Alphavbeta integrins play an essential role in BMP-2 induction of osteoblast differentiation.
Both integrins and BMP-2 exert similar effects on osteoblasts. We examined the relationship between the alphav-containing integrins (alphavbeta) and BMP-2 in osteoblast function. BMP-2 stimulates alphavbeta expression. BMP-2 receptors co-localize/overlap with alphavbeta integrins, and the intact function of alphavbeta is essential in BMP-2 activity.


BACKGROUND
Bone morphogenetic protein (BMP)-2 not only induces osteoblast differentiation and bone matrix mineralization, but also stimulates osteoblast migration on and adhesion to bone matrix proteins. The alphavbeta- and beta1- (alphabeta1) containing integrins mediate osteoblast interaction with many bone matrix proteins and play important roles in osteoblast adhesion, migration, and differentiation. Because alphavbeta integrins and BMP-2 share common effects on osteoblasts, we analyzed their relationship in osteoblast function.


METHODS
The effects of BMP-2 on integrin expression were determined by surface labeling/immunoprecipitation and cell adhesion to matrix proteins. Confocal analysis of the immunostained cells and co-immunoprecipitation of cell extracts were used to study the spatial relationship between integrins and BMP-2 receptors. A function-blocking anti-alphavbeta integrin antibody (L230) was employed to investigate the roles of alphavbeta integrins in BMP-2 function.


RESULTS
Human osteoblasts (HOBs) express alphabeta1, alphavbeta3, alphavbeta5, alphavbeta6, and alphavbeta8 integrins at focal adhesion sites. BMP-2 increases the levels of these integrins on osteoblast surface and enhances HOB adhesion to osteopontin and vitronectin. Immunoprecipitation and immunostaining analyses show that BMP-2 receptors co-localize or overlap with alphavbeta and alphabeta1 integrins. Incubation of HOBs with L230 abolishes the antiproliferative effect of BMP-2 and reduces the capacity of BMP-2 to stimulate alkaline phosphatase activity and the expression of osteocalcin, osteopontin, and bone sialoprotein. Furthermore, L230 prevents BMP-2 induction of matrix mineralization. Although BMP-2 retains its receptor-binding capability in the presence of L230, BMP-2 stimulation of Smad signaling is abolished by L230.


CONCLUSIONS
BMP-2 upregulates the expression of alphavbeta integrins, and these integrins, in turn, play a critical role in BMP-2 function in osteoblasts.
Lai
Chung-Fang
CF
Division of Bone and Mineral Diseases, Department of Internal Medicine, Washington University School of Medicine, St Louis, MO 63110, USA.
Cheng
Su-Li
SL
D000469:Alkaline Phosphatase / Q000378:metabolism; D000906:Antibodies / Q000737:chemistry; D019921:Biotinylation; D015153:Blotting, Western; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000378:metabolism*; D001842:Bone and Bones; D002448:Cell Adhesion; D002454:Cell Differentiation; D045744:Cell Line, Tumor; D002465:Cell Movement; D049109:Cell Proliferation; D002478:Cells, Cultured; D006801:Humans; D047468:Immunoprecipitation; D058574:Integrin-Binding Sialoprotein; D016023:Integrins / Q000378:metabolism / Q000502:physiology*; D008156:Luciferases / Q000378:metabolism; D018613:Microscopy, Confocal; D010006:Osteoblasts / Q000166:cytology* / Q000378:metabolism; D015675:Osteocalcin / Q000096:biosynthesis; D053495:Osteopontin; D019030:Receptors, Vitronectin / Q000502:physiology*; D011994:Recombinant Proteins / Q000737:chemistry; D012795:Sialoglycoproteins / Q000096:biosynthesis / Q000378:metabolism; D013997:Time Factors; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D015854:Up-Regulation; D019096:Vitronectin / Q000378:metabolism
D000906:Antibodies; C525718:BMP2 protein, human; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; C546871:IBSP protein, human; D058574:Integrin-Binding Sialoprotein; D016023:Integrins; D019030:Receptors, Vitronectin; D011994:Recombinant Proteins; C506895:SPP1 protein, human; D012795:Sialoglycoproteins; D016212:Transforming Growth Factor beta; D019096:Vitronectin; C094340:integrin alphavbeta1; C499469:recombinant human bone morphogenetic protein-2; D015675:Osteocalcin; D053495:Osteopontin; D008156:Luciferases; D000469:Alkaline Phosphatase