基于Wnt/β-catenin信号通路探讨复方土贝母对裸鼠乳腺癌移植瘤的影响
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摘要
[目的]研究复方土贝母对裸鼠乳腺癌移植瘤生长以及对Wnt/β-catenin信号通路相关蛋白的影响。[方法]40只4～5周龄雌性BALB/C裸鼠皮下接种1×10~7/mL MCF-7细胞悬液0.2mL/只,造模后随机分为荷瘤对照组、复方土贝母低、中、高剂量组。复方土贝母各剂量组分别以0.5、1.0、2.0g·m L~(-1)的复方土贝母连续灌胃30d,对照组予0.9%氯化钠溶液灌胃,灌胃容积均为0.6mL/20g。所有小鼠第30天处死,测量各组移植瘤体积,并计算抑瘤率。实时荧光定量PCR检测β-连环蛋白(β-catenin)、E-钙粘蛋白(E-cadherin)、波形蛋白(Vimentin)mRNA表达水平,Western blot和免疫组化检测β-catenin、E-cadherin、Vimentin蛋白表达水平。[结果]复方土贝母低、中、高剂量组抑瘤率分别为10.56%、37.06%、33.17%,中、高剂量组移植瘤体积小于荷瘤对照组,差异有统计学意义(P<0.05),复方土贝母各剂量组的β-catenin、Vimentin m RNA表达低于荷瘤对照组,E-cadherin mRNA表达高于荷瘤对照组,差异有统计学意义(P<0.05)。中、高剂量组β-catenin、Vimentin蛋白表达低于荷瘤对照组,E-cadherin蛋白表达高于荷瘤对照组,差异有统计学意义(P<0.05)。[结论]复方土贝母能抑制乳腺癌移植瘤的生长,可能与抑制肿瘤中β-catenin、Vimentin的表达,上调E-cadherin表达有关。
[Objective] To study the inhibitory effect of compound Tubeimu on MCF-7 xenografts and the effects of Wnt/β-catenin signaling pathway-related proteins. [Methods] Forty female BALB/C nude mice aged 4 to 5 weeks were subcutaneously inoculated with 1 ×10~7/mL MCF-7 cell suspension0.2 mL each. After modeling, they were randomly divided into tumor control group, compound Tubeimu low, medium and high dose group. The drug-administered group was continuously administered with 0.5 g·mL~(-1), 1.0 g·mL~(-1), 2.0 g·mL~(-1) compound Tubeimu for 30 days, and tumor control group was given normal saline, and the gastric volume was 0.6 mL/20 g. All mice were sacrificed on the 30 th day, the tumor volume of each group was measured, and the tumor inhibition rate was calculated. The expression of β-catenin, E-cadherin, Vimentin m RNA was detected by fluorescent quantitative PCR, and protein expression was detected by Western blot and immunohistochemistry.[Results] The tumor inhibition rates of compound Tubeimu low, medium and high dose groups were 10.56%, 37.06% and 33.17%, respectively. The tumor volume of medium and high dose groups was significantly different from that of tumor control group(P<0.05). The mRNA expression of β-catenin, Vimentin in compound Tubeimu groups was lower than tumor control group(P<0.05),while mRNA expression of E-cadherin in compound Tubeimu groups was higher than tumor control group( P<0.05). The protein expression of β-catenin,Vimentin in compound Tubeimu medium and high groups was lower than tumor control group(P <0.05), while protein expression of E-cadherin in compound Tubeimu medium and high groups was higher than tumor control group(P <0.05). [Conclusion]Compound Tubeimu can inhibit the growth of breast cancer xenografts, which may be related to inhibit the expression of β-catenin and Vimentin, and upregulate the expression of E-cadherin.
[Objective] To study the inhibitory effect of compound Tubeimu on MCF-7 xenografts and the effects of Wnt/β-catenin signaling pathway-related proteins. [Methods] Forty female BALB/C nude mice aged 4 to 5 weeks were subcutaneously inoculated with 1 ×10~7/mL MCF-7 cell suspension0.2 mL each. After modeling, they were randomly divided into tumor control group, compound Tubeimu low, medium and high dose group. The drug-administered group was continuously administered with 0.5 g·mL~(-1), 1.0 g·mL~(-1), 2.0 g·mL~(-1) compound Tubeimu for 30 days, and tumor control group was given normal saline, and the gastric volume was 0.6 mL/20 g. All mice were sacrificed on the 30 th day, the tumor volume of each group was measured, and the tumor inhibition rate was calculated. The expression of β-catenin, E-cadherin, Vimentin m RNA was detected by fluorescent quantitative PCR, and protein expression was detected by Western blot and immunohistochemistry.[Results] The tumor inhibition rates of compound Tubeimu low, medium and high dose groups were 10.56%, 37.06% and 33.17%, respectively. The tumor volume of medium and high dose groups was significantly different from that of tumor control group(P<0.05). The mRNA expression of β-catenin, Vimentin in compound Tubeimu groups was lower than tumor control group(P<0.05),while mRNA expression of E-cadherin in compound Tubeimu groups was higher than tumor control group( P<0.05). The protein expression of β-catenin,Vimentin in compound Tubeimu medium and high groups was lower than tumor control group(P <0.05), while protein expression of E-cadherin in compound Tubeimu medium and high groups was higher than tumor control group(P <0.05). [Conclusion]Compound Tubeimu can inhibit the growth of breast cancer xenografts, which may be related to inhibit the expression of β-catenin and Vimentin, and upregulate the expression of E-cadherin.
引文
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[18]张风波,李玲丽,江欣星,等.上皮间质转化在干细胞诱导分化及肿瘤发生中的作用[J].中国组织工程研究,2017,21(17):2753-2758.ZHANG Fengbo,LI Lingli,JIANG Xinxing,et al.Role of epithelial-mesenchymal transition in stem cell differentiation and tumorigenesis[J].J Clin Rehabil Tis Eng Res,2017,21(17):2753-2758.
[19]王雪玮,王昕,高纪东.肿瘤细胞上皮间质转化与肿瘤转移及治疗耐药机制关系的研究进展[J].中国肿瘤临床与康复,2017,24(6):765-768.WANG Xuewei,WANG Xin,GAO Jidong.Advances in research on relationship between tumor cell epithelialmesenchymal transition and tumor metastasis and treatment of drug resistance[J].Chinese Journal of Clinical Oncology and Rehabilitation,2017,24(6):765-768.
[20]许艳艳,李慧杰,李秀荣.上皮-间质转化分子机制及中医药干预的研究[J].山东中医药大学学报,2014,38(1):76-79.XU Yanyan,LI Huijie,LI Xiurong.Molecular mechanism of epithelial-mesenchymal transition and intervention of traditional Chinese medicine[J].Journal of Shandong University of Traditional Chinese Medicine,2014,38(1):76-79.
[21]燕慧,王捷.上皮间质转化与肿瘤转移的研究进展[J].中华肿瘤防治杂志,2010,17(4):311-314.YAN Hui,WANG Jie.Epithelial-mesenchymal transitions and tumor metastasis[J].Chinese Journal of Cancer Prevention and Treatment,2010,17(4):311-314.
[22]林晓芳,姚新苗,李威,等.益骨汤对去势大鼠骨组织Wnt/β-catenin经典信号通路的影响[J].浙江中医药大学学报,2018,42(2):97-104,110.LIN Xiaofang,YAO Xinmiao,LI Wei,et al.Effect of Yigu Decoction on Wnt/β-catenin signaling pathway in bone tissue of ovariectomized rats[J].Journal of Zhejiang Chinese Medical University,2018,42(2):97-104,110.
[2]周际昌.实用肿瘤内科治疗[M].2版.北京:北京科学技术出版社,2016:270-271.ZHOU Jichang.Practice of Medical Oncology Treatment[M].2nd edition.Beijing:Beijing Science and Technology Publishing House,2016:270-271.
[3]Clevers H,Loh K M,Nusse R.Stem cell signaling.An integral program for tissue renewal and regeneration:Wnt signaling and stem cell control[J].Science,2014,346(6205):1248012.
[4]Janssen R,Posnien N.Identification and embryonic expression of Wnt2,Wnt4,Wnt5,and Wnt9,in the millipede glomeris marginata(Myriapoda:Diplopoda)[J].Gene Expr Patterns,2014,14(2):55-61.
[5]Sun Y,Zhu Q,Zhou M,et al.Restoration of miRNA-148a in pancreatic cancer reduces invasion and metastasis by inhibiting the Wnt/β-catenin signaling pathway via downregulating maternally expressed gene-3[J].Exp Ther Med,2019,17(1):639-648.
[6]Zhan T,Rindtorff N,Boutros M.Wnt signaling in cancer[J].Oncogene,2017,36(11):1461-1473.
[7]崔雄,韩龙海.人乳腺癌组织中Wnt2基因的表达及靶向抑制基因表达对乳腺癌细胞MCF-7增殖、迁移和侵袭力的影响[J].实用癌症杂志,2018,33(7):1053-1056.CUI Xiong,HAN Longhai.Expression of Wnt2 gene in human breast cancer tissue and effects of targeted suppression gene expression on proliferation,migration and invasion of breast cancer cell line MCF-7[J].The Practical Journal of Cancer,2018,33(7):1053-1056.
[8]吴秉纯.医学动物实验基础及基本技术方法[M].哈尔滨:黑龙江人民出版社,2008:184.WU Bingchun.Basic Techniques and Methods of Medical Animal Experiment[M].Harbin:Heilongjiang People’s Publishing House,2008:184.
[9]Naito S,Von Eschenbach A C,Giavazzi R,et al.Growth and metastasis of tumor cells isolated from a human renal cell carcinoma implanted into different organs of nude mice[J].Cancer Res,1986,46(8):4109-4115.
[10]巢元方.诸病源候论(宋刊本)[M].北京:北京科学技术出版社,2016:314-315.CHAO Yuanfang.Source of Disease(Song edition)[M].Beijing:Beijing Science and Technology Publishing House,2016:314-315.
[11]陈实功.外科正宗[M].北京:中国医药科技出版社,2011:152-153.CHEN Shigong.Surgical Authentic[M].Beijing:China Medical Science Press,2011:152-153.
[12]朱海鸥,朱晓薇,侯文彬,等.中药土贝母的现代研究进展[J].时珍国医国药,2008,19(12):2985-2987.ZHU Haiou,ZHU Xiaowei,HOU Wenbin,et al.New research progress of Bolbostemma paniculatum[J].Lishizhen Medicine and Materia Medica Research,2008,19(12):2985-2987.
[13]刘加军,张俊峰,林东军,等.冬凌草甲素对Nalm-6细胞的增殖抑制作用及其作用机制[J].中华肿瘤防治杂志,2009,16(13):966-969.LIU Jiajun,ZHANG Junfeng,LIN Dongjun,et al.Antiproliferation effect of oridonin on Nalm-6 cells and its mechanisms of action[J].Chinese Journal of Cancer Prevention and Treatment,2009,16(13):966-969.
[14]岳静,陈如意,洪姣,等.基于Wnt信号通路抑制的冬凌草甲素抗乳腺癌研究[J].浙江中医药大学学报,2014,38(11):1315-1321.YUE Jing,CHEN Ruyi,HONG Jiao,et al.Study on oridonin for antitumor based on restraining Wnt signal pathway in breast cancer MCF-7 cells[J].Journal of Zhejiang Chinese Medical University,2014,38(11):1315-1321.
[15]邵明坤,范青.苦参碱抗肿瘤作用研究的新进展[J].实用药物与临床,2014,17(6):756-759.SHAO Mingkun,FAN Qing.New progress of antitumor research on matrine[J].Practical Pharmacy and Clinical Remedies,2014,17(6):756-759.
[16]季于彬.抗癌中药药理与应用[M].哈尔滨:黑龙江科技出版社,1999.JI Yubin.Pharmacological Action and Application of Anticancer Traditional Chinese Medicine[M].Harbin:Heilongjiang Science and Technology Press,1999.
[17]孙丹,辛彦.上皮间质转化与肿瘤侵袭转移关系的研究进展[J].现代肿瘤医学,2011,19(10):2088-2091.SUN Dan,XIN Yan.The research progress of the relation between epithelial-mesenchymal transition and tumor invasion or metastasis[J].J Mod Oncol,2011,19(10):2088-2091.
[18]张风波,李玲丽,江欣星,等.上皮间质转化在干细胞诱导分化及肿瘤发生中的作用[J].中国组织工程研究,2017,21(17):2753-2758.ZHANG Fengbo,LI Lingli,JIANG Xinxing,et al.Role of epithelial-mesenchymal transition in stem cell differentiation and tumorigenesis[J].J Clin Rehabil Tis Eng Res,2017,21(17):2753-2758.
[19]王雪玮,王昕,高纪东.肿瘤细胞上皮间质转化与肿瘤转移及治疗耐药机制关系的研究进展[J].中国肿瘤临床与康复,2017,24(6):765-768.WANG Xuewei,WANG Xin,GAO Jidong.Advances in research on relationship between tumor cell epithelialmesenchymal transition and tumor metastasis and treatment of drug resistance[J].Chinese Journal of Clinical Oncology and Rehabilitation,2017,24(6):765-768.
[20]许艳艳,李慧杰,李秀荣.上皮-间质转化分子机制及中医药干预的研究[J].山东中医药大学学报,2014,38(1):76-79.XU Yanyan,LI Huijie,LI Xiurong.Molecular mechanism of epithelial-mesenchymal transition and intervention of traditional Chinese medicine[J].Journal of Shandong University of Traditional Chinese Medicine,2014,38(1):76-79.
[21]燕慧,王捷.上皮间质转化与肿瘤转移的研究进展[J].中华肿瘤防治杂志,2010,17(4):311-314.YAN Hui,WANG Jie.Epithelial-mesenchymal transitions and tumor metastasis[J].Chinese Journal of Cancer Prevention and Treatment,2010,17(4):311-314.
[22]林晓芳,姚新苗,李威,等.益骨汤对去势大鼠骨组织Wnt/β-catenin经典信号通路的影响[J].浙江中医药大学学报,2018,42(2):97-104,110.LIN Xiaofang,YAO Xinmiao,LI Wei,et al.Effect of Yigu Decoction on Wnt/β-catenin signaling pathway in bone tissue of ovariectomized rats[J].Journal of Zhejiang Chinese Medical University,2018,42(2):97-104,110.