Analysis Of Histone Acetylation Modification Patterns And Related Immune Microenvironment In Abdominal Aortic Aneurysms

Objectives:1.To construct a scoring model that can quantify the regulatory patterns of histone acetylation in abdominal aortic aneurysms（AAA）.2.To explore the potential relationship between the regulatory patterns of histone acetylation modifications in AAA and the immune microenvironment.3.To screen for core genes that can aid in the diagnosis and treatment of AAA.Methods:1.This study utilized gene expression data from AAA tissue and normal abdominal aortic tissue from the Gene Expression Omnibus（GEO）database.A total of 63 AAA samples and 18 normal abdominal aortic tissue samples were included for differential gene expression analysis and immune cell infiltration analysis.2.Additionally,a search of existing literature on histone acetylation modification was conducted in order to screen for 35 recognized histone acetylation regulatory genes,which were then used for clustering analysis using an unsupervised consensus clustering algorithm to identify different regulatory patterns of histone acetylation modifications in AAA tissue.3.Core genes that may serve as potential targets in the diagnosis and treatment of AAA were also screened for,using three LASSO（Least Absolute Shrinkage and Selection Operator）algorithms,namely SVM-REF（Support vector machine-recursive feature elimination）and RF（Random Forest）.4.An animal model of AAA employing Apoe^-/-mice was constructed,and AAA tissue samples were assessed using HE staining,Masson staining,and EVG staining.Western Blot assays were also performed to verify the protein expression levels of BRD3,Bromodomain Containing 4（BRD4）and Histone Acetyltransferase 1（HAT1）.Results:1.Compared with the normal arterial tissue,the aneurysmal tissue showed heavy infiltration by a variety of immune cells,including neutrophils,macrophages,regulatory T cells and so on.2.Using an unsupervised cluster analysis of histone acetylation modified differentially-expressed genes（DEGs）,the AAA patients were divided into three subgroups:HAcluster-A,HAcluster-B,and HAcluster-C.Significant differences were discovered in the immune microenvironments of the three subgroups.Among them,increased infiltration of immune cells was noted in HAcluster-A,while immune cell infiltration was mostly inhibited in HAcluster-B.Enrichment analysis also revealed associations between HAcluster-A and the epidermal growth factor receptor（EGFR）signaling pathway,whereas HAcluster-B and HAcluster-C were associated with the vascular endothelial growth factor A（VEGFA）signaling pathway and lipid metabolism,respectively.3.Two phenotypically related subtypes were established:gene Cluster-A and gene Cluster-B;these were used as the basis for a scoring model that was constructed to describe and quantify the regulation patterns of histone acetylation in individual aneurysm patients.4.Machine learning was used to screen the core genes:BRD3（AUC=0.78,95%Cl:0.658-0.885）,BRD4（AUC=0.77,95%Cl:0.537-0.787）,HAT1（AUC=0.68,95%Cl:0.532-0.806）,and MYST1（AUC=0.74,95%Cl:0.612-0.853）.5.Significant differences in immune cell infiltration were demonstrated between the high and low expression groups of the core genes.The samples in the BRD3 high expression group showed high levels of infiltration by immune cells such as macrophages,mast cells,and regulatory T cells.The samples in the HAT1 high expression group showed high infiltration by natural killer cells,neutrophils,and other immune cells.The samples in the BRD4 high expression group revealed minimal infiltration by macrophages,mast cells,and other immune cells.6.Tissue proteins were validated by western blot following the successful modeling of mouse aneurysms via HE staining,Masson staining,and EVG staining（BRD3,P=0.01;BRD4,P=0.026;HAT1,P=0.5）.Conclusions:1.This study demonstrated that the HAscore,a scoring model constructed based on histone acetylation regulatory genes,can accurately quantify the level of histone acetylation modification in individual AAA patients and predict the degree of immune cell infiltration.2.The findings suggest that BRD3,BRD4,and HAT1 can potentially serve as genetic targets for the diagnosis of AAA.