Kurita T, Kiyono H, Komiyama K, Grossi C E, Mestecky J, McGhee J R
J Immunol. 1986 Jun 1;136(11):3953-60.
Several methods have been used in the present study to characterize Fc receptors (FcR) expressed on T-T hybridomas derived from mouse Peyer's patch T helper (Th) cell clones that preferentially support IgA responses. These T hybridomas (designated Th HA cells) produce IgA-binding factor (IBF alpha) which regulates antigen-dependent IgA responses. The ultrastructure of Th HA cells and the distribution of Fc alpha R on these cell lines were determined by colloidal gold (CG) immunoelectron microscopy (IEM). When Th HA cells were incubated with purified mouse IgA followed by CG-labeled anti-IgA, an even pattern of CG was distributed on the cell membrane. To ensure that binding occurred through Fc alpha R, Th HA cells were mixed with MOPC 315 IgA anti-DNP, followed by staining with CG-labeled TNP-human serum albumin. This resulted in an identical pattern of gold particle distribution, confirming expression of Fc alpha R on Th HA cells. No Fc mu R or Fc gamma 1R were detectable on Th HA cells by IEM. Immunocytoadherence with TNP-conjugated erythrocytes confirmed that Th HA cells were Fc alpha R+; however, no IgM or IgG rosettes were seen. When these cell lines were analyzed by flow cytometry (FACS) using IgA, IgM, or IgG1 and FITC-labeled anti-H chain-specific antibodies, 55 to 65% of cultured Th HA cells expressed Fc alpha R, and 11 to 18% expressed Fc mu R; however, no Fc gamma 1R was detectable on Th HA cells. The use of ELISA with Th HA cells as antigen confirmed the expression of Fc alpha R and the presence of less Fc mu R on these two cell lines. Solubilized membrane fractions derived from Th HA cells were tested for the presence of FcR by ELISA and for biologic function for support of IgA responses in Peyer's patch B cell cultures. Both Fc alpha R and Fc mu R were detected in fractions derived from Th HA cells. Furthermore, these fractions supported in vitro IgA anti-sheep erythrocyte responses, comparable to those obtained with Th HA cell culture supernatants containing IBF alpha. These studies show that Th HA cells express Fc alpha R with less Fc mu R, and the solubilized form of Fc alpha R exhibits IBF alpha-like activity. The significance of FcR expression by Th cell clones and cell lines and the relationship of soluble Fc alpha R and IBF alpha for IgA response regulation are discussed.
在本研究中，已采用多种方法来表征源自小鼠派尔集合淋巴结T辅助（Th）细胞克隆的T - T杂交瘤细胞上表达的Fc受体（FcR），这些克隆优先支持IgA反应。这些T杂交瘤细胞（命名为Th HA细胞）产生调节抗原依赖性IgA反应的IgA结合因子（IBFα）。通过胶体金（CG）免疫电子显微镜（IEM）确定了Th HA细胞的超微结构以及FcαR在这些细胞系上的分布。当将Th HA细胞与纯化的小鼠IgA孵育，然后用CG标记的抗IgA处理时，CG在细胞膜上呈均匀分布模式。为确保结合是通过FcαR发生的，将Th HA细胞与MOPC 315 IgA抗DNP混合，然后用CG标记的TNP - 人血清白蛋白染色。这导致了相同的金颗粒分布模式，证实了Th HA细胞上FcαR的表达。通过IEM在Th HA细胞上未检测到FcμR或Fcγ1R。用TNP偶联的红细胞进行免疫细胞黏附实验证实Th HA细胞是FcαR阳性；然而，未观察到IgM或IgG玫瑰花结。当使用IgA、IgM或IgG1以及异硫氰酸荧光素（FITC）标记的抗重链特异性抗体通过流式细胞术（FACS）分析这些细胞系时，55%至65%的培养Th HA细胞表达FcαR，11%至18%表达FcμR；然而，在Th HA细胞上未检测到Fcγ1R。以Th HA细胞作为抗原进行酶联免疫吸附测定（ELISA），证实了这两种细胞系上FcαR的表达以及较少的FcμR的存在。通过ELISA检测源自Th HA细胞的可溶性膜组分中FcR的存在，并检测其在派尔集合淋巴结B细胞培养中支持IgA反应的生物学功能。在源自Th HA细胞的组分中检测到了FcαR和FcμR。此外，这些组分支持体外IgA抗绵羊红细胞反应，与含有IBFα的Th HA细胞培养上清液所获得的反应相当。这些研究表明Th HA细胞表达FcαR且FcμR较少，并且可溶性形式的FcαR具有类似IBFα的活性。讨论了Th细胞克隆和细胞系中FcR表达的意义以及可溶性FcαR和IBFα在IgA反应调节中的关系。
