脑出血中锰代谢与铁死亡相关关键基因的鉴定

Identification of Key Genes Associated with Manganese Metabolism and Ferroptosis in Intracerebral Hemorrhage

摘要:

目的:识别脑出血(Intracerebral hemorrhage, ICH)中与锰代谢和铁死亡相关的共有关键基因,探索ICH潜在的发病与发展机制。方法:将从急性ICH相关的GEO数据集中得到的差异表达基因与加权基因共表达网络分析得到的模块基因、铁死亡相关基因和锰代谢相关基因进行交集分析,以识别重叠基因。对重叠基因进行功能富集分析与蛋白质-蛋白质相互作用网络构建,并使用LASSO回归和随机森林算法识别关键基因。基于关键基因,构建用于评估ICH风险的列线图模型。随后进行细胞类型鉴定、轨迹分析及免疫浸润分析。蛋白质印迹法验证了关键基因在大鼠体内的蛋白表达。结果:共鉴定出16个重叠基因。其中,Ftl1、Gdf15和Hmox1通过LASSO回归和随机森林算法被鉴定为关键基因。基于关键基因构建的列线图在风险预测方面表现出较高的准确性。这些基因富集于免疫和炎症通路,并在ICH样本的整体水平和单细胞水平上均显著上调。值得注意的是,在ICH中巨噬细胞占比显著升高,且三个基因均呈现显著的巨噬细胞特异性表达模式,提示巨噬细胞可能是这些基因共同参与ICH病理过程的关键细胞类型。此外,轨迹分析显示,在向促炎性(M1)巨噬细胞转变的过程中,这些基因的表达随时间推移而增加。免疫浸润分析表明,Gdf15和Hmox1与T细胞和NK细胞亚群存在显著关联。蛋白质印迹实验证实,在ICH模型中,Ftl1、Gdf15和Hmox1的蛋白表达均升高。结论:Ftl1、Gdf15 和 Hmox1 被鉴定为参与ICH中锰代谢和铁死亡的关键共有基因,提示锰代谢异常与铁死亡过程之间可能存在直接的分子联系,为进一步的机制研究提供了候选分子靶点。

Abstract:

Objective: To identify shared key genes associated with manganese (Mn) metabolism and ferroptosis in intracerebral hemorrhage (ICH), and to explore potential mechanisms underlying the onset and progression of ICH. Methods: Differentially expressed genes derived from acute ICH-related GEO datasets were intersected with module genes identified by weighted gene co-expression network analysis, as well as with ferroptosis-related genes and Mn metabolism-related genes, to identify overlapping candidates. Functional enrichment analyses and protein-protein interaction network construction were performed for the overlapping genes. LASSO regression and random forest algorithms were applied to identify key genes. Based on these genes, a nomogram model was established to evaluate ICH risk. Subsequently, cell type identification, trajectory analysis, and immune infiltration analysis were conducted. Protein expression of the key genes in rats was validated using western blot analysis. Results: A total of 16 overlapping genes were identified. Among them, Ftl1, Gdf15, and Hmox1 were recognized as key genes by both LASSO regression and random forest analysis. The nomogram constructed based on these genes demonstrated high accuracy in risk prediction. These genes were enriched in immune- and inflammation-related pathways and were significantly upregulated in ICH samples at both the bulk and single-cell levels. Notably, macrophage proportions were markedly increased in ICH, and all three genes exhibited pronounced macrophage-specific expression patterns, suggesting that macrophages may represent a critical cell type through which these genes jointly contribute to ICH pathology. Furthermore, trajectory analysis revealed that the expression of these genes increased over time during the transition toward pro-inflammatory (M1) macrophages. Immune infiltration analysis indicated that Gdf15 and Hmox1 were significantly associated with T-cell and natural killer cell subpopulations. Western blot experiments confirmed elevated protein expression levels of Ftl1, Gdf15, and Hmox1 in the ICH model. Conclusion: Ftl1, Gdf15, and Hmox1 were identified as shared key genes involved in Mn metabolism and ferroptosis in ICH. These findings suggest a potential direct molecular link between Mn metabolic dysregulation and ferroptosis, providing candidate molecular targets for further mechanistic investigations.