华蟾酥毒基通过调控基质金属蛋白酶抑制食管癌细胞侵袭转移的作用  

Inhibition effect of cinobufagin on invasion and metastasis of esophageal cancer cells by mediating matrix metalloproteinases

作　　者：邓旭 盛杰霞 王佳宝 代二庆[2] 邓全军 DENG Xu;SHENG Jie-xia;WANG Jia-bao;DAI Er-qing;DENG Quan-jun(The Postgraduate Culture Base of Jinzhou Medical University,The Affiliated Hospital of Logistics University of Chinese People's Armed Police Force,Tianjin 300162,China;The Affiliated Hospital of Logistics University of Chinese People's Armed Police Force,Tianjin 300162,China;Department of Military Health Care Center,The Affiliated Hospital of Logistics University of Chinese People's Armed Police Force, Tianjin 300162,China)

机构地区：[1]锦州医科大学中国人民武装警察部队后勤学院附属医院研究生培养基地,天津300162 [2]中国人民武装警察部队后勤学院附属医院军人医疗保健中心,天津300162 [3]中国人民武装警察部队后勤学院附属医院消化内科,天津300162

出　　处：《中国临床药理学杂志》2019年第2期133-136,共4页The Chinese Journal of Clinical Pharmacology

基　　金：国家自然科学基金资助项目(81273745);武警后勤学院基础研究项目面上基金资助项目(WHJ2016022)

摘　　要：目的研究华蟾酥毒基对食管癌细胞侵袭转移的影响。方法分别将食管癌EC-109和Kyse-150细胞分为4组:对照组和3个浓度实验组。对照组加入含有溶媒(0. 1%DMSO)的培养基,低、中、高3个浓度实验组给予25,50,100 nmol·L^(-1)华蟾酥毒基,处理1 d。用划痕实验和Transwell小室法检测各组细胞迁移水平和侵袭能力,用免疫印迹实验检测细胞中磷酸化黏着斑激酶Tyr397[p-FAK(Tyr397)]、P53、人第10号染色体缺失的磷酸酶及张力蛋白同源基因(PTEN)、基质金属蛋白酶2(MMP-2)和基质金属蛋白酶9(MMP-9)的蛋白表达情况,用实时荧光定量PCR检测各组细胞中上述指标mRNA的相对表达水平。结果 EC-109细胞:以25～100 nmol·L^(-1)的华蟾酥毒基处理1 d后,实验组划痕愈合面积约为(9. 96±1. 17)%～(5. 02±0. 92)%,侵袭细胞数约为(162. 33±12. 01)～(42. 33±12. 50)个;与对照组的(23. 80±1. 32)%和(254. 33±14. 01)个相比,差异均有统计学意义(均P <0. 01)。低、中、高3个浓度实验组的MMP-2蛋白表达水平分别为0. 74±0. 04,0. 54±0. 06和0. 44±0. 05。中、高2个浓度实验组的MMP-9蛋白表达分别为0. 69±0. 08,0. 52±0. 04;这2组的p-FAK (Tyr397)的表达水平分别为0. 90±0. 08,0. 62±0. 06;这2组的P53蛋白表达水平分别为1. 20±0. 06,2. 86±0. 05;这2组的PTEN蛋白表达水平分别为1. 31±0. 06,1. 53±0. 05,与对照组的(1. 00±0. 00)比较,差异均有统计学意义(均P <0. 01);在同样的处理条件下,实验组上述指标的mRNA的表达变化与蛋白表达趋势相同。Kyse-150细胞:迁移和侵袭能力的改变以及各指标的表达变化与EC-109细胞趋势相同。结论华蟾酥毒基可能通过上调P53和PTEN表达,下调FAK的磷酸化及其下游分子MMP-2和MMP-9的表达,进而抑制食管癌细胞的迁移和侵袭。Objective To observe the effect of cinobufagin on the invasion and metastasis of esophageal cancer cells. Methods EC-109 and Kyse-150 cells were divided into 4 groups: control group and three concentration experimental groups,control group was treated with medium containing vehicle( 0. 1% DMSO),and the experimental groups were treated with low,middle and high concentrations( 25,50,100 nmol· L-1) of cinobufagin for 1 d. The wound healing and Transwell chamber assay were used to detect the migration and invasion of each group cells. The protein expression levels of some important molecules were assessed by Western blot,such as phosphorylated focal adhesionkinase Tyr397 [p-FAK( Tyr397) ],P53,human chromosome 10 deleted phosphatase and of tensin homolog( PTEN),matrix metalloproteinase 2( MMP-2) and matrix metalloproteinase 9( MMP-9). The relative mRNA expressions of above molecules in each group cells were measured using real-time quantitative PCR. Results EC-109 cells: after 1 d treatment with 25-100 nmol·L-1 of cinobufagin,the scratch healing area of each experimental group approximately were( 9. 96 ± 1. 17) %-( 5. 02 ± 0. 92) %, and the number of invasive cells were( 162. 33 ±12. 01)-( 42. 33 ± 12. 50),compared with the control group( 23. 80 ± 1. 32) % and 254. 33 ± 14. 01,the differences were statistically significant( all P < 0. 01). The MMP-2 protein expression in the low,middle and high concentrations experimental groups were 0. 74 ± 0. 04,0. 54 ± 0. 06,0. 44 ± 0. 05 respectively;the MMP-9 protein expression in middle and high concentrations experimental groups were 0. 69 ± 0. 08,0. 52 ± 0. 04;the p-FAK( Tyr397) expression in the two groups were 0. 90 ± 0. 08,0. 62 ± 0. 06;P53 protein expression in the two groups were1. 20 ± 0. 06,2. 86 ± 0. 05;PTEN expression in the two groups were 1. 31 ± 0. 06,1. 53 ± 0. 05. The protein expression levels in the control group was all( 1. 00 ± 0. 00). Comparison between two concentrations experimental groups and control group,the difference of the protein

关 键 词：华蟾酥毒基 食管癌 侵袭 迁移 基质金属蛋白酶 黏着斑激酶 

分 类 号：R979[医药卫生—药品]