胃癌间质干细胞调节中性粒细胞极化的作用及分子机制  

Regulation effect of gastric cancer-derived mesenchymal stem cells on polarization of neutrophils and its molecular mechanism

作　　者：朱琼芳[1] 张徐[2] 袁潇[2] 牛健[1] 

机构地区：[1]苏州大学附属第一医院临床检测中心,江苏苏州215006 [2]江苏大学医学院分子生物学教研室,江苏镇江212013

出　　处：《肿瘤》2016年第9期959-967,共9页Tumor

摘　　要：目的 :探讨胃癌间质干细胞(mesenchymal stem cells,MSCs)条件培养液对调节中性粒细胞极化的作用及可能的分子作用机制。方法:采用免疫组织化学和HE染色法检测胃癌组织中中性粒细胞和MSCs的浸润情况。采用趋化实验检测胃癌MSCs条件培养液对中性粒细胞的趋化作用;用胃癌MSCs条件培养液处理中性粒细胞18 h后,FCM法检测中性粒细胞的凋亡情况;实时荧光定量PCR法检测中性粒细胞中抗凋亡相关基因MCL-1以及极化相关基因自杀相关因子(factor associated suicide,FAS)、细胞间黏附分子-(1intercellular cell adhesion molecule-1,ICAM-1)、单核细胞趋化蛋白1(monocyte chemotactic protein 1,MCP1,又称为CCL2)和白细胞介素8(interleukin-8,IL-8)m RNA的表达情况。将胃癌MSCs条件培养液处理的中性粒细胞作用于胃癌SGC-7901细胞,采用Transwell小室迁移实验检测对胃癌SGC-7901细胞迁移能力的影响。蛋白质印迹法检测胃癌MSCs条件培养液处理的中性粒细胞中细胞外信号调节激酶(extracellular signal-regulated kinase,ERK)信号通路的活化情况。结果 :胃癌组织中存在中性粒细胞和MSCs的浸润。胃癌MSCs条件培养液能高效诱导中性粒细胞的趋化(P<0.01);胃癌MSCs条件培养液处理后的中性粒细胞,凋亡率明显下降(P<0.05),MCL-1 m RNA的表达水平则明显上调(P<0.05);极化相关基因FAS和ICAM-1 m RNA的表达水平明显下调(P<0.05和P<0.01),CCL2和IL-8 m RNA的表达水平均明显上调(P值均<0.01)。SGC-7901细胞与胃癌MSCs条件培养液预处理的中性粒细胞共培养后,其迁移能力明显增强(P<0.01)。胃癌MSCs条件培养液预处理中性粒细胞15 min后,中性粒细胞中磷酸化ERK(phospho-ERK,p-ERK)蛋白的表达水平显著升高(P<0.05)。结论 :胃癌MSCs条件培养液能够调节中性粒细胞趋化并抑制其自发性凋亡;诱导中性粒细胞N2型极化并促进胃癌细胞迁移;这可能与中性粒细胞内ERK信号通路被激活相关。Objective： To investigate the regulation effect of conditioned medium of gastric cancer- derived mesenchymal stem cells （MSCs） on the polarization of neutrophils, and to explore its possible molecular mechanism. Methods： The infiltrations of neutrophils and MSCs in gastric cancer tissues were detected by immunohistochemistry and HE staining. The effect of gastric cancer-derived MSCs conditioned medium on the chemotaxis of neutrophil was detected by chemotaxis assay. After the conditioned medium of gastric cancer-derived MSCs was used to treat the neutrophils for 18 h, the apoptosis of neutrophils was detected by flow cytometry, while the expression levels of anti-apoptosis-related genes MCL-1 and polarization-dependent genes factor associated suicide （FAS）, intercellular cell adhesion molecule-1 （ICAM-1）, monocyte chemotactic protein 1 （MCP1, also known as CCL2） and interleukin-8 （IL-8） mRNAs in neutrophils were detected by real-time fluorescent quantitative PCR. After the neutrophils were incubated with gastric cancer-derived MSCs conditioned medium for 15 min, the neutrophils pretreated by gastric cancer-derived MSCs conditioned medium was used to incubate with gastric cancer SGC-7901 cells, then the migration ability of gastric cancer cells was evaluated by Transwell migration assay. Finally, the activation of extracellular signal- regulated kinase （ERK） signaling pathway in neutrophils was detected by Western blotting. Results： The infiltrations of neutrophils and MSCs in gastric cancer tissues were identified. The conditioned medium of gastric cancer-derived MSCs remarkably induced the chemotaxis of neutrophils （P 〈 0.01）. After treatment with gastric cancer-derived MSCs conditioned medium, the apoptosis of neutrophils was significantly decreased （P 〈 0.05）, the expression levels of MCL-1, CCL2 and IL-8 mRNAs in neutrophils were significantly increased （P 〈 0.05, P 〈 0.01 and P 〈 0.01, respectively）, and the expression levels of FAS and ICAM-1 mRNAs in neut

关 键 词：胃肿瘤 间质干细胞 中性粒细胞 极化 转移 

分 类 号：R735.2[医药卫生—肿瘤]