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  • 范晶炎,程忠平.中性粒细胞弹性蛋白酶、金属基质蛋白酶9对卵巢癌细胞膜E-cadherin、MUC16表达的影响[J].同济大学学报(医学版),2019,40(6):821-826.    [点击复制]
  • FAN Jing-yan,CHENG Zhong-ping.Effect of neutrophil elastase and matrix metalloproteinase-9 on expression of membrane protein in ovarian cancer cells[J].同济大学学报(医学版),2019,40(6):821-826.   [点击复制]
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中性粒细胞弹性蛋白酶、金属基质蛋白酶9对卵巢癌细胞膜E-cadherin、MUC16表达的影响
范晶炎,程忠平
0
(上海交通大学医学院附属第九人民医院妇产科,上海 200001;同济大学附属第十人民医院妇产科,上海 200072; 同济大学医学院妇科微创医学研究所,上海 20009)
摘要:
目的 检测中性粒细胞弹性蛋白酶(neutrophil elastase, NE)、金属基质蛋白酶9(matrix metalloproteinase-9, MMP-9)对体外卵巢癌细胞膜蛋白E-cadherin、黏蛋白16(mucin16,MUC16)表达的影响。方法 体外培养人卵巢癌OVCAR3细胞。温控4℃,MMP-9、NE、0.125%胰蛋白酶分别处理OVCAR3细胞,光镜下观察不同时间段细胞形态学变化;采用Western印迹法检测OVCAR3细胞中E-cadherin、MUC16总体表达;免疫荧光技术定位E-cadherin 、MUC16。应用酶联吸附测定技术(ELISA)定量培养基中脱落的MUC16胞外段蛋白。结果 NE、MMP9处理细胞后,细胞形态由平铺鹅卵石样呈细长/纺锤状,细胞间逐渐分离、扩散。NE、MMP9处理24h后E-cadherin表达均显著下降,与对照组相比差异有统计学意义(P<0.05),而0.125%胰酶组与对照组之间差异无统计学意义。免疫荧光检测结果检测E-cadherin、MUC16主要定位在细胞膜。ELISA结果显示,MMP-9处理细胞后,上清液中MUC16胞外段CA125的含量明显增高。结论 中性粒细胞相关蛋白酶NE、MMP9诱导细胞形态由平铺鹅卵石样呈细长/纺锤状,细胞间逐渐分离、扩散。两者通过酶解卵巢癌细胞膜上E-cadherin,促进上皮间叶转化(epithelial mesenchymal transformation, EMT),细胞间接触和极性的丧失,进一步脱落。此外,MMP-9可以降解细胞膜上MUC16蛋白,使MUC16胞外段脱落,揭示局部MMP-9酶浓度升高与血清学CA125水平有一定关联。
关键词:  中性粒细胞弹性蛋白酶  金属基质蛋白酶9  E-cadherin  黏蛋白16  卵巢癌
DOI:10.16118/j.1008-0392.2019.06.010
投稿时间:2019-03-12
基金项目:上海交通大学医学院紧缺专业硕士研究生临床研究能力提升计划(JQ201709)
Effect of neutrophil elastase and matrix metalloproteinase-9 on expression of membrane protein in ovarian cancer cells
FAN Jing-yan,CHENG Zhong-ping
(Dept. of Obstetrics and Gynecology, Shanghai Ninth People’s Hospital, Shanghai Jiao-tong University, Shanghai 200001, China;Dept. of Obstetrics and Gynecology, Tenth People’s Hospital, Tongji University School of Medicine, Shanghai 200072, China;Institude of Gynecological Minimal Invasive Medicine, Tongji University School of Medicine, Shanghai 200090, Chin)
Abstract:
Objective To investigate the effects of neutrophil elastase(NE) and matrix metalloproteinase-9(MMP-9) on the expression of E-cadherin and MUC16 in ovarian cancer cells. Methods Ovarian cancer OVCAR3 cells were treated with MMP-9, NE and 0.125% trypsin at 4℃. The morphological changes of OVCAR3 cells were observed under light microscope at different time points. Western blotting was used to detect E-cadherin and overall expression of MUC16 in OVCAR3 cells. Immunofluorescence technique was performed to observe the localization of E-cadherin and MUC16. The extracellular domain protein of MUC16 in the medium was quantified by enzyme-linked immunosorbent assay(ELISA). Results Light microscopy showed that after treatment with NE and MMP9, the OVCAR3 cells were slender/spindle-like, and the cells gradually separated and spread. Western blotting results showed that the expression of E-cadherin decreased(P<0.05) in cells treated with NE and MMP9 for 24h; while there was no significant difference between the 0.125% trypsin group and control group. The immunofluorescence assay illustrated that E-cadherin, MUC16 were mainly located on the cell membrane. ELISA results showed that CA125(the extracellular domain of MUC16) was significantly increased in the supernatant after MMP-9 treatment. Conclusion NE and MMP9 can induce the morphological changes of OVCAR3 cells. Both of them can decrease the expression of E-cadherin on the cell membrane, which may promote intraperitioneal metastasis of ovarian cancer cells. MMP-9 can degrade MUC16 on the cell membrane, which may lead to increase serum CA125 level.
Key words:  neutrophil elastase  matrix metalloproteinase-9  E-cadherin  mucin16  ovarian cancer

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