The Role of Ascorbic Acid in the Function of the Adrenal Cortex: Studies in Adrenocortical Cells in Culture*

Publisher Website
Google Scholar
Abstract
To investigate the role of ascorbic acid in the function of the adrenal cortex, the effects of ascorbate were studied on the regulation of 11.beta.-hydroxylase in culture. When primary bovine adrenocortical cells were cultured in a serum-free defined medium in the absence of ACTH 11.beta.-hydroxylase activity declined with a half-time of .apprx. 40 h. When 50 .mu.M cortisol, which acts as a pseudosubstrate for 11.beta.-hydroxylase, was added to such cultures, 11.beta.-hydroxylase activity declined with a half-time of .apprx. 6 h. Ascorbate (5 mM) markedly reduced the rate of loss of 11.beta.-hydroxylase activity in the presence of cortisol. Previous studies showed that phenolic and sulfoxide antioxidants, which also prevent loss of 11.beta.-hydroxylase activity, inhibited the enzyme at concentrations somewhat higher than those required for protective activity. Ascorbate at concentrations from 10 .mu.M to 5 mM did not inhibit 11.beta.-hydroxylase. The same range of ascorbate concentrations added to cells during a 24-h preincubation with cortisol showed increasing prevention of loss of 11.beta.-hydroxylase activity. Ascorbate and a lowered concentration of oxygen were synergistic in their protective action. At 2% oxygen, 5 mM ascorbate almost completely prevented loss of 11.beta.-hydroxylase activity in the presence of 50 .mu.M cortisol. 11.beta.-Hydroxylase activity was reinduced over a period of 5 days in 3rd passage cultures by addition of 1 .mu.M ACTH in defined lipoprotein-free medium. Addition of ascorbate enhanced the reinduction .apprx. 2-fold. The action of ascorbate in prevention of pseudosubstrate-mediated loss of activity and in enhancing reinduction of 11.beta.-hydroxylase is specific, neither .alpha.-tocopherol nor Se prevented loss of 11.beta.-hydroxylase in the presence of cortisol or enhanced reinduction of 11.beta.-hydroxylase in the presence of ACTH. As an additional test of specificity, it was shown that reinduction of 17-hydroxylase activity was completely unaffected by ascorbate, Se or .alpha.-tocopherol, and addition of cortisol to cultures with high 17-hydroxylase did not result in any loss of enzyme activity. Thus, a major function of ascorbate in the adrenal cortex is as a protective compound for cytochrome P-450.