摘要
目的研究CD4＋T淋巴细胞在调节单核细胞与血管内皮细胞相互作用中CD80的表达水平及其意义。方法建立单核细胞-血管内皮细胞（EC）共培养和单核细胞-CD4＋T淋巴细胞-EC共培养体系，于培养72h时收集细胞。采用流式细胞术（FACS）检测CD80在CD14＋单核细胞表面的表达；通过实时逆转录聚合酶链反应（RT-PCR）技术检测CD80基因转录的表达变化。建立含有或不含抗CD86、CD28和CD154单克隆抗体的单核白细胞（PBMC）-EC混合培养体系，通过FACS检测CD14＋单核细胞表面CD80的表达，采用混合淋巴细胞-EC反应（MLER）来研究CD154和CD28封闭在抑制淋巴细胞对同种异体EC增殖中的作用。结果经FACS分析确定，无论是活化还是未活化的EC膜均缺乏CDS0、CD86和CD14的表达。RT-PCR表明在缺乏CD4＋T淋巴细胞时，经同种EC刺激的单核细胞可上调CD80基因转录水平的表达，但这些CD14＋单核细胞膜表面CDS0的表达并未上调，当CD4＋T淋巴细胞加入到共培养中时，CD80的表达上调。用抗CD28和抗CD86抗体封闭CD28和CD86并不能阻止CD14＋单核细胞表面CD80表达的上调。此外，阻断CD154也不能下调CDS0的高表达。MLER证明淋巴细胞对EC的增殖反应可部分的被抗CD28和抗CD154抗体所阻断；阻断CD80可抑制经EC刺激的单核细胞诱导T淋巴细胞增殖反应。结论在缺乏T淋巴细胞的条件下，经同种EC刺激的单核细胞可在基因转录水平上调CD80，而不是在其细胞膜表面的表达。当T淋巴细胞存在时，经EC刺激的CD14＋单核细胞可在其膜表面上调CD80的表达。CD14＋单核细胞膜表面CDS0表达的上调并不能被CD154和CD28封闭所阻止，显示其上调是通过CD154和CD86非依赖性通道。
Objective To study the role of monocyte-derived CD80 in providing co-stimulation to T cells, and to determine the role of CD86/CD28 and CD40/CD154 pathways in regulating monocyte-derived CD80 expression during allogeneic immunoresponses. Methods Monocyteendothelial cell （EC） and monocyte/CD4＋ cell-EC co-cultures were established. Cells were collected after 72-h co-culture followed by FACS to detect the CD80 expression. Real-time quantitative polymerase chain reaction （RT-PCR） was performed to study monocyte-derived CD80 transcript expression. Peripheral blood mononuclear cell （PBMC）-EC co-cultures with or without anti-CD86, CD28, and CD154 antibodies were established and analyzed by FACS for the monocyte-derived CD80 expression. Mixed lymphocyte-EC reaction （MLER） was performed to determine the effects of CD154 and CD28 blockade in inhibiting lymphocyte proliferation to EC. Results FACS analysis confirmed absence of CD80, CD86 and CD14 expression in resting and activated EC. RT-PCR demonstrated that EC-stimulated monoeytes in the absence of CD4＋cells upregulated the CD80 transcript expression, and the surface expression of CD80 was undetectable as measured by FACS. The expression of CD80 was restored when CD4＋ ceils were added into the co-cultures. Up-regulation of CD80 was revealed on monocyte surface during PBMC-EC interaction. Anti-CD28 and anti-CD86 antibodies and anti- CD2 8 Fab did not prevent upregulation of monoeyte - derived CD 8 0 expression. Anti - CD 1 5 4 antibody did not inhibit CD80 up-regulation. CD28 and CD154 blockade partially inhibited lymphocyte proliferation of MLER. Conclusions Human monocytes play an important role during cell-mediated immune responses. EC-stimulated monocytes up-regulate the CD80 mRNA expression but not on their surface in the absence of T cells. The surface expression of monocyte-derived CD80 is up-regulated in EC-stimulated CD14＋ monocytes in the presence of T cells. Up-regulation of the monocyte-derived CD80 expression can not be prevented by CD154/CD40 and CD28/CD86 blockade, suggesting that CD80 upegulation is through CDl54 or CD86 independent pathway. Specific therapy directed to prevent monocyte activation may be required for allograft transplantation.
出处
《中华器官移植杂志》
CAS
CSCD
北大核心
2008年第12期741-744,共4页
Chinese Journal of Organ Transplantation
