Microglial hyperresponsiveness induced by respiratory viral infection and its effect on hippocampal transcriptome in preweaning piglets Peng Ji1, Kyle M. Schachtschneider2, Lawrence B. Schook2, Rodney W. Johnson2 1University of California Davis, CA; 2University of Illinois Urbana-Champaign, IL
Despite the high prevalence of respiratory viral infection in human infants, its impact on microglial function and neuronal development is still unclear. To study this, we artificially inoculated neonatal piglets at postnatal day 7 with either sterile saline solution or porcine reproductive and respiratory syndrome virus (PRRSV), which causes interstitial pneumonia. On day 21 post-inoculation, piglets were euthanized to collect brain tissues. We assessed global gene expression of hippocampus using Illumina RNA-sequencing. Microglial cells were identified for positive staining of Iba-1 in hippocampal tissue sections and analyzed for morphology. In a subsequent study with the same experimental design, we isolated microglial cells using CD11b antibody-conjugated MicroBeads. A portion of cells were subjected to immunophenotyping using flow cytometry for expression of CD45 and MHC-II, a marker of immune activation. To investigate microglial responsiveness to secondary immune stimuli, chemotaxis, phagocytosis, pro-inflammatory cytokine production and gene expression were analyzed in primary microglial cells challenged with LPS or Poly (I:C) in vitro. Infection altered expression of 455 genes, of which 334 were up-regulated and 121 were down-regulated. Genes associated with a spectrum of immune functions were unanimously upregulated in response to infection, whereas functional annotation revealed a set of differentially expressed genes involved in synapse as the most enriched cellular component. Respiratory viral infection induced microglial activation suggesting a neuroinflammatory response. The soma of microglial cells displayed bipolar rod-like morphology in response to PRRSV infection. Intriguingly, microglial cells isolated from infected piglets had exaggerated response to subsequent immune challenge in vitro in terms of phagocytosis and TNFα production. This was associated with enhanced expression TLR3 and TLR4 and decreased expression of BDNF. In conclusion, our study highlighted that early life respiratory viral infection cause neuroinflammation and renders microglial hyper-responsiveness to further insult.
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