褐藻多糖硫酸酯对人黑色素瘤A357系细胞增殖与凋亡功能的影响研究

发布时间：2018-05-21 08:11

本文选题：褐藻多糖硫酸酯 + 黑色素瘤　；参考：《西南医科大学》2016年硕士论文

【摘要】：背景:黑色素瘤(melanoma)是常见的皮肤恶性肿瘤之一,其发病率在皮肤恶性肿瘤中为第三,死亡率为第一。针对黑色素瘤的治疗方法较多,包括手术、放化疗、免疫治疗、个性化生物治疗、个体靶向治疗等。但这些治疗方法仍存疗效不高、副作用较大等问题。因此,研究新的有效的预防和治疗黑色素瘤的药物显得十分的重要。褐藻多糖硫酸酯(fucoidan)是从褐藻或无脊椎动物中提取的,具有多种生物学活性,大量研究发现褐藻多糖硫酸酯对肺癌、结肠癌、乳腺癌、胃癌、肝癌等多种肿瘤均有一定抗肿瘤治疗作用。褐藻多糖硫酸酯不仅可以通过多种机制促进肿瘤细胞凋亡,还与某些药物具有协同作用,能增加机体对化疗药物的敏感性及疗效并减少其毒性。国外有学者在体外细胞实验中发现褐藻多糖硫酸酯对黑色素瘤(B16)细胞有抑制增殖和促进凋亡的作用,目前国内尚无褐藻多糖硫酸酯对黑色素瘤细胞作用的研究。目的:通过体外细胞实验研究褐藻多糖硫酸酯对人恶性黑色素瘤A375系细胞增殖和凋亡的影响,以及凋亡相关蛋白Caspase-3的表达情况。为其应用于黑色素瘤的临床治疗提供实验和理论依据。方法:体外培养人黑色素瘤A375系细胞,用不同浓度褐藻多糖硫酸酯预处理细胞24h后,通过CCK8检测细胞增殖并计算出细胞增殖抑制率,绘制量效曲线,得出其半数增殖抑制浓度(IC50)。通过Annexin V-FITC/PI双标法流式细胞仪观察其对细胞凋亡的影响;以蛋白免疫印迹法(Western blot)检测凋亡蛋白Caspase-3表达水平的变化,初步探讨褐藻多糖硫酸酯对人黑色素瘤细胞增殖和凋亡功能的影响。结果:与空白对照组相比,CCK8检测结果显示褐藻多糖硫酸酯作用24h后,A375细胞的增殖抑制率随药物浓度的增加而增高,计算出其半数增殖抑制浓度为(IC50)0.51mg/ml;流式细胞计数检测细胞凋亡结果表明褐藻多糖硫酸酯作用后细胞的凋亡率明显增高;Western blot的结果表明:随着褐藻多糖硫酸酯浓度的增加,Caspase-3的表达水平增高。结论:褐藻多糖硫酸酯可以抑制人黑色素瘤A375系细胞的增殖并促进该细胞的凋亡。
[Abstract]:Background: melanoma is one of the most common malignant skin tumors. Its morbidity is the third and mortality is the first among the malignant skin tumors. There are many methods for the treatment of melanoma, including surgery, radiotherapy and chemotherapy, immunotherapy, individualized biotherapy, individual targeted therapy and so on. However, these treatments are still not high efficacy, side effects and other problems. Therefore, it is very important to study new effective drugs for the prevention and treatment of melanoma. Fucoidan is derived from brown algae or invertebrates, and has many biological activities. A large number of studies have found that fucoidan sulfates from brown algae are effective against lung, colon, breast and stomach cancer. Many kinds of tumor, such as liver cancer, have certain anti-tumor effect. Fucoidan sulfate can not only promote apoptosis of tumor cells through various mechanisms, but also have synergistic effect with some drugs. It can increase the sensitivity and efficacy of the body to chemotherapy drugs and reduce its toxicity. Some foreign scholars have found that fucoidan sulfate can inhibit proliferation and promote apoptosis of melanoma B16 cells in vitro. There is no study on the effect of fucoidan sulfate on melanoma cells. Aim: to study the effect of polysaccharide sulfate of brown algae on proliferation and apoptosis of human malignant melanoma cell line A375 in vitro and the expression of apoptosis-related protein (Caspase-3). To provide experimental and theoretical basis for the clinical treatment of melanoma. Methods: human melanoma A375 cells were cultured in vitro. After 24 h pretreatment with different concentrations of brown algae polysaccharide sulfate, the proliferation of A375 cells was measured by CCK8, the inhibition rate of cell proliferation was calculated, the dose-response curve was drawn, and the IC50 ~ (50) inhibition concentration was obtained. The effect of Annexin V-FITC/PI on apoptosis was observed by flow cytometry, and the expression of apoptotic protein Caspase-3 was detected by Western blotting. To study the effect of polysaccharide sulfate of brown algae on proliferation and apoptosis of human melanoma cells. Results: compared with the control group, the results of CCK8 detection showed that the inhibition rate of proliferation of A375 cells increased with the increase of drug concentration after 24 hours of treatment with polysaccharide sulfate of brown algae. The results of flow cytometry analysis showed that the apoptosis rate of the cells treated with polysaccharides sulfate from brown algae increased significantly. The results showed that: with the concentration of polysaccharides sulfate of brown algae, the cell apoptosis rate was increased with the concentration of fucoidan sulfate. The expression of Caspase-3 was increased. Conclusion: fucoidan sulfate can inhibit the proliferation of human melanoma cell line A375 and promote the apoptosis of A375 cell line.
【学位授予单位】：西南医科大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R739.5

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