Molecular mechanisms of isotype switch recombination.

小鼠B细胞表达CD38，抗CD38抗体可诱导其增殖并保护其免于凋亡。我们先前的研究表明，白介素5(IL-5)和抗小鼠CD38McAb CS/2共同刺激小鼠脾B细胞，诱导产生IgG1和IgM。在此，我们研究了IL-5和CS/2在生殖系γ1转录本表达和μ-γ1开关重组副产物形成DNA环的相互产物产生中的作用。CS/2单独诱导脾原始B细胞表达生殖系γ-1，而IL-5既不诱导也不增强生殖系γ-1的表达。CS/2与B细胞共培养后，细胞内μ-γ-1重组产物的细胞含量未见增加，但CS/2+IL-5培养后，B细胞的相互产物增加，分泌高水平的Ig G 1。尽管IL-4本身不能诱导μ-γ-1重排…在CS/2刺激的B细胞中结合更多的IL-4，联合CS/2和IL-5，IL-4显著促进无菌γ1的转录和产生。这些结果表明，γ-5只能诱导胚系μ-γ-1的转录，并且IL-5以非IL-4的方式促进Ig G 1的开关重组和分泌。尽管已经研究了活化B细胞中IL-5的一些受体后信号转导事件，但Jak/Stat通路在IL-5信号转导中的作用还没有得到彻底的评价。在这项研究中，我们检测了IL-5是否激活了CD38激活的小鼠脾B细胞中的JAK/STAT通路。IL-5刺激可激活Stat5a和Stat5b。在IL-5诱导的μ-γ-1开关重组和免疫球蛋白G_1的产生中，Stat5a和Stat5b的作用被证实，因为IL-5不作用于Stat5a和-/-&gt；-/-&gt；-/-&gt；在IL-5刺激下，生殖系γ-1转录产物CD38T和激活诱导型胞苷脱氨酶的表达水平与野生型B细胞相当。因此，STAT5a和STAT5b在依赖IL-5的μ-γ-1开关重组中都是必需的，它们的靶标可能不是G1和AID基因。IL-4可部分挽救B细胞受损的μ-γ1开关重组，但不能挽救B细胞受损的μ-γ1开关重组，这是因为在经IL-5和CD38刺激的B细胞培养中加入IL-4确实诱导了CD1开关重组，导致显著的免疫球蛋白1的产生。我们的数据支持这样的观点，即Stat5a和Stat5b并不是多余的，而是至少在它们在B细胞分化中的部分功能上是不同的。用5-，6-羧基荧光素二乙酸酯琥珀酰亚胺酯对B细胞分裂周期进行分析，发现CS/2和IL-5在5~6个分裂周期后均可观察到μ-γ-1开关重组和表面阳性细胞。STAT5a^&lt；-/-&gt；和STAT5b^&lt；-/-&gt；B细胞有5~6个细胞分裂周期，但既不表达μ-γ-1开关，也不表达表面免疫球蛋白1。较少

Mouse B cells express CD38 whose ligation by anti-CD38 antibody induces their proliferation and protection from apoptosis. We previously showed that stimulation of mouse splenic B cells with interleukin 5 (IL-5) together with CS/2, an anti-mouse CD38 monoclonal antibody, induces production of IgG1 and IgM.Here we examined the role of IL-5 and CS/2 in the expression of germline γ1 transcripts and the generation of reciprocal products forming DNA circles as by products of μ-γ1 switch recombination. By itself, CS/2 induced significant expression of germline γ1 transcripts in splenic naive B cells, whereas IL-5 neither induced nor enhanced germline γ1 expression. Increased cellular content of reciprocal product, which is characteristic of μ-γ1 recombination, was not observed after culturing B cells with CS/2, but increased reciprocal product along with high levels of IgG1 secretion was found when B cells were cultured with CS/2 plus IL-5. Although IL-4 did not, by itself, induce μ-γ1 recom … More bination in B cells stimulated with CS/2, in conjunction with CS/2 plus IL-5, IL-4 dramatically enhanced sterile γ1 transcription and IgG1 production. These results demonstrate that CD38 ligation induces only germline γ1 transcription and that IL-5 promotes both μ-γ1 switch recombination and IgG1 secretion in an IL-4 independent manner.Although some post-receptor signaling events of IL-5 in activated B cells have been characterized, the involvement of the Janus kinase/signal transducer and activator (Jak/Stat) pathway in IL-5 signaling has not been thoroughly evaluated. In this study, we examined whether IL-5 activates the Jak/Stat pathway in CD38-activated mouse splenic B cells. Both Stat5a and Stat5b were activated by IL-5 stimulation. The role of Stat5a and Stat5b in IL-5-induced μ-γ1 switch recombination and IgG1 production were documented, as IL-5 did not act on CD38-stimnulated splenic B cells of Stat5a^<-/-> and Stat5b^<-/-> mouse. Expression levels of germline γ1 transcripts to CD38 and of activation-induced cytidine deaminase (AID) in Stat5a^<-/-> and Stat5b^<-/-> B cells upon IL-5 stimulation were comparable to these of wild type B cells. Thus, both Stat5a and Stat5b are essential for at IL-5-dependent μ-γ1 switch recombination, and their targets may not be g1 and AID genes. The impaired μ-γ1 switch recombination by Stat5b^<-/-> B cells, but not by Stat5a^<-/-> B cells, were rescued in part by IL-4, as the addition of IL-4 to the culture of CD38- and IL-5-stimulated B cells did induce μ-γ1 switch recombination leading to significant IgG1 production. Our data support the notion that Stat5a and Stat5b are not redundant, but rather are at least partially distinctive in their function on B cell differentiation. Analysis of cell division cycle number of B cells, examined by using 5-, 6-carboxyfluorescein diacetate, succinimidyl ester (CFSE), revealed that μ-γ1 switch recombination and surface IgG1-positive cells were observed after 5 to 6 division cycles upon CS/2 and IL-5. Stat5a^<-/-> and stat5b^<-/-> B cells showed 5 to 6 cell division cycles, but they could express neither μ-γ1 switching nor surface IgG1. Less