Sevoflurane Combined With ATP Activates Caspase-1and Triggers Caspase-1Dependent Pyroptosis In Macrophage Cell Line J774

Objective:The modulation of inflammation and immunity by sevoflurane has been extensively studied, but the specific molecular mechanisms remain unclear. Caspase-1is pivotal in the new cell death mode, pyroptosis, and plays an important role in regulating of innate immunity and inflammation. Whether sevoflurane can modulate the activation of caspase-1in immume cells is unknown. Therefore this study was designed to investigate the effects of sevoflurane on the activation of caspase-1and caspase-1-dependent pyroptosis in macrophages.Methods:We used the murine macrophage cell line J774as cell model. J774cells were incubated with or without5mM ATP, and exposed to6%sevoflurane simultaneously. After0.5hours, the cells were harvested for western blotting of activated caspase-1, for Ethidium Bromide (EtBr) and Ethidium Homodimer-2(EthD-2) staining, and for detecting the levels of reactive oxygen species (ROS) by the ROS-specific fluorescent probe CM-H2DCFDA.Results:Compared with the non-sevoflurane-ATP group, the level of activated caspase-1, P10(220.4±75.6vs.44.9±24.1, P=0.019), and the percentage of EtBr uptake (87.9%± 1.3%vs.20.0%±2.2%, P<0.001) were markedly increased in the sevoflurane-ATP group, which can be inhibited by the caspase-1inhibitor Ac-YVAD-CMK (87.9%±1.3%vs.10.7%±3.3%, P<0.001). While, there was no significant difference in the EthD-2uptake between the two groups (5.0%±2.2%vs.9.1%±2.1%, P=0.10). In addition, the mean fluorescence density of ROS was significantly increased in the sevoflurane-ATP group than that in the non-sevoflurane group (1270.0±81.3vs.780.0±8.8, P<0.001).Conclusion:This study demonstrates that sevoflurane combined with ATP could activate caspase-1and trigger caspase-1-dependent pyroptosis, which might be partially attributed to the modulation of ROS production by sevoflurane. The present data suggest that sevoflurane might regulate immune response via activating caspase-1and triggering caspase-1-dependent pyroptosis under the pathological condition, such as sepsis, which ATP is released from.