MyD88-dependent influx of monocytes and neutrophils impairs lymph node B cell responses to chikungunya virus infection via Irf5, Nos2 and Nox2

McCarthy, Mary K.; Reynoso, Glennys V.; Winkler, Emma S.; Mack, Matthias; Diamond, Michael S.; Hickman, Heather D.; Morrison, Thomas E.

https://doi.org/10.1371/journal.ppat.1008292.g003 with attenuated CHIKV (Fig 6A) . However, total cell numbers in the dLN were partially restored in Nos2 -/and Nox2 -/mice inoculated with pathogenic CHIKV compared with WT mice (Fig 6A) , suggesting that NO and superoxide contribute to reduced lymphocyte numbers in the dLN during pathogenic CHIKV infection. In addition to partial restoration of lymphocyte numbers, the dLNs of Nos2 -/and Nox2 -/mice infected with pathogenic CHIKV showed improved follicular and paracortical organization, and a more clearly defined B-T cell border compared with the dLN of WT mice (Fig 6B) . These data suggest that both Nos2 and Nox2 contribute to the disruption of lymphocyte organization that occurs during pathogenic CHIKV infection. The partial restoration of cell numbers and markedly improved lymphocyte organization in the dLN were not due to a difference in early monocyte and neutrophil infiltration or localization at 24 hpi between WT, Nos2 -/-, and Nox2 -/mice (Fig 6C-6F) . To define the iNOSexpressing cell type(s) that mediate the Nos2-dependent effects on dLN cell numbers and organization, we analyzed cell populations in the dLN at 24 hpi by flow cytometry. Monocytes elicited by pathogenic CHIKV infection expressed iNOS in the dLN, but not in the blood (Fig 6G and 6H) . iNOS staining was absent in neutrophils in the dLN (Fig 6G) , and was absent in Nos2 -/mice, as expected (Fig 6G) . Together, these data indicate that expression of Nos2 and Nox2 contribute to impaired lymphocyte accumulation and dLN disorganization, and that monocytes upregulate iNOS expression following entry into the dLN.