Cytokine Profiling in Low- and High-Density Small Extracellular Vesicles from Epidermoid Carcinoma Cells

Abstract Exosomes, or small extracellular vesicles (sEVs), are membrane-bound, nanoparticles that carry various macromolecules and act as autocrine and paracrine signaling messengers. Here, sEVs from epidermoid carcinoma cells influenced by membrane presentation of the glycoprotein desmoglein 2 (Dsg2) and its palmitoylation state were investigated. In this study, sEVs were isolated by sequential ultracentrifugation followed by iodixanol density gradient separation. They were then subjected to multiplex profiling of cytokines associated on the surface of intact sEVs. The results revealed a previously undescribed active sorting of cytokines onto the surface of low-density (LD) and high-density (HD) sEV subpopulations. Specifically, an altered surface presentation of Dsg2 decreased FGF-2 and VEGF in LD-sEVs. In addition, in response to Dsg2, IL-8 and RANTES were increased in LD-sEVs but only slightly decreased in HD-sEVs. Finally, IL-6 and G-CSF were increased dramatically in HD-sEVs. This comprehensive analysis of the cytokine production profile by SCC-derived sEVs highlights their contribution to immune evasion, pro-oncogenic and -angiogenic activity and the potential to identify diagnostic disease biomarkers.