GITRL对Kupffer细胞内脂多糖诱导的吲哚胺2,3双加氧酶的作用研究

The role of GITRL on lipopolysaccharide induced indoleamine 2,3-dioxygenase in Kupffer cells

目的:研究糖皮质激素诱导的TNF受体配体(GITRL)对Kupffer细胞(KCs)内脂多糖(LPS)诱导的吲哚胺2,3双加氧酶(IDO)的影响。方法:分离小鼠KCs后分为5组:Control组,只加培养基;LPS组,加入LPS(10μg/ml);LPS+Control siRNA组,转染Control siRNA后同LPS组;LPS+GITRLsiRNA组,转染GITRLsiRNA后同LPS组;LPS+Dex组,地塞米松(100μmol/L)处理后同LPS组。处理24小时后,分别采用免疫细胞化学染色、蛋白免疫印记和ELISA法检测KCs的GITRL、IDO表达及肿瘤坏死因子(TNF)-α分泌。结果:和Control组比较,LPS刺激后KCs上GITRL的表达明显上调(P<0.05),而沉默GITRL基因或者使用地塞米松能减弱其升高(P<0.05)。LPS可以诱导IDO和TNF-α在KCs上的高表达,然而GITRL基因沉默可以抑制其诱导的IDO和TNF-α的表达(P<0.05),同样地塞米松预处理也能够减弱其诱导的IDO和TNF-α的表达(P<0.05)。结论:GITRL介导了KCs内LPS诱导的IDO,地塞米松可通过下调GITRL的表达以降低LPS诱导的IDO。

Objective：To investigate the role of glucocorticoid-induced tumor necrosis factor receptor ligand （GITRL） on lipopolysaccharide （LPS） induced indoleamine 2,3-dioxygenase （IDO） in Kupffer cells （KCs）. Methods：The mouse KCs were isolated and randomly divided into five groups：Control group, cultured in media only; LPS group, cultured in media with LPS （10 μg/ml） ;LPS ＋ Control siRNA group, after transfected with Control siRNA, cultured in media as IPS group; LPS ＋ GITRL siRNA group, after transfected with GITRL siRNA, cultured in media as LPS group; LPS ＋ Dex group, Dex （100 μmol/L） pretreatmemt, then cultured in media LPS group. After 24 h of the treatment, the expression of GITRL, IDO, and tumour necrosis factor （TNF）-α was measured by immunocytochemist,y, Western blot, and ELISA, respectively. Results： Compared with control group, the expression of GITRL on KCs was increased obviously after LPS stimulation（ P 〈 0.05 ）, where as GITRL gene silencing or dexamethasone pretreatment attenuated/he increase（ P 〈 0.05）. LPS could induce high expression of IDO and TNF-α in KCs（ P 〈 0.05, respectively）, where as the expression of IDO and TNF-α was inhibited by G1TRL gene silencing（ P 〈 0.05, respectively）, similarly, dexamethasone pretreatment attenuated the expression of IDO and TNF-α（ P 〈 0.05, respetively）. Conclusion： GITRL plays a mediated role on LPS induced IDO in KCs. Dexamethasone may reduce the LPS-stimulated IDO by suppressing the expression of GITRL.