Oral Presentation International Veterinary Immunology Symposium 2016

Comparative analysis of signature genes in PRRSV-infected porcine monocyte-derived dendritic cells at differential activation statuses. (#39)

Laura C Miller 1 , Yongming Sang 2 , Darrell O Bayles 1 , Frank Blecha 2
  1. USDA Agricultural Research Service - National Animal Disease Center, Ames, IOWA, United States
  2. Departments of Anatomy and Physiology, College of Veterinary Medicine, Kansas State University, Manhattan, Kansas, USA

The gene response pathways revealed in monocytic cells will serve as a functional framework to understand virus-host interaction in pigs. We have recently profiled signature genes and gene responsive pathways in MΦs at different activation statuses, and reported that MΦ 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 MΦs, and applied genome-wide transcriptomic analysis to compare signature genes involved in mDCs activation and response to PRRSV infection. 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. 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.