LPS通过上调Toll样受体2和4表达增强IFN-α抗病毒效应的研究

摘要

Objective To analyse lipopolysaccharide(LPS) on the expression of Toll-like receptor 2 and 4 (TLR2, TLR4 ) and to explore its function and mechanism in the antiviral effects of interferon-α ( IFN-α) . MethodsHepG2 cells were treated with 1 000 IU/ml IFN-α, 5 mg/L LPS, and LPS combined with IFN-αfor 24 hours, and the mRNA levels of TLRs, STAT1, STAT2, IRF9, MyD88 and MxA were assessed by RT-PCR. Meanwhile, the expression of MyD88 and STAT1, STAT2 was detected by Western blot. Results The mRNA levels of TLR2, TLR4 , STAT1 , MyD88 and MxA in the IFN-αgroup were significantly higher than those of the control group ( P<0.05 ) . The expression of other moleculars in the LPS group was similar to the control group except TLR2 and TLR4 . There were no significant difference of MyD88 and MxA between the combination group and the IFN-αgroup, however the mRNA levels of STAT1, STAT2, IRF9, 2'-5'OAS and ds-RNA-dependent protein kinase R (PKR) moleculars were increased significantly in the combination group (P<0.05). Western blot results indicated that, compared to the IFN-αgroup, STAT1, STAT2 protein was increased significantly (P<0.05), while MyD88 protein was not increased in the combination group. Conclusion LPS can enhance the expression of TLR2 and TLR4 as well as the antiviral proteins induced by IFN-α, suggesting that LPS may participate in the antiviral effect of IFN-α via TLR2 and TLR4 .%目的探讨细菌脂多糖( LPS)对Toll样受体2和4(TLR2和TLR4)表达的影响及其在α-干扰素(IFN-α)抗病毒效应中可能发挥的作用及相关机制。方法以1000 IU/ml IFN-α和5 mg/L LPS单独及联合作用于HepG2细胞24 h后,运用RT-PCR和Western blot法分析作用前后细胞表面TLR2、TLR4和 IFN-α JAK-STAT 途径分子 STAT1、STAT2、IRF9表达情况,并进一步分析IFN-α诱导的 MyD88、MxA、2忆-5忆寡腺苷合成酶(2忆-5忆OAS)、双链RNA依赖性蛋白激酶( PKR)抗病毒蛋白的表达。结果 RT-PCR结果显示,IFN-α作用于HepG2细胞24 h后, TLR2、TLR4和STAT1、STAT2、IRF9信号分子及MyD88、抗黏病毒A蛋白( MxA)、2忆-5忆寡腺苷合成酶(2忆-5忆OAS)、PKR抗病毒蛋白的表达上升,而单独LPS处理细胞24 h后,除TLR2、TLR4 mRNA显著升高外( P<0.05),其他分子的表达均无显著变化；联合处理组较IFN-α处理组除MyD88和MxA mRNA水平无显著变化,其他分子的表达均显著升高( P<0.05)。 Western blot结果显示,与IFN-α处理组相比,联合处理组STAT1、STAT2蛋白表达显著增加(P<0.05),而MyD88蛋白表达无显著变化。结论 LPS可能通过上调细胞表面TLR2、TLR4的表达而发挥增强IFN-α诱导的抗病毒蛋白的表达水平,从而参与IFN-α抗病毒效应。