Formulation, characterization, and expression of a recombinant MOMP Chlamydia trachomatis DNA vaccine encapsulated in chitosan nanoparticles.
Formulation, characterization, and expression of a recombinant MOMP Chlamydia trachomatis DNA vaccine encapsulated in chitosan nanoparticles.
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DOI：
10.2147/ijn.s42723
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发表时间：
2013
影响因子：
8
通讯作者：
Dennis VA
中科院分区：
文献类型：
--
作者：
Cambridge CD;Singh SR;Waffo AB;Fairley SJ;Dennis VA
沙眼衣原体是一种细菌性性传播感染，影响全世界数百万人。以前的疫苗接种尝试采用了C.尽管如此，沙眼，但成功有限，可能是由于稳定性、降解和递送问题。本研究克隆了C.沙眼衣原体重组MOMP DNA（DMOMP），并使用复凝聚技术将其包封在壳聚糖纳米粒（DMCNP）中。DMCNP的理化特性包括透射和扫描电子显微镜，傅立叶变换红外光谱和紫外-可见光谱，和zeta电位。包封的DMOMP为167-250 nm，具有均匀的球形和均匀的形态，并且包封率> 90%。包封DMOMP的缓慢释放模式，特别是在酸性溶液中，观察到超过7天。DMCNP的zeta电位约为8.80 mV，具有较高的稳定性。使用MTT测定的DMCNP（25-400 μg/mL）对Cos-7细胞的毒性研究显示在24-72小时内具有>90%活细胞的最小毒性。紫外可见（UV-vis）光谱表明封装DMOMP保护壳聚糖，而琼脂糖凝胶电泳验证其保护酶降解。MOMP蛋白在DMCNP转染的Cos-7细胞中的表达通过Western印迹和免疫荧光显微镜证实。值得注意的是，肌内注射DMCNP的BALB/c小鼠证实了包封的DMOMP的递送，以及MOMP基因转录物在大腿肌肉和脾脏中的表达。我们的数据表明，在可生物降解的壳聚糖纳米颗粒中封装DMOMP赋予稳定性和保护免受酶消化，并增强DMOMP在体外和小鼠中的递送和表达。进一步研究了纳米胶囊化的DMCNP疫苗制剂对C.小鼠沙眼的研究是有必要的。
Chlamydia trachomatis is a bacterial sexually transmitted infection affecting millions of people worldwide. Previous vaccination attempts have employed the recombinant major outer membrane protein (MOMP) of C. trachomatis nonetheless, with limited success, perhaps, due to stability, degradation, and delivery issues. In this study we cloned C. trachomatis recombinant MOMP DNA (DMOMP) and encapsulated it in chitosan nanoparticles (DMCNP) using the complex coacervation technique. Physiochemical characterizations of DMCNP included transmission and scanning electron microcopy, Fourier transform infrared and ultraviolet-visible spectroscopy, and zeta potential. Encapsulated DMOMP was 167–250 nm, with a uniform spherical shape and homogenous morphology, and an encapsulation efficiency > 90%. A slow release pattern of encapsulated DMOMP, especially in acidic solution, was observed over 7 days. The zeta potential of DMCNP was ~8.80 mV, which indicated that it was highly stable. Toxicity studies of DMCNP (25–400 μg/mL) to Cos-7 cells using the MTT assay revealed minimal toxicity over 24–72 hours with >90% viable cells. Ultra-violet visible (UV-vis) spectra indicated encapsulated DMOMP protection by chitosan, whereas agarose gel electrophoresis verified its protection from enzymatic degradation. Expression of MOMP protein in DMCNP-transfected Cos-7 cells was demonstrated via Western blotting and immunofluorescence microscopy. Significantly, intramuscular injection of BALB/c mice with DMCNP confirmed the delivery of encapsulated DMOMP, and expression of the MOMP gene transcript in thigh muscles and spleens. Our data show that encapsulation of DMOMP in biodegradable chitosan nanoparticles imparts stability and protection from enzymatic digestion, and enhances delivery and expression of DMOMP in vitro and in mice. Further investigations of the nanoencapsulated DMCNP vaccine formulation against C. trachomatis in mice are warranted.