Wermerskirchen A S, LaTocha D H, Clarke B L
Department of Medical Microbiology and Immunology, University of Minnesota at Duluth, 55812, USA.
Life Sci. 2000 Sep 22;67(18):2177-87. doi: 10.1016/s0024-3205(00)00810-9.
The initiation of DNA synthesis and secretion of Interleukin 2 (IL-2) was measured in isolated rat splenic lymphocytes following activation with Concanavalin A (ConA). The extent of 3H-thymidine incorporation into activated cells was tested when cultured with various concentrations of Adrenocorticotropic hormone (ACTH). A paradoxical dose-response curve resulted when ACTH caused a biphasic response of augmenting and inhibiting 3H-thymidine uptake in lymphocytes depending on the hormone concentration. Low levels of ACTH (0.001-1-nM) augmented 3H-thymidine uptake and high levels (10-1000 nM) reversed the effect. The optimal ACTH concentration was 10 pM ACTH in the presence of 5 ug/ml ConA and there was no ACTH effect on quiescent cells (no ConA). Conditioned media from splenic lymphocytes treated with various concentrations of ConA or ACTH was tested for increased uptake of 3H-thymidine by the IL-2 growth dependent Cytotoxic T Lymphocyte Leukemia (CTLL-2) cells. ConA conditioned medium could sustain the CTLL-2 cells indicating the presence of IL-2. Conditioned medium from splenic lymphocytes treated with both ConA and 100 pM ACTH further increased CTLL-2 cell proliferation indicating an additional increase of IL-2 secretion. The identity of IL-2 was confirmed by using an anti-rat IL-2 antibody to neutralize the growth potential of the conditioned medium. ACTH alone had no effect on the CTLL-2 cell proliferation indicating the effect is due solely to induced IL-2 found in the conditioned medium. IL-2 levels in the conditioned media were quantitated by ELISA assay; splenic lymphocytes produced 4.2 ng/ml to ConA only, 19.2 ng/ml in ConA plus 10 nM ACTH, and no detectable IL-2 at ConA plus 10 uM ACTH. These results demonstrated that ACTH modulates IL-2 secretion from activated lymphocytes, which is both biphasic and concentration dependent.
在用刀豆蛋白A（ConA）激活后，测定分离的大鼠脾淋巴细胞中DNA合成的起始和白细胞介素2（IL-2）的分泌。在用不同浓度的促肾上腺皮质激素（ACTH）培养时，测试3H-胸腺嘧啶核苷掺入活化细胞的程度。当ACTH根据激素浓度引起淋巴细胞中3H-胸腺嘧啶核苷摄取的双相反应（增强和抑制）时，得到了一条矛盾的剂量反应曲线。低水平的ACTH（0.001 - 1 nM）增强了3H-胸腺嘧啶核苷的摄取，而高水平（10 - 1000 nM）则逆转了这种作用。在存在5 μg/ml ConA的情况下，最佳ACTH浓度为10 pM，并且ACTH对静止细胞（无ConA）没有影响。测试了用不同浓度的ConA或ACTH处理的脾淋巴细胞的条件培养基对依赖IL-2生长的细胞毒性T淋巴细胞白血病（CTLL-2）细胞摄取3H-胸腺嘧啶核苷增加的情况。ConA条件培养基可以维持CTLL-2细胞，表明存在IL-2。用ConA和100 pM ACTH处理的脾淋巴细胞的条件培养基进一步增加了CTLL-2细胞增殖，表明IL-2分泌进一步增加。通过使用抗大鼠IL-2抗体中和条件培养基的生长潜力来确认IL-2的身份。单独的ACTH对CTLL-2细胞增殖没有影响，表明该作用仅归因于条件培养基中诱导产生的IL-2。通过ELISA测定法定量条件培养基中的IL-2水平；脾淋巴细胞仅对ConA产生4.2 ng/ml，在ConA加10 nM ACTH时产生19.2 ng/ml，在ConA加10 μM ACTH时未检测到IL-2。这些结果表明，ACTH调节活化淋巴细胞分泌IL-2，这是双相的且依赖于浓度。
