Effects of Th17 cytokines on microglial responses and the blood-brain barrier following recurrent intranasal Group A Streptococcus infections

Peripheral infections can result in neuropsychiatric changes in many contexts, including after recurrent Group A Streptococcus (GAS) infections in children. To explore the underlying mechanisms, here we used an intranasal inoculation model to analyze more than 100,000 cells from the mouse olfactory bulb (OB) and nasal lymphoid tissue by single-cell RNA sequencing (scRNAseq). Upon repeated GAS inoculations, endothelial cells responded by downregulating blood-brain barrier (BBB)-specific genes, and microglia upregulated interferon-response, chemokine and antigen-presentation genes. To determine whether specific Th17 cytokines play distinct roles in producing this phenotype, we administered a neutralizing antibody against interleukin 17A (IL-17A), which decreased microglial expression of interferon-response and chemokine genes, but unexpectedly exacerbated levels of antigen presentation markers. Another Th17 cytokine, granulocyte-macrophage colony-stimulating factor (GM-CSF) rescued a different subset of microglial transcripts. Olfactory bulbs were isolated from P60 mice following five repeated inoculations with either a suspension of GAS or with vehicle (PBS), and profiled at 18 hours post-inoculation. After perfusion with cold PBS, tissue was finely chopped, then processed with a papain-based neural dissociation kit to create a single-cell suspension. Following bead-based myelin removal, cells were stained for CD31, CD11b and live/dead and collected by FACS. For analysis of nasal lymphoid tissue, the nasal-associated lymphoid tissue (NALT) and olfactory epithelium were dissected and combined for analysis.