343 Exosomes containing miRNAs from keratinocytes under oxidative stress contribute to the destruction of melanocytes and CD8+ T cell activation in vitiligo

Vitiligo is a skin disease characterized by the destruction of epidermal melanocytes. As the main constituent cells in the epidermis, keratinocytes facilitate melanocytes loss under oxidative stress by inducing melanocyte death or recruiting CD8+ T cells to destroy melanocytes. However, the effect of exosomes from keratinocytes under oxidative stress on the survival of melanocyte and the activity of CD8+T cells is unknown. Exosomes (Exo.T) from culture media of H2O2-treated human keratinocytes (HaCaT) were extracted using ultracentrifugation and identified using transmission electron microscopy (TEM), Nanoparticle tracking analysis (NTA) and Western blotting. The effect of Exo.T on the activity of CD8+T cells and the survival human melanocytes (PIG1) was tested. Small RNAs-seq was performed to screen the miRNAs enriched in Exo.T. The effect of the validated miRANs on the activity of CD8+T cells, the survival of melanocytes and the underlying mechanism was explored. Finally, the role of Exo.T in the progression of vitiligo mouse model was investigated. Exo.T were 60 to 200 nm in diameter, with double membranes, and expressed CD9, CD63, TSG101, Alix, and HSP70 exosome marker proteins. Oxidative stress enhanced the secretion of Exo.T (p<0.01). Functionally, Exo.T suppressed the survival of melanocytes while promoted the activation of CD8+ T cells (p<0.05). miR-31-3p enriched in Exo.T induced the destruction of melanocytes and inhibited melanogenesis through MITF signal axis while its promotive role in the activation of CD8+ T cells may depends on multiple pathways. Exo.T drove vitiligo progression in a robust mouse model of vitiligo (p<0.05). Exosomes containing miRNAs from keratinocytes under oxidative stress induce the destruction of melanocytes and the activation of CD8+T cell, as a result, contribute to vitiligo progression.