Evidence that α5β1 integrins mediate Leydig cell binding to fibronectin and enhance Leydig cell proliferation stimulated by a Sertoli cell-secreted mitogenic factor in vitro

Naixing Wu, Eisuke P. Murono, Wayne E. Carver, Louis Terracio, Thierry Bacro

Research output: Contribution to journalArticle

Abstract

We reported previously that coculture of immature rat Sertoli cells with Leydig cells or the addition of a concentrate from Sertoli cell-conditioned medium (SCCM) stimulated Leydig cell [ 3H]-thymidine incorporation, increased cell number, and altered Leydig cell morphology (Wu and Murono, 1994). In the present studies, the effect of various extracellular matrix proteins on immature Leydig cell binding, proliferation and response to SCCM concentrate was investigated. Pretreatment of culture wells with 50 μg/mL collagen I or 10 μg/mL laminin inhibited Leydig cell binding to culture wells about 95 and 89%, respectively; however, 5 μg/mL fibronectin did not change the level of attachment. The binding of Leydig cells to fibronectin was reduced by antifibronectin or-β1 integrin antibodies (66 and 91%, respectively). Treatment of culture wells with five or 50 μg/mL fibronectin alone increased [ 3H]thymidine incorporation about twofold. When Leydig cells were cultured in wells precoated with increasing concentrations of fibronectin and then treated with SCCM concentrate for 2 d, [ 3H]-thymidine incorporation increased progressively with the concentration of fibronectin, beyond the levels observed with SCCM concentrate alone. This response was associated with increases in both Leydig cell number and labeling indices. When Leydig cells were cultured on fibronectin, their numbers increased by 3.7-and 5.1-fold following treatment with SCCM concentrates or coculture for 6 d, respectively; whereas, they increased 2.6- and 3.9-fold, respectively, when cultured on plastic. Labeling indices of Leydig cells cultured on plastic for 2 d and treated with SCCM or cocultured were 6.9 and 11.9%, respectively, while labeling indices of Leydig cells grown on fibronectin increased further to 17.6 and 26.3%, respectively. α5β1 integrin complexes and α5 integrin mRNA were expressed in Leydig cells, suggesting that binding to fibronectin may be mediated by α5β1 integrins, a fibronectin receptor. These results suggest that Leydig cell proliferation stimulated by a Sertoli cell-secreted mitogenic factor(s) is enhanced by Leydig cell binding fibronectin, and that this binding may be mediated by α5β1 integrins.

Original languageEnglish (US)
Pages (from-to)75-83
Number of pages9
JournalEndocrine
Volume5
Issue number1
DOIs
StatePublished - Aug 1996

Fingerprint

Leydig Cells
Sertoli Cells
Fibronectins
Integrins
Cell Proliferation
Conditioned Culture Medium
Thymidine
Coculture Techniques
In Vitro Techniques
Plastics
Cell Count
Integrin alpha5beta1
Extracellular Matrix Proteins
Laminin

Keywords

  • α5β1 integrins
  • fibronectin
  • Leydig cell
  • mitogen
  • proliferation
  • Sertoli cell

ASJC Scopus subject areas

  • Endocrinology, Diabetes and Metabolism
  • Endocrinology

Cite this

Evidence that α5β1 integrins mediate Leydig cell binding to fibronectin and enhance Leydig cell proliferation stimulated by a Sertoli cell-secreted mitogenic factor in vitro. / Wu, Naixing; Murono, Eisuke P.; Carver, Wayne E.; Terracio, Louis; Bacro, Thierry.

In: Endocrine, Vol. 5, No. 1, 08.1996, p. 75-83.

Research output: Contribution to journalArticle

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abstract = "We reported previously that coculture of immature rat Sertoli cells with Leydig cells or the addition of a concentrate from Sertoli cell-conditioned medium (SCCM) stimulated Leydig cell [ 3H]-thymidine incorporation, increased cell number, and altered Leydig cell morphology (Wu and Murono, 1994). In the present studies, the effect of various extracellular matrix proteins on immature Leydig cell binding, proliferation and response to SCCM concentrate was investigated. Pretreatment of culture wells with 50 μg/mL collagen I or 10 μg/mL laminin inhibited Leydig cell binding to culture wells about 95 and 89{\%}, respectively; however, 5 μg/mL fibronectin did not change the level of attachment. The binding of Leydig cells to fibronectin was reduced by antifibronectin or-β1 integrin antibodies (66 and 91{\%}, respectively). Treatment of culture wells with five or 50 μg/mL fibronectin alone increased [ 3H]thymidine incorporation about twofold. When Leydig cells were cultured in wells precoated with increasing concentrations of fibronectin and then treated with SCCM concentrate for 2 d, [ 3H]-thymidine incorporation increased progressively with the concentration of fibronectin, beyond the levels observed with SCCM concentrate alone. This response was associated with increases in both Leydig cell number and labeling indices. When Leydig cells were cultured on fibronectin, their numbers increased by 3.7-and 5.1-fold following treatment with SCCM concentrates or coculture for 6 d, respectively; whereas, they increased 2.6- and 3.9-fold, respectively, when cultured on plastic. Labeling indices of Leydig cells cultured on plastic for 2 d and treated with SCCM or cocultured were 6.9 and 11.9{\%}, respectively, while labeling indices of Leydig cells grown on fibronectin increased further to 17.6 and 26.3{\%}, respectively. α5β1 integrin complexes and α5 integrin mRNA were expressed in Leydig cells, suggesting that binding to fibronectin may be mediated by α5β1 integrins, a fibronectin receptor. These results suggest that Leydig cell proliferation stimulated by a Sertoli cell-secreted mitogenic factor(s) is enhanced by Leydig cell binding fibronectin, and that this binding may be mediated by α5β1 integrins.",
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