Vaccine-induced innate immune responses and examination of exosomes in Marek's disease virus (MDV) pathogenesis and vaccination
Date
2018
Journal Title
Journal ISSN
Volume Title
Publisher
University of Delaware
Abstract
Marek’s disease (MD) is a T cell lymphoproliferative disease of chickens caused by an alphaherpesvirus known as Marek’s Disease Virus (MDV). MD is the first neoplastic lymphoma to be successfully prevented by vaccination. MD vaccines are not sterilizing and do not limit pathogenic MDV1 transmission, but reduce early viremia and prevent tumor formation. MD vaccines are administered either via in ovo or sub-cutaneous inoculation on embryonic day (E18) and 1-day post-hatch, respectively. Currently in the US, commercial broilers are mass vaccinated in ovo with bivalent HVT (FC126) and SB-1 vaccine, whereas breeders and layers are vaccinated with either CVI988 alone, or in combination with HVT, in order to induce long term protection. Bivalent HVT/SB-1 was introduced during early 1980’s following the emergence of very virulent (vv) strains in response to near ubiquitous use of HVT as the vaccine of choice. Supplementing SB-1 to HVT was found to confer protective synergism and reduce MD condemnation rates by an average of 78% compared to HVT alone (1, 2). Despite being in use since early 1980s, the mechanisms of vaccine protection, in particular the synergistic protection conferred by bivalent HVT/SB-1 combination, still remains obscure. Furthermore, to broaden the protection provided by MD vaccines to other avian pathogens, HVT has been employed as a vector to carry immunogenic antigens belonging to a range of avian viral pathogens (e.g. IBDV VP2, NDV –HN or –F, ILTV gB). The bivalent vaccine is composed of viruses that are genomically distinct compared to CVI988 (attenuated MDV1) yet induces a high level of protection against a very virulent challenge. ☐ Putative mechanisms of vaccine-induced protection include replication in MDV1 target cells to stimulate innate immune responses during the first few days post-vaccination. Rapid induction of non-specific innate immune signaling is sufficient to polarize the immune system towards TH1 phenotype, leading to a CD8+ CTL response, as opposed to TH2 and TREG polarization seen during MDV1 infection or lymphoma formation. In line with this, early post vaccination, induction of Type I IFNs and Interferon stimulated genes (ISGs), but not pro-inflammatory cytokines was found to be a critical mechanism for MD vaccine mediated protection. ☐ Innate immune stimulation of Antigen Presenting Cells (APCs, such as macrophages, B-cells, and dendritic cells) and CD8+ CTL responses towards shared epitopes (structural glycoproteins, capsid, and tegument proteins) are implicated in protective immune responses, whereas antibodies play a limited role in protection, given the cell-associated nature of MDV1. Presumably, MD vaccines elicit continual anti-viral CD8+ CTL responses via a low-level of replication, a limited-establishment of latency, and periodic reactivation. We hypothesize that systemic TH1 patterning by MD vaccines is mediated by exosomes secreted by APCs and present in the serum of protected chickens. Antigenic peptide-complexed MHC-I+ and MHC-II+ exosomes, secreted by APCs, may function by direct or indirect antigen presentation to naïve CD8+ and CD4+ T cells, respectively, resulting in the generation of effector CD8+ CTLs and CD4+ TH1 cells. In line with our hypothesis, we identified a greater abundance of tumor suppressor-targeting miRNAs and lower-abundance MDV-1 miRNAs in serum exosomes derived from CVI-988 vaccinated and protected chickens. In addition, transcripts corresponding to structural glycoproteins, tegument proteins, and immediate-early transactivator ICP4 appeared to be transferred via exosomes in order to mediate CD8+ CTL responses. Furthermore, peptides corresponding to ICP4 were found in the exosomes suggesting a role played by the exosomal transfer of ICP4 in mediating CD8+ CTL responses. ☐ In contrast, we hypothesized that exosomes secreted by latently-infected and transformed CD4+ TREG-like cells in the serum of infected chickens are immune suppressive towards APCs, and CD8+ CTLs, resulting in a permanent phase of immune suppression. Moreover, we hypothesized that lymphoma-derived exosomes contribute to disease progression in the form of profound immune suppression, increased tumor metastasis, and overall tumor load. Serum exosomes derived from MDV-inoculated or contact-exposed, unvaccinated birds displaying MD lesions carried oncogenic miRNAs (OncomiRs) and MDV-1 miRNAs at a greater abundance. Furthermore, peptides corresponding to latent protein UL36 major tegument protein and UL47 tegument protein were found and suggest potential immune suppressive functions of these exosomes. ☐ Finally, exosomal miRNA biomarkers associated with MD vaccine-mediated protection and immunosuppression, respectively include mir-146b and mir-21, whereas exosomal protein biomarkers associated with MD-vaccine mediated protection versus pathogenesis include COL22A1, IGFBP5, and pantetheinase respectively.