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目的:研究硬脂醇半乳糖苷修饰的阿西替尼脂质体对人肝癌SMMC-7721细胞株的增殖及凋亡的诱导作用。方法:采用冷冻干燥法制备硬脂醇半乳糖苷修饰的阿西替尼脂质体,以包封率为评价指标,采用葡聚糖凝胶过滤法和高效液相色谱法测定硬脂醇半乳糖苷修饰的阿西替尼脂质体的包封率。采用Box-Behnken响应面设计法优化制备工艺,并研究CCK-8法检测硬脂醇半乳糖苷修饰的阿西替尼脂质体对人肝癌SMMC-7721细胞株的抑制情况,采用Annexin V/PI流式细胞分析法检测细胞凋亡,Western blot检测凋亡蛋白Bax,Bcl-2,P53的表达。结果:最佳工艺:药脂比为1∶20,胆脂比为1∶8,硬脂醇半乳糖苷与磷脂比为1∶15,水合温度为55℃。CCK-8法检测不同浓度的硬脂醇半乳糖苷修饰的阿西替尼脂质体对SMMC-7721细胞株的增殖具有抑制作用,呈现时间和浓度依赖性。Annexin V/PI流式细胞分析法检测硬脂醇半乳糖苷修饰的阿西替尼脂质体对人肝癌SMMC-7721细胞株的抑制率优于人肺癌A549细胞株,Western blot检测发现随着硬脂醇半乳糖苷修饰的阿西替尼脂质体浓度的升高,Bax,P53蛋白的表达逐渐升高,Bcl-2蛋白的表达逐渐降低。结论:硬脂醇半乳糖苷修饰的阿西替尼脂质体在一定浓度范围内,可在体外抑制人肝癌SMMC-7721细胞株的增殖并诱导其凋亡,且随着硬脂醇半乳糖苷修饰的阿西替尼前体脂质体的浓度升高,凋亡率显著增加,可能的机制是增强促凋亡蛋白Bax,P53的表达,降低抗凋亡蛋白Bal-2的表达。硬脂醇半乳糖苷修饰的阿西替尼脂质体诱导SMMC-7721细胞株凋亡能力比A549强,初步判定硬脂醇半乳糖苷修饰的阿西替尼脂质体具有主动肝靶向性。
OBJECTIVE: To study the induction of proliferation and apoptosis of human hepatoma SMMC-7721 cell line with stearyl alcohol galactoside modified axitinib liposome. Methods: The freeze-drying method was used to prepare stearyl alcohol galactoside modified axitinib liposomes. The entrapment efficiency was used as the evaluation index to determine the fraction of stearyl alcohol by using Sephadex LH-20 and high performance liquid chromatography Encapsulation efficiency of axitinib-modified liposomes. The Box-Behnken response surface design method was used to optimize the preparation process. The inhibitory effect of CCK-8 on the proliferation of SMMC-7721 hepatocarcinoma cells was detected by using liposomes containing stearyl galactoside modified by liposomes. Annexin V / PI flow cytometry analysis of apoptosis, Western blot detection of apoptosis protein Bax, Bcl-2, P53 expression. Results: The optimum process was as follows: the ratio of lipid to lipid was 1:20, the ratio of cholesterol to lipid was 1: 8, the ratio of stearyl galactoside to phospholipid was 1:15, and the hydration temperature was 55 ℃. CCK-8 assay of different concentrations of stearyl alcohol galactoside modified axitinib liposome SMMC-7721 cell proliferation inhibited, showing time and concentration-dependent. Annexin V / PI flow cytometry detection of stearyl galactoside modified axitinib liposomes on human hepatocellular carcinoma SMMC-7721 cell line inhibition rate is superior to human lung cancer A549 cell line, Western blot test found that with Stearyl alcohol galactoside modified axitinib liposome concentration increased, Bax, P53 protein expression gradually increased, Bcl-2 protein expression decreased. CONCLUSIONS: Stearyl alcohol galactoside modified axitinib liposomes can inhibit the proliferation and induce the apoptosis of human hepatoma SMMC-7721 cells in vitro in a certain concentration range. With the increase of stearyl alcohol galactoside The concentration of glycoside modified axitinib precursor liposomes increased, and the apoptosis rate increased significantly. The possible mechanism was to increase the expression of pro-apoptotic proteins Bax and P53 and decrease the expression of anti-apoptotic protein Bal-2. Stearyl alcohol galactoside modified axitinib liposome SMMC-7721 cells induced apoptosis than A549 strong, initially determined stearyl alcohol galactoside modified axitinib liposomes with active liver targeting Sex.