IL13修饰雷公藤红素/人参总皂苷脂质体的制备及其抑制BT325细胞作用

Preparation of IL13-modified tripterine/total ginsenosides liposomes and the inhibitory effect on BT325 cells

目的制备白介素13(IL13)修饰雷公藤红素/人参总皂苷脂质体,并评价其对人脑胶质瘤细胞BT325的抑制作用。方法薄膜分散法制备脂质体,1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐(EDC)/N-羟基琥珀酰亚胺(NHS)水相缩合法将IL13锚定在粒子表面,考察其形态、粒径、Zeta电位、IL13修饰密度的质量分数、包封率、体外释放。以药脂比(雷公藤红素/卵磷脂)、胆脂比、水合时间、IL13投料比为影响因素,粒径、Zeta电位、聚合物分散性指数(PDI)为评价指标,正交试验优化制备工艺。荧光定性和胞内药物定量法评价BT325细胞摄取行为,MTT法和流式细胞仪法分别测定脂质体抗肿瘤和诱导细胞凋亡作用。结果最佳条件为药脂比1∶20,胆脂比1∶4,水合时间45 min,IL13投料比0.6%。所得脂质体均一圆整,粒径分布较窄,平均粒径(118.3±3.2)nm,Zeta电位(-38.3±3.4)m V,IL13修饰密度的质量分数0.3%,雷公藤红素和人参总皂苷包封率分别为(82.3±2.9)%和(71.2±3.2)%,24 h累计释放度分别为(42.5±4.9)%和(24.3±2.1)%。它可显著促进BT325细胞摄取,IC50为(2.8±0.3)μg/m L(以雷公藤红素计),低于IL13未修饰脂质体和物理混合物(雷公藤红素-人参总皂苷)。当雷公藤红素质量浓度为10μg/m L时,它还能诱导84.3%的BT325细胞发生凋亡。结论 IL13修饰雷公藤红素/人参总皂苷脂质体在体外表现出明显的人脑胶质瘤细胞BT325靶向性和体外抗肿瘤作用。

AIM To prepare the interleukin-13（ IL13）-modified tripterine/total ginsenosides liposomes and to evaluate the inhibitory effect on human brain glioma BT325 cells. METHODS By 1-（ 3-dimethylaminopropyl）-3-ethylcarbodiimide hydrochloride（ EDC）/N-hydroxysuccinimide（ NHS） aqueous phase condensation method,IL13 was anchored onto the particle surface of liposomes prepared by film dispersion method. The morphology,particle size,Zeta potential,concentration of IL13-modification density,encapsulation efficiency and in vitro release were investigated. In addition to the influencing factors of drug-lipid ratio（ tripterine/lecithin）,cholesterollipid ratio,hydration time and IL13 feed ratio,the evaluation indices of particle size,Zeta potential and polydispersity index（ PDI） for the preparation were taken into account in the optimization by orthogonal test. The uptake behaviors of BT325 cells were evaluated by fluorescence qualitative method and intracellular drug quantitative method,after which the liposomes＇ anti-tumor activity and apoptosis induction were investigated by MTT method and flow cytometry method,respectively. RESULTS The optimal conditions were determined to be 1 ∶ 20 for druglipid ratio,1 ∶ 4 for cholesterol-lipid ratio,45 min for hydration time,and 0. 6% for IL13 feed ratio. The obtained liposomes were uniformly round with narrow particle size distribution,the average particle size was（ 118. 3 ± 3. 2）nm,Zeta potential was（- 38. 3 ± 3. 4） m V,the concentration of IL13-modification density was 0. 3%,and the encapsulation efficiencies of tripterine and total ginsenosides were（ 82. 3 ± 2. 9） % and（ 71. 2 ± 3. 2） % with the cumulative release rates of（ 42. 5 ± 4. 9） % and（ 24. 3 ± 2. 1） % at 24 h,respectively. It could significantly promote the uptake of BT325 cells,and the IC50 value was（ 2. 8 ± 0. 3） μg/m L（ in terms of tripterine）,which was lower than those of non-IL13-modified liposomes and physical mixture of tripterine and total ginsenosides. At10 μg/m L for tripterine concentration,the lipsomes induced 84. 3% apoptosis of BT325 cells. CONCLUSION IL13-modified tripterine/total ginsenosides liposomes exhibit obvious human brain glioma BT325 cells-targeting ability and in vitro anti-tumor effect.