Rowlinson S W, Waters M J, Lewis U J, Barnard R
Department of Physiology and Pharmacology, University of Queensland, St. Lucia, Australia.
Endocrinology. 1996 Jan;137(1):90-5. doi: 10.1210/endo.137.1.8536647.
Human GH (hGH) fragments 1-43 and 44-191 have potent in vivo effects on glucose homeostasis in rodents but cannot stimulate body growth. To assess the in vitro bioactivity of these hGH fragments we tested their activity against GH-responsive FDC-P1 cell lines expressing full-length human (h), mouse (m), or rabbit (r) GH receptors (GHR). Binding specificity and affinity was tested using GHR-containing membrane preparations from three species and recombinant hGH binding protein (hGHBP). Recombinant hGH 1-43 and recombinant 44-191 stimulated proliferation of FDC-P1-hGHR cells with half-maximal effect at approximately 2000 and 100 nM, respectively, whereas intact hGH stimulates proliferation of FDC-P1-hGHR cells with ED50 of 0.02-0.03 nM. However, these fragments had minimal effect on cells expressing mGHR or rGHR. Although 44-191 did not stimulate proliferation of FDC-P1-rGHR cells, when added to these cells in the presence of 0.23 nM hGH, it antagonized hGH action in a dose-dependent manner (ED50 at 230 nM). Binding of these GH fragments was compared using membrane preparations from rabbit liver, rabbit and mouse adipose tissue, and recombinant hGHBP. Binding competition curves were consistent, with 44-191 having at least a 10-fold lower affinity for rabbit liver GHR and rabbit adipose GHR than bovine GH and a 61-fold lower affinity for hGHBP than hGH. Binding of hGH 1-43 could not be demonstrated to GHRs in rabbit liver microsomes, adipose microsomes, or to hGHBP. HGH 1-43 did not compete for insulin binding sites in adipose microsomes. In conclusion, hGH 44-191 binds with low affinity to the GHR and at supraphysiologic levels stimulates proliferation of FDC-P1-hGHR cells. At high doses, 44-191 can also antagonize GH action in FDC-P1-rGHR cells, presumably by blocking receptor dimerization. Binding of 1-43 to GHR could not be detected, and the basis for its weak in vitro mitogenic effect remains to be elucidated. The low affinity of the fragments for cloned GHRs and low biopotency in these systems suggests that the metabolic actions of these fragments are unlikely to be mediated by the cloned GHR. This raises the possibility of a separate receptor mediating metabolic effects of these fragments.
人生长激素（hGH）片段1 - 43和44 - 191对啮齿动物的葡萄糖稳态具有强大的体内效应，但不能刺激身体生长。为了评估这些hGH片段的体外生物活性，我们测试了它们对表达全长人（h）、小鼠（m）或兔（r）生长激素受体（GHR）的GH反应性FDC - P1细胞系的活性。使用来自三种物种的含GHR的膜制剂和重组hGH结合蛋白（hGHBP）测试结合特异性和亲和力。重组hGH 1 - 43和重组44 - 191刺激FDC - P1 - hGHR细胞增殖，半数最大效应分别约为2000和100 nM，而完整的hGH刺激FDC - P1 - hGHR细胞增殖的ED50为0.02 - 0.03 nM。然而，这些片段对表达mGHR或rGHR的细胞影响极小。虽然44 - 191不刺激FDC - P1 - rGHR细胞增殖，但当在0.23 nM hGH存在下添加到这些细胞中时，它以剂量依赖性方式拮抗hGH的作用（ED50为230 nM）。使用兔肝、兔和小鼠脂肪组织的膜制剂以及重组hGHBP比较这些GH片段的结合。结合竞争曲线一致，44 - 191对兔肝GHR和兔脂肪GHR的亲和力比对牛生长激素至少低10倍，对hGHBP的亲和力比对hGH低61倍。在兔肝微粒体、脂肪微粒体或hGHBP中未证明hGH 1 - 43与GHR的结合。HGH 1 - 43不竞争脂肪微粒体中的胰岛素结合位点。总之，hGH 44 - 191以低亲和力与GHR结合，在超生理水平下刺激FDC - P1 - hGHR细胞增殖。在高剂量时，44 - 191也可拮抗FDC - P1 - rGHR细胞中的GH作用，推测是通过阻断受体二聚化。未检测到1 - 43与GHR的结合，其体外促有丝分裂作用微弱的原因仍有待阐明。这些片段对克隆的GHR的低亲和力以及在这些系统中的低生物活性表明，这些片段的代谢作用不太可能由克隆的GHR介导。这增加了存在单独受体介导这些片段代谢效应的可能性。
