Zhang Huihui, Lv Yanhe, Li Jingjing, Jiao Bingze, Fu Jiahui, Zhao Xujie, Cheng Likun, Bai Yilin, Wang Lei, Li Yanwei, Hang Bolin, Wei Xiaobing, Liu Mingcheng, Teng Zhanwei, Chang Meinan, Liao Chengshui, Bai Yueyu, Xia Xiaojing, Ding Ke, Hu Jianhe
College of Animal Science and Veterinary Medicine, Henan Institute of Science and Technology, Xinxiang, 453003, China.
Ministry of Education Key Laboratory for Animal Pathogens and Biosafety, Zhengzhou, 450002, China.
Sci Rep. 2025 Jun 2;15(1):19252. doi: 10.1038/s41598-025-03969-7.
Antimicrobial peptides (AMPs) possess vaccine adjuvant activity; however, their specific targets and molecular mechanisms remain incompletely understood, which hinders their clinical application. This study aimed to elucidate the key targets and pathways through which the antimicrobial peptide BSN-37 modulates immune responses in macrophages, providing evidence for its potential clinical translation. In this investigation, Balb/c mice were administered BSN-37 for 12 h, after which total RNA was extracted from peritoneal macrophages to assess the mRNA expression levels of cytokines and key molecules on the cell surface, followed by transcriptomic sequencing. The results demonstrated that BSN-37 significantly upregulated the mRNA expression of these molecules and cytokines. A total of 228 differentially expressed long non-coding RNAs (lncRNAs) (121 upregulated, 107 downregulated) and 149 differentially expressed mRNAs (104 upregulated, 45 downregulated) were identified. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses revealed significant enrichment of differentially expressed mRNAs in immune response pathways, PI3K-Akt signaling, and NOD-like receptor signaling. Differentially expressed lncRNA target genes were associated with T cell receptor signaling, PD-1 checkpoint regulation, and other immune regulatory pathways. Protein-protein interaction network analysis identified core genes such as CCchemokine receptor 1 (CCR1) and Toll Like Receptor 8 (TLR8). Molecular docking studies confirmed that BSN-37 exhibited strong binding affinity to TLR8 and CCR1, with binding energies less than - 5 kcal/mol. RT-qPCR validation confirmed the reliability of the sequencing data. These findings indicate that BSN-37 activates multiple immune response pathways in macrophages by targeting immune-related genes such as TLR8 and CCR1, offering theoretical support for the development of novel immune adjuvants.
抗菌肽（AMPs）具有疫苗佐剂活性；然而，它们的具体靶点和分子机制仍未完全明确，这阻碍了它们的临床应用。本研究旨在阐明抗菌肽BSN-37调节巨噬细胞免疫反应的关键靶点和途径，为其潜在的临床转化提供证据。在本研究中，给Balb/c小鼠注射BSN-37 12小时后，从腹腔巨噬细胞中提取总RNA，以评估细胞因子和细胞表面关键分子的mRNA表达水平，随后进行转录组测序。结果表明，BSN-37显著上调了这些分子和细胞因子的mRNA表达。共鉴定出228个差异表达的长链非编码RNA（lncRNAs）（121个上调，107个下调）和149个差异表达的mRNA（104个上调，45个下调）。基因本体（GO）和京都基因与基因组百科全书（KEGG）通路分析显示，差异表达的mRNA在免疫反应通路、PI3K-Akt信号通路和NOD样受体信号通路中显著富集。差异表达的lncRNA靶基因与T细胞受体信号通路、PD-1检查点调节和其他免疫调节通路相关。蛋白质-蛋白质相互作用网络分析确定了核心基因，如趋化因子受体1（CCR1）和Toll样受体8（TLR8）。分子对接研究证实，BSN-37对TLR8和CCR1表现出很强的结合亲和力，结合能小于-5千卡/摩尔。RT-qPCR验证证实了测序数据的可靠性。这些发现表明，BSN-37通过靶向TLR8和CCR1等免疫相关基因激活巨噬细胞中的多种免疫反应通路，为新型免疫佐剂的开发提供了理论支持。
