在過去的研究已經證實，不論在活體或細胞株的實驗都發現腎上腺皮質有自我抑制 (self-suppression) 的功能。糖皮質素 (glucocorticoids) 對腎上腺固醇類激素 (corticosteroid) 製造細胞的直接作用可能可以用來解釋臨床上使用糖皮質素所造成的腎上腺功能被抑制的現象。類固醇生成速控蛋白 (steroidogenic acute regulatory protein, StAR) 被認為可讓膽固醇 (cholesterol) 傳送至粒線體內膜而快速增加類固醇激素的生成。促激素 (tropic hormone) 藉由環腺嘌呤單磷酸 (cyclic AMP, cAMP) 的訊息傳導作用使StAR基因表現上升。基因的轉錄是被組蛋白 (histone) 的乙醯化 (acetylation) 及去乙醯化 (deacetylation) 所調控，利用cAMP類似物8-bromo-cAMP刺激StAR基因轉錄上升已經被證實與組蛋白H3的乙醯化相關。為了要證實糖皮質素對腎上腺固醇類激素生成的抑制作用，是否經由糖皮質素抑制StAR基因表現所產生的結果，我們研究利解痛 (dexamethasone, DEX) 對小鼠腎上腺腫瘤細胞 (mouse adrenocortical Y-1 tumor cells) StAR信息核糖核酸 (messenger RNA, mRNA) 表現量的影響，同時測試DEX對StAR基因啟動子 (promoter) 上組蛋白H3乙醯化的影響。對Y-1細胞給予漸增濃度的DEX (0.001 – 10 mg/ml) 培養24小時，發現DEX會抑制8-bromo-cAMP (0.5 mM) 誘導的StAR蛋白與mRNA的表現及黃體激素 (progesterone) 的產生，並呈現劑量相關性。Y-1細胞與8-bromo-cAMP (0.5 mM) 一起培養1至24小時，StAR蛋白與mRNA表現量隨培養時間增加而上升，且伴隨著黃體激素的產生量上升。DEX (10 mg/ml) 抑制8-bromo-cAMP誘導的StAR蛋白與mRNA的表現。以染色質免疫沉澱法 (chromatin immunoprecipitation assay) 分析乙醯化的組蛋白H3與StAR基因啟動子的接合關係，發現DEX使StAR啟動子上組蛋白H3乙醯化的程度下降。這些結果證實糖皮質素對腎上腺固醇類激素生成的自我抑制作用，至少有部分是藉由糖皮質素抑制StAR蛋白及mRNA表現所產生的結果，而糖皮質素抑制Y-1細胞StAR基因表現的作用機制，牽涉組蛋白的乙醯化。

Evidence from studies in vivo and in vitro suggests that the adrenal cortex is capable of self-suppression. The direct actions of glucocorticoid on the corticosteroid producing cells of the adrenal gland may contribute to the adrenal suppression seen with therapeutic glucocorticoid administration. The steroidogenic acute regulatory protein (StAR) is thought to mediate the rapid increase in steroid hormone biosynthesis by facilitating cholesterol transport to the inner mitochondrial membrane. Tropic hormones trigger up-regulation of StAR gene expression by cAMP signaling. Gene transcription is regulated by acetylation and deacetylation of histones. Acetylation of histone H3 is associated with 8-bromo-cAMP-stimulated StAR gene transcription. To ascertain whether the suppressive effect of glucocorticoids on corticosteroidogenesis results from direct glucocorticoid inhibition of StAR gene expression, we investigated the effect of dexamethasone (DEX) on StAR mRNA levels in mouse adrenocortical Y-1 tumor cells. Whether StAR promoter-associated histone H3 acetylation is affected by DEX was then tested. Treatment of Y-1 cells with increasing concentrations (0.001-10 mg/ml) of DEX for 24 h suppressed 8-bromo-cAMP (0.5 mM)-stimulated StAR protein and mRNA levels and progesterone production in a dose-dependent manner. Treatment of Y-1 cells with 8-bromo-cAMP (0.5 mM) for 1-24 h resulted in a marked increase in StAR protein and mRNA levels. This increase was associated with an increase in progesterone production. DEX (10 mg/ml) reduced 8-bromo-cAMP-stimulated StAR protein and mRNA levels. Using chromatin immunoprecipitation assays to assess histone H3 acetylation associated with StAR promoter, we show that DEX decreased histone H3 acetylation over a 1.3 kb region of the StAR promoter. These results demonstrate that the self-suppressive effect of glucocorticoids on corticosteroidogenesis is at least partly mediated by a decrease in StAR protein and mRNA levels. The mechanism of glucocorticoid action in suppressing StAR gene expression in Y-1 cells involves histone acetylation.

References

1. Esteban NV, Loughlin T, Yergey AL 1991 Daily cortisol production rate in man determined by stable isotope dilution/mass spectrometry. J Clin Endocrinol Metab 72:39-45.
2. Barton RN, Stoner HB, Watson SM 1987 Relationships among plasma cortisol, adrenocorticotrophin, and severity of injury in recently injured patients. J Trauma 27:384-392.
3. Chernow B, Alexander HR, Smallridge RC 1987 Hormonal responses to graded surgical stress. Arch Intern Med 147:1273-1278.
4. Perrot D, Bonneton A, Dechaud H, Motin J, Pugeat M 1993 Hypercortisolism in septic shock is not suppressible by dexamethasone infusion. Crit Care Med 21:396-401.
5. Chernow B, Rainey TR, Lake CR 1982 Endogenous and exogenous catecholamines in critical care medicine. Crit Care Med 10: 409-416.
6. Hart BB, Stanford GG, Ziegler MG 1989 Catecholamines: Study of interspecies variation. Crit Care Med 17:1203-1218.
7. Drucker D, McLaughlin J 1986 Adrenocortical dysfunction in acute medical illness. Crit Care Med 14:789-791.
8. Drucker D, Shandling M 1985 Variable adrenocortical function in acute medical illness. Crit Care Med 13:477-479.
9. Zaloga GP, Marik P 2001 Hypothalamic-pituitaryadrenal insufficiency. Crit Care Clin 17:25-42.
10. Streeten DHP 1999 What test for hypothalamicpituitary adrenocortical insufficiency? Lancet 354:179-180.
11. Lamberts SW, Bruining HA, de Jong FH 1997 Corticosteroid therapy in severe illness. N Engl J Med 337:1285-1292.
12. Melby JC, Spink WW 1958 Comparative studies of adrenal cortisol function and cortisol metabolism in healthy adults and in patients with septic shock due to infection. J Clin Invest 37:1791-1798.
13. Parker LN, Levin ER, Lifrak ET 1985 Evidence for adrenocortical adaptation to severe ill-ness. J Clin Endocrinol Metab 60:947-952.
14. Richards ML, Caplan RH, Wickus GG 1999 The rapid low-dose (1 microgram) cosyntropin test in the immediate postoperative period: Results in elderly subjects after major abdominal surgery. Surgery 125:431-440.
15. Schein RMH, Sprung CL, Marcial E 1990 Plasma cortisol levels in patients with septic shock.Crit Care Med 18:259-263.
16. Soni A, Pepper GM, Wyrwinski PM 1995 Adrenal insufficiency occurring during septic shock: Incidence, outcome, and relationship to peripheral cytokine levels. Am J Med 98:266-271.
17. Bouachour G, Tirot P, Gouello JP 1995 Adrenocortisol function during septic shock. Intensive Care Med 21:57-62.
18. Reincke M, Allolio B, Wurth G 1993 The hypothalamic-pituitary-adrenal axis in critical illness: Response to dexamethasone and corticotropin-releasing hormone. J Clin Endocrinol Metab 77:151-156.
19. Sibbald WJ, Short A, Cohen MP 1977 Variations in adrenocortical responsiveness during severe bacterial infections. Unrecognized adrenocortical insufficiency in severe bacterial infections. Ann Surg 186:29-33.
20. Briegel J, Scheelling G, Haller M 1996 A comparison of the adrenocortical response during septic shock and after complete recovery. Intensive Care Med 22:894-899.
21. Beishuizen A, Vermes I, Hylkema BS 1999 Relative eosinophilia and functional adrenal insufficiency in critically ill patients. Lancet 353:1675-1676.
22. Hill CD, Singer B 1968 Inhibition of the response to pituitary adrenocorticotrophic hormone in the hypophysectomized rat by circulatory corticosterone. J Endocrinol 42:301-309.
23. Carsia RV, MacDonald GJ, Malamed S 1983 Steroid control of steroidogenesis in isolated rat adrenocortical cells: molecular and species specificity. Steroids 41:741-746.
24. Morrow LB, Burrow GN, Mulrow PF 1967 Inhibition of adrenal protein synthesis by steroids in vitro. Endocrinology 80:883-888.
25. Carsia RV, Malamed S 1983 Glucocorticoid control of steroidogenesis in isolated rat adrenocortical cells. Biochem Biophys Acta 763:83-89.
26. Kahri AI, Voutilainen R, Salmenpera M 1979 Different biological action of corticosteroids, corticosterone, and cortisol, as a base of zonal function of the adrenal cortex. Acta Endocrinol 91:329-337.
27. Hornsby PJ 1982 Regulation of 21-hydroxylase activity by steroids in cultured bovine adrenocortical cells: possible significance for adrenocortical androgen synthesis. Endocrinology 111:1092-1101.
28. Saito E, Mukai M, Muraki T, Ichikawa Y, Homma M 1979 Inhibitory effects of corticosterone on cell proliferation and steroidogenesis in the mouse adrenal tumor cell. Endocrinology 104:487-492.
29. Carsia RV, Scanes CG, Malamed S 1984 Self-suppression of corticosteroidogenesis: evidence for a role of adrenal 5a-reductase. Endocrinology 115:2464-2472.
30. Musajo F, Neri G, Tortorella C, Mazzocchi G, Nussdorfer GG 1996 Intra-adrenal 11ß-hydroxysteroid dehydrogenase plays a role in the regulation of corticosteroid secretion: an in vitro study in the rat. Life Sci 59:1401-1406.
31. Loose DS, Do YS, Chen TL, Feldman D 1980 Demonstration of glucocorticoid receptors in the adrenal cortex: evidence for a direct dexamethasone suppressive effect on the rat adrenal gland. Endocrinology 107:137-146.
32. Trzeciak WH, LeHoux JG, Waterman MW, Simpson ER 1993 Dexamethasone inhibits corticotropin-induced accumulation of CYP11A and CYP17 messenger RNAs in bovine adrenocortical cells. Mol Endocrinol 7:206-213.
33. Picard-Hagen N, Penhoat A, Hue D, Jaillard C, Durand P 1997 Glucocorticoids enhance corticotropin receptor mRNA levels in ovine adrenocortical cells. J Mol Endocrinol 19:20-36.
34. Fassnacht M, Beuschlein F, Vay S, Mora P, Allolio B, Reincke M 1998 Aminoglutethimide suppresses adrenocorticotropin receptor expression in the NCI-h295 adrenocortical tumer cell line. J Endocrinol 159:35-42.
35. Darbeida H, Durand P 1988 Glucocorticoid enhancement of glucocorticoid production by cultured ovine adrenocortical cells. Biochim Biophys Acta 972:200-208.
36. Truss M, Beato M 1993 Steroid hormone receptors: interaction with deoxyribonucleic acid and transcription factors. Endocr Rev 14:459-479.
37. Rosewicz S, McDonald AR, Maddux BA, Goldfine ID, Miesfeld RL, Logsdon CD 1988 Mechanism of glucocorticoid receptor downregulation by glucocorticoids. J Biol Chem 263:2581-2584.
38. Yudt RM, Cidlowski JA 2002 The glucocorticoid receptor: coding a diversity of proteins and responses through a single gene. Mol Endocrinol 16:1719-1726.
39. Breslin MB, Geng CD, Vedeckis WV 2001 Multiple promoters exist in the human GR gene, one of which is activated by glucocorticoids. Mol Endocrinol 15:1381-1395.
40. Wallace AD Cidlowski JA 2001 Proteasome-mediated glucocorticoid receptor degradation restricts transcriptional signaling by glucocorticoids. J Biol chem. 276:42714-42721.
41. Jefcoate CR, McNamara BC, Artemenko I, Yamazaki T 1992 Regulation of cholesterol movement to mitochondrial cytochrome P450scc in steroid hormone biosynthesis. J Steroid Biochem Mol Biol 43:751-767.
42. Clark BJ, Well SJ, King SR, Stocco DM 1994 The purification, cloning, and expression of a novel luteinizing hormone-induced mitochondrial protein in MA-10 mouse Leydig tumor cells. J Biol Chem 269:28314-28322.
43. Stocco DM, Clark BJ 1996 Regulation of the acute production of steroids in steroidogenic cells. Endocr Rev 17:221-244.
44. Lin D, Sugawara T, Strauss III JF, Clark BJ, Stocco DM, Saenger P, Rogol A, Miller WL 1995 Role of steroidogenic acute regulatory protein in adrenal and gonadal steroidogenesis. Science 267:1828-1831.
45. Caron KM, Soo SC, Wetsel WC, Stocco DM, Clark BJ, Parker KL 1997 Targeted disruption of the mouse gene encoding steroidogenic acute regulatory protein provides insights into congenital lipoid adrenal hyperplasia. Proc Natl Acad Sci USA 94:11540-11545.
46. Furukawa A, Miyatake A, Ohnishi T, Ichikawa Y 1998 Steroidogenic acute regulatory protein (StAR) transcripts constitutively expressed in the adult rat central nervous system colocalization of StAR, cytochrome P450scc (CYP 11A1), and 3-beta-hydroxysteroid dehydrogenase in the rat brain. J Neurochem 71:2231-2238.
47. Bose HS, Sugawara T, Straus III JF, Miller WL 1996 The pathophysiology and genetics of congenital lipoid adrenal hyperplasia - International Congenital Lipoid Adrenal Hyperplasia Consortium. N Engl J Med 335:1870-1878.
48. Mcallister JM, Hornsby PJ 1988 Dual regulation of 3b-hydroxysteroid dehydrogenase, 17a-hydroxylase and dehydroepiandrosterone sufotransferase by adenosine 3’-5’-monophosphate and activators of protein kinase C in cultured human adrenocortical cells. Endocrinology 122:2012-2018.
49. Sano Y, Suzuki K, Arai K, Okinaga S, Tamaoki BI 1981 Changes in enzyme activity related to steroidogenesis in human ovaries during the menstrual cycle. J Clin Endocrinol Metab 52:994-1001.
50. Rheaume E, Lachance Y,. Zhao HF, Breton N, Dumont M, de Launoit Y, Trudel C, Luu-The V, Simard J, Labrie F 1991 Structure and expression of a new complementary DNA encoding the almost exclusive 3b-hydroxysteroid dehydrogenase/delta 5–delta 4-isomerase in human adrenals and gonads. Mol Endocrinol 5:1147-1157.
51. Feltus FA, Cote S, Simard J, Gingras S, Kovac WJ, Nicholson WE, Clark BJ, Melner MH 2002 Glucocorticoids enhance activation of the human type II
3b-hydroxysteroid dehydrogenase/△5–△4 isomerase gene. J Steroid Biochem Mol Biol 82:55-63.
52. Feltus FA, Groner B, Melner MH 1999 Stat5-mediated regulation of the human type II 3b-hydroxysteroid dehydrogenase/△5–△4 isomerase gene: activation by prolactin. Mol Endocrinol 13:1084-1093.
53. Grunstein M 1997 Histone acetylation and chromatin structure and transcription. Nature 389:349-352.
54. Kodonaga JT 1998 Eukaryotic transcription: an interlaced network of transcription factors and chromatin-modifying machines. Cell 92:307-313.
55. Christenson LK, Stouffer RL, Strauss III JF 2001 Quantitative analysis of the hormone-induced hyperacetylation of histone H3 associated with the steroidogenic acute regulatory protein gene promoter. J Biol Chem 276:27392-27399.
56. Hu MC, Gup IC, Lin JH, Chung BC 1991 Regulated expression of cytochrome P-450scc (cholesterol-side chain cleavage enzyme) in cultured cell lines detected by antibody against bacterially expressed human protein. Biochem J 274:813-817.
57. Schimmer BP 1979 Adrenocortical Y1 cells. Methods Enzymol 58:570-574.
58. Lowry DH, Rosenbrough NJ, Farr AI, Randall RJ 1951 Protein mesurement with the folin phenol reagent. J Biol Chem 193:265-275.
59. Li X, Wong J, Tsai SY, Tsai MJ, O’Malley BW 2003 Progesterone and glucocorticoid receptors recruit distinct coactivator complexes and promote distinct patterns of local chromatin modification. Mol Cell Biol 23:3763-3773.
60. Pepe GJ, Albrecht ED 1990 Regulation of the primate fetal adrenal cortex. Endocr Rev 11:151-176.
61. Ilvesmaki V, Voutilainen R 1991 Interaction of phorbol ester and adrenocorticotropin in the regulation of steroidogenic P450scc genes in human fetal and adult adrenals. Endocrinology 128:1450-1458.
62. Clark BJ, Soo SC, Caron KM, Ikeda Y, Parker KL, Stocco DM 1995 Hormonal and developmental regulation of the steroidogenic acute regulatory protein. Mol Endocrinol 9:1346-1355.
63. Sugawara T, Lin D, Holt JA, Martin KO, Javitt NB, Miller WL, Strauss III JF 1995 Structure of the human steroidogenic acute regulatory protein (StAR) gene: StAR stimulates mitochondrial cholesterol 27-hydroxylase activity. Biochemistry 34:12506-12512.
64. Latner AL, Cook DB, Solanji KU 1997 Inhibition of binding of corticotropin-(1-24)-tetracorsapeptide (synachen) to membrane receptor of adrenal cortex by cortisol. Biochem J 164:477-480.
65. Hornsby PJ, Crivello JF 1983 The role of lipid peroxidation and biological antioxidants in the function of the adrenal cortex. Mol Cell Endocrinol 30:123-147.
66. King SR, Ronen-Fuhrmann T, Timberg R, Clark BJ, Orly J, Stocco DM 1995 Steroid production after in vitro transcription, translation, and mitochondrial processing of protein products of complementary deoxyribonucleic acid for steroidogenic acute regulatory protein. Endocrinology 136:5165-5176.
67. Stocco DM, Sodeman TC 1991 The 30-kDa mitochondrial protein induced by hormone stimulation in MA10 mouse Leydig tumor cells and processed from larger precursors. J Biol Chem 266:19731-19738.