Intramuscular Immunization with Chemokine-Adjuvanted Inactive Porcine Epidemic Diarrhea Virus Induces Substantial Protection in Pigs

Abstract

Intramuscular (IM) immunization is generally considered incapable of generating a protective mucosal immune response. In the swine industry, attempts to develop a safe and protective vaccine for controlling porcine epidemic diarrhea (PED) via an IM route of administration have been unsuccessful. In the present study, porcine chemokine ligand proteins CCL25, 27, and 28 were constructed and stably expressed in the mammalian expression system. IM co-administration of inactivated PEDV (iPEDV) particles with different CC chemokines and Freund's adjuvants resulted in recruiting CCR9+ and/or CCR10+ inflammatory cells to the injection site, thereby inducing superior systemic PEDV specific IgG, fecal IgA, and viral neutralizing antibodies in pigs. Moreover, pigs immunized with iPEDV in combination with CCL25 and CCL28 elicited substantial protection against a virulent PEDV challenge. We show that the porcine CC chemokines could be novel adjuvants for developing IM vaccines for modulating mucosal immune responses against mucosal transmissible pathogens in pigs.Vaccines 2020, 8, 102 2 of 16 immunization region to mucosal sites [25][26][27][28][29][30]. On the other hand, the CCL25/TECK expressed by mucosal epithelial cells can chemo-attract CCR9+ cells and support CCR9 expression leukocytes to migrate to the small intestine [30,31]. The application of CCL27 and CCL25 as an adjuvant in DNA vaccines was proven to enhance systemic and/or mucosal immune responses following IM injection in mice or macaques [32,33]. Therefore, the incorporation of these CC chemokines as immune trafficking signals could be a potential strategy for the development of effective parenteral PEDV vaccine regimens.In the present study, three porcine CCL proteins, namely, CCL27, CCL28, and CCL25 were constructed and stably expressed in the mammalian cell expression system. Different combinations of these chemokines were intramuscularly co-administrated with inactivated PEDV (iPEDV) viral particles and Freund's adjuvant in pigs. Immunohistochemical (IHC) staining was performed to detect the infiltration of cognate chemokine receptors, CCR9+ or CCR10+, bearing immune cells, to the sites of immunization. The immunogenicity and protective efficiency of iPEDV adjuvanted with Freund's adjuvant in different combinations of CC chemokines were evaluated in 5-week-old pigs.

Parenterally Administered Porcine Epidemic Diarrhea Virus-Like Particle-Based Vaccine Formulated with CCL25/28 Chemokines Induces Systemic and Mucosal Immune Protectivity in Pigs

“…In addition, a milder clinical sign was also observed in pigs of the VLP+CCL group as compared to the VLP group. However, when compared to the animals immunized using the inactivated virus formulated with CCL25/28 in a previous study [ 46 ], pigs in the VLP+CCL group showed relatively less protection against the PEDV challenge. It might be due to the PEDV exposure age at 11 weeks old and failure to elicit the optimal immune response, as it can be observed that the mean PEDV S-specific IgG titers were relatively low, with mean S/P ratios of around 0.4 and 0.7 in the VLP and VLP-CCL groups, respectively.…”

“…In this study, PEDV VLPs were generated and characterized for developing a safe and potent immunogen against highly virulent strains in pigs. To induce effective protection via a convenient parenteral route, we incorporated CCL25 and CCL28, which have been shown to be effective in our previous study [ 46 ], as mucosal adjuvants in this strategy. Our results demonstrated that the regimen is capable of eliciting not only systemic PEDV S-specific IgG and IFN-γ-producing cells in PBMCs but also mucosal PEDV S-specific IgA.…”

“…The CC chemokines, CCL25 and CCL28, were prepared as described in a previous study [ 46 ]. The aqueous formulations comprised the immunogen with/without CC chemokines for different groups ( Table 1 ).…”

“…The clinical signs for each pig were monitored and recorded daily. Based on previous studies [ 22 , 46 ], the severity of diarrhea was graded as 0: normal consistency; 1: loose consistency; 2: semi-fluid consistency; 3: liquid consistency. The body weight (BW) of each pig was measured weekly.…”

“…Besides, the synergistic effects of CCL25 and CCL28 in trafficking IgA + cells into the intestines has been confirmed in mice [ 45 ]. Our previous work also proved that inactivated PEDV co-adjuvanted with porcine CCL25 and CCL28 is competent in enhancing mucosal and systemic antibody responses and protective efficacy in pigs [ 46 ]. Therefore, intramuscular injection of an immunogen in combination with CCL25 and CCL28 could be a potential strategy for developing vaccines against enteric diseases.…”