title: TGF-beta1-stimulated osteoblasts require intracellular calcium signaling for enhanced alpha5 integrin expression.
abstract: The osteoactive factor, transforming growth factor beta1 (TGF-beta1), influences osteoblast activity and bone function. We recently characterized a Smad-independent TGF-beta1-induced Ca(2+) signal in human osteoblasts (HOB) and demonstrated its importance in cell adhesion. Here, we further elucidate the role of the TGF-beta1 Ca(2+) signal in the mechanics of HOB adhesion. Osteoblast interaction with fibronectin (FN) through alpha5beta1 integrin is principally responsible for osteoblast-substrate adhesion. Our results show that the TGF-beta1 intracellular Ca(2+) signal is responsible, in part, for stimulation of alpha5 integrin expression, but not beta1 integrin or FN expression. Increased alpha5 integrin protein and mRNA expression was seen as early as 12 h after TGF-beta1 treatment, but was inhibited by cotreatment with nifedipine, a Ca(2+) channel blocker. TGF-beta1 increased both FN and beta1 integrin protein production within 48 h, independent of nifedipine cotreatment. Immunofluorescence observations revealed that TGF-beta1 increased alpha5 integrin staining, clustering, and colocalization with the actin cytoskeleton, effects that were blocked by nifedipine. The TGF-beta1 Ca(2+) signal, a pathway crucial for HOB adhesion, enhances alpha5 integrin expression, focal contact formation, and cytoskeleton reorganization. These early events are necessary for osteoblast adhesion; thus they determine the fate of the cell and ultimately affect bone function.
authors.affiliation: Department of Orthopaedic Surgery, 501 Curtis Building, Thomas Jefferson University, 1015 Walnut Street, Philadelphia, PA 19107, USA. leonnesti@yahoo.com
authors.forename: Leon J
authors.initials: LJ
authors.lastname: Nesti
authors.forename: E J
authors.initials: EJ
authors.lastname: Caterson
authors.forename: Mark
authors.initials: M
authors.lastname: Wang
authors.forename: Richard
authors.initials: R
authors.lastname: Chang
authors.forename: Felix
authors.initials: F
authors.lastname: Chapovsky
authors.forename: Jan B
authors.initials: JB
authors.lastname: Hoek
authors.forename: Rocky S
authors.initials: RS
authors.lastname: Tuan
mesh_terms: D000199:Actins / Q000378:metabolism; D015703:Antigens, CD / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002448:Cell Adhesion; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000378:metabolism; D005353:Fibronectins / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D039482:Integrin alpha5; D016023:Integrins / Q000378:metabolism; D009543:Nifedipine / Q000494:pharmacology; D010006:Osteoblasts / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D013997:Time Factors; D023822:Tissue Engineering; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D053773:Transforming Growth Factor beta1
chemical_list: D000199:Actins; D015703:Antigens, CD; D002121:Calcium Channel Blockers; D005353:Fibronectins; D039482:Integrin alpha5; D016023:Integrins; C508960:TGFB1 protein, human; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D009543:Nifedipine; D002118:Calcium