Graphic summary of the mechanisms underlying the effects of the combined therapy.

After pH1N1 virus infection, mTOR signaling is activated, which promotes NF-κB activity and ROS production, and leads to the activation of NLRP3 inflammasome. Two signals are needed for the activation of NLRP3 inflammasome. Signal one is associated with the activation of the transcription factor NF-κB, which upregulates the transcription and expression of NLRP3, ASC, caspase-1, pro-IL-1β, and pro-IL-18. Signal two is mediated by pH1N1 virus-induced ROS production. The two signals cooperatively mediate the assembly of NLRP3 inflammasome and the cleavage of pro-caspase-1 into active caspase-1. The active caspase-1 can further cleave pro-IL-1β and pro-IL-18 into activated IL-1β and IL-18, respectively, and induce their secretion out of the cells. The bioactive IL-1β and IL-18 produced by NLRP3 inflammasome promotes pH1N1-induced lung injury. Sirolimus inhibits the activation of NF-κB by targeting mTOR. This decreases the expression of NLRP3 inflammasome components, pro-IL-1β and pro-IL-18. Furthermore, sirolimus induces autophagy and reduces ROS production to inhibit NLRP3 inflammasome assembly, which further decreases IL-1β and IL-18 release. Subsequently, there is a decline in the levels of various cytokines and chemokines, which reduces inflammatory cell infiltration into the lungs and BALF. As a result, combined therapy with oseltamivir and sirolimus inhibits viral replication and attenuates virus-induced severe immunopathological lung injury.