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Fibronectin gene expression, synthesis and accumulation during in vitro differentiation of chicken osteoblasts.

Authors
  • Winnard, R G
  • Gerstenfeld, L C
  • Toma, C D
  • Franceschi, R T
Type
Published Article
Journal
Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research
Publication Date
Dec 01, 1995
Volume
10
Issue
12
Pages
1969–1977
Identifiers
PMID: 8619378
Source
Medline
Keywords
License
Unknown

Abstract

A well-defined chicken osteoblast culture system(18) has been used to examine fibronectin (FN) mRNA levels, synthesis, and accumulation during in vitro differentiation and matrix mineralization. Immunofluorescent staining of cells after 6 or 18 days in culture revealed that FN was initially associated with the cell surface and in partial coalignment with cytoskeletal elements while at the latter time most FN was associated with the extracellular matrix as a ubiquitous fibrillar network. Western blot analysis of total cell-associated proteins also detected FN at all culture times. However, when results were normalized to cellular DNA, FN levels increased until 12-16 and remained relatively constant thereafter. Similarly, FN synthesis as measured by [35S]-methionine labeling, and immunoprecipitation was greatest in early cultures (culture day 3) and then declined such that synthesis decreased 60% at day 18 and 94% after 24-31 days. FN mRNA levels as measured by Northern blot analysis were well correlated with FN synthesis. These results clearly show that FN is made by primary osteoblasts during their in vitro maturation. In contrast to other osteoblast markers such as alkaline phosphatase, osteocalcin, and osteopontin, whose expression increases as cells differentiate, FN accumulates in the matrix during periods of early cell growth and attachment and then remains proportional to cell number. Results with FN differ from those obtained with collagen which continues to accumulate in the extracellular matrix during osteoblast maturation. These results are consistent with FN being important for the initial attachment of early osteoblasts or osteoblast precursors to the pericellular matrix.

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