背景:脓毒症被认为是新发心房颤动(NOAF)的高风险因素,中性粒细胞胞外陷阱(NETs)与许多疾病的发病机理有关。然而,NETs和NETs相关基因(NRGs)在脓毒症NOAF发生中的确切作用仍未得到充分阐明.本研究的目的是确定连接脓毒症和房颤的中心NRGs,并研究脓毒症中NETs和NOAF之间的潜在关联。
方法:从基因表达综合(GEO)数据库中检索AF和败血症微阵列数据集,用于分析共享的病理生理机制和NRGs涉及使用生物信息学技术的败血症和AF。采用CIBERSORT算法评估免疫细胞浸润并识别这些疾病的共同免疫特征。此外,脂多糖(LPS)诱导的脓毒症大鼠模型被用来研究NETs之间的关联,NRGs,和脓毒症诱发的房颤。西方印迹,酶联免疫吸附测定,苏木精-伊红染色,免疫组织化学,和免疫荧光用于评估NRGs的表达,NET的形成,以及中性粒细胞的浸润.电生理分析和多电极阵列技术用于检查脓毒症大鼠AF的易损性和传导异质性。此外,使用DNaseI对LPS诱导的脓毒症大鼠进行干预,一种专门针对NETs的药物,为了评估其对中性粒细胞浸润的影响,NET的形成,hubNRGs蛋白表达,和AF漏洞。
结果:在脓毒症和AF的情况下,共鉴定出61个常见差异表达基因(DEG)和4个中心DE-NRG。功能富集分析显示这些DEGs主要与炎症和免疫相关的过程相关。免疫浸润分析进一步证明了免疫浸润细胞的存在,特别是中性粒细胞浸润,败血症和房颤。此外,四个中心DE-NRGs的相对表达与中性粒细胞浸润呈正相关.在LPS诱导的脓毒症大鼠中,我们观察到四个DE-NRGs的表达明显上调,NET的形成,心房组织有中性粒细胞浸润.通过电生理评估,我们发现了房颤的高度脆弱性,心房表面传导速度降低,并增加了LPS诱导的脓毒症大鼠的传导异质性。值得注意的是,这些有害影响可以通过DNaseI治疗得到部分改善。
结论:通过生物信息学分析和实验验证,我们在脓毒症和房颤中确定了4个中心NRGs.随后的实验表明,心房中NETs的形成可能有助于脓毒症中NOAF的发病机理。这些发现为预防和治疗脓毒症中NOAF提供了潜在的新目标和见解。
BACKGROUND: Sepsis is considered a high risk factor for new-onset atrial fibrillation (NOAF), with neutrophil extracellular traps (NETs) being implicated in the pathogenesis of numerous diseases. However, the precise role of NETs and NETs-related genes (NRGs) in the occurrence of NOAF in sepsis remains inadequately elucidated. The objective of this study was to identify hub NRGs connecting sepsis and AF, and to investigate the potential association between NETs and NOAF in sepsis.
METHODS: The AF and sepsis microarray datasets were retrieved from the Gene Expression Omnibus (GEO) database for analysis of shared pathophysiological mechanisms and NRGs implicated in both sepsis and AF using bioinformatics techniques. The CIBERSORT algorithm was employed to assess immune cell infiltration and identify common immune characteristics in these diseases. Additionally, a rat model of lipopolysaccharide (LPS)-induced sepsis was utilized to investigate the association between NETs, NRGs, and sepsis-induced AF. Western blotting, enzyme-linked immunosorbent assay, hematoxylin-eosin staining, immunohistochemistry, and immunofluorescence were employed to assess the expression of NRGs, the formation of NETs, and the infiltration of neutrophils. Electrophysiological analysis and multi-electrode array techniques were utilized to examine the vulnerability and conduction heterogeneity of AF in septic rats. Furthermore, intervention was conducted in LPS-induced sepsis rats using DNase I, a pharmacological agent that specifically targets NETs, in order to assess its impact on neutrophil infiltration, NETs formation, hub NRGs protein expression, and AF vulnerability.
RESULTS: A total of 61 commonly differentially expressed genes (DEGs) and four hub DE-NRGs were identified in the context of sepsis and AF. Functional enrichment analysis revealed that these DEGs were predominantly associated with processes related to inflammation and immunity. Immune infiltration analysis further demonstrated the presence of immune infiltrating cells, specifically neutrophil infiltration, in both sepsis and AF. Additionally, a positive correlation was observed between the relative expression of the four hub DE-NRGs and neutrophil infiltration. In rats with LPS-induced sepsis, we observed a notable upregulation in the expression of four DE-NRGs, the formation of NETs, and infiltration of neutrophils in atrial tissue. Through electrophysiological assessments, we identified heightened vulnerability to AF, reduced atrial surface conduction velocity, and increased conduction heterogeneity in LPS-induced sepsis rats. Notably, these detrimental effects can be partially ameliorated by treatment with DNase I.
CONCLUSIONS: Through bioinformatics analysis and experimental validation, we identified four hub NRGs in sepsis and AF. Subsequent experiments indicated that the formation of NETs in the atria may contribute to the pathogenesis of NOAF in sepsis. These discoveries offer potential novel targets and insights for the prevention and treatment of NOAF in sepsis.