纳米明胶硅氧烷的制备及其负载p53基因对肝癌抑制效果

Preparation of nano-gelatin siloxane and inhibition effect of its loaded p53 gene on the hepatocellular carcinoma

采用溶胶-凝胶法制备明胶硅氧烷纳米颗粒(GS NPs),分析反应中的不同pH值盐酸溶液对GS NPs的影响,并通过粒径、电位和形貌对GS NPs进行表征。结果表明:pH值3.0盐酸溶液为制备的最佳条件,制备的GS NPs呈无规则球状,分散性较好,平均粒径为250nm左右且分布集中,表面Zeta电位为38mV。为评估其作为基因载体的可行性及负载p53基因对肝癌的抑制效果,采用凝胶电泳实验检测GS NPs对pDNA的负载率和释放性,CCK-8法检测其生物相容性和激光共聚焦观察细胞内摄取情况以及MTT法、Western blot综合评价其携载pEGFP-C1-p53对肝癌细胞的抑制效果,结果显示:GS NPs可有效负载pDNA且最适负载量比为200∶1,在盐溶液下表现出一定的释放性;此外GS NPs具有良好的生物相容性,可携载pDNA逃逸出溶酶体的吞噬,携载pEGFPC1-p53后对肝癌细胞Hep-3b表现出显著的抑制效果,并介导GFP-p53融合蛋白的表达。研究结果表明所制备的GS NPs可作为一种基因载体,负载p53基因对肝癌细胞具有一定的抑制效果。

Gelatin siloxane nanoparticles （GS NPs） were prepared with sol gel method, and the effect of hydrochloric acid solution with different pH on GS NPs was discussed. GS NPs were characterized through particle size, potential and morphology. The results showed that, hydrochloric acid solution with the pH value of 3. 0 was the optimal condition to prepare GS NPs. The GS NPs with good dispersion presented irregular spherical. The average particle size was about 250 nm with concentrated distribution and the surface Zeta potential was 38 inV. To evaluate its feasibility as a gene carrier and the inhibition effect of loaded p53 gene to liver cancer, gel electrophoresis was used to test the load rate and release of GS NPs on pDNA. CCK 8 method was applied to detect the biocompatibility and laser scanning confocal microscopy （I.SCM） to observeintracellular uptake. The inhibition effect of GS NPsloaded with pEGFP CI 1953 on hepatocellular carcinoma cells was evaluated by using MTT and Western blot. The results showed that GS NPs could effectively load pDNA with the optimal load ratio of 200 ： 1 and showed certain release in the salt solution. In addition, GS NPs had good biocompatibility and could carry pDNA to escape from the phagocytosis of lysosomes. After carrying pEGFP-CI-p53, it showed significant inhibition effect on hepatoma cells Hep 3b and mediated expression of GFP-p53 fusion protein. The results of this study indicated that GS NPs could be used as a gene carrier and the loaded p58 gene had certain inhibition effect on liver cancer cells.