【摘要】 目的观察生殖支原体脂质相关膜蛋白(LAMP)能否诱导胎盘滋养层细胞表达血红素氧合酶1(HO-1),从而影响细胞因子的产生。方法体外培养胎盘滋养层细胞,用0.5~5μg/m L LAMP作用4~12 h。实时定量PCR和Western blot法分别检测HO-1 mRNA和蛋白的表达以及核因子相关因子2(Nrf2)的核转位;2’,7’-二氯二氢荧光黄二乙酸酯(H2DCFDA)检测活性氧(ROS)产生;采用N-乙酰半胱氨酸(NAC)或Nrf2 siRNA处理滋养层细胞,观察其对HO-1表达的影响。采用HO-1的激动剂钴原卟啉(Co PP),抑制剂锌原卟啉(Zn PP)或HO-1 siRNA处理细胞,ELISA检测处理前后LAMP对诱导肿瘤坏死因子α(TNF-α)和白细胞介素1β(IL-1β)分泌的影响。结果 LAMP能诱导滋养层细胞HO-1 mRNA和蛋白表达,并能诱导其产生ROS以及促进Nrf2核转位。ROS抑制剂NAC预处理后,可明显降低HO-1的表达水平以及细胞核内Nrf2含量。同时,Nrf2siRNA转染后,HO-1表达显著减少。采用Zn PP处理滋养层细胞,或RNA干扰HO-1表达后,可促进LAMP诱导滋养层细胞分泌TNF-α和IL-1β,而Co PP处理能进一步降低TNF-α和IL-1β水平。结论 LAMP能通过ROS/Nrf2诱导滋养层细胞表达HO-1,从而抑制细胞因子的过度分泌。更多还原

【Abstract】 Objective To observe the expression of heme oxygenase-1( HO-1) in regulation of cytokines response induced by Mycoplasma genitalium-derived lipid-associated membrane proteins( LAMPs) in placental trophoblast cells.Methods Placental trophoblast cells were cultured in vitro and stimulated by 0. 5- 5 μg / m L LAMPs for 4 to 12 hours.Expression of HO-1 mRNA and protein,and nuclear translocation of nuclear factor erythroid-2 related factor 2( Nrf2) were detected by real-time quantitative PCR and Western blotting,respectively. The intracellular formation of reactive oxygen species( ROS) was detected by the fluorescent probe H2 DCFDA. N-acetyl-cysteine( NAC) and nuclear factor erythroid-2related factor 2( Nrf2) siRNA were respectively used to analyze the roles of ROS and Nrf2 in mediating HO-1 expression.Finally,placental trophoblast cells were transfected with HO-1 siRNA,or preincubated by the HO-1 agonist cobalt protoporphyrin( Co PP) or its inhibitor zinc protoporphyrin( Zn PP),and LAMPs-induced secretion of TNF-α and IL-1β were detected by ELISA.Results M. genitalium LAMPs induced the expression of HO-1 mRNA and protein,the accumulation of ROS and the nuclear translocation of Nrf2 in placental trophoblast cells. NAC treatment inhibited LAMPs-induced HO-1 expression and Nrf2 nuclear translocation,and the transfection of Nrf2 siRNA significantly abrogated HO-1 expression. Furthermore,HO-1 siRNA and Zn PP treatment increased LAMPs-induced TNF-α and IL-1β secretion,while the HO-1 agonist Co PP treatment further decreased their production. Conclusion M. genitalium LAMPs could induce placental trophoblast cells to express HO-1through ROS / Nrf2 pathways. Up-regulation of HO-1 negatively regulates excessive production of cytokines.更多还原