Huang Shih-shiung, Li I-Hsun, Hong Po-da, Yeh Ming-kung
Biomedical Engineering Program, Graduate Institute of Engineering, Department of Materials Science and Engineering, National Taiwan University of Science and Technology, Taipei, Taiwan, Republic of China.
School of Pharmacy, National Taiwan University of Science and Technology, Taipei, Taiwan, Republic of China ; Department of Pharmacy Practice, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan, Republic of China.
Int J Nanomedicine. 2014 Feb 7;9:813-22. doi: 10.2147/IJN.S56260. eCollection 2014.
Yersinia pestis F1 antigen-loaded poly(DL-lactide-co-glycolide)/polyethylene glycol (PEG) (PLGA/PEG) microspheres were produced using a water-in-oil-in-water emulsion/solvent extraction technique and assayed for their percent yield, entrapment efficiency, surface morphology, particle size, zeta potential, in vitro release properties, and in vivo animal protect efficacy. The Y. pestis F1 antigen-loaded microspheres (mean particle size 3.8 μm) exhibited a high loading capacity (4.5% w/w), yield (85.2%), and entrapment efficiency (38.1%), and presented a controlled in vitro release profile with a low initial burst (18.5%), then continued to release Y. pestis F1 antigen over 70 days. The distribution (%) of Y. pestis F1 on the microspheres surface, outer layer, and core was 3.1%, 28.9%, and 60.7%, respectively. A steady release rate was noticed to be 0.55 μg Y. pestis F1 antigen/mg microspheres/day of Y. pestis F1 antigen release maintained for 42 days. The cumulative release amount at the 1st, 28th, and 42nd days was 8.2, 26.7, and 31.0 μg Y. pestis F1 antigen/mg microspheres, respectively. The 100 times median lethal dose 50% (LD50) of Y. pestis Yokohama-R strain by intraperitoneal injection challenge in mice test, in which mice received one dose of 40 μg F1 antigen content of PLGA/PEG microspheres, F1 antigen in Al(OH)3, and in comparison with F1 antigen in Al(OH)3 vaccine in two doses, was evaluated after given by subcutaneous immunization of BALB/c mice. The study results show that the greatest survival was observed in the group of mice immunized with one dose of F1 antigen-loaded PLGA/PEG microspheres, and two doses of F1 antigen in Al(OH)3 vaccine (100%). In vivo vaccination studies also demonstrated that F1 vaccines microspheres had a protective ability; its steady-state IgG immune protection in mice plasma dramatic increased from 2 weeks (18,764 ± 3,124) to 7 weeks (126,468 ± 19,176) after vaccination. These findings strongly suggest that F1-antigen loaded microspheres vaccine offer a new therapeutic strategy in optimizing the vaccine incorporation and delivery properties of these potential vaccine targeting carriers.
采用水包油包水乳液/溶剂萃取技术制备了负载鼠疫耶尔森菌F1抗原的聚（DL-丙交酯-共-乙交酯）/聚乙二醇（PLGA/PEG）微球，并对其产率、包封率、表面形态、粒径、zeta电位、体外释放特性及体内动物保护效果进行了测定。负载鼠疫耶尔森菌F1抗原的微球（平均粒径3.8μm）具有较高的负载量（4.5% w/w）、产率（85.2%）和包封率（38.1%），呈现出体外可控释放曲线，初始突释较低（18.5%），然后在70多天内持续释放鼠疫耶尔森菌F1抗原。鼠疫耶尔森菌F1在微球表面、外层和核心的分布（%）分别为3.1%、28.9%和60.7%。观察到鼠疫耶尔森菌F1抗原的稳定释放速率为0.55μg鼠疫耶尔森菌F1抗原/毫克微球/天，持续42天。第1天、第28天和第42天的累积释放量分别为8.2、26.7和31.0μg鼠疫耶尔森菌F1抗原/毫克微球。在对BALB/c小鼠进行皮下免疫后，通过腹腔注射攻击小鼠试验评估了横滨-R株鼠疫耶尔森菌的100倍半数致死剂量50%（LD50），其中小鼠分别接受一剂含40μg F1抗原的PLGA/PEG微球、氢氧化铝中的F1抗原，以及与两剂氢氧化铝疫苗中的F1抗原进行比较。研究结果表明，在接受一剂负载F1抗原的PLGA/PEG微球免疫的小鼠组以及两剂氢氧化铝疫苗中的F1抗原免疫的小鼠组中观察到最大存活率（100%）。体内疫苗接种研究还表明，F1疫苗微球具有保护能力；接种疫苗后，小鼠血浆中的稳态IgG免疫保护从2周时的（18,764±3,124）显著增加到7周时的（126,468±19,176）。这些发现有力地表明，负载F1抗原的微球疫苗为优化这些潜在疫苗靶向载体的疫苗掺入和递送特性提供了一种新的治疗策略。
