Involvement of the phosphoinositide 3-kinase/protein kinase B signaling pathway in insulin/IGF-I-induced chondrogenesis of the mouse embryonal carcinoma-derived cell line ATDC5

Kiyoshi Hidaka, Takashi Kanematsu, Hiroshi Takeuchi, Minoru Nakata, Ushio Kikkawa, Masato Hirata

Research output: Contribution to journalArticle

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Abstract

The embryonal carcinoma-derived cell line, ATDC5, differentiates into chondrocytes in response to insulin/insulin-like growth factor-I (IGF-I) stimulation. In the present study, we examined whether insulin/IGF-I stimulation caused activation of the phosphoinositide 3-kinase (PI3K)/protein kinase B (PKB) pathway in ATDC5 cells. We also determined whether the insulin-stimulated differentiation of ATDC5 cells into chondrocytes could be mimicked by activation of the PKB pathway alone. ATDC5 cells produced phosphatidylinositol 3,4,5-trisphosphate and the pleckstrin homology domain of PKB was recruited to the plasma membrane in response to insulin stimulation. This was probably a result of activation of PI3K because the PI3K inhibitors, wortmannin and LY294002, inhibited both responses, although the effective concentrations were as high as 10 μM. Insulin stimulation caused the chondrogenic differentiation of ATDC5 cells as assessed by chondrogenic nodule staining with alcian blue. The addition of wortmannin or LY294002, PI3K inhibitors, suppressed the staining, and the suppression was reversible, indicating the effect of the inhibitors is not toxic. Finally, we exogenously expressed a constitutively-activated from of PKB (myristoylated PKB, myr-PKB) in ATDC5 cells, and found the chondrogenic differentiation of ATDC5 cells to form nodules occurred in the absence of insulin stimulation. The kinase-negative mutant of myr-PKB did not caused differentiation, indicating that kinase activity is required. These results support the hypothesis that the PI3K/PKB signaling pathway is involved in the chondrogenic differentiation of ATDC5 cells in response to insulin/IGF-I stimulation. This is the first report that demonstrates the involvement of phosphoinositide signaling in the induction of chondrogenesis from undifferentiated cells.

Original languageEnglish
Pages (from-to)1094-1103
Number of pages10
JournalInternational Journal of Biochemistry and Cell Biology
Volume33
Issue number11
DOIs
Publication statusPublished - Sep 15 2001

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Embryonal Carcinoma Stem Cells
Chondrogenesis
Proto-Oncogene Proteins c-akt
1-Phosphatidylinositol 4-Kinase
Phosphatidylinositols
Insulin-Like Growth Factor I
Cells
Insulin
Cell Line
Cell Differentiation
Phosphotransferases
2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
Chemical activation
Chondrocytes
Staining and Labeling
Alcian Blue
Poisons
Cell membranes
Cell Membrane

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Cell Biology

Cite this

Involvement of the phosphoinositide 3-kinase/protein kinase B signaling pathway in insulin/IGF-I-induced chondrogenesis of the mouse embryonal carcinoma-derived cell line ATDC5. / Hidaka, Kiyoshi; Kanematsu, Takashi; Takeuchi, Hiroshi; Nakata, Minoru; Kikkawa, Ushio; Hirata, Masato.

In: International Journal of Biochemistry and Cell Biology, Vol. 33, No. 11, 15.09.2001, p. 1094-1103.

Research output: Contribution to journalArticle

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AU - Takeuchi, Hiroshi

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AU - Hirata, Masato

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