Expression of insulin-like growth factor I by cultured skin substitutes does not replace the physiologic requirement for insulin in vitro

Citation
Vb. Swope et al., Expression of insulin-like growth factor I by cultured skin substitutes does not replace the physiologic requirement for insulin in vitro, J INVES DER, 116(5), 2001, pp. 650-657
Citations number
50
Categorie Soggetti
Dermatology,"da verificare
Journal title
JOURNAL OF INVESTIGATIVE DERMATOLOGY
ISSN journal
0022202X → ACNP
Volume
116
Issue
5
Year of publication
2001
Pages
650 - 657
Database
ISI
SICI code
0022-202X(200105)116:5<650:EOIGFI>2.0.ZU;2-K
Abstract
Clinical efficacy of cultured skin substitutes may be increased if their ca rbohydrate metabolism is optimized by understanding whether endogenous insu lin-like growth factor I can substitute for exogenous insulin. Cultured ski n substitutes were prepared and incubated at the air-liquid interface for 4 wk in media containing 0.5 or 5 mug per mi insulin, 10 or 50 ng per mi ins ulin-like growth factor I, or 0 insulin and 0 insulin-like growth factor I (negative control). In situ hybridization showed that the epidermal and der mal cultured skin substitute components express insulin-like growth factor I mRNA throughout the 28 d interval. Immunohistochemistry confirmed the exp ression of insulin-like growth factor I protein by the human keratinocytes and fibroblasts in cultured skin substitutes. Insulin-Like growth factor I at 10 or 30 ng per mi could partially replace insulin in a clonal assay of keratinocyte growth. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assays showed significantly higher values in cultured skin su bstitutes incubated with insulin at incubation days 14 and 28 compared to n egative control or the 10 ng per mi insulin-like growth factor I condition. Cultured skin substitutes incubated in 50 ng per mi insulin-like growth fa ctor I had MTT values similar to the insulin-treated cultured skin substitu tes at day 14, but were significantly lower by day 28, Light microscopy agr eed with MTT data showing that cultured skin substitutes grown with insulin media had multiple layers of nucleated keratinocytes and stratum corneum a t days 14 and 28, The negative control and 10 ng per mi insulin-like growth factor I exhibited poor cultured skin substitute epidermal morphology thro ughout the experiment, In contrast, the cultured skin substitutes in 50 ng per mi insulin-like growth factor I were similar to the insulin-treated cul tured skin substitutes at day 14, but by day 28 had deteriorated to resembl e the negative control. Bromodeoxyuridine incorporation at day 28 was signi ficantly higher for 5 mug per mi insulin cultured skin substitutes versus a ll other treatment groups, These data suggest. that medium. containing 5 mu g per mi insulin supports greater physiologic stability in cultured skin su bstitutes over time, and that expression of insulin-like growth factor I by keratinocytes and fibroblasts in cultured skin substitutes is not sufficie nt to fully replace the requirement for exogenous insulin in vitro.