RESUMEN
Background: Myeloid-derived suppressor cells (MDSCs) can prevent allograft rejection and induce immune tolerance in transplantation models. Previous studies have demonstrated that inhibition of mTOR signaling can enhance the MDSC protective effect in heart transplantation (HTx) by promoting MDSC expansion. In addition, mTOR inhibition is related to autophagy. The present study investigated the protective mechanism of mTOR-deficient monocytic MDSCs (M-MDSCs) in mouse HTx. Methods: Myeloid-specific mTOR conditional knockout mice were generated to obtain mTOR-/- M-MDSCs. The proliferation and immunosuppressive function of mTOR-/- M-MDSCs were determined by flow cytometry and T cell proliferation assays. The mTOR-/- M-MDSC intracellular autophagy levels were determined using western blotting and electron microscopy. RNAseq analysis was performed for wild-type (WT) and mTOR-/- M-MDSCs. Allogeneic HTx mouse model was established and treated with WT or mTOR-/- M-MDSCs. Enzyme-linked immunosorbent assay, flow cytometry, and immunohistochemistry assays were performed to determine WT and mTOR-/- M-MDSC-induced immune tolerance. Results: The mTOR deficiency promoted M-MDSC differentiation and enhanced intracellular autophagy levels in vivo and in vitro. mTOR deficiency also enhanced the immunosuppressive function of M-MDSCs. In addition, infusing with WT and mTOR-/- M-MDSCs prolonged cardiac allograft survival and established immune tolerance in recipient mice by inhibiting T cell activation and inducing regulatory T cells. Conclusion: mTOR deficiency enhances the immunosuppressive function of M-MDSCs and prolongs mouse cardiac allograft survival.
Asunto(s)
Diferenciación Celular/inmunología , Trasplante de Corazón/métodos , Células Supresoras de Origen Mieloide/inmunología , Serina-Treonina Quinasas TOR/inmunología , Tolerancia al Trasplante/inmunología , Aloinjertos/inmunología , Animales , Autofagia/genética , Autofagia/inmunología , Diferenciación Celular/genética , Proliferación Celular , Expresión Génica/inmunología , Tolerancia Inmunológica/genética , Tolerancia Inmunológica/inmunología , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Microscopía Electrónica de Transmisión , Células Supresoras de Origen Mieloide/metabolismo , Células Supresoras de Origen Mieloide/ultraestructura , Linfocitos T/inmunología , Linfocitos T/metabolismo , Serina-Treonina Quinasas TOR/deficiencia , Serina-Treonina Quinasas TOR/genética , Tolerancia al Trasplante/genéticaRESUMEN
Ubiquitinated proteins carried by the extracellular vesicles (EV) released by myeloid-derived suppressor cells (MDSC) have been investigated using proteomic strategies to examine the effect of tumor-associated inflammation. EV were collected from MDSC directly following isolation from tumor-bearing mice with low and high inflammation. Among the 1092 proteins (high inflammation) and 925 proteins (low inflammation) identified, more than 50% were observed as ubiquitinated proteoforms. More than three ubiquitin-attachment sites were characterized per ubiquitinated protein, on average. Multiple ubiquitination sites were identified in the pro-inflammatory proteins S100 A8 and S100 A9, characteristic of MDSC and in histones and transcription regulators among other proteins. Spectral counting and pathway analysis suggest that ubiquitination occurs independently of inflammation. Some ubiquitinated proteins were shown to cause the migration of MDSC, which has been previously connected with immune suppression and tumor progression. Finally, MDSC EV are found collectively to carry all the enzymes required to catalyze ubiquitination, and the hypothesis is presented that a portion of the ubiquitinated proteins are produced in situ.