Hornsby P J, Gill G N
Endocrinology. 1978 Mar;102(3):926-36. doi: 10.1210/endo-102-3-926.
The characteristics of adult bovine adrenocortical cells were studied throughout their life span of 55-65 generations in monolayer culture. Over this period, the cells maintained the capacity to synthesize steroids when tested with repeated maximal doses of ACTH, prostaglandin E1, monobutyryl cAMP, or cholera toxin. Prostaglandin E1 stimulated cAMP production and steroidogenesis, and inhibited DNA synthesis, as measured by incorporation of [3H]thymidine, with dose-response characteristics that did not vary over the first 50 generations in culture. In contrast, the maximal rate of cAMP production stimulated by ACTH declined exponentially at a rate of 7% per generation. In primary and secondary cultures, ACTH stimulated steroidogenesis maximally and inhibited [3H]thymidine incorporation into DNA completely at a half-maximal effective concentration (ED50) of 0.08 nM which was two orders of magnitude less than the ED50 of 8 nM for stimulation of cAMP production. As the ACTH-stimulated maximal rate of cAMP production fell with increasing generation number, the ED50 for ACTH stimulation of steroidogenesis and inhibition of DNA synthesis increased. From about the 20th generation onward, the ability of ACTH to inhibit DNA synthesis maximally declined so that by the 40th generation, cells were completely resistant to the growth-inhibitory effects of ACTH. High-dose ACTH continued, however, to stimulate steroid production maximally over the 50 generations studied. In late passage cells, the ED50 for ACTH stimulation of steroidogenesis was 8 nM, identical to that for cAMP production. Although ACTH-stimulated cAMP production was related to both stimulation of steroidogenesis and inhibition of DNA synthesis, higher cAMP levels appeared required for inhibition of DNA synthesis than for stimulation of steroidogenesis. Mitogenic responses to fibroblast growth factor and to angiotensin II were retained throughout long term growth in culture. The progressive loss of ACTH-responsiveness was specific and a function of aging of bovine adrenocortical cells in culture.
在单层培养中，对成年牛肾上腺皮质细胞55 - 65代的整个生命周期特性进行了研究。在此期间，当用重复的最大剂量促肾上腺皮质激素（ACTH）、前列腺素E1、单丁酰环磷腺苷（monobutyryl cAMP）或霍乱毒素进行测试时，这些细胞保持了合成类固醇的能力。前列腺素E1刺激环磷腺苷（cAMP）生成和类固醇生成，并抑制DNA合成，这通过[3H]胸腺嘧啶核苷掺入来测定，其剂量 - 反应特性在培养的前50代中没有变化。相比之下，ACTH刺激产生的cAMP最大速率以每代7%的速度呈指数下降。在原代和传代培养中，ACTH在半最大有效浓度（ED50）为0.08 nM时最大程度地刺激类固醇生成，并完全抑制[3H]胸腺嘧啶核苷掺入DNA，这比刺激cAMP生成的8 nM的ED50低两个数量级。随着ACTH刺激产生的cAMP最大速率随着代数增加而下降，ACTH刺激类固醇生成和抑制DNA合成的ED50增加。从大约第20代起，ACTH最大程度抑制DNA合成的能力下降，以至于到第40代时，细胞对ACTH的生长抑制作用完全产生抗性。然而，在研究的50代中，高剂量ACTH继续最大程度地刺激类固醇生成。在传代后期细胞中，ACTH刺激类固醇生成的ED50为8 nM，与刺激cAMP生成的ED50相同。尽管ACTH刺激的cAMP生成与类固醇生成刺激和DNA合成抑制都相关，但抑制DNA合成似乎比刺激类固醇生成需要更高的cAMP水平。在整个长期培养生长过程中，对成纤维细胞生长因子和血管紧张素II的促有丝分裂反应得以保留。ACTH反应性的逐渐丧失是特异性的，并且是培养中牛肾上腺皮质细胞衰老的一个功能。
