Rainbow Trout Erythrocytes ex vivo Transfection With a DNA Vaccine Encoding VHSV Glycoprotein G Induces an Antiviral Immune Response

Abstract

Fish red blood cells (RBCs), are integral in several biologic processes relevant to immunity, such as pathogen recognition, pathogen binding and clearance, and production of effector molecules and cytokines. So far, one of the best strategies to control and prevent viral diseases in aquaculture is DNA immunization. DNA vaccines (based on the rhabdoviral glycoprotein G [gpG] gene) have been shown to be effective against fish rhabdoviruses. However, more knowledge about the immune response triggered by DNA immunization is necessary to develop novel and more effective strategies. In this study, we investigated the role of fish RBCs in immune responses induced by DNA vaccines. We show for the first time that rainbow trout RBCs express gpG of viral hemorrhagic septicaemia virus (VHSV) (GVHSV) when transfected with the DNA vaccine ex vivo and modulate the expression of immune genes and proteins. Functional network analysis of transcriptome profiling of RBCs expressing GVHSV revealed changes in gene expression related to G-protein coupled receptor (GPCR)-downstream signaling, complement activation, and RAR related orphan receptor α (RORA). Proteomic profile functional network analysis of GVHSV-transfected RBCs revealed proteins involved in the detoxification of reactive oxygen species, interferon-stimulated gene 15 (ISG15) antiviral mechanisms, antigen presentation of exogenous peptides, and the proteasome. Conditioned medium of GVHSV-transfected RBCs conferred antiviral protection and induced ifn1 and mx gene expression in RTG-2 cells infected with VHSV. In summary, rainbow trout nucleated RBCs could be actively participating in the regulation of the fish immune response to GVHSV DNA vaccine, and thus may represent a possible carrier cells for the development of new vaccine approaches.

Immune Functions of Erythrocytes in Osteichthyes

“…Expression and antiviral activity of the IFIT5, observed by Chico et al 4, may not depend on interferon type I production, which seems to be of special interest. A similar lack of dependence, increased expression of antiviral genes, ISG15 (interferon-stimulated gene 15), mx, pkr (RNA-activated protein kinase, also known as protein kinase R, PKR), and ifit5, with decreased IFN expression, have been demonstrated in the studies of rainbow trout (O. mykiss) erythrocytes exposed to glycoprotein G of the VHSV (26).…”

“…Owing to the demonstrated scope and type of RBCs' immune activity in Osteichthyes, they are currently considered mediators of immune cells ( 21 , 25 ). RBCs of Osteichthyes are capable of responding to viral infections ( 1 , 4 , 6 , 9 , 10 , 16 , 17 , 25 ), producing cytokines ( 2 , 5 , 26 ) and endocytosis of nanoparticles and viral pathogens ( 11 , 15 ).…”

Fish Red Blood Cells Modulate Immune Genes in Response to Bacterial Inclusion Bodies Made of TNFα and a G-VHSV Fragment

“…Therefore, TNFα treatment augments or blocks MHC class II induction depending on the cell type and cellular differentiation state (35). mhcII and cd83 gene expression has been previously reported for rainbow trout RBCs (3, 36) and chicken RBCs (37). However, this is the first report that shows the regulation of cd83 and mhcII gene transcripts in response to an immunostimulant.…”

Rainbow Trout Red Blood Cells Exposed to Viral Hemorrhagic Septicemia Virus Up-Regulate Antigen-Processing Mechanisms and MHC I&II, CD86, and CD83 Antigen-presenting Cell Markers

“…Given that RBCs are the most abundant cell type in the blood, this new knowledge will shed light on the design of novel vaccine targets. Potential applications of these results could imply that RBCs, which can be transfected and induce immune gene expression [32], are target of new strategies for vaccination.…”

“…Expression of MHC molecules indicates that a cell can play an APC role. MHC class I (MHCI) protein expression has been detected in rainbow trout RBCs [29,30] and MHC class II (MHCII) transcriptional expression has been recently reported in nucleated rainbow trout [31,32] and chicken [33] RBCs. However, the role of RBCs in viral antigen presentation is unknown.…”