CDC42抑制剂CASIN对食管癌细胞活力与迁移能力的抑制作用

doi:10.3969/j.issn.1004-616x.2021.02.009

癌变·畸变·突变 ›› 2021, Vol. 33 ›› Issue (2): 136-142.doi: 10.3969/j.issn.1004-616x.2021.02.009

CDC42抑制剂CASIN对食管癌细胞活力与迁移能力的抑制作用

陈婉娴^1,2, 王思梦^1,2, 陈张瑜^1,2, 王潘集^1,2, 郑淳文^1,2, 李恩民^1,2, 许丽艳¹

1. 汕头大学医学院基础医学研究所, 广东汕头 515041;
2. 汕头大学医学院生物化学与分子生物学教研室, 广东汕头 515041

收稿日期:2020-12-08 修回日期:2021-02-22 出版日期:2021-03-30 发布日期:2021-04-12
通讯作者: 许丽艳,E-mail:lyxu@stu.edu.cn;李恩民,E-mail:nmli@stu.edu.cn E-mail:lyxu@stu.edu.cn;nmli@stu.edu.cn
作者简介:陈婉娴,E-mail:17wxchen2@stu.edu.cn。
基金资助:
广东大学生科技创新培育专项资金资助重点项目（pdjh2020a0218）；国家自然科学基金面上项目（81872372）

CASIN, a CDC42 inhibitor, inhibited viability and migration of esophageal carcinoma cells in vitro

CHEN Wanxian^1,2, WANG Simeng^1,2, CHEN Zhangyu^1,2, WANG Panji^1,2, ZHENG Chunwen^1,2, LI Enmin^1,2, XU Liyan¹

1. Institute of Basic Medicine, Shantou University Medical College, Shantou 515041;
2. Department of Biochemistry and Molecular Biology, Shantou University Medical College, Shantou 515041, Guangdong, China

Received:2020-12-08 Revised:2021-02-22 Online:2021-03-30 Published:2021-04-12

摘要/Abstract

摘要： 目的：研究细胞分裂周期蛋白42（CDC42）特异性抑制剂CASIN对食管癌细胞的活力、迁移能力，以及伪足形成的影响，并初步揭示其诱导细胞凋亡的分子作用机制。方法：根据CDC42蛋白表达情况将细胞分为4组，内源性CDC42高/低表达组和外源性CDC42高/低表达组。首先采用Western blot检测9种食管癌细胞系中内源性CDC42蛋白表达情况，接着选用CDC42内源性高表达和低表达食管癌细胞系各2种，采用细胞活力分析实验和划痕实验检测CDC42抑制剂CASIN对食管癌细胞活力和迁移能力的影响，Western blot检测CASIN对凋亡酶蛋白Caspase-3的表达及其底物蛋白PARP的裂解情况。然后将构建的Flag-CDC42和GFP-CDC42重组表达载体转染到食管癌细胞中，分成外源性CDC42低表达和高表达组，采用细胞活力分析实验和划痕实验检测CASIN对食管癌细胞活力和迁移能力的影响；免疫荧光实验检测CASIN对食管癌细胞伪足形成的影响。结果：与对照组（0 μmol/L的CASIN）比较，在内源性CDC42高/低表达的细胞组中，细胞活力分析和划痕实验发现，15~30 μmol/L的CASIN能明显抑制食管癌细胞的活力和迁移能力（P<0.05）；Western blot结果显示，CASIN能引起Caspase-3表达增加，裂解PARP增多，提示可能诱导了细胞凋亡的发生。与低表达CDC42的食管癌细胞对照组相比，外源性高表达CDC42的食管癌细胞在20 μmol/L CASIN作用时抑制程度显著增强（P<0.05）；免疫荧光实验结果显示CASIN对CDC42促进细胞的伪足形成具有抑制作用。结论：CASIN能使食管癌细胞活力和迁移能力明显下降，抑制细胞伪足形成，同时还可能诱导细胞发生凋亡。因此，CASIN有望被开发为一种新型的抗食管癌分子靶向药物。

关键词: 细胞分裂周期蛋白42, CASIN, 食管癌, 抑制作用

Abstract: OBJECTIVE: To investigate effects of CASIN,a specific cell division circle 42 (CDC42) inhibitor,on viability,migration and pseudopodia of esophageal cancer cells,and its mechanism of inducing apoptosis. METHODS: According to expression of the CDC42 protein,four groups were set up:endogenous CDC42 high/low expression groups and exogenous CDC42 high/low expression groups. Western blot was used to detect expression of endogenous CDC42 protein in 9 esophageal cancer cell lines. From the analyses,two endogenous high-expression and two low-expression esophageal cancer cell lines were selected. Cell viability assay was used to detect the effect of CASIN on cell viability. Wound healing assay was to detect cell invasion. Western blot was to detect protein expression levels of Caspases-3 and PARP. After constructed the Flag-CDC42 and GFP-CDC42 plasmid successfully,they were transfected into esophageal cancer cells to distinguish exogenous CDC42 high/low expression groups. Cell viability assay and wound healing assay were used as described above. Furthermore,the immunofluorescence experiment was used to detect effects of CASIN on pseudopodia formation. RESULTS: In the endogenous CDC42 high/low expression groups,our data show that CASIN at concentrations of 15-30 μmol/L significantly inhibited cell viability and cell invasion (P<0.05),and increased the expression of Caspase-3 and lyse PARP. The latter suggests that CASIN might induce cell apoptosis. Compared with the exogenous CDC42 low expression group,20 μmol/L CASIN significantly reduced expression in the exogenous CDC42 high expression group (P<0.05). In addition,data from the immunofluorescence experiments show that CASIN suppressed the effect of CDC42 in promoting the formation of pseudopodia. CONCLUSION: CASIN reduced the viability and migration of esophageal cancer cells,inhibit pseudopodia,and induce cell apoptosis in vitro. Therefore,CASIN can be developed further as a new targeted drug against esophageal cancer.

Key words: CDC42, CASIN, esophageal cancer, inhibition

中图分类号:

R735.1

陈婉娴, 王思梦, 陈张瑜, 王潘集, 郑淳文, 李恩民, 许丽艳. CDC42抑制剂CASIN对食管癌细胞活力与迁移能力的抑制作用[J]. 癌变·畸变·突变, 2021, 33(2): 136-142.

CHEN Wanxian, WANG Simeng, CHEN Zhangyu, WANG Panji, ZHENG Chunwen, LI Enmin, XU Liyan. CASIN, a CDC42 inhibitor, inhibited viability and migration of esophageal carcinoma cells in vitro[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2021, 33(2): 136-142.

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CDC42抑制剂CASIN对食管癌细胞活力与迁移能力的抑制作用

CASIN, a CDC42 inhibitor, inhibited viability and migration of esophageal carcinoma cells in vitro

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