The Role And Mechanism Of Artemisinin In Acute Lung Injury And Pulmonary Hypertension Models

In the pathological mechanisms of many acute or chronic lung diseases,pulmonary vascular injury occupies an important position.Therefore,exploring the pathological regulation mechanism of pulmonary endothelial injury is of great significance for the treatment of related pulmonary vascular diseases.Artemisinin is widely used as a first-line anti-malaria drug.It has various effects such as anti-inflammatory,anti-tumor,and anti-fibrosis.In recent years,studies have reported that ARS plays a protective role in experimental pulmonary hypertension models and endothelial cells,suggesting that ARS may play a role in pulmonary vascular injury.This study aims to explore the role and possible mechanism of the anti-malarial drug ARS on pulmonary vascular endothelial injury and provide a theoretical basis for clinical medication.Our research has obtained the following results:（1）First,LPS was used to stimulate human lung micro vascular endothelial cells（HLMVEC）,and we found that LPS stimulation attenuated mitochondrial bioenergetics of HLMVEC,which caused an increase in the production of mito-ROS.This elevated mito-ROS levels followed by the activation of its downstream inflammatory factors such as NLRP3,caspase-1 and IL-1βand the secretion of IL-1β.（2）Our results showed that fasudil pretreatment inhibited LPS-mediated Rho A activation and reduced the transcription level of NLRP3,IL-1βand the expression level of NLRP3,p S536 p65,and caspase-1,also attenuated the generation of mito-ROS and the secretion of IL-1β.The colocalization of mitochondria and e NOS in HLMVECs showed that LPS stimulation caused e NOS to be translocated to mitochondria for redistribution,and fasudil pretreatment inhibited its translocation.And we also found that Fasudil treatment decreased the phosphorylation level of e NOS T495 caused by LPS.（3）Pretreated ARS in LPS-induced HLMVECs,the data show that compared with control group,ARS significantly inhibited the expression of NLRP3 and Caspase-1 induced by LPS,and also reduced the production of mitochondrial ROS.Moreover,ARS has a significant protective effect on LPS-induced acute lung injury in mice.（4）We also found that ARS treatment reduced the elevation of pulmonary arterial basal tension induced by MCT and improved ACh-induced endothelium-dependent relaxation,increased e NOS activity and NO levels,and reduced NADPH oxidase activity and superoxide anion levels.We used isolated perfused/ventilated rat lung system to examine the effect of ARS on pulmonary vasoconstriction induced by high-K⁺solutions or hypoxia.The results show that ARS（200μM）induced a significant decrease in PAP with high potassium（40K）.Continuous perfusion of ARS for pretreatment significantly inhibited the increase in PAP induced by 40K or hypoxia（1%O₂）.We continue to conduct in vivo experiments on ARS against pulmonary vasoconstriction.The results show that ARS intervention and treatment significantly reduced the increase of RVSP in MCT-induced PH rats.In this study,western blot,immunofluorescence staining（Immunofluorescence staining,IF）,enzyme-linked immunosorbent assay（ELISA）,mitochondrial superoxide staining and other methods were first used to show that Rho A/ROCK signal regulates LPS-stimulated mitochondrial-dependent inflammatory activation,and this process involves e NOS mitochondrial translocation.ARS improved lung injury by regulating e NOS phospholation,and had a good protective effect on MCT-induced pulmonary hypertension rats.It mainly regulates the pulmonary vasodilation function by targeting endothelium to activate e NOS to release NO and eliminate superoxide.