Extracellular Vesicles from Oral Squamous Cell Carcinoma Carry OGT/OGA with Possible Implications in Tumor O-GlcNAcylation

Abstract

Oral squamous cell carcinoma (OSCC) is a malignant epithelial neoplasm characterized by high aggressiveness and limited options for early diagnosis. In recent years, extracellular vesicles (EVs) have gained attention as mediators of intercellular communication in cancer, contributing to tumor progression and remodeling of the microenvironment. O-GlcNAcylation, regulated by O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA), participates in multiple tumor processes; however, its association with EVs in OSCC has not yet been explored. In this study, EVs were isolated from SCC-152, SCC-25, and HaCaT cell lines using differential centrifugation, and their identity was confirmed by detection of CD63 and TSG101 markers and by transmission electron microscopy (TEM). Immunocytochemistry revealed the nuclear and cytoplasmic localization of OGT and OGA in all analyzed cell lines. Importantly, both enzymes were detected in EVs cargo by Western blot analysis, with significant differences between tumor and non-tumor lines as determined by densitometric and fluorescence intensity analyses. Quantitative analysis indicated a higher relative signal for OGA compared with OGT across all cell lines (with an approximate ~1.5-2.2-fold difference depending on the cell line, p < 0.05), and cell line-derived samples showed a higher relative signal than non-tumoral HaCaT (corresponding to an approximate ~1.2-1.3-fold increase under the experimental conditions evaluated). All experiments were performed using three independent biological replicates (n = 3), and statistical significance was assessed using one-way or two-way ANOVA followed by Tukey’s post hoc test. These findings suggest that OSCC-derived EVs carry enzymatic components of the O-GlcNAcylation machinery as vesicular protein cargo, potentially influencing tumor microenvironment regulation and cancer progression. Overall, these results should be considered hypothesis-generating, opening new perspectives for their use as vesicular biomarkers.

Keywords

Oral squamous cell carcinoma / O-GlcNAcylation / O-GlcNAc transferase (OGT) / O-GlcNAcase (OGA) / Extracellular vesicles

Statement of the Use of Generative AI and AI-Assisted Technologies in the Writing Process

During the preparation of this manuscript, the authors used ChatGPT solely to improve language clarity, coherence, and readability of certain sections of the text. After using this tool, the authors reviewed and edited the content as needed and take full responsibility for the content of the published article.

Acknowledgments

The authors thank the Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Mexico, for its support and research facilities.

Author Contributions

Conceptualization, M.C.C.-S. and M.C.-C.; Formal Analysis, D.S.C.-H. and M.C.-C.; Resources, M.C.-C. and D.S.C.-H.; Writing—Original Draft Preparation, M.C.C.-S.; Writing—Review & Editing, D.S.C.-H. and M.C.-C.; Visualization, M.C.-C. and D.S.C.-H.

Ethics Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

Data sharing is not applicable.

Funding

This research received no external funding.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

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