interleukin-8 and Circoviridae-Infections

Dysfunction of endothelial cells and vascular system is one of the most important pathological changes of porcine circovirus disease (PCVD) caused by porcine circovirus type 2 (PCV2). PCV2-infected endothelial cells can upregulate the production of endothelial-derived IL-8, which can inhibit the maturation of dendritic cells. Endothelial-derived IL-8 has different structural and biological characteristics compared with monocyte-derived IL-8. However, the mechanism of endothelial-derived IL-8 production is still unclear.. Key molecules of RIG-I-like signaling pathway RIG-I, MDA-5, MAVS and a key molecule of JNK signaling pathway c-Jun in PCV2-infected porcine iliac artery endothelial cells (PIECs) were upregulated significantly detected with quantitative PCR, Western blot and fluorescence confocal microscopy, while no significant changes were found in NF-κB signaling pathway. Meanwhile, the expression of endothelial-derived IL-8 was downregulated after RIG-I, MDA-5, or MAVS genes in PIECs were knocked down and PIECs were treated by JNK inhibitor.. PCV2 can activate RIG-I/MDA-5/MAVS/JNK signaling pathway to induce the production of endothelial-derived IL-8 in PIECs, which provides an insight into the further study of endothelial dysfunction and vascular system disorder caused by PCV2.

Topics: Animals; Cells, Cultured; Circoviridae Infections; Circovirus; Endothelial Cells; Gene Knockdown Techniques; Iliac Artery; Interleukin-8; Signal Transduction; Swine; Swine Diseases

The efficiency of immune responses and host defense against pathogens largely depends on the function of dendritic cells (DCs). Porcine circovirus type 2 (PCV2) infection causes viremia and extensive modulation of immune activities in the blood. The objective of the present study was to investigate the effects of PCV2 infection in vivo on the immunological function of DCs induced from peripheral blood monocytes (MoDCs). At different points after infection with PCV2, peripheral blood monocytes from PCV2-infected pigs were used to induce differentiation of DCs in vitro. Flow cytometry and quantitative real-time reverse transcription PCR were conducted to detect mRNA expression of surface markers related to antigen presentation and inflammatory/immunosuppressive cytokines of the induced MoDCs. The ability of induced MoDCs to stimulate T cells was measured using an MTS assay. In the early phase of infection at 3 days post-inoculation (DPI), IL-10, IL-8 and MIP-1β in MoDCs were upregulated significantly. By the peak of virus proliferation at 7 DPI, antigen presentation molecules SLA-DR (MHC II) and CD80/86 together with cytokines IL-12 and IL-10 had decreased, accompanied by a rapid reduction of IL-8 and MIP-1β. The T cell stimulation index of induced MoDCs in PCV2 groups after different infection times declined to some extent, with a significant difference at 7 DPI. PCV2 infection in vivo functionally reduced the antigen presentation capability of induced MoDCs from peripheral blood and modified expression of inflammatory/immunosuppressive cytokines that may be related to PCV2-induced immunosuppression.

Topics: Animals; Antigen Presentation; Circoviridae Infections; Circovirus; Cytokines; Dendritic Cells; Inflammation; Interleukin-10; Interleukin-8; Lymphocyte Activation; Monocytes; Real-Time Polymerase Chain Reaction; Swine; T-Lymphocytes; Up-Regulation; Viral Load

Porcine circovirus type 2 (PCV2) is the essential infectious agent responsible for causing porcine circovirus-associated diseases in pigs. To date, eleven RNAs and five viral proteins of PCV2 have been detected. Here, we identified a novel viral gene within the PCV2 genome, termed ORF5, that exists at both the transcriptional and translational level during productive infection of PCV2 in porcine alveolar macrophages 3D4/2 (PAMs). Northern blot analysis was used to demonstrate that the ORF5 gene measures 180 bp in length and overlaps completely with ORF1 when read in the same direction. Site-directed mutagenesis was used to show that the ORF5 protein is not essential for PCV2 replication. To investigate the biological functions of the novel protein, we constructed a recombinant eukaryotic expression plasmid capable of expressing PCV2 ORF5. The results show that the GFP-tagged PCV2 ORF5 protein localizes to the endoplasmic reticulum (ER), is degraded via the proteasome, inhibits PAM growth and prolongs the S-phase of the cell cycle. Further studies show that the GFP-tagged PCV2 ORF5 protein induces ER stress and activates NF-κB, which was further confirmed by a significant upregulation in IL-6, IL-8 and COX-2 expression. In addition, five cellular proteins (GPNMB, CYP1A1, YWHAB, ZNF511 and SRSF3) were found to interact with ORF5 via yeast two-hybrid assay. These findings provide novel information on the identification and functional analysis of the PCV2 ORF5 protein and are likely to be of benefit in elucidating the molecular mechanisms of PCV2 pathogenicity. However, additional experiments are needed to validate the expression and function of the ORF5 protein during PCV2 infection in vitro before any definitive conclusion can be drawn.

Topics: Amino Acid Sequence; Animals; Blotting, Northern; Blotting, Western; Cell Cycle; Cell Proliferation; Cells, Cultured; Circoviridae Infections; Circovirus; Fluorescent Antibody Technique, Indirect; Green Fluorescent Proteins; Immunoenzyme Techniques; Interleukin-6; Interleukin-8; Macrophages, Alveolar; Molecular Sequence Data; NF-kappa B; Real-Time Polymerase Chain Reaction; Recombinant Fusion Proteins; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sequence Homology, Amino Acid; Swine; Swine Diseases; Two-Hybrid System Techniques; Viral Proteins

Monocyte chemoattractant protein-1 (MCP-1) but not interleukin-8 (IL-8) was detected by in situ hybridization using a nonradioactive digoxigenin-labeled complementary DNA probe in granulomatous lesions of lymph nodes from 20 pigs with naturally occurring postweaning multisystemic wasting syndrome (PMWS). Complementary DNA probes of 375 and 266 base pairs for MCP-1 and IL-8, respectively, were generated by reverse transcription-polymerase chain reaction. The 20 pigs with PMWS had distinct positive hybridization signals for MCP-1 but not for IL-8. The hybridization signals for MCP-1 were strictly confined to the cells with granulomatous lesions, including macrophages and multinucleated giant cells. A very close cell-to-cell correlation between MCP-1 and porcine circovirus 2 was seen in serial sections of lymph nodes. Results of this study indicate that MCP-1 expression may play a role in the pathogenesis of granulomatous inflammation in pigs with PMWS.

Topics: Animals; Chemokine CCL2; Circoviridae Infections; Circovirus; Granuloma; In Situ Hybridization; Interleukin-8; Lymph Nodes; Reverse Transcriptase Polymerase Chain Reaction; RNA, Viral; Swine; Swine Diseases; Wasting Syndrome