Mutations in PTEN‐induced kinase 1 (PINK1), a serine/threonine kinase linked to familial early‐onset Parkinsonism, compromise mitochondrial integrity and metabolism and impair AKT signaling. As the activation of a naïve T cell requires an AKT‐dependent reorganization of a cell's metabolic machinery, we sought to determine if PINK1‐deficient T cells lack the ability to undergo activation and differentiation. We show that CD4⁺ T cells from PINK1 knockout mice fail to properly phosphorylate AKT upon activation, resulting in reduced expression of the IL‐2 receptor subunit CD25. Following, deficient IL‐2 signaling mutes the activation‐induced increase in respiratory capacity and mitochondrial membrane potential. Under polarization conditions favoring the development of induced regulatory T cells, PINK1^−/− T cells exhibit a reduced ability to suppress bystander T‐cell proliferation despite normal FoxP3 expression kinetics. Our results describe a critical role for PINK1 in integrating extracellular signals with metabolic state during T‐cell fate determination, and may have implications for the understanding of altered T‐cell populations and immunity during the progression of active Parkinson's disease or other immunopathologies.

Background.: Experimental and preclinical evidence suggest that adoptive transfer of regulatory T (Treg) cells could be an appropriate therapeutic strategy to induce tolerance and improve graft survival in transplanted patients. The University of Kentucky Transplant Service Line is developing a novel phase I/II clinical trial with ex vivo expanded autologous Treg cells as an adoptive cellular therapy in renal transplant recipients who are using everolimus (EVR)-based immunosuppressive regimen.Methods.: The aim of this study was to determine the mechanisms of action and efficacy of EVR for the development of functionally competent Treg cell-based adoptive immunotherapy in transplantation to integrate a common EVR-based regimen in vivo (in the patient) and ex vivo (in the expansion of autologous Treg cells). CD25⁺Treg cells were selected from leukapheresis product with a GMP-compliant cell separation system and placed in 5-day (short) or 21-day (long) culture with EVR or rapamycin (RAPA). Multi-parametric flow cytometry analyses were used to monitor the expansion rates, phenotype, autophagic flux, and suppressor function of the cells. phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin signaling pathway profiles of treated cells were analyzed by Western blot and cell bioenergetic parameters by extracellular flux analysis.Results.: EVR-treated cells showed temporary slower growth, lower metabolic rates, and reduced phosphorylation of protein kinase B compared with RAPA-treated cells. In spite of these differences, the expansion rates, phenotype, and suppressor function of long-term Treg cells in culture with EVR were similar to those with RAPA.Conclusions.: Our results support the feasibility of EVR to expand functionally competent Treg cells for their clinical use.

We describe a method for generating regulatory, memory and naïve T cells from a single human blood donor. Polarized Tregs can be...