Characterization of Co-Stimulatory Ligand CD80/86 and Its Effect as a Molecular Adjuvant on DNA Vaccine Against Vibrio anguillarum in Flounder (Paralichthys olivaceus)

Abstract

The CD80/86 molecule is one of the important co-stimulatory ligands and involves antigen-specific immune responses by ligating with CD28 and then delivering the required second signal to T-cell activation. In this study, a CD80/86 homolog was identified, and its expression characteristics were studied in flounder (Paralichthys olivaceus). The open reading frame (ORF) of CD80/86 is 906 bp, encoding 301 aa, and the extracellular amino acid sequence encoded two IgV- and IgC-like structural domains; fCD80/86 is highly expressed in head kidney, peripheral blood leukocytes (PBLs), and spleen, and has relatively high expression in muscle. Antibodies specific for CD80/86 were produced, and CD80/86 was colocalized with MHCII+, CD40+, and CD83+ leukocytes but not with IgM+, CD3+, or CD4+ lymphocytes. The cloned CD80/86 in flounder shares conserved structural features with its mammalian counterparts and is mainly distributed on antigen-presenting cells. Based on these data, CD80/86 as an adjuvant to enhance the immune response of DNA vaccine was investigated. A bicistronic DNA vaccine expressing both CD80/86 and the outer membrane protein (OmpK) of Vibrio anguillarum (p-OmpK-CD80/86) was successfully constructed. After immunization, p-OmpK-CD80/86 could induce the upregulation of the proportion of IgM+ and CD4+ cells in flounder, compared to the p-OmpK- or p-CD80/86-immunized group; CD28 genes were significantly induced in the p-CD80/86 and p-OmpK-CD80/86 groups. Compared to the p-OmpK group, the higher expression of CD83, MHCI, CD4, CD8, and IL-2 was detected at the injection site. The relative percent survival (RPS) produced by p-OmpK-CD80/86 is 66.11% following the V. anguillarum challenge, while the RPS of p-OmpK or p-CD80/86 is 46.30% and 5.56%, respectively. The results revealed that CD80/86 is mainly found in antigen-presenting cells, and could help elicit humoral immune responses in teleost through the CD80/86-CD28 signaling pathway involving CD4+ lymphocytes.

Bestatin, A Pluripotent Immunomodulatory Small Molecule, Drives Robust and Long-Lasting Immune Responses as an Adjuvant in Viral Vaccines

“…For inactivated vaccines, adjuvants are required to enhance the delivery of vaccine antigens and stimulate PRRs that are difficult to induce with antigens alone [39]. Therefore, research aimed at identifying and novel and powerful adjuvants such as cytokines [40], co-stimulator molecules [41], and other immunostimulators such as PRR ligands (agonists) [42] that enhance the efficacy of FMD vaccines is ongoing.…”

Characterization of myeloperoxidase and its contribution to antimicrobial effect on extracellular traps in flounder (Paralichthys olivaceus)

“…Other markers used to differentiate different types of cells are: Zap-70 (Zeta-chain-associated protein kinase 70) a phylogenetically conserved protein used to visualize the presence of both T and NK lymphocytes ( 33 ). MHCII is a heterodimer preferentially expressed in macrophages, B cells, and dendritic cells (DCs), while slightly expressed in monocytes and immature DCs ( 31 , 52 ). GCSFR is a class I cytokine receptor superfamily member and expressed on hematopoietic cells such as monocytes, as well as some non-hematopoietic cells, neutrophils, and their precursors ( 32 , 53 ).…”

“…The peripheral blood leukocytes (PBLs), head kidney leukocytes (HKLs) and peritoneal cells (PerCs) were obtained from healthy or LPS (1mg mL -1 ) -stimulated flounder (i.p injection of LPS). After being adjusted to 5 × 10 6 cells mL -1 in PBS, the potentially unspecific staining was reduced by adding 5% BSA and incubated with antibodies (rabbit anti- Po MHCII or anti- Po GCSFR Abs) for 1 hour at room temperature (BD Cytofix/Cytoperm kit; BD Biosciences) ( 31 , 32 ). Afterwards, the cells were incubated with Abs [mixture of mouse anti-r Po MPO and rabbit anti-Zap-70 (Cell Signal Technology, Danvers, Massachusetts, USA)] for 1 hour at room temperature ( 33 ).…”

Functional Nucleic Acid-Based Immunomodulation for T Cell-Mediated Cancer Therapy

“…) − and costimulatory molecules (CD80, CD86, CD40L, etc. ), − have been developed and showed potential for cancer treatments. On the other hand, a majority of DNA vaccines faced challenges regarding their clinical performance and safety, thereby offering various opportunities for the development of other vaccine formulas …”