Whole-transcriptome sequencing reveals hypoxic esophageal squamous cell carcinoma-derived migrasomes driving...

Cancer-associated fibroblasts (CAFs) activated by intercellular communication contribute to the progression of esophageal squamous cell carcinoma (ESCC). Migrasomes represent a novel mode of intercellular communication. However, the characteristics of ESCC-derived migrasomes in tumor hypoxic microenvironments and their effects on CAFs remain unclear. Migrasomes were isolated from ESCC cells under normoxia/hypoxia, with TSPAN4-GFP labeling, transmission electron microscopy, nanoparticle tracking analysis, and western blot for validation. Whole-transcriptome sequencing analyzed hypoxic migrasome RNA profiles, and ceRNA networks were predicted via RNAhybrid and Miranda. The effect of migrasomes on CAFs was assessed using fluorescence tracing, reverse transcription‑quantitative polymerase chain reaction, Transwell migration, enzyme‑linked immunosorbent assay, and western blot. ESCC cells produced migrasomes. Although hypoxia did not alter their quantity or structure, it significantly altered their RNA cargo, changing the composition of mRNA, lncRNA, and circRNA. Differentially expressed mRNAs were enriched in "Response to hypoxia" and "HIF-1 signaling pathway." DElncRNAs were enriched in "Golgi to plasma membrane protein transport" and "Cell adhesion molecules pathway," while DEcircRNAs were enriched in "ubiquitin binding" and "chromatin remodeling." Predicted ceRNA networks were constructed using RNAhybrid and Miranda, involving 659 miRNAs, 24 lncRNAs, and 132 mRNAs. Fibroblasts internalized migrasomes and acquired a CAF-like phenotype, showing enhanced migration, elevated secretion of IL-1β/TGF-β, and increased CAF marker expression (α-SMA, COL1A1, COL3A1, FAP, PDGFRβ), with effects most pronounced under hypoxic migrasome treatment. This study characterized hypoxic migrasome whole transcriptome landscapes and suggested that hypoxic migrasomes may promote CAF-like changes in vitro, uncovering a novel ESCC-tumor microenvironment interaction mechanism and offering new perspectives for ESCC research.