A Study of the Direct Effect of Pegylated Graphene Oxide Nanoparticles and Fullerenol C60(OH)24 on the Differentiation of Regulatory T Cells In Vitro.

Regulatory T cells (Tregs) play a key role in immune tolerance and are promising targets for treating immune-mediated diseases. This study investigated the direct effects of PEGylated graphene oxide nanoparticles (LP-GO, BP-GO at 5-25 μg/mL) and fullerenol C60(OH)24 (25-200 μg/mL) on human Treg viability and differentiation in vitro. Tregs were induced from peripheral blood CD4+ T cells using IL-2, TGF-β, and CD2/CD3/CD28 activation beads for 72 h with nanoparticles. Assessments included viability, apoptosis (Zombie aqua/Annexin V), phenotype (CD45+CD4+CD25+CD127dim/-FOXP3+), nanoparticle sorption (intrinsic fluorescence), and IL-10 production. Neither PEGylated graphene oxide nor fullerenol C60(OH)24 affected T-helper (CD4+) viability (95.35-96.15%) nor early/late apoptosis levels. Despite this, we found a decrease in the percentage of CD4+ cells in cultures exposed to 50-200 μg/mL of fullerenol C60(OH)24. The percentage and absolute number of Treg cells decreased with 100-200 μg/mL of fullerenol, while IL-10 levels declined following treatment with 200 μg/mL of the same nanoparticles. Graphene oxide nanoparticles showed virtually no localization within or on cells. However, T helper and Treg cells demonstrated concentration-dependent sorption of fullerenol C60(OH)24 at concentrations of 100-200 μg/mL without a reduction in viability. These findings demonstrate good in vitro biocompatibility of the nanoparticles at pharmacological concentrations up to 25 μg/mL, alongside the inhibition of Treg differentiation with 100-200 μg/mL of fullerenol C60(OH)24.