Bone morphogenetic protein-2 promotes osteoblast apoptosis through a Smad-independent, protein kinase C-dependent signaling pathway.
Bone morphogenetic protein-2 (BMP-2), a member of the transforming growth factor-beta (TGF-beta) family, regulates osteoblast differentiation and bone formation. Here we show a novel function of BMP-2 in human osteoblasts and identify a signaling pathway involved in this function. BMP-2 promotes apoptosis in primary human calvaria osteoblasts and in immortalized human neonatal calvaria osteoblasts, as shown by terminal deoxynucleotidyl transferase-mediated nick end labeling analysis. In contrast, TGF-beta 2 inhibits apoptosis in human osteoblasts. Studies of the mechanisms of action showed that BMP-2 increases the Bax/Bcl-2 ratio, whereas TG beta-2 has a negative effect. Moreover, BMP-2 increases the release of mitochondrial cytochrome c to the cytosol. Consistent with these results, BMP-2 increases caspase-9 and caspase-3, -6, and -7 activity, and an anti-caspase-9 agent suppresses BMP-2-induced apoptosis. Overexpression of dominant-negative Smad1 effectively blocks BMP-2-induced expression of the osteoblast transcription factor Runx2 but not the activation of caspases or apoptosis induced by BMP-2, indicating that the Smad1 signaling pathway is not involved in the BMP-2-induced apoptosis. The proapoptotic effect of BMP-2 is PKC-dependent, because BMP-2 increases PKC activity, and the selective PKC inhibitor calphostin C blocks the BMP-2-induced increased Bax/Bcl-2, caspase activity, and apoptosis. In contrast, the cAMP-dependent protein kinase A inhibitor H89, the p38 MAPK inhibitor SB203580, and the MEK inhibitor PD-98059 have no effect. The results show that BMP-2 uses a Smad-independent, PKC-dependent pathway to promote apoptosis via a Bax/Bcl-2 and cytochrome c-caspase-9-caspase-3, -6, -7 cascade in human osteoblasts.
Haÿ
E
E
Laboratory of Osteoblast Biology and Pathology, INSERM U 349, Affiliated CNRS, Lariboisière Hospital, 75475 Cedex 10 Paris, France.
Lemonnier
J
J
Fromigué
O
O
Marie
P J
PJ
D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000494:pharmacology*; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D050659:Core Binding Factor Alpha 1 Subunit; D003574:Cytochrome c Group / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005047:Etoposide / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D020287:In Situ Nick-End Labeling; D007231:Infant, Newborn; D007546:Isoquinolines / Q000494:pharmacology; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009281:Naphthalenes / Q000494:pharmacology; D009363:Neoplasm Proteins; D010006:Osteoblasts / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D011493:Protein Kinase C / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D011994:Recombinant Proteins / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D012886:Skull / Q000166:cytology; D051785:Smad Proteins; D051898:Smad1 Protein; D013449:Sulfonamides; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D051028:bcl-2-Associated X Protein
C494826:BAX protein, human; C525718:BMP2 protein, human; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; D050659:Core Binding Factor Alpha 1 Subunit; D003574:Cytochrome c Group; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D005419:Flavonoids; D007546:Isoquinolines; D009281:Naphthalenes; D009363:Neoplasm Proteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D011994:Recombinant Proteins; C497399:SMAD1 protein, human; D051785:Smad Proteins; D051898:Smad1 Protein; D013449:Sulfonamides; D015534:Trans-Activators; D014157:Transcription Factors; D016212:Transforming Growth Factor beta; D051028:bcl-2-Associated X Protein; C499469:recombinant human bone morphogenetic protein-2; D005047:Etoposide; D011493:Protein Kinase C; D020928:Mitogen-Activated Protein Kinases; D020169:Caspases; C058819:calphostin C; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one