Expression of osteopontin in Meckel's cartilage cells during phenotypic transdifferentiation in vitro, as detected by in situ hybridization and immunocytochemical analysis

Kiyoto Ishizeki, Shintaro Nomura, Masaharu Takigawa, Hiroya Shioji, Tokio Nawa

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Abstract

The localization of osteopontin (OP) was examined in Meckel's cartilage cells that bipotentially expressed cartilage and bone phenotypes during cellular transformation in vitro. Cultured cells were analyzed by in situ hybridization, immunostaining followed by light and electron microscopy, electron microscopy, and electron probe microanalysis. The combination of ultrastructural analysis and immunoperoxidase staining indicated that OP- synthesizing cells were cells that were autonomously undergoing a change from chondrocytes to bone-forming cells at the top of nodules. Double immunofluorescence staining of 2-week-old cultures revealed that OP was first synthesized by chondrocytic cells at the top of nodules. After further time in culture, the distribution of OP expanded from the central toward the peripheral regions of the nodules. Electron probe microanalysis revealed that the localization of OP was associated with matrices of calcified cartilage and osteoid nodules that contained calcium and phosphorus. Immunoperoxidase electron microscopy revealed that, in addition to the intracellular immunoreactivity in chondrocytes and small round cells that were undergoing transformation, matrix foci of calcospherites and matrix vesicles, in particular, included growing crystals that were immunopositive for OP. An intense signal due to mRNA for OP in 3-week-old cultures was detected in nodule-forming round cells, while fibroblastic cells, spreading in a monolayer over the periphery of nodules, were only weakly labeled. These findings indicate that OP might be expressed sequentially by chondrocytes and by cells that are transdifferentiating further and exhibit an osteocytic phenotype, and moreover, that expression of OP is closely associated with calcifying foci in the extracellular matrix.

Original languageEnglish
Pages (from-to)457-466
Number of pages10
JournalHistochemistry and Cell Biology
Volume110
Issue number5
DOIs
Publication statusPublished - 1998

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cartilage
Osteopontin
Cartilage
Electron microscopy
In Situ Hybridization
nodules
Electron probe microanalysis
Bone
cells
Chondrocytes
Crystal growth
Optical microscopy
Phosphorus
Electron Probe Microanalysis
Calcium
Monolayers
electron microscopy
Electron Microscopy
phenotype
Cells

ASJC Scopus subject areas

  • Cell Biology
  • Instrumentation

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Expression of osteopontin in Meckel's cartilage cells during phenotypic transdifferentiation in vitro, as detected by in situ hybridization and immunocytochemical analysis. / Ishizeki, Kiyoto; Nomura, Shintaro; Takigawa, Masaharu; Shioji, Hiroya; Nawa, Tokio.

In: Histochemistry and Cell Biology, Vol. 110, No. 5, 1998, p. 457-466.

Research output: Contribution to journalArticle

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