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Effects of different cytokines on immune responses of rainbow trout in a virus DNA vaccination model.

ABSTRACT: Seven rainbow trout cytokine genes (interleukin (IL)-2, IL-8, IL-15, IL-17, IL-1?, intracellular interferon (iIFN) 1a, and IFN-?2) were evaluated for their adjuvant effects on a DNA vaccine, called pG, containing the glycoprotein gene of infectious hematopoietic necrosis virus (IHNV). Distinct DNA constructs in expression plasmid pcDNA3.1 encoding a cytokine gene were generated. Immunofluorescence assays in rainbow trout gonadal cells demonstrated successful protein expression from all these constructs. Subsequently, fish were immunized with pG alone or together with a cytokine expression plasmid. Results showed that each cytokine plasmids at an appropriate dose showed notable effects on immune gene expression. IL-17 and IFN-?2 can enhance early specific IgM response. All cytokines, except IL-8, can benefit initial neutralizing antibody (NAb) titers. At 35 days post immunization (dpi), NAb titers of fish immunized with pG and IL-2, iIFN1a, or IFN-?2 plasmids remained at high levels (1:160). NAb titers of fish immunized with pG alone decreased to 1:40. IL-8 or IL-1? can enhance antigen-specific proliferative T-cell responses at 14 dpi. At 28 dpi, coinjection of pG with IL-2, IL-8, IL-15, or IL-17 plasmids induced considerably stronger lymphocyte proliferation than that with injection of pG alone. All cytokine plasmids delivered with pG plasmid enhanced protection of trout against IHNV-mediated mortality. These results indicate that the type and dose of trout cytokine genes injected into fish affect quality of immune response to DNA vaccination.

SUBMITTER: Cao Y

PROVIDER: S-EPMC5762505 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

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Effects of different cytokines on immune responses of rainbow trout in a virus DNA vaccination model.

Cao Yongsheng Y Zhang Qiya Q Xu Liming L Li Shaowu S Wang Di D Zhao Jingzhuang J Liu Hongbai H Feng Jian J Lu Tongyan T

Oncotarget 20171211 68

Seven rainbow trout cytokine genes (interleukin (IL)-2, IL-8, IL-15, IL-17, IL-1β, intracellular interferon (iIFN) 1a, and IFN-γ2) were evaluated for their adjuvant effects on a DNA vaccine, called pG, containing the glycoprotein gene of infectious hematopoietic necrosis virus (IHNV). Distinct DNA constructs in expression plasmid pcDNA3.1 encoding a cytokine gene were generated. Immunofluorescence assays in rainbow trout gonadal cells demonstrated successful protein expression from all these con ...[more]

PMID: 29348820

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Project description:Viruses across genome types produce long dsRNA molecules during replication [viral (v-) dsRNA]. dsRNA is a potent signaling molecule and inducer of type I interferon (IFN), leading to the production of interferon-stimulated genes (ISGs), and a protective antiviral state within the cell. Research on dsRNA-induced immune responses has relied heavily on a commercially available, and biologically irrelevant dsRNA, polyinosinic:polycytidylic acid (poly I:C). Alternatively, dsRNA can be produced by in vitro transcription (ivt-) dsRNA, with a defined sequence and length. We hypothesized that ivt-dsRNA, containing legitimate viral sequence and length, would be a more appropriate proxy for v-dsRNA, compared with poly I:C. This is the first study to investigate the effects of v-dsRNA on the innate antiviral response and to compare v-dsRNA to ivt-dsRNA-induced responses in fish cells, specifically rainbow trout. Previously, class A scavenger receptors (SR-As) were found to be surface receptors for poly I:C in rainbow trout cells. In this study, ivt-dsRNA binding was blocked by poly I:C and v-dsRNA, as well as SR-A competitive ligands, suggesting all three dsRNA molecules are recognized by SR-As. Downstream innate antiviral effects were determined by measuring IFN and ISG transcript levels using qRT-PCR and antiviral assays. Similar to what has been shown previously with ivt-dsRNA, v-dsRNA was able to induce IFN and ISG transcript production between 3 and 24?h, and its effects were length dependent (i.e., longer v-dsRNA produced a stronger response). Interestingly, when v-dsRNA and ivt-dsRNA were length and sequence matched both molecules induced statistically similar IFN and ISG transcript levels, which resulted in similar antiviral states against two aquatic viruses. To pursue sequence effects further, three ivt-dsRNA molecules of the same length but different sequences (including host and viral sequences) were tested for their ability to induce IFN/ISG transcripts and an antiviral state. All three induced responses similarly. This study is the first of its kind to look at the effects v-dsRNA in fish cells as well as to compare ivt-dsRNA to v-dsRNA, and suggests that ivt-dsRNA may be a good surrogate for v-dsRNA in the study of dsRNA-induced responses and potential future antiviral therapies.

Project description:The Infectious Hematopoietic Necrosic Virus (IHNV) DNA vaccine is based on the viral glycoprotein gene (G gene) and induces a non-specific anti-viral immune response and long-term specific immunity against IHNV in salmonid fishes. This study characterized gene expression responses associated with the early anti-viral response. Homozygous rainbow trout were injected intra-muscularly (I.M.) with vector DNA or the IHNV DNA vaccine. Gene expression at the I.M. site was evaluated using a 16,000 feature salmon cDNA microarray. Eighty different transcripts were significantly modulated in the vector DNA group while 910 transcripts were modulated in the IHNV DNA vaccinated group relative to control group. Quantitative reverse transcriptase PCR was used to examine expression of selected genes at the I.M. site and in other secondary tissues. In the localized response (I.M. site), the magnitudes of gene expression changes were higher in the vaccinate group relative to the vector DNA group for the majority of genes analyzed. At secondary systemic sites (e.g. gill, kidney and spleen) evaluated with RT-PCR of specific genes, the main difference was the up-regulation of the type I IFN-related genes in only the IHNV DNA vaccinated group. Keywords: disease vaccine response Gene expression at the site of I.M injection was evaluated 7 days post-injection in clonal rainbow trout. Gene expression was compared across three treatments, with replicate clonal rainbow trout individuals (Hot Creek strain) within each treatment. Treatments included injection with 1X PBS (control group), plasmid vector only (without IHNV G gene; hereafter referred to as vector group), and the pIHNV vaccine (vaccine group). Three clonal individuals from the control, three individuals from the vector group, and four individuals from the vaccine group were labeled with Cy3 and used for individual hybridizations to ten slides, comparing hybridization of each individual to a major reference RNA pool. The major reference pool consisted of three unhandled clonal rainbow trout and constituted a pool of RNA from the anterior kidney, spleen, liver and muscle and was labelled with Cy5. Total RNA was extracted from samples, amplified, and hybridized to 16,000 feature Atlantic salmon cDNA arrays developed by the Genomic Research for Atlantic Salmon project. Differential expression was compared among control, vector, and vaccine groups.

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Effects of different cytokines on immune responses of rainbow trout in a virus DNA vaccination model.

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Effects of different cytokines on immune responses of rainbow trout in a virus DNA vaccination model.

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