Fibronectin regulates calvarial osteoblast differentiation

Amr M. Moursi, Caroline H. Damsky, Jonathan Lull, Deborah Zimmerman, Stephen B. Doty, Shin Ichi Aota, Ruth K. Globus

Research output: Contribution to journalArticlepeer-review

Abstract

The secretion of fibronectin by differentiating osteoblasts and its accumulation at sites of osteogenesis suggest that fibronectin participates in bone formation. To test this directly, we determined whether fibronectin-cell interactions regulate progressive differentiation of cultured fetal rat calvarial osteoblasts. Spatial distributions of α5 integrin subunit, fibronectin, osteopontin (bone sialoprotein I) and osteocalcin (bone Gla-protein) were similar in fetal rat calvaria and mineralized, bone-like nodules formed by cultured osteoblasts. Addition of anti-fibronectin antibodies to cultures at confluence reduced subsequent formation of nodules to less than 10% of control values, showing that fibronectin is required for normal nodule morphogenesis. Anti-fibronectin antibodies selectively inhibited steady-state expression of mRNA for genes associated with osteoblast differentiation; mRNA levels for alkaline phosphatase and osteocalcin were suppressed, whereas fibronectin, type I collagen and osteopontin were unaffected. To identify functionally relevant domains of fibronectin, we treated cells with soluble fibronectin fragments and peptides. Cell-binding fibronectin fragments (type III repeats 6-10) containing the Arg-Gly-Asp (RGD) sequence blocked both nodule initiation and maturation, whether or not they contained a functional synergy site. In contrast, addition of the RGD-containing peptide GRGDSPK alone did not inhibit nodule initiation, although it did block nodule maturation. Thus, in addition to the RGD sequence, other features of the large cell-binding fragments contribute to the full osteogenic effects of fibronectin. Nodule formation and osteoblast differentiation resumed after anti-fibronectin antibodies or GRGDSPK peptides were omitted from the media, showing that the inhibition was reversible and the treatments were not cytotoxic. Outside the central cell-binding domain, peptides from the IIICS region and antibodies to the N terminus did not inhibit nodule formation. We conclude that osteoblasts interact with the central cell-binding domain of endogenously produced fibronectin during early stages of differentiation, and that these interactions regulate both normal morphogenesis and gene expression. .

Original languageEnglish (US)
Pages (from-to)1369-1380
Number of pages12
JournalJournal of Cell Science
Volume109
Issue number6
StatePublished - Jun 1996

Keywords

  • Fetal rat calvarial osteoblast
  • Fibronectin
  • Osteoblast differentiation
  • RGD sequence

ASJC Scopus subject areas

  • Cell Biology

Fingerprint Dive into the research topics of 'Fibronectin regulates calvarial osteoblast differentiation'. Together they form a unique fingerprint.

Cite this