anti- and pro-apoptotic effects of IGF-I require the activation of phosphoinositide 3-kinase.
Niesler
C U
CU
Department of Clinical Biochemistry, Addenbrooke's Hospital, University of Cambridge, Cambridge CB2 2QR, UK.
Ursø
B
B
Prins
J B
JB
Siddle
K
K
D016475:3T3 Cells; D017667:Adipocytes / Q000166:cytology / Q000187:drug effects*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D016895:Culture Media, Serum-Free; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology*; D020935:MAP Kinase Signaling System / Q000502:physiology; D051379:Mice; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology; D011994:Recombinant Proteins / Q000494:pharmacology; D015398:Signal Transduction / Q000502:physiology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D016895:Culture Media, Serum-Free; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D007334:Insulin-Like Growth Factor I; D019869:Phosphatidylinositol 3-Kinases