Skip to main content
NCBI home page
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1994 Sep;94(3):1226–1234. doi: 10.1172/JCI117440

Gene expression, localization, and characterization of endothelin A and B receptors in the human adrenal cortex.

G Rossi 1, G Albertin 1, A Belloni 1, L Zanin 1, M A Biasolo 1, T Prayer-Galetti 1, M Bader 1, G G Nussdorfer 1, G Palù 1, A C Pessina 1
PMCID: PMC295207  PMID: 8083364

Abstract

Compelling evidence indicates that the endothelium-derived potent vasoconstrictor endothelin-1 (ET-1) stimulates aldosterone secretion by interacting with specific receptors. Although two different ET-1 receptors have been identified and cloned, the receptor subtype involved in mediating aldosterone secretion is still unknown. Accordingly, we wished to investigate whether the genes of ET-1 and of its receptors A and B are expressed in the normal human adrenal cortex. We designed specific primers for ET-1 and the ETA and ETB receptors genes and developed a reverse transcription polymerase chain reaction (RT-PCR) with chemiluminescent quantitation of the cDNA. In addition, we carried out 125I ET-1 displacement studies with cold ET-1, ET-3 and the specific ETA and ETB ligands BQ123 and sarafotoxin 6C. Localization of each receptor subtype was also investigated by autoradiography. Binding experiments were first individually analyzed by Scatchard and Hofstee plot and then coanalyzed by the nonlinear iterative curve fitting program Ligand. Histologically normal adrenal cortex tissue, obtained from kidney cancer patients (n = 7), and an aldosterone-producing adenoma (APA), which is histogenetically derived from the zona glomerulosa (ZG) cells, were studied. Results showed that the ET-1, ETA and ETB mRNA can be detected by RT-PCR in all adrenal cortices as well as in the APA. The best fitting of the 125I ET-1 displacement binding data was consistently provided by a two-site model both in the normal adrenal cortex (F = 22.1, P < 0.0001) and in the APA (F = 18.4, P < 0.0001). In the former the density (Bmax) of the ETA and ETB subtype was 2.6 +/- 0.5 pmol/mg protein (m +/- SEM) and 1.19 +/- 0.6, respectively. The dissociation constant (Kd) of ET-1, ET-3, S6C, and BQ-123 for each receptor subtype resulted to be within the range reported for human tissue for the ETA and ETB receptors. In the APA tissue the Bmax tended to be lower (1.33 and 0.8 pmol/mg protein, for the ETA and ETB, respectively) but the Kd were similar. Autoradiographic studies confirmed the presence of both receptor subtypes on the ZG as well as on APA cells. Thus, the genes of ET-1 and both its receptor subtypes ETA and ETB are actively transcribed in the human adrenal cortex. Furthermore, both receptor subtypes are translated into proteins in ZG and APA cells.

Full text

PDF
1226

Images in this article

1229Image
on p.1229
1229Image
on p.1229
1229Image
on p.1229
1229Image
on p.1229
1230Image
on p.1230
1231Image
on p.1231
1232Image
on p.1232

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Arai H., Nakao K., Takaya K., Hosoda K., Ogawa Y., Nakanishi S., Imura H. The human endothelin-B receptor gene. Structural organization and chromosomal assignment. J Biol Chem. 1993 Feb 15;268(5):3463–3470. [PubMed] [Google Scholar]
  2. Auffray C., Rougeon F. Purification of mouse immunoglobulin heavy-chain messenger RNAs from total myeloma tumor RNA. Eur J Biochem. 1980 Jun;107(2):303–314. doi: 10.1111/j.1432-1033.1980.tb06030.x. [DOI] [PubMed] [Google Scholar]
  3. Cao L. Q., Banks R. O. Cardiorenal actions of endothelin, Part I: Effects of converting enzyme inhibition. Life Sci. 1990;46(8):577–583. doi: 10.1016/0024-3205(90)90125-b. [DOI] [PubMed] [Google Scholar]
  4. Clark B. A., Halvorson L., Sachs B., Epstein F. H. Plasma endothelin levels in preeclampsia: elevation and correlation with uric acid levels and renal impairment. Am J Obstet Gynecol. 1992 Mar;166(3):962–968. doi: 10.1016/0002-9378(92)91372-h. [DOI] [PubMed] [Google Scholar]
  5. Clozel M. Endothelin sensitivity and receptor binding in the aorta of spontaneously hypertensive rats. J Hypertens. 1989 Nov;7(11):913–917. doi: 10.1097/00004872-198911000-00011. [DOI] [PubMed] [Google Scholar]
  6. Cozza E. N., Chiou S., Gomez-Sanchez C. E. Endothelin-1 potentiation of angiotensin II stimulation of aldosterone production. Am J Physiol. 1992 Jan;262(1 Pt 2):R85–R89. doi: 10.1152/ajpregu.1992.262.1.R85. [DOI] [PubMed] [Google Scholar]
  7. Cozza E. N., Gomez-Sanchez C. E. Effects of endothelin-1 on its receptor concentration and thymidine incorporation in calf adrenal zona glomerulosa cells: a comparative study with phorbol esters. Endocrinology. 1990 Aug;127(2):549–554. doi: 10.1210/endo-127-2-549. [DOI] [PubMed] [Google Scholar]
  8. Cozza E. N., Gomez-Sanchez C. E., Foecking M. F., Chiou S. Endothelin binding to cultured calf adrenal zona glomerulosa cells and stimulation of aldosterone secretion. J Clin Invest. 1989 Sep;84(3):1032–1035. doi: 10.1172/JCI114226. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Davenport A. P., Nunez D. J., Hall J. A., Kaumann A. J., Brown M. J. Autoradiographical localization of binding sites for porcine [125I]endothelin-1 in humans, pigs, and rats: functional relevance in humans. J Cardiovasc Pharmacol. 1989;13 (Suppl 5):S166–S170. doi: 10.1097/00005344-198900135-00045. [DOI] [PubMed] [Google Scholar]
  10. Delarue C., Delton I., Fiorini F., Homo-Delarche F., Fasolo A., Braquet P., Vaudry H. Endothelin stimulates steroid secretion by frog adrenal gland in vitro: evidence for the involvement of prostaglandins and extracellular calcium in the mechanism of actin of endothelin. Endocrinology. 1990 Oct;127(4):2001–2008. doi: 10.1210/endo-127-4-2001. [DOI] [PubMed] [Google Scholar]
  11. Doyle V. M., Bühler F. R., Bürgisser E. Inappropriate correction for radioactive decay in fully iodinated adrenergic radioligands. Eur J Pharmacol. 1984 Apr 6;99(4):353–356. doi: 10.1016/0014-2999(84)90146-8. [DOI] [PubMed] [Google Scholar]
  12. Gomez-Sanchez C. E., Cozza E. N., Foecking M. F., Chiou S., Ferris M. W. Endothelin receptor subtypes and stimulation of aldosterone secretion. Hypertension. 1990 Jun;15(6 Pt 2):744–747. doi: 10.1161/01.hyp.15.6.744. [DOI] [PubMed] [Google Scholar]
  13. Hechler U., Becker A., Haendler B., Schleuning W. D. Stable expression of human endothelin receptors ETA and ETB by transfected baby hamster kidney cells. Biochem Biophys Res Commun. 1993 Aug 16;194(3):1305–1310. doi: 10.1006/bbrc.1993.1966. [DOI] [PubMed] [Google Scholar]
  14. Hensen J., Levenson B., Schröder K., Jereczek M., Spielberg C., Schwenn K., Gross P. Plasmaendothelin (ET) ist bei Herzinsuffizienz erhöht--Kein Einfluss einer Kurzinfusion des atrialen natriuretischen Faktors (ANF). Z Kardiol. 1991;80 (Suppl 8):101–102. [PubMed] [Google Scholar]
  15. Hinson J. P., Kapas S., Teja R., Vinson G. P. Effect of the endothelins on aldosterone secretion by rat zona glomerulosa cells in vitro. J Steroid Biochem Mol Biol. 1991;40(1-3):437–439. doi: 10.1016/0960-0760(91)90213-o. [DOI] [PubMed] [Google Scholar]
  16. Hinson J. P., Vinson G. P., Kapas S., Teja R. The role of endothelin in the control of adrenocortical function: stimulation of endothelin release by ACTH and the effects of endothelin-1 and endothelin-3 on steroidogenesis in rat and human adrenocortical cells. J Endocrinol. 1991 Feb;128(2):275–280. doi: 10.1677/joe.0.1280275. [DOI] [PubMed] [Google Scholar]
  17. Hori S., Komatsu Y., Shigemoto R., Mizuno N., Nakanishi S. Distinct tissue distribution and cellular localization of two messenger ribonucleic acids encoding different subtypes of rat endothelin receptors. Endocrinology. 1992 Apr;130(4):1885–1895. doi: 10.1210/endo.130.4.1312429. [DOI] [PubMed] [Google Scholar]
  18. Hosoda K., Nakao K., Hiroshi-Arai, Suga S., Ogawa Y., Mukoyama M., Shirakami G., Saito Y., Nakanishi S., Imura H. Cloning and expression of human endothelin-1 receptor cDNA. FEBS Lett. 1991 Aug 5;287(1-2):23–26. doi: 10.1016/0014-5793(91)80007-p. [DOI] [PubMed] [Google Scholar]
  19. Ihara M., Noguchi K., Saeki T., Fukuroda T., Tsuchida S., Kimura S., Fukami T., Ishikawa K., Nishikibe M., Yano M. Biological profiles of highly potent novel endothelin antagonists selective for the ETA receptor. Life Sci. 1992;50(4):247–255. doi: 10.1016/0024-3205(92)90331-i. [DOI] [PubMed] [Google Scholar]
  20. Imai T., Hirata Y., Eguchi S., Kanno K., Ohta K., Emori T., Sakamoto A., Yanagisawa M., Masaki T., Marumo F. Concomitant expression of receptor subtype and isopeptide of endothelin by human adrenal gland. Biochem Biophys Res Commun. 1992 Feb 14;182(3):1115–1121. doi: 10.1016/0006-291x(92)91847-j. [DOI] [PubMed] [Google Scholar]
  21. Inoue A., Yanagisawa M., Takuwa Y., Mitsui Y., Kobayashi M., Masaki T. The human preproendothelin-1 gene. Complete nucleotide sequence and regulation of expression. J Biol Chem. 1989 Sep 5;264(25):14954–14959. [PubMed] [Google Scholar]
  22. Kohzuki M., Johnston C. I., Abe K., Chai S. Y., Casley D. J., Yasujima M., Yoshinaga K., Mendelsohn F. A. In vitro autoradiographic endothelin-1 binding sites and sarafotoxin S6B binding sites in rat tissues. Clin Exp Pharmacol Physiol. 1991 Jul;18(7):509–515. doi: 10.1111/j.1440-1681.1991.tb01485.x. [DOI] [PubMed] [Google Scholar]
  23. Kohzuki M., Johnston C. I., Chai S. Y., Casley D. J., Rogerson F., Mendelsohn F. A. Endothelin receptors in rat adrenal gland visualized by quantitative autoradiography. Clin Exp Pharmacol Physiol. 1989 Apr;16(4):239–242. doi: 10.1111/j.1440-1681.1989.tb01550.x. [DOI] [PubMed] [Google Scholar]
  24. Koseki C., Imai M., Hirata Y., Yanagisawa M., Masaki T. Autoradiographic distribution in rat tissues of binding sites for endothelin: a neuropeptide? Am J Physiol. 1989 Apr;256(4 Pt 2):R858–R866. doi: 10.1152/ajpregu.1989.256.4.R858. [DOI] [PubMed] [Google Scholar]
  25. Lerman A., Click R. L., Narr B. J., Wiesner R. H., Krom R. A., Textor S. C., Burnett J. C., Jr Elevation of plasma endothelin associated with systemic hypertension in humans following orthotopic liver transplantation. Transplantation. 1991 Mar;51(3):646–650. doi: 10.1097/00007890-199103000-00020. [DOI] [PubMed] [Google Scholar]
  26. Mazzocchi G., Malendowicz L. K., Nussdorfer G. G. Endothelin-1 acutely stimulates the secretory activity of rat zona glomerulosa cells. Peptides. 1990 Jul-Aug;11(4):763–765. doi: 10.1016/0196-9781(90)90192-8. [DOI] [PubMed] [Google Scholar]
  27. Mazzocchi G., Rebuffat P., Meneghelli V., Malendowicz L. K., Kasprzak A., Nussdorfer G. G. Effects of prolonged infusion with endothelin-1 on the function and morphology of rat adrenal cortex. Peptides. 1990 Jul-Aug;11(4):767–772. doi: 10.1016/0196-9781(90)90193-9. [DOI] [PubMed] [Google Scholar]
  28. Miller W. L., Redfield M. M., Burnett J. C., Jr Integrated cardiac, renal, and endocrine actions of endothelin. J Clin Invest. 1989 Jan;83(1):317–320. doi: 10.1172/JCI113876. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Molenaar P., O'Reilly G., Sharkey A., Kuc R. E., Harding D. P., Plumpton C., Gresham G. A., Davenport A. P. Characterization and localization of endothelin receptor subtypes in the human atrioventricular conducting system and myocardium. Circ Res. 1993 Mar;72(3):526–538. doi: 10.1161/01.res.72.3.526. [DOI] [PubMed] [Google Scholar]
  30. Molinoff P. B., Wolfe B. B., Weiland G. A. Quantitative analysis of drug-receptor interactions: II. Determination of the properties of receptor subtypes. Life Sci. 1981 Aug 3;29(5):427–443. doi: 10.1016/0024-3205(81)90208-3. [DOI] [PubMed] [Google Scholar]
  31. Morishita R., Higaki J., Ogihara T. Endothelin stimulates aldosterone biosynthesis by dispersed rabbit adreno-capsular cells. Biochem Biophys Res Commun. 1989 Apr 28;160(2):628–632. doi: 10.1016/0006-291x(89)92479-0. [DOI] [PubMed] [Google Scholar]
  32. Munson P. J., Rodbard D. Ligand: a versatile computerized approach for characterization of ligand-binding systems. Anal Biochem. 1980 Sep 1;107(1):220–239. doi: 10.1016/0003-2697(80)90515-1. [DOI] [PubMed] [Google Scholar]
  33. Nakamoto H., Suzuki H., Murakami M., Kageyama Y., Ohishi A., Fukuda K., Hori S., Saruta T. Effects of endothelin on systemic and renal haemodynamics and neuroendocrine hormones in conscious dogs. Clin Sci (Lond) 1989 Nov;77(5):567–572. doi: 10.1042/cs0770567. [DOI] [PubMed] [Google Scholar]
  34. Nakamura T., Kasai K., Sekiguchi Y., Banba N., Takahashi K., Emoto T., Hattori Y., Shimoda S. Elevation of plasma endothelin concentrations during endotoxin shock in dogs. Eur J Pharmacol. 1991 Dec 3;205(3):277–282. doi: 10.1016/0014-2999(91)90910-i. [DOI] [PubMed] [Google Scholar]
  35. Ogawa Y., Nakao K., Arai H., Nakagawa O., Hosoda K., Suga S., Nakanishi S., Imura H. Molecular cloning of a non-isopeptide-selective human endothelin receptor. Biochem Biophys Res Commun. 1991 Jul 15;178(1):248–255. doi: 10.1016/0006-291x(91)91806-n. [DOI] [PubMed] [Google Scholar]
  36. Rosolowsky L. J., Campbell W. B. Endothelin enhances adrenocorticotropin-stimulated aldosterone release from cultured bovine adrenal cells. Endocrinology. 1990 Apr;126(4):1860–1866. doi: 10.1210/endo-126-4-1860. [DOI] [PubMed] [Google Scholar]
  37. Sakurai T., Yanagisawa M., Masaki T. Molecular characterization of endothelin receptors. Trends Pharmacol Sci. 1992 Mar;13(3):103–108. doi: 10.1016/0165-6147(92)90038-8. [DOI] [PubMed] [Google Scholar]
  38. Sakurai T., Yanagisawa M., Takuwa Y., Miyazaki H., Kimura S., Goto K., Masaki T. Cloning of a cDNA encoding a non-isopeptide-selective subtype of the endothelin receptor. Nature. 1990 Dec 20;348(6303):732–735. doi: 10.1038/348732a0. [DOI] [PubMed] [Google Scholar]
  39. Shichiri M., Hirata Y., Ando K., Emori T., Ohta K., Kimoto S., Ogura M., Inoue A., Marumo F. Plasma endothelin levels in hypertension and chronic renal failure. Hypertension. 1990 May;15(5):493–496. doi: 10.1161/01.hyp.15.5.493. [DOI] [PubMed] [Google Scholar]
  40. Stewart D. J., Cernacek P., Costello K. B., Rouleau J. L. Elevated endothelin-1 in heart failure and loss of normal response to postural change. Circulation. 1992 Feb;85(2):510–517. doi: 10.1161/01.cir.85.2.510. [DOI] [PubMed] [Google Scholar]
  41. Textor S. C., Wilson D. J., Lerman A., Romero J. C., Burnett J. C., Jr, Wiesner R., Dickson E. R., Krom R. A. Renal hemodynamics, urinary eicosanoids, and endothelin after liver transplantation. Transplantation. 1992 Jul;54(1):74–80. doi: 10.1097/00007890-199207000-00013. [DOI] [PubMed] [Google Scholar]
  42. Trapnell B. C. Quantitative evaluation of gene expression in freshly isolated human respiratory epithelial cells. Am J Physiol. 1993 Mar;264(3 Pt 1):L199–L212. doi: 10.1152/ajplung.1993.264.3.L199. [DOI] [PubMed] [Google Scholar]
  43. Uchihara M., Izumi N., Sato C., Marumo F. Clinical significance of elevated plasma endothelin concentration in patients with cirrhosis. Hepatology. 1992 Jul;16(1):95–99. doi: 10.1002/hep.1840160117. [DOI] [PubMed] [Google Scholar]
  44. Wang A. M., Doyle M. V., Mark D. F. Quantitation of mRNA by the polymerase chain reaction. Proc Natl Acad Sci U S A. 1989 Dec;86(24):9717–9721. doi: 10.1073/pnas.86.24.9717. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Widimsky J., Jr, Horky K., Dvorakova J. Plasma endothelin-1,2 levels in mild and severe hypertension. J Hypertens Suppl. 1991 Dec;9(6):S194–S195. [PubMed] [Google Scholar]
  46. Williams D. L., Jr, Jones K. L., Alves K., Chan C. P., Hollis G. F., Tung J. S. Characterization of cloned human endothelin receptors. Life Sci. 1993;53(5):407–414. doi: 10.1016/0024-3205(93)90644-i. [DOI] [PubMed] [Google Scholar]
  47. Williams D. L., Jr, Jones K. L., Pettibone D. J., Lis E. V., Clineschmidt B. V. Sarafotoxin S6c: an agonist which distinguishes between endothelin receptor subtypes. Biochem Biophys Res Commun. 1991 Mar 15;175(2):556–561. doi: 10.1016/0006-291x(91)91601-8. [DOI] [PubMed] [Google Scholar]
  48. Woodcock E. A., Tanner J. K., Caroccia L. M., Little P. J. Mechanisms involved in the stimulation of aldosterone production by angiotensin II, vasopressin and endothelin. Clin Exp Pharmacol Physiol. 1990 Apr;17(4):263–267. doi: 10.1111/j.1440-1681.1990.tb01318.x. [DOI] [PubMed] [Google Scholar]
  49. Yanagisawa M., Kurihara H., Kimura S., Tomobe Y., Kobayashi M., Mitsui Y., Yazaki Y., Goto K., Masaki T. A novel potent vasoconstrictor peptide produced by vascular endothelial cells. Nature. 1988 Mar 31;332(6163):411–415. doi: 10.1038/332411a0. [DOI] [PubMed] [Google Scholar]
  50. Young W. F., Jr, Hogan M. J., Klee G. G., Grant C. S., van Heerden J. A. Primary aldosteronism: diagnosis and treatment. Mayo Clin Proc. 1990 Jan;65(1):96–110. doi: 10.1016/s0025-6196(12)62114-4. [DOI] [PubMed] [Google Scholar]
  51. Zanin L., Rossi G. P., Pauletto P., Tonello M., Cargnelli G., Pessina A. C. Decreased density of endothelin-1 binding sites in aortic smooth muscle cells of spontaneously hypertensive rats. J Hypertens Suppl. 1991 Dec;9(6):S190–S191. [PubMed] [Google Scholar]
  52. Zeng Z. P., Naruse M., Guan B. J., Naruse K., Sun M. L., Zang M. F., Demura H., Shi Y. F. Endothelin stimulates aldosterone secretion in vitro from normal adrenocortical tissue, but not adenoma tissue, in primary aldosteronism. J Clin Endocrinol Metab. 1992 Apr;74(4):874–878. doi: 10.1210/jcem.74.4.1548353. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Clinical Investigation are provided here courtesy of American Society for Clinical Investigation

RESOURCES