PLGA nanoparticles encapsulating TSHR-A and rapamycin enhance the induction of dendritic cell-specific immune tolerance in mice with Graves' disease

Abstract Dendritic cells (DCs) are the most potent antigen-presenting cells with multifaceted functions in controlling immune activation and tolerance. Graves' disease, particularly Graves' ophthalmopathy, is recognized as a refractory autoimmune thyroid disease. Therefore, DC-targeted therapies aimed at inducing specific immune tolerance are important for the treatment of Graves' disease. Therefore, we utilized polylactic acid glycolic acid polymer (PLGA) polymer nanoparticles encapsulating Graves' disease auto-antigen thyrotropin receptor A (TSHR-A) peptide and the immune tolerance inducer rapamycin (Rapa) to synthesize drug-loaded nanoparticles (NP(TSHR-A+Rapa)). We first characterized the synthesized nanodrugs using transmission electron microscopy (TEM) and dynamic light scattering (DLS) techniques and tested the uptake capacity of DCs for nanoparticles after co-culturing the nanoparticles with DCs. And the safe concentration of nanoparticles to DCs was detected using Cell counting kit-8 (CCK-8) assay. Subsequently, we tested the targeting and safety of the nanoparticles in mice. And the effects of nanoparticles on the proportion and proliferation of DCs and regulatory T (Treg) cells were examined in vivo and in vitro using flow cytometry (FCM) and 5-ethynyl-2’-deoxyuridine (EdU) method, respectively. Enzyme linked immunosorbent assay (ELISA) assays were used to detect the effect of nanoparticles on cytokine release from DCs. Finally, we tested the preventive and therapeutic effects of the synthesized nanoparticles on disease models. Our results showed that the synthesized nanoparticles were well taken up by DCs in vitro, while in vivo they were mainly targeted to the spleen of mice. The nanoparticles were able to relatively inhibit the maturation of DCs in vivo and in vitro, while affecting the release of relevant cellular functional factors from DCs, and the nanoparticles also promoted the proportion and proliferation of Treg cells in vivo and in vitro. In addition, the synthesized nanoparticles were able to prevent and improve the mouse disease model well without toxic side effects on mouse organs and other physiological indicators. Therefore, the synthesis of NP (TSHR-A+Rapa) nanoparticles using PLGA encapsulated TSHR-A and rapamycin could be used as targeting DCs to alter immune tolerance and as a new potential approach for the treatment of Graves' disease