中文摘要：

目的：建立稳定表达Rab5a及其失活突变体Rab5a N133I巨噬细胞系,并分析Rab5a对LPS刺激的RAW264.7巨噬细胞中i NOS、TNF-α和IL-6表达的影响。方法：将Rab5a及其失活突变载体Rab5a N133I转染RAW264.7细胞,通过G418筛选稳定表达细胞系。通过Real-time PCR鉴定稳定表达细胞系。MTT法分析其生长特性,LPS刺激稳定表达细胞系不同时间,检测i NOS、TNF-α和IL-6的表达水平。结果：转染Rab5a/Rab5a N133I细胞Rab5a mRNA水平显著高于对照（P〈0.05）。Rab5a过表达促进RAW264.7细胞增殖,而过表达Rab5a N133I细胞增殖显著慢于对照（P〈0.05）。Rab5a过表达后,LPS诱导的RAW264.7细胞中i NOS、TNF-α和IL-6的表达显著增高（P〈0.01）。然而,过表达Rab5a N133I对LPS诱导的i NOS、TNF-α和IL-6的表达没有显著影响。结论：Rab5a过表达显著促进了LPS活化的RAW264.7细胞中i NOS、TNF-α和IL-6的表达,这种促进作用依赖于其结合GTP的能力。

英文摘要：

Objective： To establish cell lines stably expressing RabSa and its the inactive mutant RabSaN133I, analyze the effect of Rab5a on the expression of cytokines in LPS-stimulated RAW264.7 cells . Methods：RAW264.7 cells were transfected with RabSa and its inactive mutant vector RabSa N133I separately, and then screened by G418. Rab5a stable expressing cell lines were identified by Real time-PCR. The growth of the stable cell lines was analyzed by MTT assay. After the stable cell lines were stimulated by LPS for different time periods, the expression of iNOS, TNF-α and IL-6 was detected. Results： RabSa and Rab5 aN1331 transfection resulted in elevated RabSa mRNA expression compared with the control cells （ P 〈 0. 05 ）. RabSa overexpression enhanced the proliferation of RAW264.7 cells. However,the proliferation of RabSaN133I cells was significantly slower than the control cells （P〈 0. 05）. Overexpression of RabSa promoted LPS-induced production of iNOS, TNF-α and IL-6 in RAW264. 7 cells （P〈0.01）. Conversely,overexpression of Rab5aN133I abolished the stimulating effects of RabSa. Conclusion： RabSa promoted LPS-induced expression of iNOS,TNF-α and IL-6 in RAW264.7 macrophages in a GTP-bindina ability-dependent manner.