The Role Of Vascular Smooth Muscle Cell Endogenous CSE/H₂S In The Pathogenesis Of Abdominal Aortic Aneurysm

Background:Abdominal aortic aneurysms(AAAs)are characterized by the local dilatation of an artery formed by the lesion or injury,which is linked to the degradation of the elastic media of the abdominal aorta.These aneurysms usually are typical asymptomatic until the aneurysm rupture.In severe cases,and the mortality rate exceeds 80%without urgent surgical intervention.The pathophysiological mechanisms have been determined as contributing to the development of AAA,mainly including vascular extracellular matrix(ECM)degradation and remodeling,aortic wall inflammation,and apoptosis of vascular smooth muscle cells.In the early stage of AAA,a large number of immune cells enter the aortic wall through adventitial vasa vasorum,periadventitial lymph nodes,and intraluminal thrombus.The cytokines and reactive oxygen species produced by these cells promote vascular inflammation,induce apoptosis and phenotypic changes in Vascular smooth muscle cells(VSMCs),and then partially reduce the ability of matrix repair.Proteases released by infiltrating inflammatory cells and VSMCs degrade elastin and collagen and promote aortic dilation and rupture,including matrix metalloproteinases(MMPs),a disintegrin and metalloproteinases domain,a disintegrin and metalloproteinases with thrombospondin motifs,elastase,and cathepsins etc.Hydrogen sulfide(H2S),mainly generated by cystathionine-γ-lyase(CSE)to metabolize cysteine,homocysteine and cystathionine cardiovascular tissue,is a gastransmitter with cardiovascular protective effect.Endogenous CSE/H2S inhibits the production of pro-inflammatory factors and chemokines,induces neutrophil apoptosis and macrophage transition to an anti-inflammatory phenotype,inhibits leukocyte adhesion to vascular endothelium and leukocyte migration to sites of inflammation and has a significant biological effect of antagonizing chronic inflammation.It can also reduce the expression of MMP-9 in ischemia-reperfusion kidney and the expression of collagen I and III and MMPs in hypoxia-induced pulmonary artery of rats,regulate the ratio of MMPs and TIMP and microvascular remodeling.CSE is mainly expressed in vascular smooth muscle cells in aortic vascular tissues.Whether the endogenous CSE/H2S system of VSMC is involved in the pathogenesis of AAA and its molecular mechanism remains unclear.In this study,changes in endogenous CSE/H2S expression in patients and animal models were observed,furthermore,smooth muscle cell selective CSE knockout mice were used to explore the effect of endogenous CSE deletion in VSMC on the pathogenesis of AAA,and pathophysiological mechanism of macrophage infiltration and changes of ECM degradation related enzymes was preliminarily discussed.Methods:The CSE expression in α-SMA positive cells was measured by immunofluorescence in human and mouse aoric aneurysm smples.Construction of smooth muscle cell selective CSE knockout mice,Hyperlipidemia mice induced by PCSK9 were given AngⅡ to induce AAA,or BAPN drinking water to induce different mouse AAA models.The changes of VSMC gene expression profile after CSE knockout were performed to test by highthroughput RNA sequencing.Real-time quantitative PCR was used to detect changes in gene expression.The changes of in situ expression of arterial wall protein were observed by immunohistochemical staining.Results:The abdominal aortic aneurysm slices of 6 patients,and the internal mammary artery sections of 6 non-AAA patients were collected for immunofluorescence staining.The results showed that the expression of CSE in aneurysm smooth muscle cells of AAA patients was significantly decreased.The AAA model induced by AngⅡ+hyperlipidemia or BAPN was prepared,and the expression of CSE protein in smooth muscle cells of AAA mice was also significantly down-regulated.AngⅡ+ox-LDL mimics the damage of smooth muscle cells in the onset of AAA,we demonstrated that CSEs mRNA and protein expressions are decreased in both human VSMCs and mouse VSMCs.In AngII+hyperlipidemia or BAPN induced the formation of AAA mouse models,smooth muscle cell CSE-specific knockout mice(CSEsmko)promotes AAA morbidity and mortality.VSMC CSE deficiency also promotes the degradation of the elastic plate of the aortic wall,and increases the infiltration of macrophages in the aneurysm.Bulk RNAsequencing data showed 1468 up-regulated genes and 2943 down-regulated genes in CSE deficient VSMCs.Gene Ontology enrichment analysis showed that many enzymes related to ECM formation or degradation.Quantitative real-time PCR confirmed that MMP-9,ADAMTS-4,LOXL1,and Cela2a in VSMCs dramatically up-regulation in CSE knockout VSMCs.Immunohistochemical staining also showed that the MMP9,Cela2a and ADAMTS4 proteins heightened in CSEsmko mice of both two AAA models.Conculsion:VSMC endogenous CSE/H2S is downregulated in the AAA patients.Deletion CSE in VSMCs induce local inflammatory,promote the MMP9,MMP10,ADAMTS4,LOXL1,and CELA2A expression,exacerbating the degradation of extracellular matrix,causing formation and development of AAA.Endogenous VSMC CSE/H2S may be a new target for AAA drug intervention or gene therapy.