P4HA1 facilitates SPP1+ tumor-associated macrophages by activating the hypoxia pathway in head and neck squamous cell carcinoma

Sep 29, 2026

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Tumor-associated macrophages (TAMs) with high secreted phosphoprotein 1 (SPP1) expression show immunosuppressive properties that may drive tumor metastasis, yet the regulation of SPP1+ TAMs in head and neck squamous cell carcinoma (HNSCC) remains unclear. Using single-cell and bulk RNA-seq bioinformatics, the study links SPP1+ TAM abundance with prognosis, identifies SPP1-associated key genes via weighted gene co-expression network analysis, and builds a Random Forest prognostic model.

LanguageEnglish

CategoryResearch & Report

FormatPDF

Pages17

Reading Time43 min

AccessFree to download

Background

TAMs and tumor microenvironment in HNSCC

SPP1 and hypoxia-associated macrophage roles

Methods Overview

Single-cell and bulk RNA-seq bioinformatics

WGCNA and prognostic modeling

[www. nature.com/scientificreports](www. nature.com/scientificreports) OPEN P4HA1 facilitates SPP1+ tumorassociated macrophages by activating the hypoxia pathway in head and neck squamous cell carcinoma Yiding Liu1,2,5, Bing Yan1,2,5, Hui Dong4,5, Yi Li1,2, Yuying Zhang1,2, Yi Wang1,2, Ni Kou1,3􀀍 & Lu Gao1,2􀀍 Tumor-associated macrophages (TAMs) with highly expressed secreted phosphoprotein 1 (SPP1) carry immunosuppressive property as a potential target for tumor metastasis. However, the mechanisms regulating SPP1 +TAMs in head and neck squamous cell carcinoma (HNSCC) remain poorly understood. This study employs a combination of single-cell and bulk RNA sequencing bioinformatics analysis to confirm the impact ofTAMs with high levels of SPP1 on patient prognosis. Additionally, Key genes linked to SPP1 + macrophages were identified using weighted gene co-expression network analysis. A prognostic model was built using the Random Forest algorithm. Here we show, P4HA1 is strongly correlated with SPP1 + macrophages and holds significant value in predicting patient prognosis and diagnosis. In vitro experiments demonstrated thatTAMs educated by HNSCC cells with knockdown P4HA1 expressed lower SPP1 level compared to the control group. Furthermore, Gene Set Variation Analysis and Gene Set Enrichment Analysis indicated that P4HA1 mediates the hypoxia pathway in HNSCC. In xenografts model, P4HA1 knockdown effectively suppressed tumor malignant progress, confirming that P4HA1 was positive correlation with SPP1 +TAMs and could mediate tumor hypoxia pathways. Overall, this study identified P4HA1 as a key gene involved in regulating SPP1 +TAMs through modulating hypoxia, providing a potential macrophage-centered therapeutic target in HNSCC. Keywords Tumor-associated macrophages, SPP1, P4HA1, Hypoxia, HIF-1α, HNSCC Abbreviations AUC Cocl2 GEO GSEA GSVA HNCHIF-1α P4HA1 PLS-GLMPCA PDXs ROS Area under the curve Cobalt chloride 2 Gene expression omnibus Gene set enrichment analysis Gene set variation analysis Head and neck cancers Hypoxia-inducible factor 1 alpha Prolyl 4-hydroxylase subunit alpha 1 Partial least squares generalized linear models Principal component analysis Patient-derived xenografts Reactive oxygen species 1School of Stomatology, Dalian Medical University, No. 9 West Section, Lvshun South Road, Dalian 116044, P. R. China. 2Technology Innovation Center of Immune and Oral Development & Regeneration, Dalian Medical University, Dalian 116044, China. 3School of Stomatology, The Affiliated Stomatological Hospital of Dalian Medical University, Dalian 116027, China. 4Department of Stomatology, Second Affiliated Hospital of Dalian Medical University, Dalian 116011, China. 5Yiding Liu, BingYan and Hui Dong contributed equally to this work. 􀀍 email: [kouni1984@163.com](kouni1984@163.com); [dygaolu@dmu.edu.cn](dygaolu@dmu.edu.cn) [www. nature.com/scientificreports/](www. nature.com/scientificreports/) SVM SPP1 SGLM TAMs TME TCGA UMAP WGCNAPHDs α-KG CA9 CM ROC Support vector machine Secreted phosphoprotein 1 Stepwise generalized linear model Tumor-associated macrophages Tumor microenvironment The cancer genome atlas Uniform manifold approximation and projection algorithm Weighted gene co-expression network analysis Oxygen-dependent dioxygenases Alpha-ketoglutarate Carbonic anhydrase 9 Confusion matrices Receiver operating characteristic curve Head and neck squamous cell carcinoma (HNSCC) accounts for approximately 90% of head and neck cancers (HNC) and is recognized as the sixth most common type of cancer worldwide1,2. Despite multidisciplinary treatment of locally advanced HNSCC by surgery combined with adjuvant chemoradiation or platinumbased concurrent chemoradiation, less than 50% of patients can be cured3. HNSCC is characterized by the heterogeneity of tumor tissues and the complexity of tumor microenvironment (TME)4. Tumor-associated macrophages (TAMs), the

FAQs

What is the study’s main question about SPP1+ TAMs in HNSCC?

The study investigates how SPP1+ tumor-associated macrophages are regulated in head and neck squamous cell carcinoma and how this relates to patient prognosis and diagnosis.

How does the research identify key genes related to SPP1+ macrophages?

It uses single-cell and bulk RNA sequencing bioinformatics, including weighted gene co-expression network analysis, to pinpoint genes linked to SPP1+ macrophages and prognosis.

What experimental evidence supports the role of P4HA1 and hypoxia signaling?

In vitro assays show that knocking down P4HA1 lowers SPP1 expression in TAMs educated by HNSCC cells, while gene set analyses indicate P4HA1 mediates hypoxia pathways; xenograft models further confirm reduced malignant progression after P4HA1 knockdown.

Sep 29, 2026

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