{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,28]],"date-time":"2026-09-28T22:36:44Z","timestamp":1790635004956,"version":"4.1.0"},"reference-count":33,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2026,8,4]],"date-time":"2026-08-04T00:00:00Z","timestamp":1785801600000},"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:sec>\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>Laryngeal squamous cell carcinoma (LSCC) remains a major clinical challenge with limited therapeutic options and unsatisfactory outcomes. Hypoxia-induced stress responses are known to contribute to tumor progression, immune remodeling, and therapeutic resistance. However, the cell type-specific hypoxia-associated transcriptional programs within the immune and stromal microenvironment of LSCC remain incompletely understood.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>We analyzed an immune-enriched LSCC single-cell RNA sequencing dataset (GSE150321, GSM4546858), generated by CD45-based magnetic bead sorting, to characterize hypoxia-associated stress response signatures across immune and stromal cell populations. The workflow included quality control, dimensionality reduction, unsupervised clustering, cell type annotation, pathway enrichment, hypoxia score assessment, robust rank aggregation, pseudotime trajectory inference, and cell\u2013cell communication analysis. Representative hypoxia-associated genes were further validated experimentally in LSCC-related cell models.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      Nine major cell populations were identified, including T cells, epithelial cells, B cells, macrophages, dendritic cells, cycling cells, fibroblasts, mast cells, and endothelial cells. Hypoxia-associated pathway activity was broadly enriched, with particularly strong activation in myeloid populations. Slingshot analysis using macrophages as the trajectory root revealed a monocyte-to-macrophage maturation trajectory, along which hypoxia scores were highest in macrophages and decreased toward lymphoid cells (Spearman R = \u22120.271, P\u00a0=\u00a01.78 \u00d7 10\n                      <jats:sup>-<\/jats:sup>\n                      \u00b9\n                      <jats:sup>7<\/jats:sup>\n                      ). Myeloid-specific hypoxia-associated genes, including PLA2G7, CD163, and APOC1, were identified. Robust rank aggregation further highlighted candidate hypoxia-associated genes such as HIF1A, CXCL8, S100A8, and ENO1. Experimental validation in THP-1 cells confirmed hypoxia-induced upregulation of these genes, and ELISA showed increased CXCL8 secretion under hypoxia (985 \u00b1 87 vs. 215 \u00b1 34 pg\/mL, P &amp;lt; 0.001). In contrast, the novel myeloid-specific genes showed negligible expression in Hep-2 epithelial cells, supporting their cell-type specificity.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>This study characterizes hypoxia-associated transcriptional programs within the immune and stromal microenvironment of LSCC rather than providing a comprehensive malignant epithelial tumor atlas. The findings highlight myeloid-centered hypoxia adaptation as potential sources of biomarker candidates and therapeutic insight for LSCC. THP-1 monocytic cell validation provides functional support for myeloid-centered hypoxia adaptation.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.3389\/fimmu.2026.1850654","type":"journal-article","created":{"date-parts":[[2026,8,4]],"date-time":"2026-08-04T05:37:53Z","timestamp":1785821873000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":1,"title":["Single-cell RNA sequencing reveals hypoxia-associated stress response signatures in the immune and stromal microenvironment of laryngeal squamous cell carcinoma"],"prefix":"10.3389","volume":"17","author":[{"given":"Huihui","family":"Du","sequence":"first","affiliation":[{"name":"Otorhinolaryngology, Wenzhou Hospital of Integrated Traditional Chinese and Western Medicine","place":["Wenzhou, Zhejiang, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yating","family":"Wang","sequence":"additional","affiliation":[{"name":"Department of Otolaryngology, The First Affiliated Hospital of Wenzhou Medical University","place":["Wenzhou, Zhejiang, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Renyu","family":"Lin","sequence":"additional","affiliation":[{"name":"Department of Otolaryngology, The First Affiliated Hospital of Wenzhou Medical University","place":["Wenzhou, Zhejiang, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jiaqi","family":"Yu","sequence":"additional","affiliation":[{"name":"Department of Otolaryngology, The First Affiliated Hospital of Wenzhou Medical University","place":["Wenzhou, Zhejiang, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kuaiquan","family":"Zhu","sequence":"additional","affiliation":[{"name":"Department of Otolaryngology, The First Affiliated Hospital of Wenzhou Medical University","place":["Wenzhou, Zhejiang, 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