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Interleukin-2 and Interleukin-2 Receptor Expression in Human Corticotrophic Adenoma and Murine Pituitary Cell Cultures

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Interleukin-2 and Interleukin-2 Receptor Expression in Human Corticotrophic Adenoma and Murine Pituitary Cell Cultures Interleukin-2 and interleukin-2 receptor expression in human corticotrophic adenoma and murine pituitary cell cultures. E Arzt, … , O A Müller, G K Stalla J Clin Invest. 1992;90(5):1944-1951. https://doi.org/10.1172/JCI116072. Research Article The production of IL-1 and IL-6 by pituitary cells has recently been demonstrated. In this study we investigated the expression of IL-2 and its receptor (IL-2R) by pituitary cells of different species. In Northern blots, a single hybridizing band of 1 kb, identical to that in normal stimulated lymphocytes, was obtained with specific IL-2 probes. In the mouse AT- 20 pituitary tumor cell line, IL-2 mRNA expression was detected after stimulation with corticotropin-releasing hormone or phorbol myristate acetate. In human corticotrophic adenoma cells, basal IL-2 mRNA expression as well as IL-2 secretion were further stimulated by phorbol myristate acetate. Both adenoma and AtT-20 cells showed detectable amounts of IL- 2R mRNA and by immunofluorescence, IL-2R membrane expression. In addition, dual immunofluorescence studies in rat anterior pituitary cells demonstrated colocalization of IL-2R with ACTH-positive cells and other cell types expressing the receptor. In addition to the action of lymphocyte-produced IL-2, this cytokine may have a paracrine or autocrine regulatory role within the pituitary. It remains to be established whether IL-2 production occurs in the normal pituitary or is intrinsic to the process of tumor development of these cells. IL-2 may be involved in the growth control of pituitary cells. Find the latest version: https://jci.me/116072/pdf Interleukin-2 and Interleukin-2 Receptor Expression in Human Corticotrophic Adenoma and Murine Pituitary Cell Cultures Eduardo Arzt, Gertraud Steizer, Ulrich Renner, Manfred Lange,* 0. Albrecht Moiler,* and Ginter K. Stalla Max-Planck-Institute ofPsychiatry, Clinical Institute, 8000 Munich 40; and Departments of *Neurosurgery and Medicine, University ofMunich, 8000 Munich 2, Federal Republic ofGermany Abstract duction by pituitary cells has been demonstrated. IL- Ij3 immu- The production of IL-1 and IL-6 by pituitary cells has recently noreactive material and mRNA was found in rat pituitaries been demonstrated. In this study we investigated the expres- and shown to increase after bacterial lipopolysaccharide treat- sion of IL-2 and its receptor cells of differ- ment of the rats (11). IL-I receptors were characterized in (IL-2R) by pituitary mouse ent species. In Northern blots, a single hybridizing band of 1 pituitary cells and AtT-20 corticotrophs (12). IL-6 pro- kb, identical to that in normal stimulated lymphocytes, was duction and stimulation by IL- I has been demonstrated in rat IL-2 In the mouse AT-20 anterior pituitary cells (13-17). The expression of IL-6 mRNA obtained with specific probes. pitu- in rat itary tumor cell line, IL-2 mRNA expression was detected after anterior pituitary (10) and corticotrophic adenoma (18) stimulation with corticotropin-releasing hormone or phorbol cell cultures and the release of IL-6 from human pituitary ade- myristate acetate. In human adenoma cells, ba- noma cultures (19) have also been reported. The stimulatory corticotrophic IL- 1 sal IL-2 mRNA as well as IL-2 secretion were fur- action of on the hypothalamo-pituitary-adrenal axis has expression been ther stimulated by phorbol myristate acetate. Both adenoma extensively studied. Despite discrepancies in the litera- and AtT-20 cells showed detectable amounts of IL-2R mRNA ture, it seems to act at the hypothalamic, pituitary, and adrenal and by immunofluorescence, IL-2R membrane expression. In levels to induce corticotropin-releasing hormone (nCRHe), cor- addition, dual immunofluorescence studies in rat anterior pitu- ticotropin (ACTH), and cortisol production (20, reviewed in itary cells demonstrated colocalization of IL-2R with ACHI- 21 and 22). Similar results were found for IL-6 (21 and 22). It positive cells and other cell types expressing the receptor. In was postulated, therefore, that IL- I and IL-6 may be involved addition to the action of lymphocyte-produced IL-2, this cyto- in the paracrine or autocrine regulation of pituitary function kine may have a paracrine or autocrine regulatory role within (11, 14). IL-2 also acts on the hypothalamo-pituitary-adrenal the pituitary. It remains to be established whether IL-2 produc- axis. It has been shown to enhance proopiomelanocortin gene tion occurs in the normal pituitary or is intrinsic to the process expression (23, 24) and to induce ACTH release in the AtT-20 of tumor development of these cells. IL-2 may be involved in murine cell line and in rat anterior pituitary cell cultures (25- the growth control of pituitary cells. (J. Clin. Invest. 1992. 27). In the latter, it also induces the release of other hormones, such as prolactin (25, 28). When administered to human 90:1944-1951.) Key words: neuroimmunology * cytokines - hy- pothalamo-pituitary-adrenal axis * anterior pituitary * interleu- cancer patients, it increases f3-endorphin, ACTH, and cortisol kin-2 pathway levels (29, 30). Further, it stimulates f3-endorphin and ACTH levels in allografted (31) and normal (32) rats, respectively. Introduction Several reports, however, failed to find effects of IL-2 in these systems, though more recently, a report suggests that this may It has been described that the central nervous system is a target be related to species-specific IL-2 forms (32). Here it was or source of lymphokines ( 1-5). Receptors for IL- I and IL-2 shown that while rat IL-2 enhances ACTH release in rats, hu- have been found in the central nervous system, mainly local- man (hu) IL-2 has no effect on rat ACTH levels (32). As IL-I ized in the hippocampus (5-8). IL- I(# immunoreactive fibers and IL-6 are localized in pituitary cells, and as a protein related were found in the hypothalamus, innervating key endocrine to the a-chain of the IL-2 receptor (IL-2R) has been shown in cell groups (9). Rat medial basal hypothalamic cultures release these cells (27), we have investigated whether IL-2 and its re- IL-6 and express its mRNA (10). Recently, IL- 1 and IL-6 pro- ceptor could be synthesized by pituitary cells. Methods Portions of this work were presented at the 1 1th Meeting of the Euro- pean Federation of Immunological Societies, Helsinki, Finland, 9-12 Patients. Nine patients (eight women and one man) aged 16-55 yr, June 1991, and at the 5th Meeting of the European Neuroendocrine with typical features of chronic hypercortisolism, including facial and Association, Budapest, Hungary, 25-28 August 1991, and were pub- truncal obesity and hypertension, were studied. Of these patients, eight lished in abstract form (1991. J. Endocrinol. Invest. 14 [Suppl. 4- had hypogonadism and two had hypothyroidism. The presence of a 6]:188). corticotrophic adenoma was suspected by endocrine evaluation. Mean Address reprint requests to Dr. Eduardo Arzt, Max-Planck-Insti- plasma cortisol concentrations, derived from five measurements at tute of Psychiatry, Clinical Institute, Kraepelinstrasse 10, 8000 Munich 4-5-h intervals within a 24-h period, were determined in the immedi- 40, FRG. ate preoperative period and showed a canceled dtUrnal rhythm of secre- Receivedfor publication 10 September 1991 and in revisedform 12 tion. The range of mean plasma cortisol concentrations was 486-1,242 May 1992. nmol/liter (normal: 250-650 nmol/liter). A low dose dexamethasone (2 mg) suppression test failed to produce suppression of cortisol secre- J. Clin. Invest. © The American Society for Clinical Investigation, Inc. 1. Abbreviations used in this paper: CRH, corticotropin-releasing hor- 0021-9738/92/11/1944/08 $2.00 mone; hu, human; IL-2R, IL-2 receptor; POMC, proopiomelanocor- Volume 90, November 1992, 1944-1951 tin. 1944 E. Arzt, G. Stelzer, U. Renner, M. Lange, 0. A. Miller, and G. K Stalla tion. In the insulin-hypoglycemia test (0.15 IU/kg body wt), patients guanidine isothiocyanate followed by phenol-chloroform method did not show an appropriate response ofcortisol secretion. The range of (37). RNA extraction, electrophoresis, blotting, radiolabeling of basal plasma ACTH concentrations was 10-50 pmol/liter (normal: probes, and hybridization were performed as described (33, 34). 4-17 pmol/liter). All patients responded to a large dose ofdexametha- Briefly, 5-10 sg RNA was denatured with glyoxal, electrophoresed on a sone (8 mg) with a significant decrease in plasma cortisol levels. In 1.2% agarose gel, and transferred overnight to a nylon membrane: Hy- addition, every patient was subjected to a sinus-petrosus-inferior cathe- bond-N (Amersham International, Amersham, UK) or Nytran-N terization in combination with a CRH stimulation test for the determi- (Schleicher and Schuell, Dassel, FRG). Filters baked for 2 h at 80'C nation of pituitary origin and localization of ACTH secretion. Radio- were prehybridized for 4 h at 60C (50% formamide, 5X sodium chlo- logical investigation consisted of a nuclear magnetic resonance scan. ride sodium phosphate EDTA buffer [SSPE], 5X Denhardt's solution, All patients were persistently cured after transsphenoidal microsurgery. 0.1% SDS, 100 Ag/ml denatured salmon sperm DNA) and then hybrid- Tumors. After surgery the adenoma tissue was transported in the ized with the addition of the probe at the same temperature for 12 h. sterile culture medium described below. Routinely, the tumor trans- Blots were washed at increasing salt and temperature stringency with a port medium and the conditioned medium obtained after 24 h of cul- final wash of 30 min at 60C in 0.1X standard saline citrate (SSC) ture as described below, were assayed for all anterior pituitary hor- containing 1% SDS. Dried filters were exposed to Kodak XAR5 film at mones. The nine tumors were selected for these studies because they -70'C with intensifying screens for 6 h-4 d.
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