Adrenal Cells in Tissue Culture

Abstract

Abstract We have previously reported (Kowal, J., Biochemistry 8, 1821 (1969)) that stimulation of monolayer cultures of mouse adrenal cortex tumors with adrenocorticotropic hormone (ACTH) for periods of 24 to 72 hours results in increased levels of mitochondrial 11β-hydroxylase activity. This report describes some additional observations on the nature of this effect. Concomitant with the increases in enzyme activity, the levels of cytochrome P-450 and adrenal nonheme iron protein (adrenodoxin) are also increased. On the other hand, ACTH has no effect on the levels of mitochondrial respiratory chain cytochromes. The content of these latter pigments was found to be the same as those reported in bovine adrenal tissue. Long term culture, for periods up to 2 years, did not produce any alteration in the levels of these cytochromes. Cytochrome P-450 is diminished by long term culture, but its content can be restored by passage of the cells into recipient mice. The concentration of cytochrome P-450 in all culture preparations is markedly lower than that reported for normal bovine adrenals. The effect of ACTH on 11β-hydroxylase occurs in the absence of any stimulation of mitochondrial protein synthesis. Similarly, there is little or no effect of ACTH on the activities of such mitochondrial enzymes as succinic dehydrogenase, cytochrome c oxidase, DPN-dependent malate dehydrogenase, adenosine 5'-triphosphatase, and 5'-nucleotidase. Small (l50%) but consistent increases in TPNH- and DPNH-cytochrome c reductases were seen in cultures which responded to ACTH with increases in 11β-hydroxylase which averaged 500%. The data suggest that ACTH can induce the synthesis of specific enzymes in the absence of a generalized effect on growth of the adrenal cell.