Hornsby P J, Harris S E, Aldern K A
Endocrinology. 1985 Sep;117(3):1264-71. doi: 10.1210/endo-117-3-1264.
To investigate the role of ascorbic acid in the function of the adrenal cortex, we studied the effects of ascorbate 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 about 40 h. When 50 microM 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 about 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. However, ascorbate at concentrations from 10 microM 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 microM cortisol. 11 beta-Hydroxylase activity was reinduced over a period of 5 days in third passage cultures by addition of 1 microM ACTH in defined lipoprotein-free medium. Addition of ascorbate enhanced the reinduction about 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 selenium 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, selenium, 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.
为了研究抗坏血酸在肾上腺皮质功能中的作用，我们研究了抗坏血酸盐对培养物中11β-羟化酶调节的影响。当原代牛肾上腺皮质细胞在无促肾上腺皮质激素（ACTH）的无血清限定培养基中培养时，11β-羟化酶活性以约40小时的半衰期下降。当将50μM皮质醇（其作为11β-羟化酶的假底物）添加到此类培养物中时，11β-羟化酶活性以约6小时的半衰期下降。在存在皮质醇的情况下，抗坏血酸盐（5mM）显著降低了11β-羟化酶活性的丧失速率。先前的研究表明，酚类和亚砜类抗氧化剂也能防止11β-羟化酶活性丧失，它们在略高于保护活性所需浓度时会抑制该酶。然而，浓度为10μM至5mM的抗坏血酸盐不会抑制11β-羟化酶。在与皮质醇预孵育24小时期间向细胞中添加相同范围的抗坏血酸盐浓度，显示出对11β-羟化酶活性丧失的预防作用增强。抗坏血酸盐和较低浓度的氧气在其保护作用中具有协同性。在2%氧气条件下，5mM抗坏血酸盐在存在50μM皮质醇的情况下几乎完全防止了11β-羟化酶活性的丧失。在第三代培养物中，通过在限定的无脂蛋白培养基中添加1μM促肾上腺皮质激素，11β-羟化酶活性在5天内被重新诱导。添加抗坏血酸盐使重新诱导增强约2倍。抗坏血酸盐在预防假底物介导的活性丧失以及增强11β-羟化酶的重新诱导方面的作用是特异性的；在存在皮质醇的情况下，α-生育酚和硒都不能防止11β-羟化酶活性丧失，在存在促肾上腺皮质激素的情况下也不能增强11β-羟化酶的重新诱导。作为特异性的额外测试，结果表明，17-羟化酶活性的重新诱导完全不受抗坏血酸盐、硒或α-生育酚的影响，并且向具有高17-羟化酶的培养物中添加皮质醇不会导致酶活性的任何丧失。因此，抗坏血酸在肾上腺皮质中的一个主要功能是作为细胞色素的保护化合物。
