负载膜联蛋白A2(annexin A2)的间充质干细胞来源外泌体减少M2巨噬细胞极化抑制裸鼠前列腺癌细胞生长  

Exosomes derived from mesenchymal stem cells loaded with annexin A2 reduce the polarization of M2 macrophages to inhibit the growth of prostate cancer cells in nude mice

作　　者：田野[1] 郭小鹏[1] 成俊[1] 王鹏[1] TIAN Ye;GUO Xiaopeng;CHENG Jun;WANG Peng(Department of Urology,The Fourth Affiliated Hospital of Xinjiang Medical University(Xinjiang Uygur Autonomous Region Traditional Chinese Medicine Hospital),Urumqi 830000,China)

机构地区：[1]新疆医科大学第四附属医院(新疆维吾尔自治区中医医院)泌尿外科,新疆乌鲁木齐830000

出　　处：《细胞与分子免疫学杂志》2023年第2期109-116,共8页Chinese Journal of Cellular and Molecular Immunology

基　　金：新疆维吾尔自治区自然科学基金(2020D01C108)。

摘　　要：目的探究负载膜联蛋白A2(ANXA2)的骨髓间充质干细胞(BMSC)来源的外泌体(Exo-ANXA2)对前列腺癌细胞增殖、侵袭与迁移以及前列腺癌移植瘤生长的影响,并研究巨噬细胞是否参与该过程。方法分离与培养BALB/c裸鼠BMSC,采用负载ANXA2的慢病毒质粒感染BMSC,并分离外泌体;加入外泌体处理THP-1巨噬细胞,ELISA检测细胞上清培养液肿瘤坏死因子α(TNF-α)、白细胞介素1β(IL-1β)、IL-6和IL-10的水平;将外泌体处理的巨噬细胞与前列腺癌细胞共培养后,CCK-8法检测细胞增殖活性,Transwell^(TM)小室检测细胞侵袭与迁移。通过注射PC-3人前列腺癌细胞构建前列腺癌裸鼠移植瘤模型,将建模后的裸鼠随机分为对照组和实验组,每组8只,实验组裸鼠尾静脉注射1 mL Exo-ANXA2,对照组注射等量PBS,于注射后第0、3、6、9、12、15、18、21天,使用游标卡尺测量计算肿瘤体积,21 d处死裸鼠并测量肿瘤组织质量,免疫组织化学染色法检测肿瘤组织中抗原KI-67(ki67)和CD163表达。结果骨髓分离的细胞表面CD90和CD44呈高表达,CD34和CD45呈低表达,且成骨成脂分化能力较强,成功获得BMSC;负载ANXA2的慢病毒质粒感染后,BMSC中有较强的绿色荧光蛋白表达,并成功分离到Exo-ANXA2;经Exo-ANXA2处理后,THP-1细胞中TNF-α和IL-6的水平显著升高而IL-10和IL-13的水平显著下降;Exo-ANXA2处理巨噬细胞后,显著抑制Exo-ANXA2促进PC-3细胞增殖活性、侵袭与迁移的作用;经过给予前列腺癌移植瘤裸鼠注射Exo-ANXA2后,裸鼠在第6、9、12、15、18、21天的肿瘤组织体积显著减小,21 d时裸鼠肿瘤体质量也显著减少,此外,肿瘤组织内ki67和CD163的阳性表达率均显著降低。结论Exo-ANXA2能够抑制前列腺癌细胞增殖、侵袭与迁移,并抑制裸鼠前列腺癌移植瘤生长,M2型巨噬细胞减少。Objective To identify the effect of exosomes derived from bone marrow mesenchymal stem cells(BMSCs)loaded with annexin A2(ANXA2)on the proliferation,migration,invasion of prostate cancer cells,and the transplanted tumor of prostate cancer in nude mice growth,as well as the role of macrophages in this process.Methods BMsCs from BALB/c nude mice were isolated and cultured.BMSCs were infected with lentiviral plasmids loaded with ANXA2.Exosomes were isolated and then added to treat THP-1 macrophages.ELISA was used to detect the levels of tumor necrosis factor-α(TNF-α),interleukin 1β(IL-1β),IL-6 and IL-10 in the cell supernatant culture fluid;After co-culturing the exosomes-treated macrophages and prostate cancer cells,CCK-8 assay was used to detect the cell prolferation activity.Also,Transwell^(TM) chamber were used to detect the cell invasion and migration.A nude mouse xenograft model of prostate cancer was constructed by injecting PC-3 human prostate cancer cells,the modeled nude mice were randomly divided into a control group and an experimental group,with 8 mice in each group.The nude mice in experimental group were injected with 1 mL of Exo-ANXA2 through the tail vein,while the control group was injected with the same amount of PBS,on days 0,3,6,9,12,15,18,and 21 after injection.Then the tumor volume was measured and calculated with vernier calipers.The nude mice were sacrificed at 21 days with their the tumor mass measured.The immunohistochemical staining method was used to detect the expression of antigen Kl-67(ki67)and CD163 expression in tumor tissue.Results The cells isolated from bone marrow showed high expression of CD90 and CD44 on the surface,and low expression of CD34 and CD45,and had strong osteogenic adipogenic differentiation ability,indicating that BMsCs were successfully obtained.After infection with the lentiviral plasmid carrying ANXA2,a strong green fluorescent protein expression in BMSCs,and Exo-ANXA2 was isolated.After Exo-ANXA2 treatment,the levels of TNF-αand IL-6 in THP-1 cells increa

关 键 词：前列腺癌 膜联蛋白A2(ANXA2) 外泌体 巨噬细胞极化 

分 类 号：R737.25[医药卫生—肿瘤] R392.12[医药卫生—临床医学] Q279[生物学—细胞生物学] Q25