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. 1996 Apr 1;97(7):1723–1731. doi: 10.1172/JCI118599

Expression of p21(WAF1/CIP1/SDI1) and p53 in apoptotic cells in the adrenal cortex and induction by ischemia/reperfusion injury.

V V Didenko 1, X Wang 1, L Yang 1, P J Hornsby 1
PMCID: PMC507237  PMID: 8601638

Abstract

p21(WAF1/CIP1/SDI1), an inhibitor of cyclin-dependent kinases, is expressed at varying levels in human adrenal glands removed during surgery or organ recovery. In glands with p21 mRNA, nuclear p21 immunoreactivity, which was occasionally extensive, colocalized with p53 immunoreactivity and DNA damage, as evidenced by in situ end-labeling. Many cells showed morphological features of apoptosis when observed by fluorescent DNA dye staining and electron microscopy. This pattern was also associated with high levels of cytoplasmic heat shock protein 70. To address the question of the origin of p21 expression in some human adrenal glands, rat adrenal glands were subjected to 30 min of ischemia followed by 8 h of reperfusion. Cells with nuclear p21 and p53 appeared in the adrenal cortex together with DNA damage detected by in situ end-labeling. Nuclear p21 immunoreactivity was also produced in adrenal tissue fragments incubated at 37 degrees C in vitro. However, in this case, p21 expression was confined to the cut edge of the tissue. In contrast, p21 in human adrenal glands, as in ischemic rat glands, was within the inner regions of the cortex, supporting an origin of the protein in vivo rather than postmortem. The p53/p21 pathway of reaction to cellular injury, potentially leading to apoptosis, may play a role in tissue damage such as that resulting from ischemia/reperfusion. In the human adrenal cortex this process may be a precursor of adrenal failure.

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Selected References

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  1. Aharoni D., Dantes A., Oren M., Amsterdam A. cAMP-mediated signals as determinants for apoptosis in primary granulosa cells. Exp Cell Res. 1995 May;218(1):271–282. doi: 10.1006/excr.1995.1156. [DOI] [PubMed] [Google Scholar]
  2. Blake M. J., Udelsman R., Feulner G. J., Norton D. D., Holbrook N. J. Stress-induced heat shock protein 70 expression in adrenal cortex: an adrenocorticotropic hormone-sensitive, age-dependent response. Proc Natl Acad Sci U S A. 1991 Nov 1;88(21):9873–9877. doi: 10.1073/pnas.88.21.9873. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bradshaw K. D., Waterman M. R., Couch R. T., Simpson E. R., Zuber M. X. Characterization of complementary deoxyribonucleic acid for human adrenocortical 17 alpha-hydroxylase: a probe for analysis of 17 alpha-hydroxylase deficiency. Mol Endocrinol. 1987 May;1(5):348–354. doi: 10.1210/mend-1-5-348. [DOI] [PubMed] [Google Scholar]
  4. Brown E. R., Coker G. T., 3rd, O'Malley K. L. Organization and evolution of the rat tyrosine hydroxylase gene. Biochemistry. 1987 Aug 11;26(16):5208–5212. doi: 10.1021/bi00390a046. [DOI] [PubMed] [Google Scholar]
  5. Brown I. R. Induction of heat shock (stress) genes in the mammalian brain by hyperthermia and other traumatic events: a current perspective. J Neurosci Res. 1990 Nov;27(3):247–255. doi: 10.1002/jnr.490270302. [DOI] [PubMed] [Google Scholar]
  6. Canman C. E., Chen C. Y., Lee M. H., Kastan M. B. DNA damage responses: p53 induction, cell cycle perturbations, and apoptosis. Cold Spring Harb Symp Quant Biol. 1994;59:277–286. doi: 10.1101/sqb.1994.059.01.032. [DOI] [PubMed] [Google Scholar]
  7. Ceccatelli S., Diana A., Villar M. J., Nicotera P. Adrenocortical apoptosis in hypophysectomized rats is selectively reduced by ACTH. Neuroreport. 1995 Jan 26;6(2):342–344. doi: 10.1097/00001756-199501000-00029. [DOI] [PubMed] [Google Scholar]
  8. Chopp M. The roles of heat shock proteins and immediate early genes in central nervous system normal function and pathology. Curr Opin Neurol Neurosurg. 1993 Feb;6(1):6–10. [PubMed] [Google Scholar]
  9. Deng C., Zhang P., Harper J. W., Elledge S. J., Leder P. Mice lacking p21CIP1/WAF1 undergo normal development, but are defective in G1 checkpoint control. Cell. 1995 Aug 25;82(4):675–684. doi: 10.1016/0092-8674(95)90039-x. [DOI] [PubMed] [Google Scholar]
  10. Di Leonardo A., Linke S. P., Clarkin K., Wahl G. M. DNA damage triggers a prolonged p53-dependent G1 arrest and long-term induction of Cip1 in normal human fibroblasts. Genes Dev. 1994 Nov 1;8(21):2540–2551. doi: 10.1101/gad.8.21.2540. [DOI] [PubMed] [Google Scholar]
  11. Dobbie J. W. Adrenocortical nodular hyperplasia: the ageing adrenal. J Pathol. 1969 Sep;99(1):1–18. doi: 10.1002/path.1710990102. [DOI] [PubMed] [Google Scholar]
  12. Dobbie J. W., Symington T. The human adrenal gland with special reference to the vasculature. J Endocrinol. 1966 Apr;34(4):479–489. doi: 10.1677/joe.0.0340479. [DOI] [PubMed] [Google Scholar]
  13. Donehower L. A., Harvey M., Slagle B. L., McArthur M. J., Montgomery C. A., Jr, Butel J. S., Bradley A. Mice deficient for p53 are developmentally normal but susceptible to spontaneous tumours. Nature. 1992 Mar 19;356(6366):215–221. doi: 10.1038/356215a0. [DOI] [PubMed] [Google Scholar]
  14. Dulić V., Kaufmann W. K., Wilson S. J., Tlsty T. D., Lees E., Harper J. W., Elledge S. J., Reed S. I. p53-dependent inhibition of cyclin-dependent kinase activities in human fibroblasts during radiation-induced G1 arrest. Cell. 1994 Mar 25;76(6):1013–1023. doi: 10.1016/0092-8674(94)90379-4. [DOI] [PubMed] [Google Scholar]
  15. Elledge S. J., Harper J. W. Cdk inhibitors: on the threshold of checkpoints and development. Curr Opin Cell Biol. 1994 Dec;6(6):847–852. doi: 10.1016/0955-0674(94)90055-8. [DOI] [PubMed] [Google Scholar]
  16. Floyd R. A., Carney J. M. Free radical damage to protein and DNA: mechanisms involved and relevant observations on brain undergoing oxidative stress. Ann Neurol. 1992;32 (Suppl):S22–S27. doi: 10.1002/ana.410320706. [DOI] [PubMed] [Google Scholar]
  17. Gannon J. V., Greaves R., Iggo R., Lane D. P. Activating mutations in p53 produce a common conformational effect. A monoclonal antibody specific for the mutant form. EMBO J. 1990 May;9(5):1595–1602. doi: 10.1002/j.1460-2075.1990.tb08279.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Gavrieli Y., Sherman Y., Ben-Sasson S. A. Identification of programmed cell death in situ via specific labeling of nuclear DNA fragmentation. J Cell Biol. 1992 Nov;119(3):493–501. doi: 10.1083/jcb.119.3.493. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Giampaolo C., Gray A. T., Olshen R. A., Szabo S. Predicting chemically induced duodenal ulcer and adrenal necrosis with classification trees. Proc Natl Acad Sci U S A. 1991 Jul 15;88(14):6298–6302. doi: 10.1073/pnas.88.14.6298. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Gottlieb R. A., Burleson K. O., Kloner R. A., Babior B. M., Engler R. L. Reperfusion injury induces apoptosis in rabbit cardiomyocytes. J Clin Invest. 1994 Oct;94(4):1621–1628. doi: 10.1172/JCI117504. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Gu Y., Turck C. W., Morgan D. O. Inhibition of CDK2 activity in vivo by an associated 20K regulatory subunit. Nature. 1993 Dec 16;366(6456):707–710. doi: 10.1038/366707a0. [DOI] [PubMed] [Google Scholar]
  22. Harper J. W., Adami G. R., Wei N., Keyomarsi K., Elledge S. J. The p21 Cdk-interacting protein Cip1 is a potent inhibitor of G1 cyclin-dependent kinases. Cell. 1993 Nov 19;75(4):805–816. doi: 10.1016/0092-8674(93)90499-g. [DOI] [PubMed] [Google Scholar]
  23. Healy E., Reynolds N. J., Smith M. D., Harrison D., Doherty E., Campbell C., Rees J. L. Up-regulation of p21WAF1/CIP1 in psoriasis and after the application of irritants and tape stripping. J Invest Dermatol. 1995 Aug;105(2):274–279. doi: 10.1111/1523-1747.ep12318430. [DOI] [PubMed] [Google Scholar]
  24. Hornsby P. J. Biosynthesis of DHEAS by the human adrenal cortex and its age-related decline. Ann N Y Acad Sci. 1995 Dec 29;774:29–46. doi: 10.1111/j.1749-6632.1995.tb17370.x. [DOI] [PubMed] [Google Scholar]
  25. Hornsby P. J., Harris S. E. Oxidative damage to DNA and replicative lifespan in cultured adrenocortical cells. Exp Cell Res. 1987 Jan;168(1):203–217. doi: 10.1016/0014-4827(87)90429-0. [DOI] [PubMed] [Google Scholar]
  26. Hornsby P. J., McAllister J. M. Culturing steroidogenic cells. Methods Enzymol. 1991;206:371–380. doi: 10.1016/0076-6879(91)06107-e. [DOI] [PubMed] [Google Scholar]
  27. Huppi K., Siwarski D., Dosik J., Michieli P., Chedid M., Reed S., Mock B., Givol D., Mushinski J. F. Molecular cloning, sequencing, chromosomal localization and expression of mouse p21 (Waf1). Oncogene. 1994 Oct;9(10):3017–3020. [PubMed] [Google Scholar]
  28. Johnson M., Dimitrov D., Vojta P. J., Barrett J. C., Noda A., Pereira-Smith O. M., Smith J. R. Evidence for a p53-independent pathway for upregulation of SDI1/CIP1/WAF1/p21 RNA in human cells. Mol Carcinog. 1994 Oct;11(2):59–64. doi: 10.1002/mc.2940110202. [DOI] [PubMed] [Google Scholar]
  29. King K. L., Cidlowski J. A. Cell cycle and apoptosis: common pathways to life and death. J Cell Biochem. 1995 Jun;58(2):175–180. doi: 10.1002/jcb.240580206. [DOI] [PubMed] [Google Scholar]
  30. Kovács K., Carroll R., Tapp E. Temporary ischaemia of the rat adrenal gland. J Pathol Bacteriol. 1966 Jan;91(1):235–240. doi: 10.1002/path.1700910127. [DOI] [PubMed] [Google Scholar]
  31. Lane D. P. Cancer. p53, guardian of the genome. Nature. 1992 Jul 2;358(6381):15–16. doi: 10.1038/358015a0. [DOI] [PubMed] [Google Scholar]
  32. Li Y., Chopp M., Zhang Z. G., Zaloga C., Niewenhuis L., Gautam S. p53-immunoreactive protein and p53 mRNA expression after transient middle cerebral artery occlusion in rats. Stroke. 1994 Apr;25(4):849–856. doi: 10.1161/01.str.25.4.849. [DOI] [PubMed] [Google Scholar]
  33. Li Y., Jenkins C. W., Nichols M. A., Xiong Y. Cell cycle expression and p53 regulation of the cyclin-dependent kinase inhibitor p21. Oncogene. 1994 Aug;9(8):2261–2268. [PubMed] [Google Scholar]
  34. Lindh A., Carlström K., Eklund J., Wilking N. Serum steroids and prolactin during and after major surgical trauma. Acta Anaesthesiol Scand. 1992 Feb;36(2):119–124. doi: 10.1111/j.1399-6576.1992.tb03436.x. [DOI] [PubMed] [Google Scholar]
  35. Lovis C., Mach F., Donati Y. R., Bonventre J. V., Polla B. S. Heat shock proteins and the kidney. Ren Fail. 1994;16(2):179–192. doi: 10.3109/08860229409044859. [DOI] [PubMed] [Google Scholar]
  36. MacManus J. P., Hill I. E., Huang Z. G., Rasquinha I., Xue D., Buchan A. M. DNA damage consistent with apoptosis in transient focal ischaemic neocortex. Neuroreport. 1994 Jan 12;5(4):493–496. doi: 10.1097/00001756-199401120-00031. [DOI] [PubMed] [Google Scholar]
  37. McAllister J. M., Hornsby P. J. Improved clonal and nonclonal growth of human, rat and bovine adrenocortical cells in culture. In Vitro Cell Dev Biol. 1987 Oct;23(10):677–685. doi: 10.1007/BF02620980. [DOI] [PubMed] [Google Scholar]
  38. Mehta H. B., Popovich B. K., Dillmann W. H. Ischemia induces changes in the level of mRNAs coding for stress protein 71 and creatine kinase M. Circ Res. 1988 Sep;63(3):512–517. doi: 10.1161/01.res.63.3.512. [DOI] [PubMed] [Google Scholar]
  39. Meikrantz W., Schlegel R. Apoptosis and the cell cycle. J Cell Biochem. 1995 Jun;58(2):160–174. doi: 10.1002/jcb.240580205. [DOI] [PubMed] [Google Scholar]
  40. Michieli P., Chedid M., Lin D., Pierce J. H., Mercer W. E., Givol D. Induction of WAF1/CIP1 by a p53-independent pathway. Cancer Res. 1994 Jul 1;54(13):3391–3395. [PubMed] [Google Scholar]
  41. Naseeruddin S. A., Hornsby P. J. Regulation of 11 beta- and 17 alpha-hydroxylases in cultured bovine adrenocortical cells: 3', 5'-cyclic adenosine monophosphate, insulin-like growth factor-I, and activators of protein kinase C. Endocrinology. 1990 Oct;127(4):1673–1681. doi: 10.1210/endo-127-4-1673. [DOI] [PubMed] [Google Scholar]
  42. Neville A. M. The nodular adrenal. Invest Cell Pathol. 1978 Jan-Mar;1(1):99–111. [PubMed] [Google Scholar]
  43. Noda A., Ning Y., Venable S. F., Pereira-Smith O. M., Smith J. R. Cloning of senescent cell-derived inhibitors of DNA synthesis using an expression screen. Exp Cell Res. 1994 Mar;211(1):90–98. doi: 10.1006/excr.1994.1063. [DOI] [PubMed] [Google Scholar]
  44. Oonk R. B., Krasnow J. S., Beattie W. G., Richards J. S. Cyclic AMP-dependent and -independent regulation of cholesterol side chain cleavage cytochrome P-450 (P-450scc) in rat ovarian granulosa cells and corpora lutea. cDNA and deduced amino acid sequence of rat P-450scc. J Biol Chem. 1989 Dec 25;264(36):21934–21942. [PubMed] [Google Scholar]
  45. Parker S. B., Eichele G., Zhang P., Rawls A., Sands A. T., Bradley A., Olson E. N., Harper J. W., Elledge S. J. p53-independent expression of p21Cip1 in muscle and other terminally differentiating cells. Science. 1995 Feb 17;267(5200):1024–1027. doi: 10.1126/science.7863329. [DOI] [PubMed] [Google Scholar]
  46. Pines J. Cell cycle. p21 inhibits cyclin shock. Nature. 1994 Jun 16;369(6481):520–521. doi: 10.1038/369520a0. [DOI] [PubMed] [Google Scholar]
  47. Pontén F., Berne B., Ren Z. P., Nistér M., Pontén J. Ultraviolet light induces expression of p53 and p21 in human skin: effect of sunscreen and constitutive p21 expression in skin appendages. J Invest Dermatol. 1995 Sep;105(3):402–406. doi: 10.1111/1523-1747.ep12321071. [DOI] [PubMed] [Google Scholar]
  48. Rao R. H. Bilateral massive adrenal hemorrhage. Med Clin North Am. 1995 Jan;79(1):107–129. doi: 10.1016/s0025-7125(16)30087-6. [DOI] [PubMed] [Google Scholar]
  49. Ryzhavskii B. Ia. Izmenenie kory nadpochechnikov krys pri stresse posle udaleniia gipofiza, shchitovidnoi zhelezy i iaichka. Arkh Anat Gistol Embriol. 1978 Apr;74(4):40–46. [PubMed] [Google Scholar]
  50. Sato T., Koyama K., Takemasa T., Yoshida S., Sato Y., Omokawa S., Lygidakis N. J. Damage and repair of hepatocyte nuclear DNA after hepatic inflow occlusion. Hepatogastroenterology. 1992 Jun;39(3):252–256. [PubMed] [Google Scholar]
  51. Schumer M., Colombel M. C., Sawczuk I. S., Gobé G., Connor J., O'Toole K. M., Olsson C. A., Wise G. J., Buttyan R. Morphologic, biochemical, and molecular evidence of apoptosis during the reperfusion phase after brief periods of renal ischemia. Am J Pathol. 1992 Apr;140(4):831–838. [PMC free article] [PubMed] [Google Scholar]
  52. Sheikh M. S., Li X. S., Chen J. C., Shao Z. M., Ordonez J. V., Fontana J. A. Mechanisms of regulation of WAF1/Cip1 gene expression in human breast carcinoma: role of p53-dependent and independent signal transduction pathways. Oncogene. 1994 Dec;9(12):3407–3415. [PubMed] [Google Scholar]
  53. Shivji M. K., Grey S. J., Strausfeld U. P., Wood R. D., Blow J. J. Cip1 inhibits DNA replication but not PCNA-dependent nucleotide excision-repair. Curr Biol. 1994 Dec 1;4(12):1062–1068. doi: 10.1016/s0960-9822(00)00244-x. [DOI] [PubMed] [Google Scholar]
  54. Slebos R. J., Lee M. H., Plunkett B. S., Kessis T. D., Williams B. O., Jacks T., Hedrick L., Kastan M. B., Cho K. R. p53-dependent G1 arrest involves pRB-related proteins and is disrupted by the human papillomavirus 16 E7 oncoprotein. Proc Natl Acad Sci U S A. 1994 Jun 7;91(12):5320–5324. doi: 10.1073/pnas.91.12.5320. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Smith J. R. Inhibitors of DNA synthesis derived from senescent human diploid fibroblasts. Exp Gerontol. 1992 Jul-Aug;27(4):409–412. doi: 10.1016/0531-5565(92)90073-9. [DOI] [PubMed] [Google Scholar]
  56. Wyllie A. H. Apoptosis. Death gets a brake. Nature. 1994 May 26;369(6478):272–273. doi: 10.1038/369272a0. [DOI] [PubMed] [Google Scholar]
  57. Wyllie A. H., Kerr J. F., Currie A. R. Cell death: the significance of apoptosis. Int Rev Cytol. 1980;68:251–306. doi: 10.1016/s0074-7696(08)62312-8. [DOI] [PubMed] [Google Scholar]
  58. Wyllie A. H., Kerr J. F., Macaskill I. A., Currie A. R. Adrenocortical cell deletion: the role of ACTH. J Pathol. 1973 Oct;111(2):85–94. doi: 10.1002/path.1711110203. [DOI] [PubMed] [Google Scholar]
  59. Xiong Y., Hannon G. J., Zhang H., Casso D., Kobayashi R., Beach D. p21 is a universal inhibitor of cyclin kinases. Nature. 1993 Dec 16;366(6456):701–704. doi: 10.1038/366701a0. [DOI] [PubMed] [Google Scholar]
  60. Yang L., Didenko V. V., Noda A., Bilyeu T. A., Darlington D. J., Smith J. R., Hornsby P. J. Increased expression of p21Sdi1 in adrenocortical cells when they are placed in culture. Exp Cell Res. 1995 Nov;221(1):126–131. doi: 10.1006/excr.1995.1359. [DOI] [PubMed] [Google Scholar]
  61. Zhu L., Harlow E., Dynlacht B. D. p107 uses a p21CIP1-related domain to bind cyclin/cdk2 and regulate interactions with E2F. Genes Dev. 1995 Jul 15;9(14):1740–1752. doi: 10.1101/gad.9.14.1740. [DOI] [PubMed] [Google Scholar]
  62. el-Deiry W. S., Harper J. W., O'Connor P. M., Velculescu V. E., Canman C. E., Jackman J., Pietenpol J. A., Burrell M., Hill D. E., Wang Y. WAF1/CIP1 is induced in p53-mediated G1 arrest and apoptosis. Cancer Res. 1994 Mar 1;54(5):1169–1174. [PubMed] [Google Scholar]
  63. el-Deiry W. S., Tokino T., Velculescu V. E., Levy D. B., Parsons R., Trent J. M., Lin D., Mercer W. E., Kinzler K. W., Vogelstein B. WAF1, a potential mediator of p53 tumor suppression. Cell. 1993 Nov 19;75(4):817–825. doi: 10.1016/0092-8674(93)90500-p. [DOI] [PubMed] [Google Scholar]
  64. el-Deiry W. S., Tokino T., Waldman T., Oliner J. D., Velculescu V. E., Burrell M., Hill D. E., Healy E., Rees J. L., Hamilton S. R. Topological control of p21WAF1/CIP1 expression in normal and neoplastic tissues. Cancer Res. 1995 Jul 1;55(13):2910–2919. [PubMed] [Google Scholar]

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