ABSTRACT
The immune system contains a series of checks and balances that maintain tolerance and prevent autoimmunity. Sialic acid-binding Ig-type lectins (Siglecs) are cell surface receptors found on immune cells and inhibit inflammation by recruiting protein tyrosine phosphatases to ITIMs. Islet-resident macrophages express Siglec-E, and Siglec-E expression decreases on islet-resident macrophages as insulitis progresses in the NOD mouse. The sialyltransferase ST8Sia6 generates α-2,8-disialic acids that are ligands for Siglec-E in vivo. We hypothesized that engaging Siglec-E through ST8Sia6-generated ligands may inhibit the development of immune-mediated diabetes. Constitutive overexpression of ST8Sia6 in pancreatic ß cells mitigated hyperglycemia in the multiple low-dose streptozotocin model of diabetes, demonstrating that engagement of this immune receptor facilitates tolerance in the setting of inflammation and autoimmune disease.
Subject(s)
Diabetes Mellitus/chemically induced , Diabetes Mellitus/metabolism , Sialyltransferases/metabolism , Streptozocin/pharmacology , Animals , Antigens, Differentiation, B-Lymphocyte/immunology , Antigens, Differentiation, B-Lymphocyte/metabolism , Autoimmunity/immunology , Diabetes Mellitus/immunology , Diabetes Mellitus, Experimental/chemically induced , Diabetes Mellitus, Experimental/immunology , Diabetes Mellitus, Experimental/metabolism , Disease Models, Animal , Female , Humans , Hyperglycemia/immunology , Hyperglycemia/metabolism , Immune Tolerance/immunology , Inflammation/immunology , Inflammation/metabolism , Insulin-Secreting Cells/immunology , Insulin-Secreting Cells/metabolism , Ligands , Macrophages/metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Inbred NOD , Receptors, Immunologic/immunology , Receptors, Immunologic/metabolism , Sialic Acid Binding Ig-like Lectin 2/immunology , Sialic Acid Binding Ig-like Lectin 2/metabolism , Sialyltransferases/immunologyABSTRACT
Recent thymic emigrants that fail postpositive selection maturation are targeted by complement proteins. T cells likely acquire complement resistance during maturation in the thymus, a complement-privileged organ. To test this, thymocytes and fresh serum were separately obtained and incubated together in vitro to assess complement deposition. Complement binding decreased with development and maturation. Complement binding decreased from the double-positive thymocyte to the single-positive stage, and within single-positive thymocytes, complement binding gradually decreased with increasing intrathymic maturation. Binding of the central complement protein C3 to wild-type immature thymocytes required the lectin but not the classical pathway. Specifically, MBL2 but not MBL1 was required, demonstrating a unique function for MBL2. Previous studies demonstrated that the loss of NKAP, a transcriptional regulator of T cell maturation, caused peripheral T cell lymphopenia and enhanced complement susceptibility. To determine whether complement causes NKAP-deficient T cell disappearance, both the lectin and classical pathways were genetically ablated. This blocked C3 deposition on NKAP-deficient T cells but failed to restore normal cellularity, indicating that complement contributes to clearance but is not the primary cause of peripheral T cell lymphopenia. Rather, the accumulation of lipid peroxides in NKAP-deficient T cells was observed. Lipid peroxidation is a salient feature of ferroptosis, an iron-dependent nonapoptotic cell death. Thus, wild-type thymocytes naturally acquire the ability to protect themselves from complement targeting by MBL2 with maturation. However, NKAP-deficient immature peripheral T cells remain scarce in complement-deficient mice likely due to ferroptosis.
Subject(s)
Cell Differentiation/immunology , Complement C3/immunology , Mannose-Binding Lectin/immunology , Repressor Proteins/immunology , T-Lymphocytes/immunology , Animals , Lymphopenia/immunology , Mice , Mice, Inbred C57BL , Mice, Knockout , Thymocytes/immunology , Thymus Gland/immunology , Transcription, Genetic/immunologyABSTRACT
To generate functional peripheral T cells, proper gene regulation during T cell development is critical. In this study, we found that histone deacetylase (HDAC) 3 is required for T cell development. T cell development in CD2-icre HDAC3 conditional knockout (cKO) mice (HDAC3-cKO) was blocked at positive selection, resulting in few CD4 and CD8 T cells, and it could not be rescued by a TCR transgene. These single-positive thymocytes failed to upregulate Bcl-2, leading to increased apoptosis. HDAC3-cKO mice failed to downregulate retinoic acid-related orphan receptor (ROR) γt during positive selection, similar to the block in positive selection in RORγt transgenic mice. In the absence of HDAC3, the RORC promoter was hyperacetylated. In the periphery, the few CD4 T cells present were skewed toward RORγt(+) IL-17-producing Th17 cells, leading to inflammatory bowel disease. Positive selection of CD8 single-positive thymocytes was restored in RORγt-KO Bcl-xL transgenic HDAC3-cKO mice, demonstrating that HDAC3 is required at positive selection to downregulate RORγt.
Subject(s)
Cell Differentiation/immunology , Gene Expression Regulation/immunology , Histone Deacetylases/immunology , Nuclear Receptor Subfamily 1, Group F, Member 3/immunology , Thymocytes/cytology , Animals , Chromatin Immunoprecipitation , Down-Regulation , Flow Cytometry , Histone Deacetylases/metabolism , Mice , Mice, Knockout , Mice, Transgenic , Nuclear Receptor Subfamily 1, Group F, Member 3/metabolism , Thymocytes/immunologyABSTRACT
Recent thymic emigrants are newly generated T cells that need to undergo postthymic maturation to gain functional competency and enter the long-lived naive T cell pool. The mechanism of T cell maturation remains incompletely understood. Previously, we demonstrated that the transcriptional repressor NKAP is required for T cell maturation. Because NKAP associates with histone deacetylase 3 (HDAC3), we examined whether HDAC3 is also required for T cell maturation. Although thymic populations are similar in CD4-cre HDAC3 conditional knockout mice compared with wild-type mice, the peripheral numbers of CD4(+) and CD8(+) T cells are dramatically decreased. In the periphery, the majority of HDAC3-deficient naive T cells are recent thymic emigrants, indicating a block in T cell maturation. CD55 upregulation during T cell maturation is substantially decreased in HDAC3-deficient T cells. Consistent with a block in functional maturation, HDAC3-deficient peripheral T cells have a defect in TNF licensing after TCR/CD28 stimulation. CD4-cre HDAC3 conditional knockout mice do not have a defect in intrathymic migration, thymic egress, T cell survival, or homeostasis. In the periphery, similar to immature NKAP-deficient peripheral T cells, HDAC3-deficient peripheral T cells were bound by IgM and complement proteins, leading to the elimination of these cells. In addition, HDAC3-deficient T cells display decreases in the sialic acid modifications on the cell surface that recruit natural IgM to initiate the classical complement pathway. Therefore, HDAC3 is required for T cell maturation.
Subject(s)
Cell Differentiation , Histone Deacetylases/metabolism , T-Lymphocytes/cytology , T-Lymphocytes/metabolism , Animals , Bone Marrow Cells/metabolism , Cell Differentiation/genetics , Cell Differentiation/immunology , Cell Movement/genetics , Cell Movement/immunology , Complement Activation/immunology , Complement System Proteins/immunology , Histone Deacetylases/genetics , Homeostasis , Interleukin-7/metabolism , Lymphocyte Count , Mice , Mice, Knockout , Mice, Transgenic , T-Lymphocyte Subsets/cytology , T-Lymphocyte Subsets/immunology , T-Lymphocyte Subsets/metabolism , T-Lymphocytes/immunology , Thymus Gland/immunology , Thymus Gland/metabolism , Tumor Necrosis Factors/metabolismABSTRACT
For the past century, insulin injections have saved millions of lives, but glycemic instability is still a persistent challenge for people with diabetes, leading to tremendous morbidity and premature mortality. Research in the field of islet transplantation has demonstrated that replacing insulin-producing ß cells can restore euglycemia comparable to individuals without diabetes. However, a short supply of cadaveric islet donors, the technically challenging process of isolating islets, and the requirement for chronic immune suppression have impeded widespread clinical adoption. Rather than relying on cadaveric cells, pluripotent stem cells could serve as a virtually unlimited supply of insulin-producing ß cells. Protocols have been developed that mimic the normal in vivo development of the human pancreas to generate pancreatic progenitor cells in vitro. Ongoing investigations have yielded progressively more mature ß-like cells in vitro that produce insulin but do not yet fully mimic healthy mature ß cells. Alongside development of differentiation protocols, other work has provided insight into potential implantation sites for stem cell-derived islet cells including the subcutaneous space, portal vein, and omentum. To optimize implanted cell survival and function, development of immune modulation therapies is ongoing, including selection of immunomodulatory medications and genetic modification of implanted cells to evade immune responses. Further, macroencapsulation or microencapsulation devices could be used to contain and/or immunoprotect implanted cells from the immune response including by using 3-dimensional bioprinting to facilitate the process. Remarkably, ongoing clinical trials have now yielded the first patient relying on differentiated stem cells rather than syringes as their insulin replacement therapy.
Subject(s)
Insulin-Secreting Cells , Islets of Langerhans , Humans , Insulin , Stem Cells , Cell Differentiation , CadaverABSTRACT
The transcription factor Runx1 has essential roles throughout hematopoiesis. Here, we demonstrate that Runx1 is critical for T cell maturation. Peripheral naïve CD4(+) T cells from CD4-cre Runx1 cKO mice are phenotypically and functionally immature as shown by decreased production of TNF-α upon TCR stimulation. The loss of peripheral CD4(+) T cells in CD4-cre Runx1 cKO mice is not due to defects in homeostasis or decreased expression of IL-7Rα, as transgenic expression of IL-7Rα does not rescue the loss of CD4(+) T cells. Rather, immature Runx1-deficient CD4(+) T cells are eliminated in the periphery by the activation and fixation of the classical complement pathway. In the thymus, there is a severe block in all aspects of intrathymic T cell maturation, although both positive and negative selection are unaltered. Thus, loss of Runx1 leads to the earliest characterized block in post-positive selection intrathymic maturation of CD4 T cells.