Eeva J, Nuutinen U, Ropponen A, Mättö M, Eray M, Pellinen R, Wahlfors J, Pelkonen J
Department of Clinical Microbiology, University of Kuopio, Kuopio, Finland.
Scand J Immunol. 2009 Dec;70(6):574-83. doi: 10.1111/j.1365-3083.2009.02331.x.
During the germinal centre reaction (GC), B cells with non-functional or self-reactive antigen receptors are negatively selected by apoptosis to generate B cell repertoire with appropriate antigen specificities. We studied the molecular mechanism of Fas/CD95- and B cell receptor (BCR)-induced apoptosis to shed light on the signalling events involved in the negative selection of GC B cells. As an experimental model, we used human follicular lymphoma (FL) cell line HF1A3, which originates from a GC B cell, and transfected HF1A3 cell lines overexpressing Bcl-x(L), c-FLIP(long) or dominant negative (DN) caspase-9. Fas-induced apoptosis was dependent on the caspase-8 activation, since the overexpression of c-FLIP(long), a natural inhibitor of caspase-8 activation, blocked apoptosis induced by Fas. In contrast, caspase-9 activation was not involved in Fas-induced apoptosis. BCR-induced apoptosis showed the typical characteristics of mitochondria-dependent (intrinsic) apoptosis. Firstly, the activation of caspase-9 was involved in BCR-induced DNA fragmentation, while caspase-8 showed only marginal role. Secondly, overexpression of Bcl-x(L) could block all apoptotic changes induced by BCR. As a novel finding, we demonstrate that caspase-9 can enhance the cytochrome-c release and collapse of mitochondrial membrane potential (DeltaPsi(m)) during BCR-induced apoptosis. The requirement of different signalling pathways in apoptosis induced by BCR and Fas may be relevant, since Fas- and BCR-induced apoptosis can thus be regulated independently, and targeted to different subsets of GC B cells.
在生发中心反应（GC）期间，具有无功能或自身反应性抗原受体的B细胞通过凋亡被阴性选择，以产生具有适当抗原特异性的B细胞库。我们研究了Fas/CD95和B细胞受体（BCR）诱导凋亡的分子机制，以阐明参与GC B细胞阴性选择的信号事件。作为实验模型，我们使用了源自GC B细胞的人滤泡性淋巴瘤（FL）细胞系HF1A3，并转染了过表达Bcl-x(L)、c-FLIP（长型）或显性负性（DN）caspase-9的HF1A3细胞系。Fas诱导的凋亡依赖于caspase-8的激活，因为caspase-8激活的天然抑制剂c-FLIP（长型）的过表达阻断了Fas诱导的凋亡。相反，caspase-9的激活不参与Fas诱导的凋亡。BCR诱导的凋亡表现出线粒体依赖性（内在）凋亡的典型特征。首先，caspase-9的激活参与了BCR诱导的DNA片段化，而caspase-8仅起边缘作用。其次，Bcl-x(L)的过表达可以阻断BCR诱导的所有凋亡变化。作为一个新发现，我们证明caspase-9可以增强BCR诱导凋亡期间细胞色素c的释放和线粒体膜电位（ΔΨm）的崩溃。BCR和Fas诱导凋亡中不同信号通路的需求可能是相关的，因为Fas和BCR诱导的凋亡因此可以独立调节，并靶向GC B细胞的不同亚群。
