Gogolák P, Réthy B, Horváth A, Tóth G K, Cervenák L, László G, Rajnavölgyi E
Department of Immunology, L. Eotvos University, God, Hungary.
Immunol Lett. 1996 Dec;54(2-3):135-44. doi: 10.1016/s0165-2478(96)02663-6.
A previously developed experimental system was applied to obtain qualitative and quantitative data on the contribution of TCR-, CD4- and CD28-mediated signalling in the activation of an antigen specific T-cell hybridoma. All the three signal transducing receptors were stimulated by their natural ligands, and intermediate and late responses of an I-Ed restricted, CD4 +, influenza HA specific murine T-hybridoma (IP-12-7) were monitored by measuring the concentration of intracellular calcium [Ca2+]i and secreted IL-2. This type of analysis of T-cell activation revealed: (i) calcium mobilization induced by peptide loaded APC requires rapid conjugate formation; (ii) a direct correlation between the magnitude of the intermediate and the late responses was observed as a consequence of differential TCR ligation modulated by peptide dose or by the presence CD4; (iii) considering the APC/peptide and T/APC ratios, the concentration dependence of the intermediate and late responses was similar in both assays but a substantial difference in the sensitivity of the two methods was observed; (iv) CD4 mediated signalling has a co-stimulatory effect predominantly at suboptimal in vitro conditions; and (v) sustained increase of [Ca2+]i as well as the production of high concentrations of IL-2 is highly dependent on the CD28-B7 interaction. These results demonstrate that distinct peptide doses and the presence or absence of CD4 result in quantitative changes in T-cell responses, while the degree of CD28 mediated signalling has a qualitative affect on the outcome of T-cell activation, revealed by complete or partial inhibition of IL-2 secretion as a result of limited CD28-B7 interaction as well as by alteration in the duration and time kinetics of the calcium response.
应用先前开发的实验系统来获取关于TCR、CD4和CD28介导的信号传导在抗原特异性T细胞杂交瘤激活中的作用的定性和定量数据。所有这三种信号转导受体均由其天然配体刺激，并通过测量细胞内钙[Ca2+]i浓度和分泌的IL-2来监测I-Ed限制性、CD4 +、流感HA特异性小鼠T杂交瘤（IP-12-7）的中期和晚期反应。这种对T细胞激活的分析类型揭示：（i）由负载肽的抗原呈递细胞（APC）诱导的钙动员需要快速形成结合物；（ii）由于肽剂量或CD4的存在调节的差异性TCR连接，观察到中期和晚期反应的幅度之间存在直接相关性；（iii）考虑到APC/肽和T/APC比率，两种测定中中期和晚期反应的浓度依赖性相似，但观察到两种方法的灵敏度存在实质性差异；（iv）CD4介导的信号传导主要在次优体外条件下起共刺激作用；（v）[Ca2+]i的持续增加以及高浓度IL-2的产生高度依赖于CD28-B7相互作用。这些结果表明，不同的肽剂量以及CD4的存在与否导致T细胞反应的定量变化，而CD28介导的信号传导程度对T细胞激活的结果具有定性影响，这通过有限的CD28-B7相互作用导致的IL-2分泌的完全或部分抑制以及钙反应的持续时间和时间动力学的改变得以揭示。
