Maintenance of the Keratocyte Phenotype during Cell Proliferation Stimulated by Insulin
| Autor: | Bridgette Alexandrou, Bradley Kane, Kurt Musselmann, John R. Hassell |
|---|---|
| Rok vydání: | 2005 |
| Předmět: |
Keratinocytes
Time Factors medicine.medical_treatment Blotting Western Immunoblotting Becaplermin Biology Cell morphology Biochemistry Culture Media Serum-Free Cytosol medicine Animals Insulin Microscopy Phase-Contrast Fibroblast Molecular Biology Cell Proliferation Platelet-Derived Growth Factor Wound Healing Cell growth Temperature DNA Proto-Oncogene Proteins c-sis Cell Biology Aldehyde Dehydrogenase Culture Media Cell biology Blot Phenotype medicine.anatomical_structure Chromatography Gel Cattle Fibroblast Growth Factor 2 Proteoglycans Wound healing Keratocan Fetal bovine serum |
| Zdroj: | Journal of Biological Chemistry. 280:32634-32639 |
| ISSN: | 0021-9258 |
| DOI: | 10.1074/jbc.m504724200 |
| Popis: | Keratocytes normally express high levels of aldehyde dehydrogenase and keratocan. They proliferate and lose their keratocyte markers when they become fibroblastic during corneal wound healing. Keratocytes cultured in fetal bovine serum also become fibroblastic, proliferate, and lose these markers. In this report, we studied the effects of three serum growth factors, fibroblast growth factor-2, insulin, and platelet-derived growth factor-BB, on keratocyte proliferation and the maintenance of the keratocyte markers in 7-day cultures in cells plated at low (5,000 cells/cm2) and high (20,000 cells/cm2) density in serum-free medium. Keratocyte proliferation was measured by [3H]thymidine incorporation and by DNA content of the cultures. Cytosolic aldehyde dehydrogenase and keratocan accumulated in the medium were quantified by Western blot. The results showed that all the growth factors stimulated proliferation, but insulin stimulated proliferation more consistently. The keratocyte markers aldehyde dehydrogenase and keratocan were maintained after 7 days in culture in all growth factors, but keratocyte cell morphology was only maintained in medium containing insulin. Most of the proteoglycans were degraded in cultures of keratocytes plated at low density and cultured in the absence of growth factors. This degradation was prevented when keratocytes were cultured in the presence of the growth factors or when keratocytes were plated at high density. The results of this study show that insulin can expand keratocytes in vitro, maintain their phenotype, and prevent proteoglycan degradation. |
| Databáze: | OpenAIRE |
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