Andreis P G, Markowska A, Champion H C, Mazzocchi G, Malendowicz L K, Nussdorfer G G
Department of Human Anatomy and Physiology, University of Padua, Italy.
Endocrinology. 2000 Jun;141(6):2098-104. doi: 10.1210/endo.141.6.7508.
The effect of adrenomedullin (ADM) on the proliferative activity of the rat adrenal cortex has been investigated in vivo, using an in situ perfusion technique of the intact left gland. ADM and other chemicals were dissolved in the perfusion medium, and the perfusion was continued for 180 min. ADM infusion concentration dependently increased the mitotic index and [3H]thymidine incorporation into DNA in the zona glomerulosa (ZG; the maximal effective concentration was 10(-8) M), but not in inner adrenocortical layers, where basal proliferative activity was negligible. The effect of 10(-8) M ADM was equipotently counteracted by both the calcitonin gene-related peptide (CGRP) type 1 receptor antagonist CGRP-(8-37) and ADM-(22-52). The adenylate cyclase inhibitor SQ-22536 (10(-4) M), the cAMP blocker Rp-cAMP-S (10(-3) M), and the protein kinase A inhibitor H-89 (10(-5) M), although counteracting the ZG proliferogenic action of 10(-9) M ACTH, did not affect the 10(-8) M ADM-elicited increase in ZG DNA synthesis. Similar results were obtained using the phospholipase C inhibitor U-73122 (10(-5) M), the inositol-1,4,5-trisphosphate antagonist D,L-myo-inositol-1,4,5-trisphosphothiate (10(-4) M), and the protein kinase C inhibitor calphostin C (10(-5) M), which, however, significantly inhibited the ZG proliferogenic effect of 10(-9) M angiotensin II. The growth-promoting action of 10(-8) M ADM was not affected by the phospholipase A2 inhibitor AACOCF3 (10(-5) M), the cyclooxygenase (COX) inhibitor indomethacin (10(-5) M), or the mixed COX/lipoxygenase inhibitor phenidone (10(-5) M). In contrast, the ZG proliferogenic effect of 10(-8) M ADM was abolished by either the tyrosine kinase (TK) inhibitor tyrphostin-23 (10(-5) M) or the mitogen-activated protein kinase (MAPK) antagonists PD-98059 and U0216 (10(-4) M). ADM (10(-8) M) stimulated TK and p42/p44 MAPK activity in dispersed ZG, but not ZF, cells, and the effect was reversed by either 10(-6) M CGRP-(8-37) and ADM-(22-52) or preincubation with 10(-5) M tyrphostin-23. Collectively, our findings indicate that 1) ADM stimulates cell proliferation in the rat ZG, through CGRP-(8-37)- and ADM-(22-52)-sensitive receptors, probably of the CGRP1 subtype; and 2) the mitogenic effect of ADM is mediated by activation of the TK-MAPK cascade, without any involvement of the adenylate cyclase/protein kinase A-, phospholipase C/protein kinase C-, and COX- or lipoxygenase-dependent signaling pathways.
采用完整左肾上腺原位灌注技术，在体内研究了肾上腺髓质素（ADM）对大鼠肾上腺皮质增殖活性的影响。将ADM和其他化学物质溶解于灌注介质中，持续灌注180分钟。ADM灌注浓度依赖性地增加了肾小球带（ZG）中的有丝分裂指数以及[3H]胸苷掺入DNA的量（最大有效浓度为10^(-8) M），但在内层肾上腺皮质中未出现这种情况，因为其基础增殖活性可忽略不计。10^(-8) M ADM的作用被降钙素基因相关肽（CGRP）1型受体拮抗剂CGRP-(8 - 37)和ADM-(22 - 52)等效拮抗。腺苷酸环化酶抑制剂SQ - 22536（10^(-4) M）、cAMP阻滞剂Rp - cAMP - S（10^(-3) M）以及蛋白激酶A抑制剂H - 89（10^(-5) M），虽然能拮抗10^(-9) M促肾上腺皮质激素（ACTH）对ZG的促增殖作用，但不影响10^(-8) M ADM引起的ZG中DNA合成增加。使用磷脂酶C抑制剂U - 73122（10^(-5) M）、肌醇 - 1,4,5 - 三磷酸拮抗剂D,L - 肌醇 - 1,4,5 - 三磷酸硫代物（10^(-4) M）以及蛋白激酶C抑制剂钙泊三醇C（10^(-5) M）也得到了类似结果，然而，这些物质显著抑制了10^(-9) M血管紧张素II对ZG的促增殖作用。10^(-8) M ADM的促生长作用不受磷脂酶A2抑制剂AACOCF3（10^(-5) M）、环氧化酶（COX）抑制剂吲哚美辛（10^(-5) M）或COX/脂氧合酶混合抑制剂非那吡啶（10^(-5) M）的影响。相反，10^(-8) M ADM对ZG的促增殖作用被酪氨酸激酶（TK）抑制剂 tyrphostin - 23（10^(-5) M）或丝裂原活化蛋白激酶（MAPK）拮抗剂PD - 98059和U0216（10^(-4) M）消除。ADM（10^(-8) M）刺激了分散的ZG细胞而非束状带（ZF）细胞中的TK和p42/p44 MAPK活性，并且该作用被10^(-6) M CGRP-(8 - 37)和ADM-(22 - 52)或与10^(-5) M tyrphostin - 23预孵育所逆转。总体而言，我们的研究结果表明：1）ADM通过CGRP-(8 - 37)和ADM-(22 - 52)敏感的受体（可能是CGRP1亚型）刺激大鼠ZG中的细胞增殖；2）ADM的促有丝分裂作用是由TK - MAPK级联的激活介导的，而腺苷酸环化酶/蛋白激酶A、磷脂酶C/蛋白激酶C以及COX或脂氧合酶依赖性信号通路均未参与其中。
