WISP-1/CCN4 regulates osteogenesis by enhancing BMP-2 activity.
Wnt-induced secreted protein 1 (WISP-1/CCN4) is a member of the CCN family that is highly expressed in skeletal tissue and in osteoprogenitor cells induced to differentiate in vitro. To determine the function of WISP-1 during osteogeneis, osteogenic bone marrow stromal cells (BMSCs) were transduced with WISP-1 adenovirus (adWISP-1) in the presence or absence of bone morphogenetic protein 2 (BMP-2) adenovirus (adBMP-2). WISP-1 overexpression enhanced the ability of BMP-2 to direct BMSCs toward osteogenic differentiation and appeared to work by stimulating Smad-1/5/8 phosphorylation and activation. The ability of WISP-1 to enhance BMP-2 activity also was shown in vivo using an ectopic osteogenesis assay with BMSCs transduced with WISP-1, BMP-2, or both. When BMSCs were infected with lentivirus containing human WISP1 shRNA, they formed less bone in vivo and were less responsive to BMP-2, confirming that WISP-1 and BMP-2 have a functional interaction. Immunoprecipitation (IP) and Western blot analysis showed that WISP-1 bound directly to BMP-2 and showed that WISP-1 increased BMP-2 binding to hBMSCs in a dose-dependent fashion. To understand how WISP-1 enhanced BMP-2 signaling, the influence of WISP-1 on integrin expression was analyzed. WISP-1 induced the mRNA and protein levels of α(5)-integrin and, further, was found to bind to it. Antibody-blocking experiments showed that the BMP-2 binding to BMSCs that was enhanced by WISP-1 was completely neutralized by treatment with anti-integrin α(5)β(1) antibody. Pilot studies and the use of transgenic mice that overexpressed human WISP-1 in preosteoblasts had increased bone mineral density (BMD), trabecular thickness, and bone volume (BV/TV) over wild-type controls, supporting observations using human osteoprogenitors that WISP-1 has a positive influence on osteogenesis in vivo. In conclusion, these studies show, for the first time, that WISP-1 has a positive influence on bone cell differentiation and function and may work by enhancing the effects of BMP-2 to increase osteogenesis through a mechanism potentially involving binding to integrin α(5)β(1).
Ono
Mitsuaki
M
Craniofacial and Skeletal Diseases Branch, National Institutes of Craniofacial and Dental Research, National Institutes of Health, Bethesda, MD 20892, USA.
Inkson
Colette A
CA
Kilts
Tina M
TM
Young
Marian F
MF
D000818:Animals; D001854:Bone Marrow Cells / Q000378:metabolism; D055396:Bone Morphogenetic Protein 2 / Q000378:metabolism*; D001842:Bone and Bones / Q000000981:diagnostic imaging / Q000378:metabolism; D055511:CCN Intercellular Signaling Proteins; D002454:Cell Differentiation; D002478:Cells, Cultured; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008822:Mice, Transgenic; D015513:Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D010006:Osteoblasts / Q000166:cytology / Q000378:metabolism; D010012:Osteogenesis; D010641:Phenotype; D011485:Protein Binding; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D011859:Radiography; D019030:Receptors, Vitronectin / Q000235:genetics / Q000378:metabolism
C525718:BMP2 protein, human; C525719:Bmp2 protein, mouse; D055396:Bone Morphogenetic Protein 2; D055511:CCN Intercellular Signaling Proteins; C116396:CCN4 protein, human; C492067:CCN4 protein, mouse; D047908:Intracellular Signaling Peptides and Proteins; D015513:Oncogene Proteins; D011518:Proto-Oncogene Proteins; D019030:Receptors, Vitronectin; C094340:integrin alphavbeta1