Cadmium Induces NLRP3 Inflammasome-dependent Pyroptosis In Vascular Endothelial Cells

Background: Cadmium(Cd) is an important occupational and environmental pollutant which easy to accumulate in the body and may exert a wide variety of damage effects on humans through bioconcentration and biomagnification processes. It was cleared that the cardiovascular was supposed to be one of most sensitive targets of Cd. Previous study demonstrates that Cd impairs the cardiovascular system by causing inflammation and cell death, but the underlying mechanisms remain obscure.Objective: The present study was designed to explore whether cadmium could induce vascular endothelial cell pyroptosis and the underlying mechanisms.Methods: The HUVECs were exposure to different concentrations of cadmium chloride(0, 20, 40 and 80 μM) for 12 h and 24 h. The toxicity of vascular endothelial induced by Cd was evaluated by CCK-8 and LDH detection. Expressions of NLRP3, pro-caspase-1(Pro-Casp1), pro-IL-1β, caspase-1 and IL-1β were estimated by western blot. Pyroptosis in HUVECs was determined by active caspase-1(Casp1)/SYTOX Green double staining, PI/Hoechst double staining and LDH release. The levels of ROS and mitochondrial ROS( mtROS) were detected by ROS and mtROS kits, respectively. Moreover, the roles of NLRP3 inflammasome and mtROS in Cd-induced HUVECs pyroptosis were investigated using siRNA against NLRP3 and scavenging of mtROS by Mito-TEMPO and NAC.Results: In human umbilical vein endothelial cells(HUVECs), we observed that Cd treatment led to significantly increased inflammation and cell death. Cadmium induces vascular endothelial cell pyroptosis: the expression of caspase-1, the number of double positive cells for caspase-1 and SYTOX Green, the ratio of PI-positive HUVECs and LDH release were increased significantly in Cd group compared with control, which were reduced by caspase-1 inhibitor Z-YVAD-FMK pre-treatment. Further experiments found that NLRP3 inflammasome activation mediates Cd-induced pyroptosis in vascular endothelial cells: Cd treatment resulted in a significant dose-dependent increase in the protein expression of Casp1 and IL-1β; with the pretreatment of NLRP3 siRNA before cadmium expose, we observed a decrease of Casp1 and IL-1β protein expression as well as the aforementioned indicators of pyroptosis. Mitochondrial ROS mediate Cd-induced NLRP3 activation and pyroptosis in HUVECs: Cd treatment promoted intracellular ROS and mtROS generation in a dose-dependent manner; NAC and Mito-TEMPO markedly reduced the expression of Casp1 and IL-1β, the number of caspase-1 and SYTOX Green double-positive cells, PI uptake and LDH release induced by Cd exposure.Conclusion: Cadmium induced NLRP3 inflammasome-dependent pyroptosis in vascular endothelial cells through the generation of ROS and mt ROS. We demonstrated that Cd induced vascular endothelial cells pyroptosis in a ROS-NLRP3-dependent manner. Our results provide novel insights into Cd-induced cytotoxicity and the underlying mechanism by which Cd induces endothelial injury.