Systemic autoimmune diseases can be modeled in transgenic mice harboring defects in DC apoptosis 10 but not in mice with apoptosis defects in T and B cells 11–13. Our study shows that in addition to the dogma of DC apoptosis as a mechanism to eliminate activated DC to prevent hyperactivation of the immune response, DC apoptosis also plays an

active role in induction and maintenance of tolerance through induction of Treg, whereby defects in DC apoptosis may trigger autoimmunity. High levels of spontaneous DC apoptosis have also been observed in breast cancer patients, with its significance being unclear 15, 16. Our study indicates that DC apoptosis in cancer patients may play a role in suppressing immune responses against the tumor by inducing immunosuppression and tolerance. Therefore, prevention Metformin research buy of DC apoptosis may enhance the therapeutic

effects of chemotherapy in tumor CH5424802 datasheet eradication 15, 16. Our findings may also represent a therapeutic approach in the prevention of unwanted immune responses in autoimmune diseases and transplantation along with inhibition of DC apoptosis to assist in tumor eradication. C57BL/6 mice were purchased from Charles River Laboratories (St. Constant, QC) and maintained as per guidelines of SickKids animal facilities. All the animal studies were reviewed and approved by the SickKids Institutional Committee for humane use of laboratory animals. OT-II mice were purchased from Jackson Laboratories (Bar Harbor, ME). The following antibodies were purchased from eBioscience (San Diego, CA): CD11c PE, CD86 PE, CD80 PE, MHC II PE, IL-10 Alexa647, IL-12 APC, IL-17 PE, Foxp3 PE along with neutralizing

IL-4 and IFN-γ Ab, and the following from BD Biosciences (Mississauga, ON): CD11c-FITC, CD4-FITC and CD3-PE. Anti-TGF-β neutralizing Ab (MAB1835) was obtained from R&D Systems (Minneapolis, MN). Isotype control IgG were obtained from eBioscience and/or PLEKHM2 Serotec (Raleigh, NC). CFSE was obtained from Molecular Probes (Burlington, ON); BrdU, OVA, cytochalasin D, rapamycin and PI were obtained from Sigma-Aldrich (Oakville, ON). GM-CSF was obtained from R&D Systems. IL-6 and TGF-β were obtained from Peprotech (Rocky Hill, NJ). Bone marrow cells were isolated from tibia and femurs of adult mice and cultured in the presence of GM-CSF for 7 days as described previously 34. DC were harvested and stained with 1 μM CFSE as described previously 35. Naïve CD4+CD25–CD62L+ T cells were isolated from spleens of mice using CD4+CD62L+ naïve T-cell isolation kit in conjunction with MACS columns from Miltenyi Biotec (Auburn, CA), following the manufacturer’s instructions. DC were cultured on a six-well dish and irradiated for 2 min with a UV transilluminator, with a peak intensity of 9000 mW/cm2 at the filter surface and a peak emission of 313 nm.