Amidi Maryam, Romeijn Stefan G, Verhoef J Coos, Junginger Hans E, Bungener Laura, Huckriede Anke, Crommelin Daan J A, Jiskoot Wim
Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, P.O. Box 80082, 3508 TB Utrecht, The Netherlands.
Vaccine. 2007 Jan 2;25(1):144-53. doi: 10.1016/j.vaccine.2006.06.086. Epub 2006 Aug 4.
In this study, the potential of N-trimethyl chitosan (TMC) nanoparticles as a carrier system for the nasal delivery of a monovalent influenza subunit vaccine was investigated. The antigen-loaded nanoparticles were prepared by mixing a solution containing TMC and monovalent influenza A subunit H3N2 with a tripolyphosphate (TPP) solution, at ambient temperature and pH 7.4 while stirring. The nanoparticles had an average size of about 800 nm with a narrow size distribution and a positive surface charge. The nanoparticles showed a loading efficiency of 78% and a loading capacity of 13% (w/w). It was shown that more than 75% of the protein remained associated with the TMC nanoparticles upon incubation of the particles in PBS for 3h. The molecular weight and antigenicity of the entrapped hemagglutinin was maintained as shown by polyacrylamide gel electrophoresis and Western blotting, respectively. Single i.n. or i.m. immunization with antigen-loaded TMC nanoparticles resulted in strong hemagglutination inhibition and total IgG responses. These responses were significantly higher than those achieved after i.m. administration of the subunit antigen, whereas the IgG1/IgG2a profile did not change substantially. The i.n. administered antigen-TMC nanoparticles induced higher immune responses compared to the other i.n. antigen formulations, and these responses were enhanced by i.n. booster vaccinations. Moreover, among the tested formulations only i.n. administered antigen-containing TMC nanoparticles induced significant IgA levels in nasal washes of all mice. In conclusion, these findings demonstrate that TMC nanoparticles are a potent new delivery system for i.n. administered influenza antigens.
在本研究中，对N-三甲基壳聚糖（TMC）纳米颗粒作为单价流感亚单位疫苗鼻腔给药载体系统的潜力进行了研究。通过在室温及pH 7.4条件下搅拌，将含有TMC和单价甲型流感H3N2亚单位的溶液与三聚磷酸钠（TPP）溶液混合，制备出负载抗原的纳米颗粒。这些纳米颗粒的平均尺寸约为800 nm，粒径分布窄，表面带正电荷。纳米颗粒的负载效率为78%，负载量为13%（w/w）。结果表明，将颗粒在PBS中孵育3小时后，超过75%的蛋白质仍与TMC纳米颗粒结合。聚丙烯酰胺凝胶电泳和蛋白质免疫印迹分别显示，包封的血凝素的分子量和抗原性得以维持。用负载抗原的TMC纳米颗粒进行单次鼻腔或肌肉注射免疫，可产生强烈的血凝抑制和总IgG反应。这些反应显著高于肌肉注射亚单位抗原后所产生的反应，而IgG1/IgG2a谱没有明显变化。与其他鼻腔抗原制剂相比，鼻腔给药的抗原-TMC纳米颗粒诱导的免疫反应更高，并且这些反应通过鼻腔加强免疫得以增强。此外，在所有测试制剂中，只有鼻腔给药的含抗原TMC纳米颗粒在所有小鼠的鼻腔灌洗液中诱导出显著的IgA水平。总之，这些发现表明，TMC纳米颗粒是一种用于鼻腔给药流感抗原的有效新型递送系统。
