COX-2、MMP-14的相互作用在食管癌细胞增殖、侵袭及凋亡中的研究

发布时间：2018-04-10 02:42

本文选题：食管癌ECa109细胞　切入点：塞来昔布　出处：《石河子大学》2017年硕士论文

【摘要】：目的:使用不同浓度COX-2抑制剂塞来昔布作用于食管癌ECa109细胞,测定细胞内环氧合酶-2(cyclooxygenase-2,COX-2)和基质金属蛋白酶14(matrix metalloproteinase 14,MMP-14)的蛋白表达,同时测定细胞增殖、侵袭、凋亡能力变化,从而进一步研究COX-2、MMP-14相互作用在食管癌发生发展中的作用及机制。方法:1.体外培养人食管癌ECa109细胞;2.使用0(对照组)、20(低)、60(中)、100(高)μmol/L四个塞来昔布浓度对ECa109细胞进行不同时间(24、48和72h)处理,MTT法检测细胞的增殖能力,流式细胞仪测凋亡,Transwell侵袭实验检测细胞的侵袭能力,酶联免疫法检测COX-2和MMP-14的蛋白表达情况;3.数据进行统计分析。结果:1.ELISA结果提示在塞来昔布浓度在0-100μmol/L之间,随着药物浓度递增,细胞内COX-2、MMP-14蛋白表达逐渐减少,均呈浓度依赖性,COX-2蛋白表达水平分别为1.581±0.116、1.226±0.089、0.846±0.076和0.521±0.082,四个组间差异有统计学意义(P0.05);MMP-14蛋白表达水平分别为11.517±0.881、7.905±0.724、5.788±0.750、和3.663±0.601,四个组间差异有统计学意义(P0.05);Pearson相关分析结果显示两者表达水平具有显著正相关性(r=0.952,P0.05)。2.MTT检测结果示不同浓度的药物作用24、48和72h后,药物对细胞增殖抑制率分别为:处理24 h后,较对照组(6.92±1.92)相比,20、60、100μmol/L组增殖率分别为25.52±2.39、40.60±4.06和49.42±2.67;处理48h后,较对照组(7.10±1.74)相比,20、60、100μmol/L组增殖率分别为43.70±5.20、55.77±4.18和66.44±4.03;处理72h后,较对照组(7.07±1.22)相比,20、60、100μmol/L组增殖率分别为60.34±4.64、72.09±3.93和84.31±3.21;随着药物浓度的提高,药物对EC109细胞的增殖抑制率逐渐增强,且随着药物作用时间的增加,抑制作用也逐渐加强,抑制作用呈时间和浓度依赖性(P0.05);3 Transwell侵袭实验结果显示在不同浓度的药物作用ECa109细胞24h后,细胞的侵袭能力下降,计数穿过基质胶的细胞个数分别为300.654±12.558、271.041±10.569、184.003±10.865和92.667±7.567(F=237.182,P0.05),并呈浓度依赖性;4.流式细胞仪测定结果显示在不同浓度的药物作用ECa109细胞24h后,细胞凋亡率分别为1.700±0.557、13.400±1.735、18.767±1.301和28.300±71.988(F=161.843,P0.05)细胞的凋亡率逐渐增加,呈药物浓度依赖性。结论:1.COX-2抑制剂塞来昔布可抑制食管癌ECa109细胞增殖,此作用具有浓度及时间依赖性;同时可促进细胞的凋亡,抑制细胞的侵袭能力,此作用具有浓度依赖性。2.COX-2抑制剂塞来昔布可能通过抑制COX-2表达,下调MMP-14表达,从而抑制食管癌细胞的增殖、侵袭能力,促进细胞凋亡,从而起到抗食管癌作用。
[Abstract]:Aim: to observe the expression of cyclooxygenase-2cyclooxygenase-2 (COX-2) and matrix metalloproteinase (14(matrix metalloproteinase 14) in esophageal carcinoma ECa109 cells treated with celecoxib at different concentrations, and to detect the changes of cell proliferation, invasion and apoptosis.To further study the role and mechanism of COX-2 MMP-14 interaction in the carcinogenesis and development of esophageal carcinoma.Method 1: 1.Human esophageal carcinoma ECa109 cells were cultured in vitro.The proliferation ability of ECa109 cells was detected by MTT assay with 0 (control group) (control group 0 (control group) with 4 celecoxib concentrations of 4 celecoxib at different time points, 24 渭 mol/L and 72 h), and the ability of cell invasion was detected by flow cytometry with apoptosis transwell invasion assay, and the cell proliferation was detected by flow cytometry (FCM), and the cell proliferation was detected by flow cytometry (FCM), and the proliferation of ECa109 cells was detected by flow cytometry.The protein expression of COX-2 and MMP-14 was detected by enzyme linked immunosorbent assay (Elisa).The data are analyzed statistically.Results 1. The results of Elisa showed that when the concentration of celecoxib ranged from 0 to 100 渭 mol/L, the expression of COX-2 MMP-14 protein gradually decreased with the increase of drug concentration.The results of MTT assay showed that the drug of different concentrations had a significant positive correlation after 24 h and 72 h, and the results of MTT assay showed that different concentrations of the drug could be used for 24 h, 48 h and 72 h, respectively.The inhibitory rates of the drug on cell proliferation were 25.52 卤2.39 渭 mol/L and 49.42 卤2.67, respectively, compared with the control group (6.92 卤1.92) and the control group (43.70 卤5.2055.77 卤4.18 and 66.44 卤4.03, respectively, 48 h after treatment), compared with the control group (7.10 卤1.74), the proliferative rates were 43.70 卤5.2055.77 卤4.18 and 66.44 卤4.03, respectively, after 48 hours of treatment, the cell proliferation inhibition rates were 43.70 卤5.2055.77 卤4.18 and 66.44 卤4.03respectively.Compared with the control group (7.07 卤1.22), the proliferative rates of the control group were 60.34 卤4.64 渭 mol/L, 72.09 卤3.93 and 84.31 卤3.21, respectively. With the increase of the drug concentration, the inhibitory rate of the drug on EC109 cells was gradually increased, and the inhibitory effect was gradually strengthened with the increase of the time of the drug treatment.The results of flow cytometry showed that the apoptosis rates of ECa109 cells treated with different concentrations of drugs for 24 h were 1.700 卤0.557 (13.400 卤1.735N) 18.767 卤1.301 and 28.300 卤71.988FN 161.843 (P0.05), respectively, in a dose-dependent manner.Conclusion 1. Celecoxib, a COX-2 inhibitor, can inhibit the proliferation of esophageal carcinoma ECa109 cells in a concentration and time dependent manner, promote cell apoptosis and inhibit cell invasion.The effect is concentration-dependent. 2. Celecoxib, a COX-2 inhibitor, may inhibit the proliferation, invasion and apoptosis of esophageal cancer cells by inhibiting the expression of COX-2 and down-regulating the expression of MMP-14.
【学位授予单位】：石河子大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R735.1

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