水杨酸影响人胃癌细胞系体外侵袭转移的实验研究
发布时间：2019-01-08 10:16
【摘要】：目的研究不同浓度水杨酸处理对胃癌细胞增殖和侵袭转移能力的影响,并初步探讨水杨酸抗肿瘤转移的可能机制。方法采用一系列浓度梯度(0.5~3mmol/L)的水杨酸处理胃癌细胞株MGC-803,MTS比色法观察不同浓度水杨酸处理24 h后细胞增殖能力的变化,筛选出最适的水杨酸处理浓度;细胞黏附实验观察0.5mmol/L和1mmol/L水杨酸处理后细胞黏附能力的变化;Transwell观察0.5mmol/L和1mmol/L水杨酸处理后细胞迁移和侵袭的能力变化;荧光定量PCR检测0.5mmol/L和1mmol/L水杨酸处理后细胞黏附分子CD44、E钙粘素(CDH1)、连环蛋白A1(CTNNA1)、纤连蛋白(FN1)、核纤层蛋白(Lamin)、核纤层蛋白B1(Lamin B1)、核纤层蛋白C1(Lamin C1)和整合素A9(ITGA9)m RNA表达水平的变化;Western-blot观察细胞黏附分子CD44、CTNNA1、FN1以及Lamin B1蛋白表达水平的差异。结果MTS结果示0.5mmol/L和1mmol/L的水杨酸处理24h后细胞抑制率分别为(32±4.5)%和(38±4.1)%,低于50%,因此选取0.5mmol/L和1mmol/L的水杨酸作为后续实验的研究。细胞黏附实验显示相比于对照组的细胞粘附数量(121.00±2.65)个,0.5mmol/L组和1mmol/L组水杨酸处理后黏附的细胞数量明显减少,分别是(88.00±1.53)个和(44.00±2.00)个;细胞迁移实验示0.5mmol/L和1mmol/L组水杨酸处理后细胞迁移率明显受到抑制,分别是(10.00±3.85)%和(12.00±3.61)%;细胞侵袭的结果示0.5mmol/L和1mmol/L水杨酸处理后,细胞OD值分别是(0.18±0.00)和(0.15±0.00),较正常组OD值(0.22±0.01)低。荧光定量PCR表明0.5mmol/L组和1mmol/L组水杨酸处理明显下调细胞黏附分子CD44、CTNNA1、FN1、ITGA9、Lamin、Lamin B1和Lamin C1 m RNA水平的转录,上调CDH1 m RNA的表达;Western-blot结果同样证实0.5mmol/L和1mmol/L组水杨酸处理后细胞黏附分子CD44、CTNNA1、FN1和Lamin B1蛋白的表达水平明显降低(P0.05)。结论1.0.5mmol/L和1mmol/L水杨酸可明显抑制胃癌细胞株的体外增殖、黏附、迁移和侵袭,并且其抑制效应随着浓度的增加而逐渐增强。2.0.5mmol/L和1mmol/L水杨酸下调细胞黏附分子CD44、CTNNA1、FN1、ITGA9、Lamin、Lamin B1和Lamin C1的表达和上调CDH1的表达,其抗肿瘤转移的机制可能与抑制细胞黏附分子的表达和抑制上皮间质转化相关。
[Abstract]:Objective to study the effects of salicylic acid treatment on the proliferation, invasion and metastasis of gastric cancer cells, and to explore the possible mechanism of salicylic acid against tumor metastasis. Methods A series of concentration gradient (0.5~3mmol/L) salicylic acid treated gastric cancer cell line MGC-803,MTS colorimetry was used to observe the change of cell proliferation ability after 24 h treatment with different concentrations of salicylic acid, and the optimal concentration of salicylic acid was selected. Cell adhesion assay was used to observe the changes of cell adhesion ability after 0.5mmol/L and 1mmol/L salicylic acid treatment and Transwell to observe the ability of cell migration and invasion after 0.5mmol/L and 1mmol/L salicylic acid treatment. 0.5mmol/L and 1mmol/L salicylic acid treated cell adhesion molecule CD44,E cadherin (CDH1), catenin A1 (CTNNA1), fibronectin (FN1) and nuclear laminin (Lamin), (Lamin B1) were detected by fluorescence quantitative PCR. Changes of expression of nuclear laminin C1 (Lamin C1) and integrin A9 (ITGA9) m RNA); The expression levels of CD44,CTNNA1,FN1 and Lamin B1 were observed by Western-blot. Results MTS results showed that the inhibitory rates of 0.5mmol/L and 1mmol/L were (32 卤4.5)% and (38 卤4.1)% after 24 h treatment, respectively, which were lower than 50%. Therefore, the salicylic acid of 0.5mmol/L and 1mmol/L was selected as the follow-up study. Cell adhesion assay showed that compared with the control group (121.00 卤2.65), the number of cell adhesion in 0.5mmol/L group and 1mmol/L group was significantly decreased after salicylic acid treatment. They were (88.00 卤1.53) and (44.00 卤2.00); Cell migration assay showed that the cell migration rate of 0.5mmol/L and 1mmol/L groups was (10.00 卤3.85)% and (12.00 卤3.61)%, respectively. The results of cell invasion showed that the OD values of cells treated with 0.5mmol/L and 1mmol/L salicylic acid were (0.18 卤0.00) and (0.15 卤0.00), respectively, which were lower than those of normal group (0.22 卤0.01). Fluorescence quantitative PCR showed that salicylic acid treatment in 0.5mmol/L group and 1mmol/L group significantly down-regulated the transcription of cell adhesion molecules CD44,CTNNA1,FN1,ITGA9,Lamin,Lamin B1 and Lamin C 1m RNA, and up-regulated the expression of CDH1 m RNA. Western-blot results also confirmed that the expression levels of CD44,CTNNA1,FN1 and Lamin B1 were significantly decreased in 0.5mmol/L and 1mmol/L groups after salicylic acid treatment (P0.05). Conclusion 1.0.5mmol/L and 1mmol/L salicylic acid can significantly inhibit the proliferation, adhesion, migration and invasion of gastric cancer cell lines in vitro. 2.0.5mmol/L and 1mmol/L salicylic acid down-regulated the expression of CD44,CTNNA1,FN1,ITGA9,Lamin,Lamin B1 and Lamin C1 and up-regulated the expression of CDH1. The mechanism of its anti-tumor metastasis may be related to the inhibition of the expression of cell adhesion molecules and the inhibition of epithelial mesenchymal transformation.
【学位授予单位】：宁波大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R735.2
[Abstract]:Objective to study the effects of salicylic acid treatment on the proliferation, invasion and metastasis of gastric cancer cells, and to explore the possible mechanism of salicylic acid against tumor metastasis. Methods A series of concentration gradient (0.5~3mmol/L) salicylic acid treated gastric cancer cell line MGC-803,MTS colorimetry was used to observe the change of cell proliferation ability after 24 h treatment with different concentrations of salicylic acid, and the optimal concentration of salicylic acid was selected. Cell adhesion assay was used to observe the changes of cell adhesion ability after 0.5mmol/L and 1mmol/L salicylic acid treatment and Transwell to observe the ability of cell migration and invasion after 0.5mmol/L and 1mmol/L salicylic acid treatment. 0.5mmol/L and 1mmol/L salicylic acid treated cell adhesion molecule CD44,E cadherin (CDH1), catenin A1 (CTNNA1), fibronectin (FN1) and nuclear laminin (Lamin), (Lamin B1) were detected by fluorescence quantitative PCR. Changes of expression of nuclear laminin C1 (Lamin C1) and integrin A9 (ITGA9) m RNA); The expression levels of CD44,CTNNA1,FN1 and Lamin B1 were observed by Western-blot. Results MTS results showed that the inhibitory rates of 0.5mmol/L and 1mmol/L were (32 卤4.5)% and (38 卤4.1)% after 24 h treatment, respectively, which were lower than 50%. Therefore, the salicylic acid of 0.5mmol/L and 1mmol/L was selected as the follow-up study. Cell adhesion assay showed that compared with the control group (121.00 卤2.65), the number of cell adhesion in 0.5mmol/L group and 1mmol/L group was significantly decreased after salicylic acid treatment. They were (88.00 卤1.53) and (44.00 卤2.00); Cell migration assay showed that the cell migration rate of 0.5mmol/L and 1mmol/L groups was (10.00 卤3.85)% and (12.00 卤3.61)%, respectively. The results of cell invasion showed that the OD values of cells treated with 0.5mmol/L and 1mmol/L salicylic acid were (0.18 卤0.00) and (0.15 卤0.00), respectively, which were lower than those of normal group (0.22 卤0.01). Fluorescence quantitative PCR showed that salicylic acid treatment in 0.5mmol/L group and 1mmol/L group significantly down-regulated the transcription of cell adhesion molecules CD44,CTNNA1,FN1,ITGA9,Lamin,Lamin B1 and Lamin C 1m RNA, and up-regulated the expression of CDH1 m RNA. Western-blot results also confirmed that the expression levels of CD44,CTNNA1,FN1 and Lamin B1 were significantly decreased in 0.5mmol/L and 1mmol/L groups after salicylic acid treatment (P0.05). Conclusion 1.0.5mmol/L and 1mmol/L salicylic acid can significantly inhibit the proliferation, adhesion, migration and invasion of gastric cancer cell lines in vitro. 2.0.5mmol/L and 1mmol/L salicylic acid down-regulated the expression of CD44,CTNNA1,FN1,ITGA9,Lamin,Lamin B1 and Lamin C1 and up-regulated the expression of CDH1. The mechanism of its anti-tumor metastasis may be related to the inhibition of the expression of cell adhesion molecules and the inhibition of epithelial mesenchymal transformation.
【学位授予单位】：宁波大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R735.2
【参考文献】
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1 李文;胡U，
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