Effect of Chitosan (Chi)-C Terminal 30 Amino Acids of Clostridium Perfringens Enterotoxin (CPE30)-pVP1 Nanoparticles on Rats with Viral Myocarditis

Abstract

Coxsackie B3 virus (CVB3) is the most common pathogen of viral myocarditis (VMC), and it is necessary to study an efficient vaccine to prevent the VMC. In this research, chitosan (chi)-C-terminal 30 amino acid (CPE30) was prepared by chemical coupling, and then chi-CPE30-pcDNA3.1-VP1 plasmid (pVP) complex particles were formed by co-aggregation method. The biological characteristics of the chi-CPE30-pVP1 complex particles were analyzed. It was immunized into SD rats intranasally at different time points as a vaccine together with other by-products (such as chi-pVP1, chi-CPE30-pcDNA3.1, and chi-pcDNA3.1). 100 μg of plasmid was inoculated each time, with 4 times in total, and the specific antibody level and cellular immune response of all rats were detected. It was revealed that based on the coupling effect of ethylcarbodiimide hydrochloride and N-hydroxysuccinimide (EDC/NHS) chemical coupling reagent, nearly 70% of CPE30 was coupled to chi, and the efficiency of chi-CPE30 to wrap DNA was close to 100%. After a certain concentration of pVP1 solution was added, the chi-CPE30-pVP1 composite particles were obtained, and the surface of the chi-CPE30-pVP1 composite was scanned as spherical particles. When used as a vaccine, the composite particles can induce high serum immunoglobulin G (IgG) and mucosal IgA antibody levels in rats. Meantime, the specific lymphocyte proliferation test confirmed that chi-CPE30-pVP1 effectively induced the proliferative response of CVB specific lymphocytes in the spleen and mesenteric lymph nodes (MLN). After the rats were infected with 3LD50CVB3, it was found that the weight of rats changed slightly under the action of chi-CPE30-pVP1 vaccine (P < 0.05). The creatine kinase and creatine kinase-myoglobin binding (CK-MB) levels of rats in this group were lower than those of chi-pVP1 rats and control group (P < 0.05). Applying the prepared chi-CPE30-pVP1 vaccine to immunize rats in this research could provide a new immune method for the molecular design of new vaccines and the prevention and treatment of CVB3 infection.