四色流式分析冠心病患者外周血单核细胞亚群及单核细胞-血小板聚集体的变化  

A four-color flow cytometric assay for the analysis of monocyte subsets and monocyte-platelet aggregates in patients with coronary heart disease

作　　者：杨国红[1,2] 卢芮伊[2] 孙海英[2] 张玲[2] 姬文婕[3] 周欣[2] 石蕊[2] 姜铁民[2] 李玉明[2] 

机构地区：[1]天津医科大学研究生院,天津300070 [2]武警后勤学院附属医院心血管病研究所心脏中心 [3]武警后勤学院附属医院呼吸与重症医学科

出　　处：《临床心血管病杂志》2013年第1期29-33,共5页Journal of Clinical Cardiology

基　　金：国家自然科学基金课题(No:81070121;81170238);天津市自然科学基金(No:09ZCZDSF04200;11JCYBJC12000)

摘　　要：目的:建立基于四色的流式细胞术检测外周血单核细胞亚群及各亚群单核细胞-血小板聚集体(MPA)的方法,并观察冠心病患者外周血单核细胞亚群及各亚群MPA的变化特点。方法:本研究纳入经冠状动脉造影证实的冠心病患者和健康对照者各25例,记录所有研究对象的一般临床资料,并采集外周静脉血,依据单核细胞和血小板的标志性分子CD86、CD14、CD16和CD41对单核细胞亚群和MPA进行四色的流式细胞术(FCM)分析。利用抗CD86-PE-Cy5圈出总的单核细胞(总Mon)群,利用抗CD14-FITC和抗CD16-PE将总Mon分为:经典型(CD14++CD16-,Mon1)、中间型(CD14++CD16+,Mon2)和非经典型(CD14+CD16++,Mon3)3个亚群,最后应用CD41-PE-Cy7分析总Mon和各个亚群的MPA,并应用Flow-count TM荧光微球对各个亚群及MPA的绝对量进行计数,比较两组研究对象外周血单核细胞各亚群及各亚群MPA的差异。结果:冠心病组患者外周血Mon2和Mon3所占百分比和绝对计数均显著高于对照组(P<0.05),而Mon1和总Mon两组间比较差异无统计学意义;与对照组相比,冠心病患者总MPA和Mon2相关MPA所占百分比和绝对计数亦显著增高(P<0.05),而Mon1相关MPA和Mon3相关MPA的百分比及绝对计数两组间比较差异无统计学意义。结论:基于CD14、CD16、CD86和CD41的四色流式分析能够有效定量分析外周血单核细胞各亚群及各亚群MPA。中间型单核细胞及其形成的MPA水平显著升高,为临床上冠心病患者的诊治策略提供了新思路。Objective：To construct a new four-color flow cytometric assay for analysis of peripheral blood monocyte subsets and monocyte-platelet aggregates（MPA） and observe their characteristics in patients with coronary heart disease（CHD）. Method：The 25 coronary angiography documented CHD patients and 25 healthy volunteers were included in the study.General characteristics were recorded and blood samples were collected.Monocyte subsets and MPA were analyzed using a four-color flow cytometric assay based on CD86,CD14,CD16 and CD41.we distinguished total monocytes using CD86-PE-Cy5 antibody and three subsets： classical（CD14＋＋CD16-,Mon1）,intermediate（CD14＋＋CD16＋,Mon2）,and non-classical（CD14＋CD16＋＋,Mon3） monocytes using CD14-FITC and CD16-PE antibodies,then MPA were distinguished using CD41-PE-Cy7 antibody.The absolute number of monocytes and MPA were calculated with reference to Flow-countTM Fluorospheres.At last we compared the percentage and absolute cell numbers of monocyte subsets and MPA in the two groups. Result：Compared with control group,percentage and absolute cell numbers of Mon2 and Mon3 were significantly increased in CHD group（P0.05）,while there were no significantly differences between two groups in total Mon and Mon1.Meanwhile,percentage and absolute cell numbers of total Mon MPA and Mon2 MPA in CHD group were significantly increased compared with control group（P0.05）,but no significant differences were found in the numbers and percentage of Mon1 MPA and Mon3 MPA in two groups. Conclusion：Four-color flow cytometric assay is an useful tool for analysis monocyte subsets and MPA.And we find that intermediate monocytes and MPA increase significantly which may be a potential therapeutic and diagnostic target for CHD patients.

关 键 词：单核细胞 亚群 血小板 单核细胞-血小板聚集体 流式细胞术 

分 类 号：R541.4[医药卫生—心血管疾病]