Bovine trophoblast-derived extracellular vesicles modulate the gene expression profile of peripheral leukocytes

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This study aimed to determine whether trophoblast-derived extracellular vesicles (EVs) alter the gene expression and secretion profiles of inflammatory mediators in circulating leukocyte populations. Circulating leukocyte populations (CD14 + , CD4 + CD25 - , CD4 + CD25 + , CD8 + , and TCR-γδ + cells) were exposed to either isolated bovine trophoblast-derived EVs (tEV), EV-free trophoblast cell culture supernatant (nEV), or whole trophoblast culture supernatant (wSP) and assessed the transcription level and secretion of immune mediators. Extracellular vesicle concentrations measured by nanoparticle tracking analysis in the nEV, tEV, and wSP treatments were 1.78 × 10⁸, 9.81 × 10⁸, and 3.22 × 10⁸ particles/mL, respectively. Scanning electron microscopy confirmed the presence and morphology of EVs, which ranged in diameter from 25 to 308 nm, with most particles falling between 72 and 134 nm. Trophoblast-derived extracellular vesicles significantly altered gene expression in multiple immune cell populations, including CD8⁺ cytotoxic T cells, CD4⁺CD25⁻ resting T helper cells, and CD4⁺CD25⁺ activated T helper cells. In CD8⁺ cells, tEVs upregulated IL13 and showed trends toward increased FOXP3 and IL23 expression. CD4⁺CD25⁻ cells treated with tEVs exhibited increased IL6 and trends toward reduced IL1B and IL13 , while CD4⁺CD25⁺ cells showed elevated CXCL8 and tendencies for higher IL10, IFNG , and TNFA expression. No significant changes in gene expression were observed in TCR-γ/δ⁺ cells across treatments. Trophoblast-derived extracellular vesicles decreased the secretion of CCL4 and NCAM1. Trophoblast-derived EVs contain miRNAs and proteins involved in many biological processes including cell proliferation, stem cell biology, cell migration, and immune response. Among proteins were the EV markers and trophoblast markers. • Trophoblast extracellular vesicles modulate gene expression in leukocytes • Trophoblast extracellular vesicles enhance secretion of immune mediators in leukocytes • Immune signaling is regulated by trophoblast extracellular vesicles in pregnancy