Effect of external cues on clock-driven time-of-day specific protection from Influenza A infection
Influenza and other respiratory viral pathogens are leading causes of mortality and morbidity. We have previously shown that circadian rhythms confer temporal protection from influenza infection. Here, we investigate whether the endogenous circadian clocks need optimal cycling of entraining cues to mediate this protection. We used two key entraining factors-- light-dark cycling and meal timing-- to determine their relative importance in allowing the host to benefit from its endogenous circadian rhythms. First, we demonstrate that disrupting environmental lighting cues, within a critical window of vulnerability, abrogated the time-of-day specific protection; this poor outcome is mediated by a dysregulated immune response evidenced by the accumulation of inflammatory monocytes and CD8+ cells in the lungs and transcriptomic profile with an exaggerated immune response. Importantly, rhythmic meal timing mitigated the adverse effects of disrupted light cycles. Our findings highlight the crucial interplay between peripheral and central clocks in shaping influenza outcomes, offering significant translational potential for improving the care of critically ill patients with respiratory viral infections. Overall design: Specific pathogen-free 8-12 week-old C57bl/6J mice  were placed in circadian cabinets on reverse light-dark cycles. After 4-6 weeks of acclimatization in these cabinets, mice were anesthetized lightly with isoflurane prior to being infected intranasally (i.n.) with mouse-adapted influenza virus (PR8 ). Given the use of circadian cabinets, mice were simultaneously infected at either ZT23 or ZT11 and maintained in 12 hr light-dark cycle. The dose chosen was likely to cause >50% mortality when mice were infected at ZT11. On day four p.i. a subset of ZT23-LD mice were moved to constant light until the end of the study. Lungs were collected at day 8 p.i., at the same ZT as the initial infection, and RNA was extracted.