|SANG, YOUNG-MING - Kansas State University|
|BLECHA, FRANK - Kansas State University|
Submitted to: North American Porcine Reproductive and Respiratory Syndrome (NA-PRRS) Symposium
Publication Type: Abstract Only
Publication Acceptance Date: 9/1/2015
Publication Date: N/A
Technical Abstract: Activation statuses of monocytic cells including monocytes, macrophages and dendritic cells (DCs) are critically important for antiviral immunity. In particular, some devastating viruses, including porcine reproductive and respiratory syndrome virus (PRRSV), are capable of directly infecting these cells to subvert host immunity. We have recently profiled signature genes and gene responsive pathways in macrophages at different activation statuses, and reported that macrophage polarization is crucial for antiviral regulation. Monocyte-derived DCs (mDCs) are major target cells in PRRSV pathogenesis; however, the plasticity of mDCs in response to activation stimuli and PRRSV infection remains unstudied. In this study, we polarized mDCs using the framework established in macrophages, and applied genome-wide transcriptomic analysis to compare signature genes involved in mDCs activation and response to PRRSV infection. Our long-term goal is to integrate activation status with antiviral responses in these cells and to functionally modulate them for a prototypic cellular adjuvant/vaccine that is ideal for potentiating antiviral immunity. Porcine mDCs were polarized with mediators for 30 hours, then mock-infected, infected with PRRSV strain VR2332, or highly pathogenic strain (HP-PRRSV), for 5 h. Total RNA was extracted from the pooled cells of four replicates, and used to construct sequencing libraries for RNA-Seq procedures previously optimized. Comparisons were made between each polarized and unpolarized groups (i.e. mediator vs. PBS), and between PRRSV-infected and uninfected cells stimulated with the same mediator. The overall similarity between samples was assessed in heat map plots calculating the Euclidean distance between regularized log transformed data to allow equal contribution from all genes. Principal component analysis, Poisson distance and DESeq2 dispersion estimates emphasized variations in comparisons. Clustering of samples was by virus strain and then by mediator. We then asked which genes showed the most variability across all treatments as these are likely to be the genes that will provide resolution for clustering the samples. Many of the genes showing the most variability were related to cellular structure and innate immune response. The magnitude of differentially expressed gene profiles detected in HP-PRRSV rJXwn06 infected mDCs as compared to VR-2332 infected mDCs was consistent with the increased pathogenicity of the HP-PRRSV in vivo.