CAJ | 학술논문

目的：探讨抗霉素 A（AMA）诱导血小板凋亡及其分子机制。方法取健康志愿者单采血小板，离心洗涤得到洗涤血小板，将洗涤血小板与不同浓度的 AMA 孵育后，流式细胞术检测线粒体跨膜电位（ΔΨm）去极化、磷脂酰丝氨酸（PS）暴露、细胞内活性氧（ROS）、线粒体 ROS、P-选择素表达和整合素αⅡbβ3活化；Western blot 法检测半胱氨酸天冬氨酸蛋白酶3（Caspase-3）的活化。在抑制试验中，先将洗涤血小板与线粒体靶向的 ROS 拮抗剂 Mito-TEMPO 预孵育，然后再与 AMA 孵育，流式细胞术检测相关凋亡指标。结果AMA 剂量依赖性诱导血小板ΔΨm 去极化、PS 暴露、细胞内ROS 和线粒体 ROS 升高以及 Caspase-3活化；但是 AMA 不能诱导血小板活化。线粒体靶向的 ROS 拮抗剂抑制 AMA诱导的血小板ΔΨm 去极化、PS 暴露、Caspase-3活化以及线粒体 ROS 的生成。结论 AMA 能够诱导血小板发生凋亡，线粒体 ROS 可能在 AMA 诱导的血小板凋亡过程中起重要作用。
목적：탐토항매소 A（AMA）유도혈소판조망급기분자궤제。방법취건강지원자단채혈소판，리심세조득도세조혈소판，장세조혈소판여불동농도적 AMA 부육후，류식세포술검측선립체과막전위（ΔΨm）거겁화、린지선사안산（PS）폭로、세포내활성양（ROS）、선립체 ROS、P-선택소표체화정합소αⅡbβ3활화；Western blot 법검측반광안산천동안산단백매3（Caspase-3）적활화。재억제시험중，선장세조혈소판여선립체파향적 ROS 길항제 Mito-TEMPO 예부육，연후재여 AMA 부육，류식세포술검측상관조망지표。결과AMA 제량의뢰성유도혈소판ΔΨm 거겁화、PS 폭로、세포내ROS 화선립체 ROS 승고이급 Caspase-3활화；단시 AMA 불능유도혈소판활화。선립체파향적 ROS 길항제억제 AMA유도적혈소판ΔΨm 거겁화、PS 폭로、Caspase-3활화이급선립체 ROS 적생성。결론 AMA 능구유도혈소판발생조망，선립체 ROS 가능재 AMA 유도적혈소판조망과정중기중요작용。
Objective To investigate the effects and molecular mechanism of platelet apoptosis induced by Antimy-cin A (AMA).Methods Washed healthy volunteers platelets were pre-incubated with different concentration AMA,and then depolarization of mitochondrial membrane potential (ΔΨm),phosphatidylserine (PS)externaliza-tion assay,the surface expression of P-selectin,the activation of integrin αIIβ3 and the production of intracellular reactive oxygen species (ROS),mitochondrial ROS were detected by flow cytometry.The activation of Caspase-3 was analyzed by Western blot.In inhibition experiment,washed platelets were pre-incubated mitochondrial ROS targeted antagonists Mito-TEMPO,and then stimulated with different concentration AMA.The apoptosis associated indicators were detected by flow cytometry.Results AMA dose-dependently induces depolarization of ΔΨm,PS exposure,Caspase-3 activation,and the production of intracellular ROS and mitochondrial ROS,but AMA does not induce platelet activation.Mito-TEMPO effectively reduces the depolarization of ΔΨm,PS exposure and the pro-duction of mitochondrial ROS which induced by the AMA.Conclusion AMA can induce platelet apoptosis,and mitochondria ROS may play an important role in the platelets apoptosis induced by the AMA.