Glucocorticoids, stress, and fertility

Minerva Endocrinologica

Volumn 35, Issue 2, 2010, Pages 109-125

  Whirledge, S ^a   Cidlowski, J A ^a  

^a NATIONAL INSTITUTE OF ENVIRONMENTAL HEALTH SCIENCES   (United States)

Author keywords

Glucocorticoids; Hypothalamus; Ovary; Pituitary gland; Uterus

Indexed keywords

AMINO ACID; GLUCOCORTICOID; GLUCOSE;

ENVIRONMENTAL EXPOSURE; FERTILITY; HORMONAL REGULATION; HORMONE ACTION; HORMONE INHIBITION; HUMAN; HYPOTHALAMUS HYPOPHYSIS ADRENAL SYSTEM; NONHUMAN; OVARY; PRENATAL EXPOSURE; REVIEW; STRESS; TESTIS;

FEMALE; FERTILITY; FOLLICLE STIMULATING HORMONE; GLUCOCORTICOIDS; GONADOTROPIN-RELEASING HORMONE; HUMANS; HYPOTHALAMO-HYPOPHYSEAL SYSTEM; MALE; OVARY; PITUITARY-ADRENAL SYSTEM; PREGNANCY; STRESS, PHYSIOLOGICAL; STRESS, PSYCHOLOGICAL; TESTIS;

EID: 77957708536     PISSN: 03911977     EISSN: 18271634     Source Type: Journal    
DOI: None     Document Type: Review

References (105)

1. Rabin D, Gold PW, Margioris AN, Chrousos GP. Stress and reproduction: physiologic and pathophysiologic interactions between the stress and reproductive axes. Adv Exp Med Biol 1988;245:377-87.
2. Collu R, Gibb W, Ducharme JR. Effects of stress on the gonadal function. J Endocrinol Invest 1984;7:529-37.
3. Dubey AK, Plant TM. A suppression of gonadotropin secretion by cortisol in castrated male rhesus monkeys (Macaca mulatta) mediated by the interruption of hypothalamic gonadotropin-releasing hormone release. Biol Reprod 1985;33:423-31.
4. Kamel F, Kubajak CL. Modulation of gonadotropin secretion by corticosterone: interaction with gonadal steroids and mechanism of action. Endocrinology 1987;121:561-8. (Pubitemid 17101999)
5. Briski KP, Sylvester PW. Acute inhibition of pituitary LH release in the male rat by the glucocorticoid agonist decadron phosphate. Neuroendocrinology 1991;54:313-20.
6. Nicolaides NC, Galata Z, Kino T, Chrousos GP, Charmandari E. The human glucocorticoid receptor: molecular basis of biologic function. Steroids 2010;75:1-12.
7. Schultz R, Isola J, Parvinen M, Honkaniemi J, Wikström AC, Gustafsson JA, Pelto-Huikko M. Localization of the glucocorticoid receptor in testis and accessory sexual organs of male rat. Mol Cell Endocrinol 1993;95:115-20.
8. Tetsuka M, Milne M, Simpson GE, Hillier SG. Expression of 11beta-hydroxysteroid dehydrogenase, glucocorticoid receptor, and mineralocorticoid receptor genes in rat ovary. Biol Reprod 1999;60:330-5.
9. Bambino TH, Hsueh AJ. Direct inhibitory effect of glucocorticoids upon testicular luteinizing hormone receptor and steroidogenesis in vivo and in vitro. Endocrinology 1981;108:2142-8.
10. Hsueh AJ, Erickson GF. Glucocorticoid inhibition of FSH-induced estrogen production in cultured rat granulosa cells. Steroids 1978;32:639-48.
11. Gharib SD, Wierman ME, Shupnik MA, Chin WW. Molecular biology of the pituitary gonadotropins. Endocr Rev 1990;11:177-99.
12. Richards JS, Pangas SA. The ovary: basic biology and clinical implications. J Clin Invest 2010;120:963-72.
13. McCullagh DR. Dual endocrine activity of the testes. Science 1932;76:19-20.
14. Michel U, Farnworth P, Findlay JK. Follistatins: more than follicle-stimulating hormone suppressing proteins. Mol Cell Endocrinol 1993;91:1-11.
15. Rosenfeld P, Van Eekelen JA, Levine S, De Kloet ER. Ontogeny of the type 2 glucocorticoid receptor in discrete rat brain regions: an immunocytochemical study. Brain Res 1988;470:119-27.
16. McEwen BS, De Kloet ER, Rostene W. Adrenal steroid receptors and actions in the nervous system. Physiol Rev 1986;66:1121-88. (Pubitemid 17181300)
17. Chandran UR, Attardi B, Friedman R, Dong KW, Roberts JL, DeFranco DB. Glucocorticoid receptor-mediated repression of gonadotropin-releasing hormone promoter activity in GT1 hypothalamic cell lines. Endocrinology 1994;134:1467-74. (Pubitemid 24085606)
18. C Chandran UR, DeFranco DB. The glucocorticoid receptor is tethered to DNA-bound Oct-1 at the mouse gonadotropin-releasing hormone distal negative glucocorticoid response element. J Biol Chem 1999;274:2372-8.
19. Tsutsui K, Saigoh E, Ukena K, Teranishi H, Fujisawa Y, Kikuchi M et al. A novel avian hypothalamic peptide inhibiting gonadotropin release. Biochem Biophys Res Commun 2000;275:661-7.
20. Kriegsfeld LJ, Mei DF, Bentley GE, Ubuka T, Mason AO, Inoue K et al. Identification and characterization of a gonadotropin-inhibitory system in the brains of mammals. Proc Natl Acad Sci U S A 2006;103:2410-5.
21. Kirby ED, Geraghty AC, Ubuka T, Bentley GE, Kaufer D. Stress increases putative gonadotropin inhibitory hormone and decreases luteinizing hormone in male rats. Proc Natl Acad Sci U S A 2009;106:11324-9.
22. Matsuwaki T, Kayasuga Y, Yamanouchi K, Nishihara M. Maintenance of gonadotropin secretion by glucocorticoids under stress conditions through the inhibition of prostaglandin synthesis in the brain. Endocrinology 2006;147:1087-93.
23. Matsuwaki T, Watanabe E, Suzuki M, Yamanouchi K, Nishihara M. Glucocorticoid maintains pulsatile aecretion of luteinizing hormone under infectious stress condition. Endocrinology 2003;144:3477-2. (Pubitemid 36904425)
24. Matsuwaki T, Suzuki M, Yamanouchi K, Nishihara M. Glucocorticoid counteracts the suppressive effect of tumor necrosis factor-alpha on the surge of luteinizing hormone secretion in rats. J Endocrinol 2004;181:509-13.
25. Konsman JP, Vigues S, Mackerlova L, Bristow A, Blomqvist A. Rat brain vascular distribution of interleukin-1 type-1 receptor immunoreactivity: relationship to patterns of inducible cyclooxygenase expression by peripheral inflammatory stimuli. J Comp Neurol 2004;472:113-9.
26. Kotitschke A, Sadie-Van Gijsen H, Avenant C, Fernandes S, Hapgood JP. Genomic and nongenomic cross talk between the gonadotropin-releasing hormone receptor and glucocorticoid receptor signaling pathways. Mol Endocrinol 2009;23:1726-45.
27. Suter DE, Schwartz NB. Effects of glucocorticoids on secretion of luteinizing hormone and follicle-stimulating hormone by female rat pituitary cells in vitro. Endocrinology 1985;117:849-54.
28. Stackpole CA, Clarke IJ, Breen KM, Turner AI, Karsch FJ, Tilbrook AJ. Sex difference in the suppressive effect of cortisol on pulsatile secretion of luteinizing hormone in sheep. Endocrinology 2006;147:5921-31.
29. Kumar TR, Wang Y, Lu N, Matzuk MM. Follicle stimulating hormone is required for ovarian follicle maturation but not male fertility. Nat Genet 1997;15:201-4.
30. McGillivray SM, Thackray VG, Coss D, Mellon PL. Activin and glucocorticoids synergistically activate follicle-stimulating hormone beta-subunit gene expression in the immortalized LbetaT2 gonadotrope cell line. Endocrinology 2007;148:762-73. (Pubitemid 46143177)
31. Ringstrom SJ, McAndrews JM, Rahal JO, Schwartz NB. Cortisol in vivo increases FSH beta mRNA selectively in pituitaries of male rats. Endocrinology 1991;129:2793-5.
32. Tohei A, Kogo H. Dexamethasone increases follicle-stimulating hormone secretion via suppression of inhibin in rats. Eur J Pharmacol 1999;38669-74.
33. Kawakami S, Fujii Y, Winters SJ. Follistatin production by skin fibroblasts and its regulation by dexamethasone. Mol Cell Endocrinol 2001;172:157-67. (Pubitemid 32101396)
34. Smals AG, Kloppenborg PW, Benraad TJ. Plasma testosterone profiles in Cushing's syndrome. J Clin Endocrinol Metab 1977;45:240-5.
35. Aono T, Kurachi K, Miyata M, Nakasima A, Koshiyama K. Influence of surgical stress under general anesthesia on serum gonadotropin levels in male and female patients. J Clin Endocrinol Metab 1976;42:144-8.
36. Bernton E, Hoover D, Galloway R, Popp K. Adaptation to chronic stress in military trainees. Adrenal androgens, testosterone, glucocorticoids, IGF-1, and immune function. Ann N Y Acad Sci 1995;774:217-31.
37. Orr TE, Mann DR. Effects of restraint stress on plasma LH and testosterone concentrations, Leydig cell LH/hCG receptors, and in vitro testicular steroidogenesis in adult rats. Horm Behav 1990;24:324-41.
38. Saez JM, Morera AM, Haour F, Evain D. Effects of in vivo administration of dexamethasone, corticotropin and human chorionic gonadotropin on steroidogenesis and protein and DNA synthesis of testicular interstitial cells in prepuberal rats. Endocrinology 1977;101:1256-63.
39. Hales DB, Payne AH. Glucocorticoid-mediated repression of P450scc mRNA and de novo synthesis in cultured Leydig cells. Endocrinology 1989;124:2099-104.
40. Wang X, Walsh LP, Reinhart AJ, Stocco DM. The role of arachidonic acid in steroidogenesis and steroidogenic acute regulatory (StAR) gene and protein expression. J Biol Chem 2000;275:20204-9.
41. Payne AH, Sha LL. Multiple mechanisms for regulation of 3 beta-hydroxysteroid dehydrogenase/delta 5 - delta 4-isomerase, 17 alpha-hydroxylase/C17-20 lyase cytochrome P450, and cholesterol side-chain cleavage cytochrome P450 messenger ribonucleic acid levels in primary cultures of mouse Leydig cells. Endocrinology 1991;129:1429-35.
42. Martin LJ, Tremblay JJ. Glucocorticoids antagonize cAMP-induced Star transcription in Leydig cells through the orphan nuclear receptor NR4A1. J Mol Endocrinol 2008;41:165-75.
43. Dong Q, Salva A, Sottas CM, Niu E, Holmes M, Hardy MP. Rapid glucocorticoid mediation of suppressed testosterone biosynthesis in male mice subjected to immobilization stress. J Androl 2004;25:973-81. (Pubitemid 39425604)
44. Norfleet AM, Thomas ML, Gametchu B, Watson CS. Estrogen receptor-alpha detected on the plasma membrane of aldehyde-fixed GH3/B6/F10 rat pituitary tumor cells by enzyme-linked immunocytocnemistry. Endocrinology 1999;140:3805-14.
45. Ge RS, Hardy DO, Catterall JF, Hardy MP. Developmental changes in glucocorticoid receptor and 11beta-hydroxysteroid dehydrogenase oxidative and reductive activities in rat Leydig cells. Endocrinology 1997;138:5089-95.
46. Ge RS, Dong Q, Niu EM, Sottas CM, Hardy DO, Catterall JF et al. 11{beta}-Hydroxysteroid dehydrogenase 2 in rat leydig cells: its role in blunting glucocorticoid action at physiological levels of substrate. Endocrinology 2005;146:2657-64.
47. Hu GX, Lian QQ, Lin H, Latif SA, Morris DJ, Hardy MP et al. Rapid mechanisms of glucocorticoid signaling in the Leydig cell. Steroids 2008;73:1018-24.
48. Ge RS, Gao HB, Nacharaju VL, Gunsalus GL, Hardy MP. Identification of a kinetically distinct activity of 11beta-hydroxysteroid dehydrogenase in rat Leydig cells. Endocrinology 1997;138:2435-42.
49. Gao HB, Tong MH, Hu YQ, Guo QS, Ge R, Hardy MP. Glucocorticoid induces apoptosis in rat leydig cells. Endocrinology 2002;143:130-8.
50. Yazawa H, Sasagawa I, Nakada T. Apoptosis of testicular germ cells induced by exogenous glucocorticoid in rats. Hum Reprod 2000;15:1917-20.
51. Compton MM, Cidlowski JA. Identification of a glucocorticoid-induced nuclease in thymocytes. A potential "lysis gene" product. J Biol Chem 1987;262:8288-92.
52. Castedo M, Macho A, Zamzami N, Hirsch T, Marchetti P, Uriel J et al. Mitochondrial perturbations define lymphocytes undergoing apoptotic depletion in vivo. Eur J Immunol 1995;25:3277-84. (Pubitemid 26004929)
53. Gao HB, Tong MH, Hu YQ, You HY, Guo QS et al. Mechanisms of glucocorticoid-induced Leydig cell apoptosis. Mol Cell Endocrinol 2003;199:153-63.
54. Zorn B, Auger J, Velikonja V, Kolbezen M, Meden-Vrtovec H. Psychological factors in male partners of infertile couples: relationship with semen quality and early miscarriage. Int J Androl 2008;31:557-64.
55. Yazawa H, Sasagawa I, Ishigooka M, Nakada T. Effect of immobilization stress on testicular germ cell apoptosis in rats. Hum Reprod 1999;14:1806-10.
56. Mahmoud H, Mahmoud O, Layasadat K, Naeim A. Dexamethasone effects on Bax expression in the mouse testicular germ cells. Folia Histochem Cytobiol 2009;47:237-41.
57. Lim HY, Müller N, Herold MJ, van den Brandt J, Reichardt HM. Glucocorticoids exert opposing effects on macrophage function dependent on their concentration. Immunology 2007;122:47-53.
58. Alourfi Z, Donn RP, Stevens A, Berry A, McMaster A, Ray DW. Glucocorticoids suppress macrophage migration inhibitory factor (MIF) expression in a cell-type-specific manner. J Mol Endocrinol 2005;34:583-95. (Pubitemid 40585203)
59. Calandra T, Bernhagen J, Metz CN, Spiegel LA, Bacher M, Donnelly T et al. MIF as a glucocorticoid-induced modulator of cytokine production. Nature 1995;377:68-71.
60. Meinhardt A, Bacher M, McFarlane JR, Metz CN, Seitz J, Hedger MP et al. Macrophage migration inhibitory factor production by Leydig cells: evidence for a role in the regulation of testicular function. Endocrinology 1996;137:5090-5.
61. Lescoat G, Lescoat D, Garnier DH. Influence of adrenalectomy on maturation of gonadotrophin function in the male rat. J Endocrinol 1982;95:1-6.
62. Sugino Y, Usui T, Okubo K, Nagahama K, Takahashi T, Okuno H et al. Genotyping of congenital adrenal hyperplasia due to 21-hydroxylase deficiency presenting as male infertility: case report and literature review. J Assist Reprod Genet 2006;23:377-80. (Pubitemid 44922383)
63. Parthasarathy C, Balasubramanian K. Effects of corticosterone deficiency and its replacement on Leydig cell steroidogenesis. J Cell Biochem 2008;104:1671-83.
64. Albiston AL, Obeyesekere VR, Smith RE, Krozowski ZS. Cloning and tissue distribution of the human 11 beta-hydroxysteroid dehydrogenase type 2 enzyme. Mol Cell Endocrinol 1994;105:R11-7.
65. Benediktsson R, Yau JL, Low S, Brett LP, Cooke BE, Edwards CR et al. 11 beta-Hydroxysteroid dehydrogenase in the rat ovary: high expression in the oocyte. J Endocrinol 1992;135:53-8.
66. Condon J, Ricketts ML, Whorwood CB, Stewart PM. Ontogeny and sexual dimorphic expression of mouse type 2 11beta-hydroxysteroid dehydrogenase. Mol Cell Endocrinol 1997;127:121-8. (Pubitemid 27148372)
67. Michael AE, Evagelatou M, Norgate DP, Clarke RJ, Antoniw JW, Stedman BA et al. Isoforms of 11beta-hydroxysteroid dehydrogenase in human granulosa-lutein cells. Mol Cell Endocrinol 1997;132:43-52.
68. Michael AE, Thurston LM, Rae MT. Glucocorticoid metabolism and reproduction: a tale of two enzymes. Reproduction 2003;126:425-41.
69. Thurston LM, Norgate DP, Jonas KC, Chandras C, Kloosterboer HJ et al. Ovarian modulators of 11beta-hydroxysteroid dehydrogenase (11betaHSD) activity in follicular fluid from gonadotrophin-stimulated assisted conception cycles. Reproduction 2002;124:801-12.
70. Schreiber JR, Nakamura K, Erickson GF. Rat ovary glucocorticoid receptor: identification and characterization. Steroids 1982;39:569-84.
71. Rae MT, Niven D, Critchley HO, Harlow CR, Hillier SG. Antiinflammatory steroid action in human ovarian surface epithelial cells. J Clin Endocrinol Metab 2004;89:4538-44. (Pubitemid 39244466)
72. Yong PY, Harlow C, Thong KJ, Hillier SG. Regulation of 11beta-hydroxysteroid dehydrogenase type 1 gene expression in human ovarian surface epithelial cells by interleukin-1. Hum Reprod 2002;17:2300-6.
73. Rae MT, Price D, Harlow CR, Critchley HO, Hillier SG. Glucocorticoid receptor-mediated regulation of MMP9 gene expression in human ovarian surface epithelial cells. Fertil Steril 2009;92:703-8.
74. Michael AE, Pester LA, Curtis P, Shaw RW, Edwards CR, Cooke BA. Direct inhibition of ovarian steroidogenesis by cortisol and the modulatory role of 11 beta-hydroxysteroid dehydrogenase. Clin Endocrinol (Oxf) 1993;38:641-4.
75. Schoonmaker JN, Erickson GF. Glucocorticoid modulation of follicle-stimulating hormone-mediated granulosa cell differentiation. Endocrinology 1983;113:1356-63.
76. Yang JG, Chen WY, Li PS. Effects of glucocorticoids on maturation of pig oocytes and their subsequent fertilizing capacity in vitro. Biol Reprod 1999;60:929-36.
77. González R, Ruiz-León Y, Gomendio M, Roldan ER. The effect of glucocorticoids on ERK-1/2 phosphorylation during maturation of lamb oocytes and their subsequent fertilization and cleavage ability in vitro. Reprod Toxicol 2010;29:198-205.
78. Jimena P, Castilla JA, Peran F, Ramirez JP, Vergara F Jr, Molina R et al. Adrenal hormones in human follicular fluid. Acta Endocrinol (Copenh) 1992;127:403-6.
79. Myers M, Lamont MC, van den Driesche S, Mary N, Thong KJ, Hillier SG et al. Role of luteal glucocorticoid metabolism during maternal recognition of pregnancy in women. Endocrinology 2007;148:5769-79. (Pubitemid 350146185)
80. Thurston LM, Chin E, Jonas KC, Bujalska IJ, Stewart PM, Abayasekara DR et al. Expression of 11beta-hydroxysteroid dehydrogenase (11betaHSD) proteins in luteinizing human granulosa-lutein cells. J Endocrinol 2003;178:127-35.
81. Harlow CR, Jenkins JM, Winston RM. Increased follicular fluid total and free cortisol levels during the luteinizing hormone surge. Fertil Steril 1997;68:48-53.
82. Bever AT, Hisaw FL, Velardo JT. Inhibitory action of desoxycorticosterone acetate, cortisone acetate, and testosterone on uterine growth induced by estradiol-17beta. Endocrinology 1956;59:165-9.
83. Bitman J, Cecil HC. Differential inhibition by cortisol of estrogen-stimulated uterine responses. Endocrinology 1967;80:423-9.
84. Johnson DC, Dey SK. Role of histamine in implantation: dexamethasone inhibits estradiol-induced implantation in the rat. Biol Reprod 1980;22:1136-41.
85. Rhen T, Grissom S, Afshari C, Cidlowski JA. Dexamethasone blocks the rapid biological effects of 17beta-estradiol in the rat uterus without antagonizing its global genomic actions. Faseb J 2003;17:1849-70. (Pubitemid 37214779)
86. Rhen T, Cidlowski JA. Estrogens and glucocorticoids have opposing effects on the amount and latent activity of complement proteins in the rat uterus. Biol Reprod 2006;74:265-74.
87. Gong H, Jarzynka MJ, Cole TJ, Lee JH, Wada T, Zhang B et al. Glucocorticoids antagonize estrogens by glucocorticoid receptor-mediated activation of estrogen sulfotransferase. Cancer Res 2008;68:7386-93.
88. Levine S, Alpert M, Lewis GW. Infantile experience and the maturation of the pituitary adrenal axis. Science 1957;126:1347.
89. Kapoor A, Dunn E, Kostaki A, Andrews MH, Matthews SG. Fetal programming of hypothalamo-pituitary-adrenal function: prenatal stress and glucocorticoids. J Physiol 2006;572(Pt 1):31-44.
90. Lingas RI, Matthews SG. A short period of maternal nutrient restriction in late gestation modifies pituitary-adrenal function in adult guinea pig offspring. Neuroendocrinology 2001;73:302-11.
91. Kapoor A, Matthews SG. Short periods of prenatal stress affect growth, behaviour and hypothalamopituitary-adrenal axis activity in male guinea pig offspring. J Physiol 2005;566(Pt 3):967-77.
92. Kapoor A, Matthews SG. Prenatal stress modifies behavior and hypothalamic-pituitary-adrenal function in female guinea pig offspring: effects of timing of prenatal stress and stage of reproductive cycle. Endocrinology 2008;149:6406-15.
93. Austin MP, Leader LR, Reilly N. Prenatal stress, the hypothalamic- pituitary-adrenal axis, and fetal and infant neurobehaviour. Early Hum Dev 2005;81:917-26.
94. O'Connor TG, Ben-Shlomo Y, Heron J, Golding J, Adams D, Glover V. Prenatal anxiety predicts individual differences in cortisol in pre-adolescent children. Biol Psychiatry 2005;58:211-7. (Pubitemid 41112269)
95. Entringer S, Kumsta R, Hellhammer DH, Wadhwa PD, Wüst S. Prenatal exposure to maternal psychosocial stress and HPA axis regulation in young adults. Horm Behav 2009;55:292-8.
96. Painter RC, Roseboom TJ, Bleker OP. Prenatal exposure to the Dutch famine and disease in later life: an overview. Reprod Toxicol 2005;20:345-52.
97. Yehuda R, Engel SM, Brand SR, Seckl J, Marcus SM, Berkowitz GS. Transgenerational effects of posttraumatic stress disorder in babies of mothers exposed to the World Trade Center attacks during pregnancy. J Clin Endocrinol Metab 2005;90:4115-8.
98. Burton PJ, Waddell BJ. Dual function of 11beta-hydroxysteroid dehydrogenase in placenta: modulating placental glucocorticoid passage and local steroid action. Biol Reprod 1999;60:234-40.
99. Thompson A, Han VK, Yang K. Spatial and temporal patterns of expression of 11beta-hydroxysteroid dehydrogenase types 1 and 2 messenger RNA and glucocorticoid receptor protein in the murine placenta and uterus during late pregnancy. Biol Reprod 2002;67:1708-18.
100. Liggins GC, Fairclough RJ, Grieves SA, Kendall JZ, Knox BS. The mechanism of initiation of parturition in the ewe. Recent Prog Horm Res, 1973. 29: p. 111-59.
101. Challis, J.R.G., et al., Endocrine and paracrine regulation of birth at term and preterm. Endocr Rev 2000;21:514-50.
102. Liggins GC. Premature delivery of foetal lambs infused with glucocorticoids. J Endocrinol 1969;45:515-23.
103. Moss TJ, Doherty DA, Nitsos I, Harding R, Newnham JP. Pharmacokinetics of betamethasone after maternal or fetal intramuscular administration. Am J Obstet Gynecol 2003;189:1751-7. (Pubitemid 38056455)
104. Waddell BJ, Hisheh S, Dharmarajan AM, Burton PJ. Apoptosis in rat placenta is zone-dependent and stimulated by glucocorticoids. Biol Reprod 2000;63:1913-7.
105. Cole TJ, Blendy JA, Monaghan AP, Krieglstein K, Schmid W, Aguzzi A et al. Targeted disruption of the glucocorticoid receptor gene blocks adrenergic chromaffin cell development and severely retards lung maturation. Genes Dev 1995;9:1608-21.