title: Osteogenesis requires FAK-dependent collagen synthesis by fibroblasts and osteoblasts.
abstract: Focal adhesion kinase (FAK) is critical in adhesion-dependent signaling, but its role in osteogenesisin vivois ill defined. We deletedFakin fibroblasts and osteoblasts inFloxed-Fakmice bred with those expressing Cre-recombinase driven by 3.6-kb α1(I)-collagen promoter. Compared with wild-type (WT), conditional FAK-knockout (CFKO) mice were shorter (2-fold;P< 0.0001) and had crooked, shorter tails (50%;P< 0.0001). Microcomputed tomography analysis showed reduced bone volume (4-fold in tails;P< 0.0001; 2-fold in mandibles;P< 0.0001), whereas bone surface area/bone volume increased (3-fold in tails;P< 0.0001; 2.5-fold in mandibles;P< 0.001). Collagen density and fiber alignment in periodontal ligament were reduced by 4-fold (P< 0.0001) and 30% (P< 0.05), respectively, in CFKO mice. In cultured CFKO osteoblasts, mineralization at d 7 and mineralizing colony-forming units at d 21 were 30% (P< 0.0001) and >3-fold less than WT, respectively. Disruptions of FAK function in osteoblasts by conditional knockout, siRNA-knockdown, or FAK inhibitor reduced mRNA and protein expression of Runx2 (>30%), Osterix (>25%), and collagen-1 (2-fold). Collagen synthesis was abrogated in WT osteoblasts with Runx2 knockdown and inFak-null fibroblasts transfected with an FAK kinase domain mutant or a kinase-impaired mutant (Y397F). These data indicate that FAK regulates osteogenesis through transcription factors that regulate collagen synthesis.-Rajshankar, D., Wang, Y., McCulloch, C. A. Osteogenesis requires FAK-dependent collagen synthesis by fibroblasts and osteoblasts.
authors.affiliation: Matrix Dynamics Group, Faculty of Dentistry, University of Toronto, Toronto, Ontario, Canada dhaarmini.rajshankar@utoronto.ca.
authors.forename: Dhaarmini
authors.initials: D
authors.lastname: Rajshankar
authors.affiliation: Matrix Dynamics Group, Faculty of Dentistry, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Yongqiang
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Matrix Dynamics Group, Faculty of Dentistry, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Christopher A
authors.initials: CA
authors.lastname: McCulloch
mesh_terms: D000818:Animals; D002113:Calcification, Physiologic; D002478:Cells, Cultured; D024042:Collagen Type I / Q000235:genetics / Q000378:metabolism*; D050659:Core Binding Factor Alpha 1 Subunit / Q000235:genetics / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D051417:Focal Adhesion Kinase 1 / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D010006:Osteoblasts / Q000166:cytology / Q000378:metabolism*; D010012:Osteogenesis; D000076182:Sp7 Transcription Factor; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
chemical_list: D024042:Collagen Type I; D050659:Core Binding Factor Alpha 1 Subunit; C493716:Runx2 protein, mouse; D000076182:Sp7 Transcription Factor; C479466:Sp7 protein, mouse; D014157:Transcription Factors; D051417:Focal Adhesion Kinase 1; C496051:Ptk2 protein, mouse
keywords: focal adhesions; periodontal ligament; picrosirius red stain; polarized light microscopy; quantitative RT-PCR