Mechanic Of Hydrogen Sulfide In Homocysteine-induced Endothelial Dysfunction

Part one Hydrogen sulfide improves homocysteine-induced endothelial dysfunction in rat thoracic aortaObjectives:Homocysteine?Hcy?is a cytotoxic sulfur-containing amino acid which may induce endothelial dysfunction.Its mechanism is related to the activation of NLRP3 inflammasome and the subsequent promotion of inflammatory response.Hydrogen sulfide?H₂S?,as the third gasotransmitter after nitric oxide?NO?and carbon monoxide?CO?,has anti-inflammatory or anti-oxidative effects in cardiovascular system.In this part of the study,Hcy-incubated rat thoracic aorta was used to determine whether exogenous H₂S supplementation could improve endothelial dysfunction through the inhibition of NLRP3 inflammasome..Methods:1.Experimental group:Eight-week-old male Wistar rats weighing 280±10g were used in this study.The thoracic aorta were obtain and randomly divided into 8 groups:Control,Hcy?50?mol/L?group,Hcy?50?mol/L?+NaHS?50?mol/L?group,Hcy?50?mol/L?+MCC950?NLRP3 inhibitor,1?mol/L?group,Hcy?50?mol/L?+VX765?caspase-1 inhibitor,10?mol/L?group,NaHS?50?mol/L?group,MCC950?1?mol/L?group,VX765?10?mol/L?group.2.The rat thoracic aorta were isolated and incubated in the CO₂ incubator for 24h.The vascular tone was measured to test the endothelium-dependent vasodilation and endothelium-independent vasodilation.3.Western blot was used to determine the protein expression of NLRP3,caspase-1 and IL-1?in rat thoracic aorta.Result:1.The acetylcholine?ACh?-induced vasodilation is decreased in thoracic aorta with Hcy stimulation for 24h.Administration of MCC950,VX765 and H₂S donor NaHS significantly improves ACh-induced endothelium-dependent vasodilation.Sodium nitroprusside?SNP?-mediated endothelium-independent vasodilation shows no difference among these groups.2.The protein expression of NLRP3,caspase-1 and IL-1?are down-regulated in thoracic aorta with Hcy incubation,which are restored by NaHS supplementation.Conclusion:Exogenous administration of H₂S may ameliorate Hcy-induced endothelial dysfunction in rat thoracic aorta,and the mechanism may be related to the inhibition of NLRP3/caspase1/IL-1?inflammatory signaling pathway.Part two Hydrogen sulfide protects Hcy-induced endothelial injury by inhibiting TXNIPObjective:Thioredoxin interacting protein?TXNIP?is a multifunctional induction protein involved in the regulation of various biological processes.Studies have shown that TXNIP could activate NLRP3 inflammasome and promote the production of inflammatory factors such as caspase-1 and IL-1?.Whether TXNIP is involved in Hcy-induced endothelial injury has not been reported.In this part of the study,human umbilical vein endothelial cells?HUVECs?were used with Hcy incubation to investigate whether exogenous H₂S administration could inhibit Hcy-induced endothelial cell injury by inhibiting TXNIP-NLRP3 signaling pathway.Methods:1.Experimental group:HUVECs were randomly divided into 6 groups:Control,Hcy?50?mol/L?group,Hcy?50?mol/L?+NaHS?50?mol/L?group,NaHS?50?mol/L?group,Hcy?50?mol/L?+Verapamil?TXNIP inhibitor,50?mol/L?,Verapamil group?50?mol/L?.2.Determination of HUVECs activity by CCK-8.3.The protein expression of TXNIP,NLRP3,caspase-1 and IL-1?in HUVECs were determined by western blot.Result:1.Hcy could reduce the bioactivity of HUVECs,and exogenous administration of H₂S improves the Hcy-induced cell injury.2.The protein expression of TXNIP,NLRP3,caspase-1 and IL-1?are up-regulated in HUVECs with Hcy incubation.TXNIP inhibitor Verapamil blocks the protein expression of TXNIP and NLRP3.Exogenous administration of NaHS down-regulates the expression of TXNIP and NLRP3/Caspase-1/IL-1?inflammatory protein.Conclusion:Hcy causes HUVECs injury through TXNIP-NLRP3 pathway.H₂S protects Hcy-induced endothelial cell injury by inhibiting TXNIP and NLRP3/caspase-1/IL-1?inflammatory pathway.