{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,3]],"date-time":"2026-09-03T08:12:32Z","timestamp":1788423152519,"version":"build-2803163510"},"reference-count":47,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2026,9,3]],"date-time":"2026-09-03T00:00:00Z","timestamp":1788393600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Immunol."],"abstract":"<jats:p>Peripheral artery disease (PAD) is a chronic ischemic condition characterized by vascular occlusion and metabolic-immune dysregulation. To elucidate the molecular mechanisms underlying PAD and the therapeutic effects of Kaempferol, we integrated bulk transcriptomics, single-cell RNA sequencing, network pharmacology, and experimental validation. Transcriptomic analysis identified 1,144 differentially expressed genes in PAD, highlighting extensive transcriptional remodeling. WGCNA revealed distinct gene modules significantly correlated with PAD, suggesting key networks associated with vascular inflammation and immune dysregulation. Network pharmacology identified kaempferol as the core active component of Ruan Jian Qing Mai formula that is clinically effective in treating PAD, targeting 87 PAD-related genes enriched in immune and inflammatory pathways, predominantly expressed in macrophages. Integration of kaempferol-associated targets from RNA-seq and multi-database prediction uncovered 70 overlapping targets implicated in macrophage activation, lipid metabolism, and atherosclerosis signaling. CIBERSORTx and single-cell analysis further pinpointed macrophages as major cellular mediators, with kaempferol-target genes (e.g., TNF, SYK, LCK, IL4I1) specifically enriched in macrophages. Macrophages were stratified into high-inflammatory and low-inflammatory subsets; PAD lesions exhibited prominent enrichment of high-inflammatory macrophages, which concurrently displayed elevated inflammatory signatures and enhanced ferroptosis transcriptional activity, and represented the primary cellular population responsive to kaempferol. In silico gene knockout analysis further verified that SYK and SNX5 serve as core immune signaling regulators in macrophages, while TNF and LCK lack macrophage-specific immune regulatory functions. Functional experiments confirmed that kaempferol significantly suppressed LPS-induced M1 macrophage polarization and inflammatory cytokine expression in RAW264.7 cells. Ferroptosis scoring revealed enhanced ferroptosis susceptibility in PAD macrophages, with kaempferol-target genes correlating positively with ferroptosis drivers. Molecular docking further validated strong binding affinities between kaempferol and key macrophage-related targets (IL4I1, LCK, TNF, SYK). Using a zebrafish vascular regeneration injury model, we further verified that high-dose kaempferol rescues impaired angiogenesis under vascular ischemic damage. Collectively, these multi-omics and experimental findings uncover macrophage polarization and ferroptosis modulation as central mechanisms mediating kaempferol\u2019s therapeutic efficacy in PAD, providing mechanistic insights and potential molecular targets for precision intervention.<\/jats:p>","DOI":"10.3389\/fimmu.2026.1733284","type":"journal-article","created":{"date-parts":[[2026,9,3]],"date-time":"2026-09-03T07:17:35Z","timestamp":1788419855000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":0,"title":["Multi-omics integration reveals macrophage polarization and ferroptosis as key mechanisms underlying kaempferol\u2019s therapeutic efficacy in peripheral artery disease"],"prefix":"10.3389","volume":"17","author":[{"given":"Zi-jun","family":"Qiao","sequence":"first","affiliation":[{"name":"Guangdong Cardiovascular Institute, Guangdong Provincial People's Hospital (Guangdong Academy of Medical Sciences), Southern Medical University","place":["Guangzhou, Guangdong, China"]},{"name":"Guangdong Provincial Key Laboratory of Pathogenesis, Targeted Prevention and Treatment of Heart Disease,Guangzhou Key Laboratory of Cardiac Pathogenesis and Prevention","place":["Guangzhou, Guangdong, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Miao-miao","family":"Wang","sequence":"additional","affiliation":[{"name":"Institute of Vascular Anomalies, Shanghai TCM-Integrated Hospital, Shanghai University of Traditional Chinese Medicine","place":["Shanghai, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wang-sheng","family":"Luo","sequence":"additional","affiliation":[{"name":"Department of Cardiology, the First Affiliated Hospital of University of South China","place":["Hengyang, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bo-lan","family":"Yu","sequence":"additional","affiliation":[{"name":"School of Medicine South China University of Technology, Guangdong Provincial People's Hospital, Guangdong Academy of Medical Sciences, Southern Medical University","place":["Guangzhou, Guangdong, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tao","family":"Liu","sequence":"additional","affiliation":[{"name":"Department of Hepatobiliary Surgery, Shandong Provincial Hospital Heze Branch","place":["Heze, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ping","family":"Zhu","sequence":"additional","affiliation":[{"name":"Guangdong Cardiovascular Institute, Guangdong Provincial People's Hospital (Guangdong Academy of Medical Sciences), Southern Medical University","place":["Guangzhou, Guangdong, China"]},{"name":"Guangzhou Xiangxue Pharmaceutical Co., Ltd","place":["Guangzhou, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Can","family":"Hu","sequence":"additional","affiliation":[{"name":"Institute of Vascular Anomalies, Shanghai TCM-Integrated Hospital, Shanghai University of Traditional Chinese Medicine","place":["Shanghai, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ji","family":"Li","sequence":"additional","affiliation":[{"name":"Institute of Vascular Anomalies, Shanghai TCM-Integrated Hospital, Shanghai University of Traditional Chinese Medicine","place":["Shanghai, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zhi-qiang","family":"Liang","sequence":"additional","affiliation":[{"name":"Institute of Vascular Anomalies, Shanghai TCM-Integrated Hospital, Shanghai University of Traditional Chinese Medicine","place":["Shanghai, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jian","family":"Chen","sequence":"additional","affiliation":[{"name":"Institute of Vascular Anomalies, Shanghai TCM-Integrated Hospital, Shanghai University of Traditional Chinese Medicine","place":["Shanghai, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2026,9,3]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","DOI":"10.1093\/eurheartj\/ehae867","article-title":"Lifetime risk of lower extremity peripheral arterial disease: a Danish nationwide longitudinal study","volume":"46","author":"Sogaard","year":"2025","journal-title":"Eur Heart J"},{"key":"B2","doi-asserted-by":"publisher","first-page":"76","DOI":"10.2337\/dc24-1644","article-title":"Trends in peripheral artery disease, lower-extremity revascularization, and lower-extremity amputation in incident type 2 diabetes: a Danish population-based cohort study","volume":"48","author":"Gyldenkerne","year":"2025","journal-title":"Diabetes Care"},{"key":"B3","doi-asserted-by":"publisher","first-page":"103","DOI":"10.1186\/s12951-025-03140-4","article-title":"Biomaterial-based vascularization strategies for enhanced treatment of peripheral arterial disease","volume":"23","author":"Wang","year":"2025","journal-title":"J Nanobiotechnology"},{"key":"B4","doi-asserted-by":"publisher","DOI":"10.1161\/cir.0000000000001310","article-title":"Sex differences in peripheral vascular disease: a scientific statement from the American Heart Association","volume":"151","author":"Kim","year":"2025","journal-title":"Circulation"},{"key":"B5","doi-asserted-by":"publisher","DOI":"10.1113\/ep091901","article-title":"Unilateral hindlimb ischaemia-induced systemic inflammation is associated with non-ischaemic skeletal muscle inflammation","volume":"109","author":"Evans","year":"2024","journal-title":"Exp Physiol"},{"key":"B6","doi-asserted-by":"publisher","DOI":"10.3390\/ijms25105457","article-title":"FGF23 as a potential pathophysiological factor in peripheral arterial disease associated with chronic kidney disease","volume":"25","author":"Donate-Correa","year":"2024","journal-title":"Int J Mol Sci"},{"key":"B7","doi-asserted-by":"publisher","first-page":"76","DOI":"10.1186\/s40360-025-00908-0","article-title":"Kaempferol inhibits cardiomyocyte pyroptosis via promoting O-GlcNAcylation of GSDME and improved acute myocardial infarction","volume":"26","author":"Zhou","year":"2025","journal-title":"BMC Pharmacol Toxicol"},{"key":"B8","doi-asserted-by":"publisher","DOI":"10.1016\/j.cyto.2024.156846","article-title":"Kaempferol: unveiling its anti-inflammatory properties for therapeutic innovation","volume":"186","author":"Herrera","year":"2025","journal-title":"Cytokine"},{"key":"B9","doi-asserted-by":"publisher","DOI":"10.3389\/fphar.2024.1411273","article-title":"Integrative transcriptome-proteome approach reveals key hypoxia-related features involved in the neuroprotective effects of Yang Xue oral liquid on Alzheimer's and Parkinson's disease","volume":"15","author":"Chen","year":"2024","journal-title":"Front Pharmacol"},{"key":"B10","doi-asserted-by":"publisher","DOI":"10.1172\/jci.insight.177334","article-title":"Myeloid DRP1 deficiency limits revascularization in ischemic muscles via inflammatory macrophage polarization and metabolic reprogramming","volume":"10","author":"Yadav","year":"2025","journal-title":"JCI Insight"},{"key":"B11","doi-asserted-by":"publisher","DOI":"10.1186\/1471-2105-9-559","article-title":"WGCNA: an R package for weighted correlation network analysis","volume":"9","author":"Langfelder","year":"2008","journal-title":"BMC Bioinf"},{"key":"B12","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkae1037","article-title":"HERB 2.0: an updated database integrating clinical and experimental evidence for traditional Chinese medicine","volume":"53","author":"Gao","year":"2025","journal-title":"Nucleic Acids Res"},{"key":"B13","doi-asserted-by":"publisher","DOI":"10.1021\/acs.jcim.0c00291","article-title":"NetInfer: a web server for prediction of targets and therapeutic and adverse effects via network-based inference methods","volume":"60","author":"Wu","year":"2020","journal-title":"J Chem Inf Model"},{"key":"B14","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkz382","article-title":"SwissTargetPrediction: updated data and new features for efficient prediction of protein targets of small molecules","volume":"47","author":"Daina","year":"2019","journal-title":"Nucleic Acids Res"},{"key":"B15","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkz240","article-title":"NetworkAnalyst 3.0: a visual analytics platform for comprehensive gene expression profiling and meta-analysis","volume":"47","author":"Zhou","year":"2019","journal-title":"Nucleic Acids Res"},{"key":"B16","doi-asserted-by":"publisher","DOI":"10.1038\/s41592-019-0619-0","article-title":"Fast, sensitive and accurate integration of single-cell data with Harmony","volume":"16","author":"Korsunsky","year":"2019","journal-title":"Nat Methods"},{"key":"B17","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkac935","article-title":"FerrDb V2: update of the manually curated database of ferroptosis regulators and ferroptosis-disease associations","volume":"51","author":"Zhou","year":"2023","journal-title":"Nucleic Acids Res"},{"key":"B18","doi-asserted-by":"publisher","DOI":"10.1016\/j.jare.2024.11.016","article-title":"Kaempferol regulating macrophage foaming and atherosclerosis through Piezo1-mediated MAPK\/NF-kappaB and Nrf2\/HO-1 signaling pathway","volume":"75","author":"Chu","year":"2024","journal-title":"J Adv Res"},{"key":"B19","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkac394","article-title":"CB-Dock2: improved protein-ligand blind docking by integrating cavity detection, docking and homologous template fitting","volume":"50","author":"Liu","year":"2022","journal-title":"Nucleic Acids Res"},{"key":"B20","doi-asserted-by":"publisher","DOI":"10.1080\/07391102.2021.1936179","article-title":"Identification of potential inhibitors for LLM of Staphylococcus aureus: structure-based pharmacophore modeling, molecular dynamics, and binding free energy studies","volume":"40","author":"Kumari","year":"2022","journal-title":"J Biomol Struct Dyn"},{"key":"B21","doi-asserted-by":"publisher","DOI":"10.1016\/j.molstruc.2022.132476","article-title":"Structural-based virtual screening and identification of novel potent antimicrobial compounds against YsxC of Staphylococcus aureus","volume":"1255","author":"Kumari","year":"2022","journal-title":"J Mol Struct"},{"key":"B22","doi-asserted-by":"publisher","first-page":"57","DOI":"10.1016\/j.niox.2025.03.003","article-title":"Exploring the therapeutic potential of beetroot juice in patients with peripheral artery disease: a narrative review","volume":"156","author":"Wang","year":"2025","journal-title":"Nitric Oxide"},{"key":"B23","doi-asserted-by":"publisher","DOI":"10.1016\/j.jvs.2025.03.394","article-title":"The incidence and significance of delayed bleeding events after lower extremity revascularization in patients with advanced peripheral arterial disease","volume":"82","author":"Hundito","year":"2025","journal-title":"J Vasc Surg"},{"key":"B24","doi-asserted-by":"publisher","DOI":"10.1016\/b978-1-4377-2930-6.00016-1","article-title":"Epidemiology of peripheral artery disease","volume":"116","author":"Criqui","year":"2015","journal-title":"Circ Res"},{"key":"B25","doi-asserted-by":"publisher","first-page":"95","DOI":"10.1186\/s13073-023-01250-y","article-title":"Skeletal muscle regeneration failure in ischemic-damaged limbs is associated with pro-inflammatory macrophages and premature differentiation of satellite cells","volume":"15","author":"Southerland","year":"2023","journal-title":"Genome Med"},{"key":"B26","doi-asserted-by":"publisher","DOI":"10.26355\/eurrev_202009_22822","article-title":"LncRNA HMGA1P4 promotes cisplatin-resistance in gastric cancer","volume":"24","author":"Qiao","year":"2020","journal-title":"Eur Rev Med Pharmacol Sci"},{"key":"B27","doi-asserted-by":"publisher","DOI":"10.1002\/jcb.29058","article-title":"Identification of functional lncRNAs in gastric cancer by integrative analysis of GEO and TCGA data","volume":"120","author":"Zhang","year":"2019","journal-title":"J Cell Biochem"},{"key":"B28","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0141397","article-title":"MARCKS signaling differentially regulates vascular smooth muscle and endothelial cell proliferation through a KIS-, p27kip1- dependent mechanism","volume":"10","author":"Yu","year":"2015","journal-title":"PloS One"},{"key":"B29","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.m110.196022","article-title":"The MARCKS protein plays a critical role in phosphatidylinositol 4,5-bisphosphate metabolism and directed cell movement in vascular endothelial cells","volume":"286","author":"Kalwa","year":"2011","journal-title":"J Biol Chem"},{"key":"B30","doi-asserted-by":"publisher","DOI":"10.3390\/biology10070686","article-title":"MYOC promotes the differentiation of C2C12 cells by regulation of the TGF-beta signaling pathways via CAV1","volume":"10","author":"Zhang","year":"2021","journal-title":"Biol (Basel)"},{"key":"B31","doi-asserted-by":"publisher","DOI":"10.1080\/10408398.2022.2121261","article-title":"Kaempferol and atherosclerosis: from mechanism to medicine","volume":"64","author":"Chen","year":"2024","journal-title":"Crit Rev Food Sci Nutr"},{"key":"B32","doi-asserted-by":"publisher","DOI":"10.1093\/jpp\/rgae130","article-title":"Activation of HSPA5 contributes to pazopanib-induced hepatotoxicity through l-ornithine metabolism pathway and endoplasmic reticulum stress","volume":"77","author":"Chen","year":"2025","journal-title":"J Pharm Pharmacol"},{"key":"B33","doi-asserted-by":"publisher","DOI":"10.1016\/j.metabol.2019.153954","article-title":"The metabolic regulator small heterodimer partner contributes to the glucose and lipid homeostasis abnormalities induced by hepatitis C virus infection","volume":"100","author":"Chen","year":"2019","journal-title":"Metabolism"},{"key":"B34","doi-asserted-by":"publisher","DOI":"10.3390\/jcm14072352","article-title":"Beyond cholesterol: emerging risk factors in atherosclerosis","volume":"14","author":"Bekbossynova","year":"2025","journal-title":"J Clin Med"},{"key":"B35","doi-asserted-by":"publisher","first-page":"76","DOI":"10.1186\/s12944-024-02049-5","article-title":"Nobiletin alleviates atherosclerosis by inhibiting lipid uptake via the PPARG\/CD36 pathway","volume":"23","author":"Wang","year":"2024","journal-title":"Lipids Health Dis"},{"key":"B36","doi-asserted-by":"publisher","first-page":"23","DOI":"10.1007\/s10565-024-09973-3","article-title":"Exosomal NEDD4L derived from HG+oxLDL-induced vascular endothelial cells accelerates macrophage M1 polarization and oxLDL uptake by ubiquitinating IkappaBalpha and PPARgamma","volume":"41","author":"Chen","year":"2025","journal-title":"Cell Biol Toxicol"},{"key":"B37","doi-asserted-by":"publisher","DOI":"10.1016\/j.bcp.2023.115965","article-title":"Is the suppression of CD36 a promising way for atherosclerosis therapy","volume":"219","author":"Wen","year":"2024","journal-title":"Biochem Pharmacol"},{"key":"B38","doi-asserted-by":"publisher","DOI":"10.1016\/j.heliyon.2024.e32073","article-title":"Macrophage polarisation and inflammatory mechanisms in atherosclerosis: implications for prevention and treatment","volume":"10","author":"Yang","year":"2024","journal-title":"Heliyon"},{"key":"B39","doi-asserted-by":"publisher","DOI":"10.3390\/biomedicines11082284","article-title":"Micro- and macrovascular effects of inflammation in peripheral artery disease-pathophysiology and translational therapeutic approaches","volume":"11","author":"Poledniczek","year":"2023","journal-title":"Biomedicines"},{"key":"B40","doi-asserted-by":"publisher","first-page":"26","DOI":"10.1016\/j.clim.2018.06.006","article-title":"Involvement of CD8+ T cell subsets in early response to vascular injury in patients with peripheral artery disease in vivo","volume":"194","author":"Maga","year":"2018","journal-title":"Clin Immunol"},{"key":"B41","doi-asserted-by":"publisher","DOI":"10.1016\/j.molmet.2024.102092","article-title":"Deficiency of the mitochondrial transporter SLC25A47 minimally impacts hepatic lipid metabolism in fasted and diet-induced obese mice","volume":"92","author":"Attema","year":"2025","journal-title":"Mol Metab"},{"key":"B42","doi-asserted-by":"publisher","DOI":"10.1038\/s41569-021-00663-9","article-title":"Update on the pathophysiology and medical treatment of peripheral artery disease","volume":"19","author":"Golledge","year":"2022","journal-title":"Nat Rev Cardiol"},{"key":"B43","doi-asserted-by":"publisher","DOI":"10.1016\/j.jacc.2025.05.062","article-title":"Paclitaxel-coated balloon for the treatment of multilayer in-stent restenosis: AGENT IDE subgroup analysis","volume":"86","author":"Kirtane","year":"2025","journal-title":"J Am Coll Cardiol"},{"key":"B44","doi-asserted-by":"publisher","DOI":"10.4103\/hm.hm-d-24-00106","article-title":"Effects of a diet rich in polyphenols on lipid metabolism: an updated narrative review","volume":"9","author":"Scavuzzi","year":"2025","journal-title":"Heart Mind"},{"key":"B45","doi-asserted-by":"publisher","first-page":"5077","DOI":"10.1038\/s41467-023-40824-7","article-title":"CD36 mediates SARS-CoV-2-envelope-protein-induced platelet activation and thrombosis","volume":"14","author":"Tang","year":"2023","journal-title":"Nat Commun"},{"key":"B46","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2024.1447901","article-title":"Spleen tyrosine kinase: a novel pharmacological target for sepsis-induced cardiac dysfunction and multi-organ failure","volume":"15","author":"Verra","year":"2024","journal-title":"Front Immunol"},{"key":"B47","doi-asserted-by":"publisher","DOI":"10.1016\/j.cbi.2024.111010","article-title":"Inhibition of PARP1 improves cardiac function after myocardial infarction via up-regulated NLRC5","volume":"395","author":"Luo","year":"2024","journal-title":"Chem Biol Interact"}],"container-title":["Frontiers in Immunology"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fimmu.2026.1733284\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,9,3]],"date-time":"2026-09-03T07:17:37Z","timestamp":1788419857000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fimmu.2026.1733284\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,9,3]]},"references-count":47,"alternative-id":["10.3389\/fimmu.2026.1733284"],"URL":"https:\/\/doi.org\/10.3389\/fimmu.2026.1733284","relation":{},"ISSN":["1664-3224"],"issn-type":[{"value":"1664-3224","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,9,3]]},"article-number":"1733284"}}