Yu Yeqing, Yu Chengwei, Dai Shanshan, Liu Yixu, Hu Lanlan, Lu Weimin
Affiliated Hospital of Nanjing University of Chinese Medicine, Nanjing, China.
Front Genet. 2025 Sep 5;16:1617834. doi: 10.3389/fgene.2025.1617834. eCollection 2025.
Ulcerative colitis (UC) is a type of inflammatory bowel disease (IBD) marked by persistent inflammation and ulceration of the colonic mucosal lining. Macrophages regulate intestinal inflammation through distinct polarization profiles. Emerging evidence indicates that the transcription factor SPI1 is a critical regulator of macrophage activity and contributes to both the initiation and progression of UC.
In this study, single-cell RNA sequencing (scRNA-seq) was conducted to profile the transcriptomic landscape of macrophages in the intestinal tissues of UC patients. A gene regulatory network (GRN) was constructed using pySCENIC, which identified SPI1 as a distinct transcriptional regulator involved in macrophage activation. To pinpoint key downstream targets of SPI1, microarray data were analyzed through a combination of weighted gene co-expression network analysis (WGCNA), differential expression (DE) analysis, and several machine learning algorithms, including LASSO, Recursive feature elimination with a random forest classifier (RFE-RF), and Support Vector Machine-Recursive Feature Elimination (SVM-RFE). An model of M1-polarized macrophages was then established, and Western blot (WB) was used to assess the protein expression of SPI1. SPI1 was then silenced using siRNA, and its impact on macrophage polarization was evaluated using flow cytometry and ELISA.
GRN analysis results suggest that the SPI1(+) regulon regulates macrophage activation in UC. Using WGCNA on microarray data, we identified key downstream regulatory target genes, specifically IRAK3, IL1RN, CD55 and PEA15, based on microarray data. Their potential as biomarkers was subsequently validated through several machine learning algorithms. experiments demonstrated elevated expression of SPI1 in M1-polarized macrophages, as confirmed by WB analysis. Flow cytometry and ELISA analyses revealed that SPI1 silencing inhibited M1 macrophage polarization.
This study identified SPI1 as a potential key transcription factor involved in macrophage polarization in UC, possibly exerting its regulatory effects through IRAK3, IL1RN, CD55 and PEA15. These findings offer a novel perspective on the molecular mechanisms underlying intestinal inflammation in UC.
溃疡性结肠炎（UC）是一种炎症性肠病（IBD），其特征为结肠黏膜持续炎症和溃疡。巨噬细胞通过不同的极化模式调节肠道炎症。新出现的证据表明，转录因子SPI1是巨噬细胞活性的关键调节因子，在UC的发生和发展中均起作用。
在本研究中，进行了单细胞RNA测序（scRNA-seq）以分析UC患者肠道组织中巨噬细胞的转录组图谱。使用pySCENIC构建基因调控网络（GRN），该网络确定SPI1是参与巨噬细胞激活的独特转录调节因子。为了确定SPI1的关键下游靶点，通过加权基因共表达网络分析（WGCNA）、差异表达（DE）分析以及包括套索回归（LASSO）、基于随机森林分类器的递归特征消除（RFE-RF）和支持向量机递归特征消除（SVM-RFE）在内的几种机器学习算法对微阵列数据进行分析。然后建立M1极化巨噬细胞模型，并用蛋白质免疫印迹法（WB）评估SPI1的蛋白表达。随后使用小干扰RNA（siRNA）沉默SPI1，并通过流式细胞术和酶联免疫吸附测定（ELISA）评估其对巨噬细胞极化的影响。
GRN分析结果表明，SPI1（+）调节子在UC中调节巨噬细胞激活。基于微阵列数据，通过WGCNA分析，我们确定了关键的下游调控靶基因，特别是IRAK3、IL1RN、CD55和PEA15。随后通过几种机器学习算法验证了它们作为生物标志物的潜力。WB分析证实，实验表明M1极化巨噬细胞中SPI1表达升高。流式细胞术和ELISA分析显示，SPI1沉默抑制了M1巨噬细胞极化。
本研究确定SPI1是UC中参与巨噬细胞极化的潜在关键转录因子，可能通过IRAK3、IL1RN、CD55和PEA15发挥其调节作用。这些发现为UC肠道炎症的分子机制提供了新的视角。
