BACKGROUND Helicobacter pylori (H. pylori) infect more than half of the world population, and they cause different serious diseases such as gastric carcinomas. This study aims to design a vaccine on the basis of cagW against H. pylori infection. After pcDNA3.1 (+)-cagW-CS-NPs complex is produced, it will be administered into the muscles of healthy BALB/c mice in order to study the effect of this DNA vaccine on the interleukin status of mice, representing its effect on the immune system. After that, the results will be compared with the control groups comprising the administration of cagW-pCDNA3.1 (+) vaccine, the administration of chitosan and the administration of PBS in the muscles of mice. METHODS The cagW gene of H. pylori was amplified by employing PCR, whose product was then cloned into the pcDNA3.1 (+) vector, and this cloning was confirmed by PCR and BamHI/EcoRV restriction enzyme digestion. CagW gene DNA vaccine was encapsulated in chitosan nanoparticles (pcDNA3.1 (+)-cagW-CS-NPs) using a complex coacervation method. The stability and in vitro expression of chitosan nanoparticles were studied by DNase I digestion and transfection, and the immune responses elicited in specific pathogen-free (SPF) mice by the pcDNA3.1 (+)-cagW-CS-NPs were evaluated. Apart from that, the protective potential pcDNA3.1 (+)-cagW-CS-NPs was evaluated by challenging with H. pylori. RESULTS The pcDNA3.1 (+)-cagW-CS-NPs comprises cagW gene of H. pylori that is encapsulated in chitosan nanoparticles, produced with good morphology, high stability, a mean diameter of 117.7 nm, and a zeta potential of + 5.64 mV. Moreover, it was confirmed that chitosan encapsulation protects the DNA plasmid from DNase I digestion, and the immunofluorescence assay showed that the cagW gene could express in HDF cells and maintain good bioactivity at the same time. In comparison to the mice immunized with the control plasmid, in vivo immunization revealed that mice immunized with pcDNA3.1 (+)-cagW-NPs showed better immune responses and prolonged release of the plasmid DNA. CONCLUSIONS This research proves chitosan-DNA nanoparticles as potent immunization candidates against H. pylori infection and paves the way for further developments in novel vaccines encapsulated in chitosan nanoparticles.

中文翻译：

---

基于cagW的基因疫苗对BALB / c小鼠免疫学特性的影响：幽门螺杆菌DNA疫苗的有效候选者。

背景技术幽门螺杆菌（H.pylori）感染世界一半以上的人口，它们引起不同的严重疾病，例如胃癌。这项研究旨在基于cagW设计针对幽门螺杆菌感染的疫苗。产生pcDNA3.1（+）-cagW-CS-NPs复合物后，将其施用到健康的BALB / c小鼠的肌肉中，以研究该DNA疫苗对小鼠白介素状态的影响，以代表其作用在免疫系统上。之后，将结果与对照组进行比较，这些对照组包括在小鼠肌肉中施用cagW-pCDNA3.1（+）疫苗，壳聚糖的施用和PBS的施用。方法采用PCR扩增幽门螺杆菌cagW基因，并将其产物克隆到pcDNA3.1（+）载体中，并通过PCR和BamHI / EcoRV限制酶消化证实了该克隆。使用复杂凝聚法将CagW基因DNA疫苗封装在壳聚糖纳米颗粒（pcDNA3.1（+）-cagW-CS-NPs）中。通过DNase I消化和转染研究了壳聚糖纳米粒的稳定性和体外表达，并评估了由pcDNA3.1（+）-cagW-CS-NPs在特定的无病原体（SPF）小鼠中引起的免疫应答。除此之外，通过用幽门螺杆菌攻击来评估保护性潜力pcDNA3.1（+）-cagW-CS-NPs。结果pcDNA3.1（+）-cagW-CS-NPs包含幽门螺杆菌的cagW基因，该基因封装在壳聚糖纳米颗粒中，具有良好的形态，高稳定性，平均直径为117.7 nm，ζ电位为+ 5.64。毫伏 此外，证实了壳聚糖的封装保护了DNA质粒免于DNase I消化，并且免疫荧光分析表明cagW基因可以在HDF细胞中表达并同时保持良好的生物活性。与用对照质粒免疫的小鼠相比，体内免疫显示，用pcDNA3.1（+）-cagW-NP免疫的小鼠表现出更好的免疫反应和质粒DNA的延长释放。结论这项研究证明壳聚糖DNA纳米颗粒是针对幽门螺杆菌感染的有效免疫候选物，为进一步封装在壳聚糖纳米颗粒中的新型疫苗铺平了道路。与用对照质粒免疫的小鼠相比，体内免疫显示，用pcDNA3.1（+）-cagW-NP免疫的小鼠表现出更好的免疫反应和质粒DNA的延长释放。结论这项研究证明壳聚糖DNA纳米颗粒是针对幽门螺杆菌感染的有效免疫候选物，为进一步封装在壳聚糖纳米颗粒中的新型疫苗铺平了道路。与用对照质粒免疫的小鼠相比，体内免疫显示，用pcDNA3.1（+）-cagW-NP免疫的小鼠表现出更好的免疫反应和质粒DNA的延长释放。结论这项研究证明壳聚糖DNA纳米颗粒是针对幽门螺杆菌感染的有效免疫候选物，为进一步封装在壳聚糖纳米颗粒中的新型疫苗铺平了道路。