CAJ | 학술논문

目的：研究重组创伤弧菌溶细胞素（rVvhA）对人单核细胞白血病细胞（THP-1）的损伤机制及其钙离子通道的研究。方法采用倒置显微镜、CCK-8法、Fluo3/AM法、caspase 活性检测等方法测定rVvhA对THP-1细胞的影响，并研究胞外Ca2＋的内流途径。结果 rVvhA作用后细胞形态学变化明显，细胞皱缩、变形甚至裂解，且呈剂量依赖性。12μg/ml rVvhA 作用THP-1细胞1 h后胞外K＋浓度升高，作用6 h后胞外LDH浓度显著升高。 Verapamil、Mibefradil和SKF-96365均不能抑制rVvhA引起的Ca2＋内流。同时rVvhA可导致THP-1细胞的caspase-3和caspase-9活性升高，并呈时间依赖性。结论 rVvhA对THP-1细胞具有明显的损伤作用，且可能通过caspase-9/3途径诱导细胞凋亡，并能引起细胞内Ca2＋浓度升高，升高的Ca2＋可能通过细胞膜上形成的孔道被动入胞。
목적：연구중조창상호균용세포소（rVvhA）대인단핵세포백혈병세포（THP-1）적손상궤제급기개리자통도적연구。방법채용도치현미경、CCK-8법、Fluo3/AM법、caspase 활성검측등방법측정rVvhA대THP-1세포적영향，병연구포외Ca2＋적내류도경。결과 rVvhA작용후세포형태학변화명현，세포추축、변형심지렬해，차정제량의뢰성。12μg/ml rVvhA 작용THP-1세포1 h후포외K＋농도승고，작용6 h후포외LDH농도현저승고。 Verapamil、Mibefradil화SKF-96365균불능억제rVvhA인기적Ca2＋내류。동시rVvhA가도치THP-1세포적caspase-3화caspase-9활성승고，병정시간의뢰성。결론 rVvhA대THP-1세포구유명현적손상작용，차가능통과caspase-9/3도경유도세포조망，병능인기세포내Ca2＋농도승고，승고적Ca2＋가능통과세포막상형성적공도피동입포。
Objective To investigate the role of recombinant Vibrio vulnificus cytolysin (rVvhA) in inducing THP-1 cells damage and study the pathway of associated calcium influx .Methods Inverted mi-croscope, CCK-8 cell proliferation kit, Fluo3/AM staining and caspase activity detection were performed to analyze the damage of THP-1 cells induced by rVvhA and the pathway of calcium influx .Results rVvhA had cytotoxic effects on THP-1 cells in a dose-dependent manner .The concentrations of extracellular K +and LDH were respectively up-regulated after 1 h and 6 h of 12 μg/ml rVvhA intervention .Verapamil , Mibe-fradil and SKF-96365 could not prevent the influx of free Ca 2+induced by rVvhA .The activities of caspase-3 and caspase-9 were singanificantly enhanced by rVvhA in a time-dependant manner .Conclusion rVvhA can induce THP-1 cells damage through triggering extracellular calcium influx via porous channel on cell membrane.Moreover, rVvhA might induce THP-1 cell apoptosis through activating caspase-9/3-dependent pathway .