中国免疫学杂志2016，Vol.32Issue(4)：470-475,479,7.DOI:10.3969/j.issn.1000-484X.2016.04.004
IRF3基因干扰对LPS刺激原代枯否细胞早期细胞因子分泌动态变化的影响
Effects of IRF3 gene interference on dynamic changes of cytokine secretions in early of LPS stimulation in primary Kupffer cells
摘要
Abstract
Objective:To investigate the effects of IRF 3 shRNA adenovirus on dynamic changes of early cytokines in LPS-stimulated primary Kupffer cells ( KCs ).Methods: Rat KCs were isolated and purified by means of in situ perfusion.After being infected with adenovirus carrying IRF 3 shRNA for 48 h, KCs were stimulated with LPS.Cell culture supernatants were collected respectively at 0,2,4 and 6 h after LPS stimulation as well as cells at 6 h.Supernatant cytokine secretion levels were detected by enzyme-linked immunosorbent assay ( ELISA).Intracellular gene expressions were tested by RT-PCR and Westeron blot.Results:IRF3 mRNA and protein were induced by LPS ,but suppressed by IRF 3 shRNA adenovirus in LPS-stimulated or non-stimulated KCs.IFN-βsecretions rose in the very early stage ( at 2 h) ,reached the peak at 4 h,and began to reduce but still remained high levels at 6 h after LPS stimulation in KCs.Interference adenovirus pretreatment suppressed IFN-βsecretions ( especially the secretion peak ) at each time point after LPS stimulation.IFN-βsecretions reached normal levels at 6 h after the stimulation in adenovirus-pretreated cells;TNF-αse-cretions rapidly increased in the very early stage and reached the peak at 2 h,then began to decrease gradually ,but remained high levels at 6 h after LPS stimulation in KCs.Interference adenovirus pretreatment inhibited LPS-induced TNF-αsecretions, especially the secretion peak;IL-1βsecretions did not increase untill 4 h, but reached a higher level at 6 h after LPS stimulation.Interference adenovirus suppressed IL-1βsecretions in the early stage of LPS stimulation;IL-10 secretions began to rise in the very early stage ,and gradually increased over time after LPS stimulation in KCs.Pretreatment of adenovirus with IRF 3 shRNA promoted upregulations of IL-10 secretions at each time point of the early of LPS stimulation.Conclusion:IRF3 gene expression can be silenced by IRF 3 shRNA ad-enovirus.IRF3 can promote its downstream signaling molecule IFN-βand pro-inflammatory cytokines including TNF-αand IL-1β,and block anti-inflammation cytokine IL-10 secretions in LPS-stimulated primary KCs.Therefore,IRF3 may play a central role in immune inflammatory injury of liver tissues.关键词
原代枯否细胞/干扰素调节因子3/基因干扰/固有免疫/炎性反应Key words
Primary kupffer cells/IRF3/Gene interference/Innate immune/Inflammatory reaction分类
医药卫生引用本文复制引用
朱彤,涂文娟,谈志丽,刘亮明..IRF3基因干扰对LPS刺激原代枯否细胞早期细胞因子分泌动态变化的影响[J].中国免疫学杂志,2016,32(4):470-475,479,7.基金项目
本文为国家自然科学基金项目（81070357，30660066）和上海市松江区科学技术攻关项目（14SJGGYY22）。 （）
