叶酸对宫颈癌SiHa细胞生物学特性及βeclin-1蛋白表达影响的初步研究

发布时间：2018-04-01 12:11

本文选题：宫颈癌　切入点：叶酸　出处：《山西医科大学》2016年硕士论文

【摘要】：目的:宫颈癌作为全球女性最为常见的妇科恶性肿瘤之一,发病率和死亡率呈逐年上升趋势。叶酸是机体必需的微量营养素,相关流行病学证据亦表明叶酸与宫颈癌的发生有着密切的关系。自噬是细胞死亡的一种独特方式,细胞自噬在对抗肿瘤生成方面也扮演了作用的角色。但对于宫颈癌、自噬、叶酸这三者间关系的研究却尚未涉及,因此它们的关系未明确。本实验旨在探讨叶酸对宫颈癌SiHa细胞生物学功能及对自噬基因βeclin-1表达的影响。方法:1.常规条件下培养SiHa细胞,利用1ug/mL(对照组)、10ug/ml、100ug/ml、500ug/ml、1000ug/ml的叶酸进行干预,观察并记录细胞的形态变化和生长速度。2.利用1ug/mL(对照组)、10ug/ml、100ug/ml、500ug/ml、1000ug/ml的叶酸进行干预后以CCK8检测宫颈癌SiHa细胞的增殖活性,划痕实验观察细胞的迁移能力及流式细胞仪分析细胞的凋亡情况,采用Western bloting检测自噬蛋白βeclin-1表达水平。结果:1.与对照组比较,叶酸浓度达到1000ug/ml时细胞形态发生明显变化,当实验组叶酸浓度在100ug/ml及以上时,与对照组相比,实验组细胞总数明显降低。2.CCK8细胞增殖实验显示在不同叶酸浓度干预下,除了10ug/ml叶酸的实验组与对照组相比P=0.650无差异无统计学意义外,100ug/ml、500ug/ml、1000ug/ml浓度的叶酸细胞增殖与对照组相比均有统计学意义,P值分别为0.041、0.001、0.001。3.划痕实验表明10ug/ml的叶酸实验组划痕愈合面积与对照组比较差异不明显。100ug/ml、500ug/ml、1000ug/ml浓度的叶酸实验组与对照组相比划痕愈合面积减少,且随着叶酸浓度的增加愈合面积减少愈发明显。4.流式细胞仪分析叶酸对宫颈癌细胞凋亡的影响,结果显示10ug/ml的叶酸实验组细胞的凋亡率与对照组比较P=0.580,差异无统计学意义外,100ug/ml、500ug/ml、1000ug/ml浓度的叶酸细胞的凋亡率与对照组相比均有统计学意义,P值分别为0.020、0.001、0.001。5.Wesrern Bloting检测叶酸对宫颈癌细胞自噬基因βeclin-1蛋白表达的作用。实验组10ug/ml、100ug/ml、500ug/ml、1000ug/ml浓度的叶酸实验组βeclin-1蛋白表达水平与对照组相比差异均有统计学意义P值分别为0.014、0.003、0.001、0.001。结论:1.高浓度叶酸抑制宫颈癌SiHa细胞的增殖、迁移,促进宫颈癌SiHa细胞的凋亡。2.高浓度叶酸促进宫颈癌自噬蛋白βeclin-1的表达。
[Abstract]:Objective: cervical cancer is one of the most common gynecological malignancies in the world.Folic acid is an essential micronutrient in the body. Relevant epidemiological evidence also shows that folic acid is closely related to the occurrence of cervical cancer.Autophagy is a unique way of cell death. Autophagy also plays a role in fighting tumor formation.However, the relationship among cervical cancer, autophagy and folic acid has not been studied, so their relationship is not clear.The aim of this study was to investigate the effects of folic acid on the biological function of cervical cancer SiHa cells and the expression of autophagy gene 尾 eclin-1.Method 1: 1.SiHa cells were cultured under conventional conditions and treated with folic acid of 10ugrmL (control group). The morphological changes and growth rate of the cells were observed and recorded.The result is 1: 1.When the concentration of folic acid reached 1000ug/ml, the morphology of the cells changed obviously. When the concentration of folic acid in the experimental group was 100ug/ml and above, it was compared with the control group.The total number of cells in the experimental group was significantly decreased. 2. CCK8 cell proliferation assay showed that the cells were treated with different folic acid concentrations.There was no significant difference in the proliferation of folic acid cells between the experimental group and the control group except that of the control group. The proliferation of folic acid cells at the concentration of 100U / ml / ml / ml / ml was significantly higher than that of the control group (P = 0.041 / 0.001 / ml, P = 0.001.3respectively).Scratch test showed that there was no significant difference in scratch healing area between the folic acid experimental group of 10ug/ml and the control group. Compared with the control group, the area of scratch healing in the folic acid experimental group with the concentration of 100ugml / ml was less than that in the control group, and with the increase of folic acid concentration, the healing area of folic acid group was decreased more obviously than that of the control group.The effect of folic acid on apoptosis of cervical cancer cells was analyzed by flow cytometry.The results showed that the apoptotic rate of the folic acid experimental group of 10ug/ml was 0.580 compared with that of the control group. There was no significant difference between the two groups. The apoptosis rate of the folic acid cells at the concentration of 100U / ml / ml was 0.020 卤0.001 / ml and 0.001.5.Wesrern Bloting was used to detect the apoptosis rate of the folic acid cells in the control group and the control group respectively (P = 0.020, P = 0.020, P = 0.001.5.Wesrern Bloting).Effect of 尾 eclin-1 protein on autophagy gene in cervical cancer cells.Compared with the control group, the expression level of 尾 eclin-1 protein in the experimental group (10ugr / ml) was significantly higher than that in the control group (P = 0.014, 0.003, P = 0.001, P = 0.001, respectively), and the expression level of 尾 eclin-1 protein in the experimental group was significantly higher than that in the control group (P < 0.05).Conclusion 1.High concentration folic acid inhibited the proliferation and migration of cervical cancer SiHa cells and promoted the apoptosis of cervical cancer SiHa cells.High concentration folic acid promoted the expression of autophagy protein 尾 eclin-1 in cervical cancer.
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R737.33

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