卵巢癌细胞来源的外泌体对成纤维细胞分化的影响  

Effect of exosome derived from ovarian cancer cell on the differentiation of fibroblast

作　　者：张丹[1] 宋克娟[1] 任远中 隋磊 姚勤[1] Zhang Dan;Song Kejuan;Ren Yuanzhong;Sui Lei;Yao Qin(Department of Gynecology,the Affiliated Hospital of Qingdao University,Qingdao 266555,China;Department of Emergency Trauma Surgery,Luoyang Central Hospital Affiliated to Zhengzhou University,Luoyang 471000,China)

机构地区：[1]青岛大学附属医院妇科,青岛266555 [2]郑州大学附属洛阳中心医院急诊创伤外科,洛阳471000

出　　处：《中华肿瘤杂志》2022年第7期737-742,共6页Chinese Journal of Oncology

基　　金：山东省自然科学基金(ZR2016HM08);青岛市民生科技计划(19-6-46-nsh)。

摘　　要：目的探讨卵巢癌细胞分泌的外泌体对卵巢间质正常成纤维细胞(NFs)分化和迁移能力的影响。方法NFs来自2019年5—12月于青岛大学附属医院行子宫肌瘤切除的患者。使用超高速离心方法从卵巢癌SKOV3细胞培养上清中提取外泌体,将条件培养基(CM)、SKOV3-exo、磷酸盐缓冲液分别与NFs细胞共培养,分别记为CM组、SKOV3-exo组和空白组。采用实时荧光定量聚合酶链反应和Western blot法检测NFs中成纤维细胞活化蛋白(FAP)、α-平滑肌肌动蛋白(α-SMA)的mRNA和蛋白表达水平,采用Transwell实验检测NFs细胞的迁移能力。结果透射电镜下可观察到SKOV3的培养上清液中所提取到的细胞外囊泡为长径为30~100 nm,呈杯托样双层膜结构。外泌体中TSG101和HSP27蛋白表达水平分别为1.00±0.05和1.12±0.13,均高于卵巢癌SKOV3细胞(分别为0.22±0.21和0.36±0.14,均P<0.05)。PKH67荧光标记的外泌体可被NFs摄取。CM组细胞中α-SMA、FAP mRNA表达水平分别为2.91±0.15和3.21±0.33,SKOV3-exo组分别为3.50±0.21和4.63±0.24,均高于空白组(分别为1.00±0.06和1.00±0.13,均P<0.05)。CM组和SKOV3-exo组细胞中α-SMA(分别为0.89±0.11和1.25±0.09)、FAP蛋白表达水平(分别为0.81±0.09和1.20±0.12)均高于空白组(分别为0.12±0.31和0.11±0.19,均P<0.05)。CM组和SKOV3-exo组细胞迁移数分别为(215.01±14.80)个和(389.72±19.43)个,均高于空白组[(113.73±4.70)个,均P<0.05]。结论SKOV3分泌的外泌体可被NFs摄取,使其激活分化为肿瘤相关成纤维细胞(CAFs)并提高其迁移能力,说明卵巢癌细胞可能通外泌体的途径促进卵巢间质NFs向CAFs的转化,进而促进卵巢癌的发展。Objective To study the effects of exosome secreted by ovarian cancer(OC)cell on the differentiation and metastasis of normal fibroblasts(NFs).Methods NFs were collected from patients who underwent hysteromyoma resection in the Affiliated Hospital of Qingdao University from May to December 2019.Exosome was extracted from the culture supernatant of SKOV3 cells by using ultra-high-speed centrifugation.The NFs were co-cultured with condition medium(CM),exosome of SKOV3(SKOV3-exo)and control medium.The expression levels of fibroblast activation protein(FAP)andα-smooth muscle actin(α-SMA)were detected by real-time quantitative reverse transcription polymerase chain reaction(qRT-PCR)and western blot.The metastatic ability of NFs was detected by Transwell array.Results Under the transmission electron microscope,the extracellular vesicles extracted from the culture supernatant of SKOV3 were 30-100 nm in diameter with cup holder-like bilayer membrane structure,and the protein expression levels of TSG101 and HSP27 in exosomes(1.00±0.05 and 1.12±0.13)were higher than those of ovarian cancer SKOV3 cells(0.22±0.21 and 0.36±0.14,respectively,P<0.05).PKH67 fluorescently labeled exosomes could be taken up by NFs.The expression levels ofα-SMA and FAP mRNA in CM group(2.91±0.15 and 3.21±0.33)and SKOV3-exo group(3.50±0.21 and 4.63±0.24,respectively)were higher than that in blank group(1.00±0.06 and 1.00±0.13,P<0.05).The protein expression levels ofα-SMA and FAP in CM group and SKOV3-exo group(0.89±0.11 and 1.25±0.09,0.81±0.09 and 1.20±0.12)were higher than those in the blank group(0.12±0.31 and 0.11±0.19,respectively,P<0.05).The migrated numbers of cells in the CM group and SKOV3-exo group[(215.01±14.80)and(389.72±19.43),respectively]were higher than that in the blank group[(113.73±4.70),P<0.05].Conclusion The exosome secreted by SKOV3 cells can be taken up by NFs,which makes it to differentiate into cancer associated fibroblasts(CAFs)and significantly enhances its metastatic ability,indicating that OC cells m

关 键 词：卵巢肿瘤 肿瘤相关成纤维细胞 外泌体 

分 类 号：R737.31[医药卫生—肿瘤]