脂多糖经MyD88／NF-KB信号途径抑制Bewo细胞中乙型肝炎病毒复制

目的探讨Toll样受体4（TLR4）配体脂多糖（LPS）抑制人滋养层细胞Bewo中乙型肝炎病毒（rtBv）复制的作用机制，为防治HBV宫内感染提供依据。方法首先将2txg1．3倍HBV全基因重组质粒pcDNA3．1（＋）-HBV1．3转染Bewo细胞12h后，以TLR4配体LPS处理3d。尔后用LPS处理Bewo细胞，观察IFN—β、TNF-a表达的动力学、NF—KB拮抗剂二硫代氨基甲酸吡咯烷（PDTC）对LPS诱导Bewo细胞产生细胞因子的作用。采用微粒子酶免疫分析法（MEIA）和荧光定量PCR法分别检测HBsAg、HBeAg和HBVDNA水平，并以ELISA和RT—PCR分别检测IFN-β、TNF—a水平及TIR结构域的转接蛋白（TRIF）、髓样分化蛋白（MyD88）表达。结果与对照组比较，LPS可显著抑制Bewo细胞中HBV复制（P〈0．01），且LPS可显著诱导Bewo细胞产生TNF-a（P〈0．05），呈时间和剂量依赖性。PDTC可抑制LPS诱导细胞产生TNF-a，显著低于对照组（P〈0．01），但对IFN—β无显著作用（P〉0．05）。与对照组比较，LPS可诱导HBV重组质粒转染的Bewo细胞表达MyD88（P〈0．01）。结论通过MyD88／NF—KB信号途径诱导Bewo细胞产生TNF—a，TLR4配体LPS可显著抑制HBV复制。

LPS-mediated inhibition hepatitis B virus replication in Bewo cells via the NF-kB/MyD88 pathway

Objective To explore the effect and mechanism of Toll-like receptor 4 （TLR4） ligand LPS-mediated inhibition hepatitis B virus （HBV） replication in Bewo cells. Methods First of all, 2 ug 1.3-fold HBV recombinant vector pcDNA3.1 （ ＋ ）-HBV1.3 were transfected into Bewo cells, after 12 h, the ceils were treated with LPS for 3 d. To observe the kinetics of IFN-β and TNF-α expression in Bewo cells, the Bewo cells were exposed to TLR4 ligand LPS. And the effect of pyrrolidine dithiocarbamate （PDTC） , an inhibitor of NF-kB, on LPS-induced cytokines was also observed. The HBsAg, HBeAg and HBV DNA level in the culture supernatant were detected by Microparticle Enzyme Immunoassay （MEIA） and fluorescence quantitative PCR, respectively, and the expression of IFN-β,TNF-α, TRIF and MyD88 was detected by ELISA and RT-PCR,respectively. Results Compared with control group, LPS could significantly suppress HBV replication in Bewo cells （ P 〈 0. 01 ） , and it could induce the production of TNF- α in Bewo cells （ P 〈 0. 05 ） , in time-and dose-dependent manners. PDTC strongly inhibited LPS and induced TNF-α production, but had no much effect on IFN-β in Bewo cells （ P 〈 0. 001 ）. Compared with control group, the mRNA levels of MyD88 were significantly induced by LPS in the Bewo cells transfected with this recombinant vector （ P 〈 0. 001 ）. Conclusions TLR4 ligand LPS could significantly suppress HBV replication by inducing TNF-α production in Bewo cells mainly via the MyD88/ NF-kB signal pathway.