Li Wei, Yang Jun
The Second Clinical Medical College of Bin Zhou Medical College, Shandong, China.
Yantai Yuhuangding Hospital, Shandong, China.
Medicine (Baltimore). 2024 Nov 22;103(47):e40590. doi: 10.1097/MD.0000000000040590.
The neutrophil-mediated generation of neutrophil extracellular traps (NETs) results in an augmented inflammatory response and cellular tissue injury during acute myocardial infarction (AMI). Through the analysis of public database information, we discovered and confirmed putative critical genes involved in NETs-mediated AMI.
The AMI dataset GSE66360 and the single-cell dataset GSE163465 were downloaded from the Gene Expression Omnibus database. Key genes were screened by bioinformatics. Quantitative real-time PCR (qRT-PCR) was used to verify the key genes, and then a Mendelian randomization (MR) study was conducted on the basis of the genome-wide association study to determine the causal relationship between key genes and AMI. Dimensionality reduction clustering, pseudo-time series, and cell communication were performed on the single-cell dataset to analyze the key genes screened by bulk RNA sequencing and the dynamic evolution of NETs in the AMI process. Immunohistochemistry and Western blot were used to verify the key genes.
Six key genes, IL1β, S100A12, TLR2, CXCL1, CXCL2, and CCL4, were screened out through bioinformatics. qRT-PCR results showed that compared with the control group, the expression of 5 key genes was upregulated in the AMI group. In the MR study, CXCL1 and CCL4 were observed to have a causal relationship with AMI. Single-cell analysis showed that NETs-related genes CCL4, CXCL2, and IL1β were highly expressed. Combining single cells, qRT-PCR and MR, gene CCL4 was selected as the focus of the study. H9c2 cardiomyocytes simulated myocardial infarction under hypoxia, and the results showed that the expression of gene CCL4 was increased. The immunohistochemical results of gene CCL4 showed that the expression was upregulated in the AMI group.
We found 6 key genes related to NETs-mediated cell damage during AMI. The results of MR showed that CXCL1 and CCL4 were causally related to AMI. Combining single cells, qRT-PCR and MR, gene CCL4 may play an important role in the AMI process. Our results may provide some insights into neutrophil-mediated cell damage during AMI.
中性粒细胞介导的中性粒细胞胞外陷阱（NETs）的产生会在急性心肌梗死（AMI）期间导致炎症反应增强和细胞组织损伤。通过对公共数据库信息的分析，我们发现并确认了参与NETs介导的AMI的假定关键基因。
从基因表达综合数据库下载AMI数据集GSE66360和单细胞数据集GSE163465。通过生物信息学筛选关键基因。采用定量实时PCR（qRT-PCR）验证关键基因，然后在全基因组关联研究的基础上进行孟德尔随机化（MR）研究，以确定关键基因与AMI之间的因果关系。对单细胞数据集进行降维聚类、伪时间序列分析和细胞通讯分析，以分析通过批量RNA测序筛选出的关键基因以及NETs在AMI过程中的动态演变。采用免疫组织化学和蛋白质印迹法验证关键基因。
通过生物信息学筛选出6个关键基因，即IL1β、S100A12、TLR2、CXCL1、CXCL2和CCL4。qRT-PCR结果显示，与对照组相比，AMI组5个关键基因的表达上调。在MR研究中，观察到CXCL1和CCL4与AMI存在因果关系。单细胞分析表明，NETs相关基因CCL4、CXCL2和IL1β高表达。综合单细胞、qRT-PCR和MR结果，选择基因CCL4作为研究重点。H9c2心肌细胞在缺氧条件下模拟心肌梗死，结果显示基因CCL4的表达增加。基因CCL4的免疫组织化学结果显示，AMI组表达上调。
我们发现了6个与NETs介导的AMI期间细胞损伤相关的关键基因。MR结果显示CXCL1和CCL4与AMI存在因果关系。综合单细胞、qRT-PCR和MR结果，基因CCL4可能在AMI过程中起重要作用。我们的结果可能为AMI期间中性粒细胞介导的细胞损伤提供一些见解。
