柠檬苦素抑制胃癌细胞转移的作用及机制研究  

Inhibition of STAT3 Signaling Pathway by Limonin Suppressed Tumor Metastasis of Gastric Cancer Cells

作　　者：刘伯霞 申琴 焦海燕[1,2] 李涛[3] 陈静[1,2] LIU Boxia;SHEN Qin;JIAO Haiyan;LI Tao;CHEN Jing(School of Basic Medicine,Ningxia Medical University,Yinchuan 750004,China;Key Laboratory of Reproduction and Genetics in Ningxia,Yinchuan 750004,China;Tumor Surgical Department,the General Hospital of Ningxia Medical University,Yinchuan 750004,China)

机构地区：[1]宁夏医科大学基础医学院,银川750004 [2]宁夏回族自治区生育力保持教育部重点实验室,银川750004 [3]宁夏医科大学总医院肿瘤外科,银川750004

出　　处：《宁夏医科大学学报》2020年第4期340-345,362,共7页Journal of Ningxia Medical University

基　　金：国家自然科学基金(816660500,81560485,81260339);宁夏基础医学“西部一流”学科人才培养项目(NXYLXK2017B07,NXYLXK2017A05);宁夏自然科学基金(NZ16142)。

摘　　要：目的探究柠檬苦素(Limonin)抑制胃癌细胞转移的作用及机制。方法首先,对胃癌细胞(BGC-823、SGC-7901、AGS、MGC-803)进行体外培养。随后,利用细胞增殖实验(MTS)检测柠檬苦素(0、180、260、340、420、500μmol·L-1)对胃癌细胞增殖的影响;利用划痕愈合实验研究柠檬苦素(0、40、60μmol·L-1)对胃癌细胞迁移能力的影响。进而建立裸鼠胃癌细胞BGC-823腹腔种植转移模型,给予柠檬苦素[0、12.5、25 mg·(kg·d)-1]腹腔注射治疗28 d后,检测其对胃癌转移的抑制作用。最后,利用Western blot初步研究柠檬苦素与信号转导和转录因子(STAT3)信号通路之间的关系。结果 MTS检测结果显示,柠檬苦素在500μmol·L-1浓度下对多种胃癌细胞增殖的半数抑制率不足50%。但划痕愈合实验发现,在40μmol·L-1浓度下可抑制多种胃癌细胞(BGC-823、SGC-7901、AGS、MGC-803)的迁移。体内实验显示,给药处理21 d,25 mg·kg-1柠檬苦素能预防并抑制裸鼠胃癌移植瘤转移灶的数量,且在该剂量下对小鼠体质量影响不明显。上述结果表明:柠檬苦素剂量依赖性地抑制小鼠肿瘤组织中的STAT3磷酸化和MMP-9蛋白的表达,但对增殖标记物Ki-67的抑制效果不明显。通过免疫组化及Western blot发现,该单体化合物能降低BGC-823细胞STAT3酪氨酸705位点的磷酸化(p Tyr705-STAT3)。结论柠檬苦素能有效预防并抑制胃癌细胞的转移,其作用机制与抑制STAT3的活化有关。Objective To investigate the role and mechanism of limonin in inhibiting the metastasis of gastric cancer cells.Methods First,gastric cancer cells(BGC-823,SGC-7901,AGS,MGC-803)were cultured in vitro.Subsequently,the effect of limonin(0,180,260,340,420,500μmol·L^-1)on the proliferation of gastric cancer cells was examined using cell proliferation assay(MTS).The effect of limonin(0,40,60μmol·L-1)on the migration ability of gastric cancer cells was studied using scratch healing experiments.After four weeks intraperitoneal injection of limonin[0,12.5,25mg·(kg·d)^-1],the growth and metastasis of gastric cancer were detected.In addition,the relationship between limonin and the signal transducer and activator of transcription 3(STAT3)signaling pathway was studied in the gastric cancer cell line BGC-823 by western blot.Results MTS test results showed that the half-inhibition rate of limonin on the proliferation of various gastric cancer cells was less than 50%at the concentration of 500μmol·L^-1.However,limonoids could significantly inhibit the migration of various gastric cancer cells(BGC-823,SGC-7901,AGS,MGC-803)at the concentration of 40μmol·L^-1 in the scratch healing experiment.In vivo experiments,it was found that intra-peritoneal injection of limonin with doses of 25 mg·kg^-1 for 21 d could significantly inhibit the numbers of transplanted gastric cancer metastases in nude mice,and had no significant effect on the body weight of nude mice.Furthermore,both immunohistochemistry and western blot assays were applied to clarify the potential mechanism of limonin on tumor metastasis inhibition.It was found that limonin inhibited the phosphorylation of STAT3 and the expression of MMP-9 protein in tumor tissues of mice in a dose-dependent manner.However,the inhibitory effect of limonin on the proliferation marker Ki-67 was not obvious.Additionally,it was confirmed that the compound could decrease the phosphorylation of STAT3 tyrosine 705 site(pTyr705-STAT3)in BGC-823 cells by western blot assay.Conclusion Limon

关 键 词：柠檬苦素 胃癌 转移 信号转导和转录因子 

分 类 号：R735.2[医药卫生—肿瘤]