血清外泌体miR-21靶向PTEN激活PI3K/AKT信号通路诱导肺癌细胞的生长和转移的机制分析  

Serum exosomes miR-21 targets PTEN to activate PI3K/AKT signaling pathway to induce the growth and metastasis of lung cancer cells

作　　者：张茜[1] 同立宏[1] 马红霞[1] ZHANG Qian;TONG Lihong;MA Hongxia(Department of Respiratory and Critical Care Medicine,Affiliated Hospital of Traditional Chinese Medicine,Xinjiang Medical University,Urumqi,Xinjiang 830002,China)

机构地区：[1]新疆医科大学附属中医医院呼吸与危重症医学科,新疆乌鲁木齐830002

出　　处：《国际检验医学杂志》2022年第12期1479-1484,1490,共7页International Journal of Laboratory Medicine

基　　金：新疆维吾尔自治区重点实验室开放课题(2020D04030)。

摘　　要：目的探讨肺癌患者血清外泌体miR-21对肺癌细胞生长和转移的影响及机制。方法收集18例健康志愿者和20例肺癌患者的血清,提取并鉴定血清外泌体(记为Normal exo和LCA exo),qRT-PCR检测Normal exo和LCA exo中miR-21的表达水平,双荧光素酶报告基因实验检测miR-21与PTEN的靶向关系,肺癌细胞A549中转染miR-NC(miR-NC组)、miR-21 mimic(miR-21组)、miR-21 mimic+PTEN质粒(miR-21+PTEN组)后,CCK8法检测细胞增殖,Transwell小室法检测细胞迁移和侵袭能力,Western blot检测PTEN蛋白表达。将10μg/mL时Normal exo和LCA exo及等体积的PBS(Normal exo组、LCA exo组、PBS组)与A549细胞共孵育,检测细胞中miR-21表达,PTEN蛋白表达,细胞增殖、迁移和侵袭能力。A549细胞与10μg/mL时LCA exo共孵育后转染PTEN质粒(LCA exo+PTEN组),然后检测细胞增殖、迁移和侵袭能力。Western blot检测PBS组、Normal exo组、LCA exo组和LCA exo+PTEN组A549细胞中磷酸酰肌醇3-激酶(PI3K)和蛋白激酶B(AKT)磷酸化水平。结果Normal exo和LCA exo符合外泌体的结构及生物学特征。LCA exo组中miR-21表达显著高于Normal exo组(P<0.001)。PTEN为miR-21的靶基因,与miR-NC组比较,miR-21组A549细胞PTEN表达下调(P<0.001),细胞增殖、迁移、侵袭能力显著增加(P<0.05)。与miR-21组比较,miR-21+PTEN组细胞增殖、迁移、侵袭能力显著降低(P<0.05)。相比于PBS组,LCA exo组细胞PTEN表达显著下调(P<0.001),细胞增殖、迁移、侵袭能力显著增加(P<0.05),PI3K和AKT磷酸化水平显著增加(P<0.001);与LCA exo组比较,LCA exo+PTEN组A549细胞增殖、迁移、侵袭能力显著降低(P<0.05),PI3K和AKT磷酸化水平显著降低(P<0.001)。结论肺癌患者血清外泌体传递miR-21靶向PTEN促进肺癌细胞的生长和转移。Objective To explore the effect and mechanism of serum exosomes miR-21 on the growth and metastasis of lung cancer cells.Methods The serum of 18 healthy volunteers and 20 patients with lung cancer were collected.The serum exosomes(Normal exo and LCA exo)were extracted and identified.The expression levels of miR-21 in Normal exo and LCA exo were detected by qRT-PCR.The targeted relationship between miR-21 and PTEN was detected by dual luciferase reporter gene assay.Lung cancer cell line A549 were transfected miR-21 NC(miR-NC group),miR-21 mimic(miR-21 group)and miR-21 mimic+PTEN plasmids(miR-21+PTEN group),cell proliferation was detected by CCK8 method,cell migration and invasion were detected by Transwell chamber method,and PTEN protein expression was detected by Western blot.In addition,10μg/mL Normal exo,LCA exo and PBS(Normal exo group,LCA exo group,PBS group)were incubated with A549,the expression of miR-21,PTEN protein,cell proliferation,migration and invasion were detected.After incubating with 10μg/mL LCA exo,A549 were transfected with PTEN plasmid(LCA exo+PTEN group),cell proliferation,migration and invasion were detected.Western blot was used to detect the phosphorylation levels of phosphatidylinositol 3-kinase(PI3K)and protein kinase(AKT)in A549 cells of PBS group,normal exo group,LCA exo group and LCA exo+PTEN group.Results Normal exo and LCA exo were consistent with the structure and biological characteristics of exosomes.The expression of miR-21 in LCA exo was significantly higher than that in Normal exo group(P<0.001).PTEN was the target gene of miR-21.Compared with miR-NC group,PTEN expression was down regulated in miR-21 group(P<0.001),and cell proliferation,migration and invasion were significantly increased(P<0.05).Compared with miR-21 group,the proliferation,migration and invasion ability of miR-21+PTEN group were significantly decreased(P<0.05).Compared with PBS group,the expression of PTEN in LCA exo group was significantly down regulated(P<0.001),the proliferation,migration and invasion of

关 键 词：肺癌 血清外泌体 MIR-21 生长 转移 磷酸酰肌醇3-激酶/蛋白激酶B信号通路 

分 类 号：R734.2[医药卫生—肿瘤]