鱼精蛋白-siRNA不同形貌复合体的制备、结构及药效学分析
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摘要
利用涡旋混匀及室温孵育的方法制备了不同形貌的鱼精蛋白-siRNA复合体,并利用聚丙烯酰胺凝胶电泳结合银染、原子力显微镜和透射电子显微镜等手段进行了结构表征.结果表明,鱼精蛋白可以和不同的siRNA分子以不同的质量比形成球形和纤维状鱼精蛋白-siRNA复合物.在此基础上,利用Mg~(2+)等金属离子探针,确认鱼精蛋白与siRNA的磷酸-戊糖骨架之间通过静电力发生作用.利用原子力和透射电子显微镜表征了鱼精蛋白-siRNA复合体的形貌和结构特征;利用激光纳米粒度仪测量了该复合体的粒径;并进一步比较了黑色素瘤细胞对不同形貌复合体的吞噬及药效学特征.
Protamine-siRNA complexes of different morphologies were prepared by vortex mixing different mass ratios of protamine and siRNA at room temperature,which could be resolved by polyacrylamide gel electrophoresis with silver staining. The protamine-siRNA complexes were observed under atomic force and transmission electron microscopy,respectively. It was found that similarly shaped protamine-siRNA complexes could be formed by protamine with different siRNA molecules,indicated that the interaction between protamine and siRNA was independent on the base composition of siRNA. Moreover,the interactions between the protamine and the siRNA molecules were characterized by ionic probes,including Mg~(2+) etc. And the morphologies,the particle sizes as well as the structural properties of the protamine-siRNA complexes were further characterized using atomic force microscope,dynamic light scattering and transmission electron microscope,respectively. The results show that protamine formed toroid complexes and fibrous composites with siRNA molecules respectively in the light of the different mass ratios between protamine and siRNA. In the meantime,the phagocytosis and the pharmacodynamics of protamine-siRNA complexes of different morphologies were compared and measured using melanoma cells.
Protamine-siRNA complexes of different morphologies were prepared by vortex mixing different mass ratios of protamine and siRNA at room temperature,which could be resolved by polyacrylamide gel electrophoresis with silver staining. The protamine-siRNA complexes were observed under atomic force and transmission electron microscopy,respectively. It was found that similarly shaped protamine-siRNA complexes could be formed by protamine with different siRNA molecules,indicated that the interaction between protamine and siRNA was independent on the base composition of siRNA. Moreover,the interactions between the protamine and the siRNA molecules were characterized by ionic probes,including Mg~(2+) etc. And the morphologies,the particle sizes as well as the structural properties of the protamine-siRNA complexes were further characterized using atomic force microscope,dynamic light scattering and transmission electron microscope,respectively. The results show that protamine formed toroid complexes and fibrous composites with siRNA molecules respectively in the light of the different mass ratios between protamine and siRNA. In the meantime,the phagocytosis and the pharmacodynamics of protamine-siRNA complexes of different morphologies were compared and measured using melanoma cells.
引文
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