LPS、PLA_2及氧自由基对线粒体跨膜质子转运和H^+ -ATPase的影响

Effects of LPS, PLA_2 and oxygen free radicals on proton translocation across inner mitochondrial membrane and H^+ -ATPase

目的：研究细菌内毒素（LPS）、磷脂酶A2（PLA2）及自由基对质子跨膜转运及H+－ATP酶的影响。方法：采用正常大鼠，制备线粒体、亚线粒体（SMP），用SMP与LPS（100μg／ml）、PLA2（10u／ml）、FeSO4；／VitC（30／900μmol／L）分别共同温育（30C，30min），用ACMA荧光淬灭法测定质子跨膜转运能力。线粒体与不同浓度LPS、PLA2和FeSO4／VitC共同孵育，测定H+－ATP酶、PLA2、MDA。结果：LPS、PLA2、FeSO4／VitC可致亚线粒体ACMA荧光淬灭显著改变，LPS可引起线粒体膜PLA2活性、MDA含量升高（P＜0．05），LPS、PLA2可引起H+－ATP酶的活性进行性下降，小剂量FeSO4／VitC引起H+－ATP酶活性升高，大剂量FeSO4／VitC引起H+－ATP酶活性降低。结论：LPS、PLA2、LPO是造成质子转运能力下降的重要原因．自由基与H+－ATP酶损伤密切相关。

Objective: To elucidate the effects of LPS, PLA2 and oxygen free radical (OFR ) on proton transmembrane translocation and H+-ATPase. Methods: Normal rats were sacrificed to prepare liver mitochondria and SMPs in vitro. After the SMPs were incubated with LPS (100 μg/ml ), PLA2 (10 μg/ml )and FeSO4/Vit C (30/900 μmol/L) respectively at 30 C for 30 min, the proton translocation of SMPs was assayed with fluorescent probe ACMA. The mitochondria were incubated in different concentrations of LPS,PLA2 and FeSO4/Vit C to determine H+-ATPase, PLA2 and MDA. Results: ① The fluorescent quenching of ACMA and H +-ATPase were significantly decreased after the treatment with LPS2, PLA2 and FeSO4/Vit C at high dosages (P<O. 05). ② The mitochondrial PLA2 activity and MDA content were significantly increased after the treatment with LPS (P <0. 01 ). Conclusion: LPS, PLA2 and LPO are the important causes for the decrease of proton translocation across the membrane and OFR is closely related to the damage of H+-ATPase.