二至丸对人结肠癌细胞的侵袭转移作用
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摘要
目的:探讨二至丸(Erzhiwan,EZW)对人属结肠癌细胞株HCT116侵袭及迁移的作用。方法:以人结肠癌细胞株HCT116为研究对象,以化疗药奥沙利铂(oxaliplatin,L-OHP)作为阳性对照,以20%,10%,5%EZW含药血清做为高、中、低剂量,运用Transwell法检测EZW及联用L-OHP后对HCT116细胞侵袭的影响,划痕实验观察细胞迁移情况,蛋白免疫印迹法(Western blot,WB)检测其对细胞血管内皮生长因子(vascular endothelial growth factor,VEGF),E-钙黏蛋白(E-cadherin)表达的影响。结果:(1)Transwell实验显示,与空白组比较,各用药组穿模细胞数均减少(P<0.05),以联用后穿模细胞数最少。(2)划痕实验显示,与空白组比较,各用药组均能抑制HCT116细胞的迁移(P<0.05)。12 h EZW高浓度组抑制率高于L-OHP组(P<0.05),其余时间段EZW各剂量组抑制率低于L-OHP组(P<0.05),各时间段联用组抑制率均高于EZW高、中、低剂量组(P<0.05),12 h联用组抑制率高于L-OHP组(P<0.05),48 h联用组抑制率低于L-OHP组(P<0.05)。(3)WB实验显示,与空白组比较,10%,20%EZW组,VEGF蛋白表达量均明显降低(P<0.05),联用组与L-OHP组相比蛋白表达量降低(P<0.05);与空白组相比,EZW含药血清各组降低了E-cadherin蛋白的表达(P<0.05),联用组和L-OHP组E-cadherin蛋白表达量均显著升高,且联用后E-cadherin蛋白表达量最高(P<0.05)。结论:二至丸能抑制结肠癌HCT116细胞侵袭转移,并以协同L-OHP后效果最佳,可能与其降低VEGF蛋白表达,协同L-OHP升高E-cadherin蛋白表达有关。
Objective:To investigate the effect of Erzhiwan(EZW) on the invasion and metastasis of human colon cancer HCT116 cells.Method:The abilities of invasion(number of transmembrane cells) and migration(relative width of 48 h scratch) were observed by Transwell assay.Western blot was used to detect vascular endothelial growth factor(VEGF) and E-cadherin protein,respectively.Result:(1) Transwell results showed that compared with the blank control group,the number of model piercing cells in each drug group was decreased(P<0.05),and the number of perforating model cells was the least after combined use of L-OHP.(2)Scratch test results showed that compared with the blank control group,the migration of HCT116 cells was inhibited in each treatment group(P<0.05).The inhibitory rate of high-dose EZW group was higher than that of L-OHP group at 12 h(P<0.05),and the inhibition rate of EZW in other time groups was lower than that of L-OHP group(P<0.05).The inhibition rate of combined treatment group was higher than that of EZW groups(P<0.05).The figure of combined group was higher than that of L-OHP group at 12 h(P<0.05),and combined group was lower than L-OHP group at 48 h(P<0.05).(3) Western blot results showed that compared with the control group,the expressions of VEGF protein in 10% and 20% EZW group were significantly decreased(P<0.05),and the expression of VEGF protein in 5.19 mg·L-1 L-OHP combined with 10% EZW group was lower than that in L-OHP group(P<0.05).Compared with the blank control group,EZW groups decreased the expression of E-cadherin protein(P<0.05).And 5.19 mg·L-1 L-OHP combined with 10% EZW group and L-OHP group increased the expression of E-cadherin protein significantly.Compared with L-OHP group,the expression of E-cadherin protein was the highest(P<0.05).Conclusion:EZW can inhibit the invasion and metastasis of colon cancer HCT116 cells,and the best effect is achieved after combined with L-OHP.This may be related to the decrease of VEGF protein expression and the increase of E-cadherin protein expression after combination with L-OHP.
Objective:To investigate the effect of Erzhiwan(EZW) on the invasion and metastasis of human colon cancer HCT116 cells.Method:The abilities of invasion(number of transmembrane cells) and migration(relative width of 48 h scratch) were observed by Transwell assay.Western blot was used to detect vascular endothelial growth factor(VEGF) and E-cadherin protein,respectively.Result:(1) Transwell results showed that compared with the blank control group,the number of model piercing cells in each drug group was decreased(P<0.05),and the number of perforating model cells was the least after combined use of L-OHP.(2)Scratch test results showed that compared with the blank control group,the migration of HCT116 cells was inhibited in each treatment group(P<0.05).The inhibitory rate of high-dose EZW group was higher than that of L-OHP group at 12 h(P<0.05),and the inhibition rate of EZW in other time groups was lower than that of L-OHP group(P<0.05).The inhibition rate of combined treatment group was higher than that of EZW groups(P<0.05).The figure of combined group was higher than that of L-OHP group at 12 h(P<0.05),and combined group was lower than L-OHP group at 48 h(P<0.05).(3) Western blot results showed that compared with the control group,the expressions of VEGF protein in 10% and 20% EZW group were significantly decreased(P<0.05),and the expression of VEGF protein in 5.19 mg·L-1 L-OHP combined with 10% EZW group was lower than that in L-OHP group(P<0.05).Compared with the blank control group,EZW groups decreased the expression of E-cadherin protein(P<0.05).And 5.19 mg·L-1 L-OHP combined with 10% EZW group and L-OHP group increased the expression of E-cadherin protein significantly.Compared with L-OHP group,the expression of E-cadherin protein was the highest(P<0.05).Conclusion:EZW can inhibit the invasion and metastasis of colon cancer HCT116 cells,and the best effect is achieved after combined with L-OHP.This may be related to the decrease of VEGF protein expression and the increase of E-cadherin protein expression after combination with L-OHP.
引文
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[19]刘海晔.中药逆转肿瘤多药耐药性的研究进展[J].中草药,2015,46(7):1096-1102.
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[21]朱文君,戴建国.结肠癌的中医药治疗研究进展[J].吉林中医药,2013,33(5):535-537.
[22]王馨,刘端勇,赵海梅,等.二至丸临床应用综述[J].江西中医药,2015,46(4):67-70.
[23]马莉.VEGF及其受体的生物学特性及在肿瘤血管生成中的作用[J].中国优生与遗传杂志,2016,24(5):146-148.
[24]张贵彪,朱元章,朱国福.软坚散结颗粒对人胃腺癌SGC-7901细胞EMT的抑制作用及相关机制[J].中国实验方剂学杂志,2017,23(18):112-117.
[25]孙琳,张文静,吴靖芳.E-钙黏素、β-连环蛋白与肿瘤关系的研究进展[J].中国老年学杂志,2017,37(11):2832-2834.
[2]Connell L C,Mota J M,Braghiroli M I,et al.The rising incidence of younger patients with colorecta cancer:questions about screening,biology,and treatment[J].Curr Treat Options Oncol,2017,18(4):23.
[3]伊日贵,徐晓艳,李时荣.肿瘤侵袭转移机制研究进展[J].中华实用诊断与治疗杂志,2014,28(10):937-939.
[4]段泽星,谢立群.VEGF在肿瘤生长和血管生成中的作用[J].世界华人消化杂志,2010,18(27):2894-2900.
[5]David J M,Rajasekaran A K.Dishonorable discharge:the oncogenic roles of cleaved E-cadherin fragments[J].Cancer Res,2012,72(12):2917-2923.
[6]闫冰,蔡秀江,姚卫峰,等.二至丸保肝活性成分群对CCl4致小鼠急性肝损伤保护作用研究[J].中国中药杂志,2012,37(9):1303-1306.
[7]赵海梅,周步高,王馨,等.二至丸预防和治疗性给药对大鼠损伤后肝细胞再生障碍的保护作用[J].中国实验方剂学杂志,2017,23(16):128-132.
[8]胡科.二至丸对骨质疏松性股骨颈骨折术后愈合及髋关节功能的影响[J].现代中西医结合杂志,2017,26(4):384-386.
[9]赵益,罗蓉,尚广彬,等.二至汤含药血清对肿瘤细胞抑制作用的研究[J].新中医,2013,45(4):156-159.
[10]郑里翔,张玉仁,邓科穗,等.二至汤对阴虚阳虚荷瘤小鼠肿瘤生长的影响[J].陕西中医,2008,29(12):1678-1680.
[11]尚广彬,曾莉萍,赵益,等.二至丸对诱发性乳腺癌组织中VEGF和MMP-9表达的影响[J].中国实验方剂学杂志,2013,19(13):270-273.
[12]王国娟,余文燕.大肠癌中医证型规律研究[J].中华中医药杂志,2016,31(3):837-840.
[13]余文燕,王国娟,王桦影,等.二至丸对结肠癌细胞增殖及凋亡作用的实验研究[J].中药药理与临床,2018,34(4):11-15.
[14]国家药典委员会.中华人民共和国药典.一部[M].北京:中国医药科技出版社,2010:861-862.
[15]黄继汉,黄晓晖,陈志扬,等.药理试验中动物间和动物与人体间的等效剂量换算[J].中国临床药理学与治疗学,2004,9(9):1069-1072.
[16]张军平,张伯礼,山本清高.中药药物血清的制作方法探讨[J].天津中医药,2004,21(4):274-276.
[17]Hato S V,Khong A,de Vries I J,et al.Molecular pathways:the immunogenic effects of platinum-based chemotherapeutics[J].Clin Cancer Res,2014,20(11):2831-2837.
[18]王浩,朱莹杰.中医药防治奥沙利铂所致周围神经毒性研究进展[J].解放军医药杂志,2015,27(4):76-80.
[19]刘海晔.中药逆转肿瘤多药耐药性的研究进展[J].中草药,2015,46(7):1096-1102.
[20]YE L,JIA Y,JI K E,et al.Traditional Chinese medicine in the prevention and treatment of cancer and cancer metastasis[J].Oncol Lett,2015,10(3):1240-1250.
[21]朱文君,戴建国.结肠癌的中医药治疗研究进展[J].吉林中医药,2013,33(5):535-537.
[22]王馨,刘端勇,赵海梅,等.二至丸临床应用综述[J].江西中医药,2015,46(4):67-70.
[23]马莉.VEGF及其受体的生物学特性及在肿瘤血管生成中的作用[J].中国优生与遗传杂志,2016,24(5):146-148.
[24]张贵彪,朱元章,朱国福.软坚散结颗粒对人胃腺癌SGC-7901细胞EMT的抑制作用及相关机制[J].中国实验方剂学杂志,2017,23(18):112-117.
[25]孙琳,张文静,吴靖芳.E-钙黏素、β-连环蛋白与肿瘤关系的研究进展[J].中国老年学杂志,2017,37(11):2832-2834.