去白细胞悬浮红细胞来源外泌体的释放及其对血液肿瘤细胞的调控  

Release of Exosomes Derived from Leukocyte⁃Depleted Red Cell Suspension and Its Regulation on Hematological Tumor Cells

作　　者：黄豪博[1] 范丽萍[1] 林秋燕[1] 黄惠炆[1] 付丹晖[1,2] HUANG Hao-Bo;FAN Li-Ping;LIN Qiu-Yan;HUANG Hui-Wen;FU Dan-Hui(Department of Blood Transfusion,Fujian Medical University Union Hospital,Fuzhou 350001,Fujian Province,China;Department of Hematology,Fujian Medical University Union Hospital,Fuzhou 350001,Fujian Province,China)

机构地区：[1]福建医科大学附属协和医院输血科,福建福州350001 [2]福建医科大学附属协和医院血液科,福建福州350001

出　　处：《中国实验血液学杂志》2022年第4期1188-1192,共5页Journal of Experimental Hematology

基　　金：福建省医学创新课题资助(2019-CX-15,2018-CXB-7)。

摘　　要：目的:探讨不同储存时间去白细胞悬浮红细胞(LDRCS)中外泌体(exosome,Exo)的释放及其对血液肿瘤细胞增殖的调控与可能机制。方法:应用超高速离心方法获得不同储存时间的LDRCS中Exo(RBC⁃Exo),采用透射电镜术、蛋白免疫印迹法分别对RBC⁃Exo的形态、免疫标记进行检测。采用动态光散射检测不同储存时间LDRCS中RBC⁃Exo的粒径分布变化。采用CCK⁃8法检测RBC⁃Exo对血液肿瘤细胞增殖的影响。采用蛋白免疫印迹法检测与RBC⁃Exo共培养后血液肿瘤细胞内增殖相关蛋白的表达变化。结果:分离出呈类杯状、粒径分布于20-200 nm、CD63/TSG101富集、Calnexin阴性、CD235a阳性、CD41阴性的RBC⁃Exo。储存中晚期(>14 d)LDRCS中RBC⁃Exo粒径分布无显著差异,但显著大于储存早期(储存≤14 d)的RBC⁃Exo。与对照组相比,RBC⁃Exo可以明显促进血液肿瘤细胞HBL1、U2932和Jurkat的增殖。与RBC⁃Exo共培养3 d的HBL1、U2932和Jurkat细胞中周期相关蛋白P21的表达较对照组明显下调,而抗凋亡蛋白BCL⁃2并无明显变化。结论:储存中晚期的LDRCS中RBC⁃Exo形态与储存早期存在差异。RBC⁃Exo可以促进血液肿瘤细胞的增殖,其机制可能是通过调节周期相关蛋白P21的表达。Objective:To investigate the release of exosome(Exo)from leukocyte⁃depleted red cell suspension(LDRCS)at different storage time and its regulation on proliferation of hematological tumor cells and possible mechanism.Methods:The Exo(RBC⁃Exo)in LDRCS at different storage time was obtained by ultracentrifugation,and the morphology and immunological marker of RBC⁃Exo were detected by transmission electron microscopy and Western blot,respectively.The particle size distribution of RBC⁃Exo in LDRCS at different storage time was detected by Dynamic Light Scattering.CCK⁃8 assay was used to explore the effect of RBC⁃Exo on hematological tumor cell proliferation.Western blot was used to detect the expression of proliferation⁃related proteins in hematological tumor cells after co⁃culture with RBC⁃Exo.Results:RBC⁃Exo was isolated,which was characterized by cup⁃like shape,particle size distribution ranged from 20 to 200 nm,CD63/TSG101 enriched,Calnexin negative,CD235a positive and CD41 negative.The particle size distribution of RBC⁃Exo from LDRCS between middle was not significantly different and late stored stage.But the particle size distribution of RBC⁃Exo at middle⁃late stored stage(>14 d)was larger than that at early stored stage(≤14 days).Compared with the control group,RBC⁃Exo could significantly promote the proliferation of HBL1,U2932 and Jurkat cells.Compared with the control group,the cycle⁃related protein P21 was significantly down⁃regulated in HBL1,U2932 and Jurkat cells after co⁃culture with RBC⁃Exo for 3 days,while the anti⁃apoptotic protein BCL⁃2 was not changed significantly.Conclusion:The morphology of RBC⁃Exo from LDRCS at middle⁃late stored stage was different from that at early stored stage.RBC⁃Exo could promote the proliferation of hematological tumor cells,possibly by regulating the expression of cycle⁃associated protein P21.

关 键 词：去白细胞悬浮红细胞 外泌体 肿瘤细胞 增殖 

分 类 号：R457.1[医药卫生—治疗学] R733[医药卫生—临床医学]