Expression of ACTH-induced corticosteroid biosynthesis in newborn rat adrenocortical cells cultured in serum-free and carrier protein-free medium containing a cholesterol-alpha-cyclodextrin complex

Abstract

Newborn rat adrenocortical cells were successfully cultured in a serum-free carrier protein-free medium (SPFM) by using alpha-cyclodextrin as a cholesterol carrier and have expressed corticosteroid biosynthesis in this medium. A stable inclusion complex of cholesterol-alpha-cyclodextrin with a molar ratio of almost 1 was obtained for a 5 X 10(-5) mol/1 alpha-cyclodextrin concentration. Cell cultures incubated with [4-14C] cholesterol-alpha-cyclodextrin in SPFM produced, under ACTH stimulation, various 14C labeled steroids with a predominance of corticosterone and 18-hydroxy-11-deoxycorticosterone. As measured by gas chromatography and mass spectrometry, the ratio between corticosteroids (21-hydroxylated steroids) and 20 alpha-reduced steroids produced in SPFM with cholesterol-alpha-cyclodextrin was equal to 1.8. This corresponds to a value of 3.6 times higher than that found in the serum-free medium with cholesterol-albumin. Consequently, the chemically defined SPFM with cholesterol-alpha-cyclodextrin used in this study is more suitable for corticosteroidogenesis by adrenal cells in culture than a serum-free medium with cholesterol-albumin.