RESUMEN
The immune system of healthy individuals is capable of regulating autoimmunity through multiple mechanisms. In Type 1 Diabetes (T1D) we recently discovered natural IgM, although present at normal levels, is unable to perform its normal immunoregulatory function. Treating diabetic mice with IgM from healthy donors led to reversal of disease without immune depletion. To investigate the therapeutic potential of a human preparation of IgM, we administered an IgM-enriched preparation of immunoglobulin called Pentaglobin. Administration of Pentaglobin therapy reversed disease in diabetic NOD mice and boosted CD4 + Foxp3 + Tregs. Importantly, the impact of Pentaglobin on the immune system was limited to inhibiting beta cell destruction but was not immune depleting nor did it inhibit the immunization response to an irrelevant antigen. These findings indicate that inhibition of deleterious autoimmunity in T1D is possible while leaving protective immunity fully intact.
Asunto(s)
Diabetes Mellitus Experimental , Diabetes Mellitus Tipo 1 , Animales , Diabetes Mellitus Experimental/tratamiento farmacológico , Humanos , Inmunoglobulina A , Inmunoglobulina M , Ratones , Ratones Endogámicos NODRESUMEN
Autoimmune disease has presented an insurmountable barrier to restoration of durable immune tolerance. Previous studies indicate that chronic therapy with metabolic inhibitors can reduce autoimmune inflammation, but it remains unknown whether acute metabolic modulation enables permanent immune tolerance to be established. In an animal model of lupus, we determined that targeting glucose metabolism with 2-deoxyglucose (2DG) and mitochondrial metabolism with metformin enables endogenous immune tolerance mechanisms to respond to tolerance induction. A 2-week course of 2DG and metformin, when combined with tolerance-inducing therapy anti-CD45RB, prevented renal deposition of autoantibodies for 6 months after initial treatment and restored tolerance induction to allografts in lupus-prone mice. The restoration of durable immune tolerance was linked to changes in T cell surface glycosylation patterns, illustrating a role for glycoregulation in immune tolerance. These findings indicate that metabolic therapy may be applied as a powerful preconditioning to reinvigorate tolerance mechanisms in autoimmune and transplant settings that resist current immune therapies.
Asunto(s)
Antimetabolitos/farmacología , Linfocitos T CD4-Positivos/inmunología , Desoxiglucosa/farmacología , Hipoglucemiantes/farmacología , Tolerancia Inmunológica/efectos de los fármacos , Riñón/efectos de los fármacos , Antígenos Comunes de Leucocito/inmunología , Lupus Eritematoso Sistémico/inmunología , Lupus Eritematoso Sistémico/metabolismo , Metformina/farmacología , Animales , Anticuerpos/farmacología , Modelos Animales de Enfermedad , Glucólisis/efectos de los fármacos , Glicosilación/efectos de los fármacos , Tolerancia Inmunológica/inmunología , Riñón/inmunología , Antígenos Comunes de Leucocito/antagonistas & inhibidores , Ratones , Fosforilación Oxidativa/efectos de los fármacos , Tolerancia al Trasplante/efectos de los fármacos , Tolerancia al Trasplante/inmunología , Trasplante HomólogoRESUMEN
The purpose of this study was to assess insulin-stimulated gene expression in canine skeletal muscle with a particular focus on NPPC, the gene that encodes C-type natriuretic peptide, a key hormonal regulator of cardiometabolic function. Four conscious canines underwent hyperinsulinemic, euglycemic clamp studies. Skeletal muscle biopsy and arterial plasma samples were collected under basal and insulin-stimulated conditions. Bulk RNA sequencing of muscle tissue was performed to identify differentially expressed genes between these 2 steady-state conditions. Our results showed that NPPC was the most highly expressed gene in skeletal muscle in response to insulin infusion, rising 4-fold between basal and insulin-stimulated conditions. In support of our RNA sequencing data, we found that raising the plasma insulin concentration 15-fold above basal elicited a 2-fold (P = 0.0001) increase in arterial plasma concentrations of N-terminal prohormone C-type natriuretic peptide. Our data suggest that insulin may play a role in stimulating secretion of C-type natriuretic peptide by skeletal muscle. In this context, C-type natriuretic peptide may act in a paracrine manner to facilitate muscle-vascular bed crosstalk and potentiate insulin-mediated vasodilation. This could serve to enhance insulin and glucose delivery, particularly in the postprandial absorptive state.
RESUMEN
Dinuclear metallodrugs offer much potential in the development of novel anticancer chemotherapeutics as a result of the distinct interactions possible with bio-macromolecular targets and the unique biological activity that can result. Herein, we describe the development of isostructural homo-dinuclear OsII -OsII and hetero-dinuclear OsII -RuII organometallic complexes formed from linking the arene ligands of [M(η6 -arene)(C2 O4 )(PTA)] units (M=Os/Ru; PTA=1,3,5-triaza-7-phosphaadamantane). Using these complexes together with the known RuII -RuII analogue, a chromatin-modifying agent, we probed the impact of varying the metal ions on the structure, reactivity and biological activity of these complexes. The complexes were structurally characterised by X-ray diffraction experiments, their stability and reactivity were examined by using 1 H and 31 Pâ NMR spectroscopy, and their biological activity was assessed, alongside that of mononuclear analogues, through MTT assays and cell-cycle analysis (HT-29 cell line). The results revealed high antiproliferative activity in each case, with cell-cycle profiles of the dinuclear complexes found to be similar to that for untreated cells, and similar but distinct profiles for the mononuclear complexes. These results indicate these complexes impact on cell viability predominantly through a non-DNA-damaging mechanism of action. The new OsII -OsII and OsII -RuII complexes reported here are further examples of a family of compounds operating via mechanisms of action atypical of the majority of metallodrugs, and which have potential as tools in chromatin research.
Asunto(s)
Antineoplásicos , Compuestos Organometálicos , Osmio , Rutenio , Antineoplásicos/farmacología , Línea Celular Tumoral , Células HEK293 , Células HT29 , Humanos , Ligandos , Espectroscopía de Resonancia Magnética , Compuestos Organometálicos/farmacología , Difracción de Rayos XRESUMEN
Unchecked collaboration between islet-reactive T and B lymphocytes drives type 1 diabetes (T1D). In the healthy setting, CD8 T regulatory cells (Tregs) terminate ongoing T-B interactions. We determined that specific CD8 Tregs from NOD mice lack suppressive function, representing a previously unreported regulatory cell deficit in this T1D-prone strain. NOD mice possess 11-fold fewer Ly-49+ CD8 Tregs than nonautoimmune mice, a deficiency that worsens as NOD mice age toward diabetes and leaves them unable to regulate CD4 T follicular helper cells. As IL-15 is required for Ly-49+ CD8 Treg development, we determined that NOD macrophages inadequately trans-present IL-15. Despite reduced IL-15 trans-presentation, NOD Ly-49+ CD8 Tregs can effectively transduce IL-15-mediated survival signals when they are provided. Following stimulation with an IL-15/IL-15Ra superagonist complex, Ly-49+ CD8 Tregs expanded robustly and became activated to suppress the Ag-specific Ab response. IL-15/IL-15Ra superagonist complex-activated CD8+CD122+ T cells also delayed diabetes transfer, indicating the presence of an underactivated CD8 T cell subset with regulatory capacity against late stage T1D. We identify a new cellular contribution to anti-islet autoimmunity and demonstrate the correction of this regulatory cell deficit. Infusion of IL-15-activated CD8 Tregs may serve as an innovative cellular therapy for the treatment of T1D.
Asunto(s)
Linfocitos B/inmunología , Diabetes Mellitus Tipo 1/inmunología , Inmunoterapia Adoptiva/métodos , Macrófagos/inmunología , Linfocitos T Reguladores/inmunología , Traslado Adoptivo , Animales , Antígenos CD8/metabolismo , Células Cultivadas , Modelos Animales de Enfermedad , Humanos , Interleucina-15/metabolismo , Ratones , Ratones Endogámicos NOD , Subfamilia A de Receptores Similares a Lectina de Células NK/metabolismo , Linfocitos T Reguladores/trasplanteRESUMEN
Imatinib (Gleevec) reverses type 1 diabetes (T1D) in NOD mice and is currently in clinical trials in individuals with recent-onset disease. While research has demonstrated that imatinib protects islet ß cells from the harmful effects of ER stress, the role the immune system plays in its reversal of T1D has been less well understood, and specific cellular immune targets have not been identified. In this study, we demonstrate that B lymphocytes, an immune subset that normally drives diabetes pathology, are unexpectedly required for reversal of hyperglycemia in NOD mice treated with imatinib. In the presence of B lymphocytes, reversal was linked to an increase in serum insulin concentration, but not an increase in islet ß cell mass or proliferation. However, improved ß cell function was reflected by a partial recovery of MafA transcription factor expression, a sensitive marker of islet ß cell stress that is important to adult ß cell function. Imatinib treatment was found to increase the antioxidant capacity of B lymphocytes, improving reactive oxygen species (ROS) handling in NOD islets. This study reveals a novel mechanism through which imatinib enables B lymphocytes to orchestrate functional recovery of T1D ß cells.
Asunto(s)
Linfocitos B/efectos de los fármacos , Linfocitos B/inmunología , Diabetes Mellitus Tipo 1/tratamiento farmacológico , Diabetes Mellitus Tipo 1/inmunología , Mesilato de Imatinib/farmacología , Islotes Pancreáticos/efectos de los fármacos , Islotes Pancreáticos/inmunología , Animales , Autoinmunidad/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Proteínas de Homeodominio/genética , Hiperglucemia , Mesilato de Imatinib/uso terapéutico , Insulina/sangre , Células Secretoras de Insulina/metabolismo , Factores de Transcripción Maf de Gran Tamaño/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Ratones NoqueadosRESUMEN
Autoimmune diseases such as type 1 diabetes (T1D) arise from unrestrained activation of effector lymphocytes that destroy target tissues. Many efforts have been made to eliminate these effector lymphocytes, but none has produced a long-term cure. An alternative to depletion therapy is to enhance endogenous immune regulation. Among these endogenous alternatives, naturally occurring Igs have been applied for inflammatory disorders but have lacked potency in antigen-specific autoimmunity. We hypothesized that naturally occurring polyclonal IgMs, which represent the majority of circulating, noninduced antibodies but are present only in low levels in therapeutic Ig preparations, possess the most potent capacity to restore immune homeostasis. Treatment of diabetes-prone NOD mice with purified IgM isolated from Swiss Webster (SW) mice (nIgMSW) reversed new-onset diabetes, eliminated autoreactive B lymphocytes, and enhanced regulatory T-cell (Treg) numbers both centrally and peripherally. Conversely, IgM from prediabetic NOD mice could not restore this endogenous regulation, which represents an unrecognized component of T1D pathogenesis. Of note, IgM derived from healthy human donors was similarly able to expand human CD4 Tregs in humanized mice and produced permanent diabetes protection in treated NOD mice. Overall, these studies demonstrate that a potent, endogenous regulatory mechanism, nIgM, is a promising option for reversing autoimmune T1D in humans.
Asunto(s)
Linfocitos B/inmunología , Diabetes Mellitus Tipo 1/tratamiento farmacológico , Inmunoglobulina M/uso terapéutico , Linfocitos T Reguladores/inmunología , Animales , Diabetes Mellitus Tipo 1/inmunología , Inmunoglobulina M/inmunología , Ratones , Ratones Endogámicos NODRESUMEN
Disruption of the non-classical Major Histocompatibility Complex (MHC) Ib molecule Qa-1 impairs CD8 Treg and natural killer (NK) cell function and promotes a lupus-like autoimmune disease. This immune perturbation would be expected to enhance anti-transplant responses and impair tolerance induction, but the effect of Qa-1 deficiency on the transplant response has not been previously reported. Qa-1 deficiency enhanced CD4 TFH and germinal center (GC) B cell numbers in naïve mice and hastened islet allograft rejection. Despite enhanced immunity in B6.Qa-1-/- mice, these mice did not generate an excessive primary CD4 TFH cell response nor an enhanced alloantibody reaction. Both CD8 Tregs and NK cells, which often regulate other cells through host Qa-1 expression, were targets of anti-CD45RB therapy that had not been previously recognized. However, B6.Qa-1-/- mice remained susceptible to anti-CD45RB mediated suppression of the alloantibody response and transplant tolerance induction to mismatched islet allografts. Overall, despite enhanced immunity as demonstrated by augmented CD4 TFH/GC B cell numbers and hastened islet allograft rejection in naïve 12-week old Qa-1 deficient mice, the CD8 Treg/NK cell restriction element Qa-1 does not regulate the primary cellular or humoral alloresponse and is not required for long-term transplant tolerance.
Asunto(s)
Antígenos de Histocompatibilidad Clase I/metabolismo , Tolerancia Inmunológica , Células Asesinas Naturales/inmunología , Linfocitos T Reguladores/inmunología , Trasplante , Animales , Linfocitos B/inmunología , Linfocitos T CD4-Positivos/inmunología , Antígenos de Histocompatibilidad Clase I/genética , Ratones Endogámicos C57BL , Ratones NoqueadosRESUMEN
Overcoming the immune response to establish durable immune tolerance in type 1 diabetes remains a substantial challenge. The ongoing effector immune response involves numerous immune cell types but is ultimately orchestrated and sustained by the hematopoietic stem cell (HSC) niche. We therefore hypothesized that tolerance induction also requires these pluripotent precursors. In this study, we determined that the tolerance-inducing agent anti-CD45RB induces HSC mobilization in nonautoimmune B6 mice but not in diabetes-prone NOD mice. Ablation of HSCs impaired tolerance to allogeneic islet transplants in B6 recipients. Mobilization of HSCs resulted in part from decreasing osteoblast expression of HSC retention factors. Furthermore, HSC mobilization required a functioning sympathetic nervous system; sympathectomy prevented HSC mobilization and completely abrogated tolerance induction. NOD HSCs were held in their niche by excess expression of CXCR4, which, when blocked, led to HSC mobilization and prolonged islet allograft survival. Overall, these findings indicate that the HSC compartment plays an underrecognized role in the establishment and maintenance of immune tolerance, and this role is disrupted in diabetes-prone NOD mice. Understanding the stem cell response to immune therapies in ongoing human clinical studies may help identify and maximize the effect of immune interventions for type 1 diabetes.
Asunto(s)
Movilización de Célula Madre Hematopoyética , Trasplante de Células Madre Hematopoyéticas/métodos , Aloinjertos/inmunología , Aloinjertos/metabolismo , Animales , Femenino , Citometría de Flujo , Trasplante de Islotes Pancreáticos/inmunología , Antígenos Comunes de Leucocito/metabolismo , Ratones , Ratones Endogámicos NOD , Osteoblastos/metabolismoRESUMEN
BACKGROUND: Interactions between genetic risk factors and the environment drive type 1 diabetes (T1D). The system of Toll-like receptors (TLR) detects these environmental triggers; however, the target cell that intermediates these interactions to drive T1D remains unknown. METHODS: We investigated the effect of TLR pathway activation (myeloid differentiation primary response 88 [MyD88] vs TIR-domain-containing adapter-inducing interferon-ß [TRIF]) on B cell subsets via flow cytometry, including their activation, survival, proliferation, and cytoskeletal mobilization. The effect of polyinosinic-polycytidylic acid (poly(I:C)) on diabetes development was addressed, including the B cell-dependent activation of diabetes-protective DX5+ cells, using genetic models and adoptive transfer. RESULTS: B lymphocytes from non-obese diabetic (NOD) mice expressed enhanced levels of TLR-responsive proteins. Ex vivo analysis of B lymphocyte subsets demonstrated that TLR3 stimulation via TRIF deletes cells exhibiting a marginal zone phenotype, whereas MyD88-dependent ligands enhance their survival. In vivo, marginal zone B cells were activated by poly(I:C) and were unexpectedly retained in the spleen of NOD mice, in contrast with the mobilization of these cells in non-autoimmune mice, a phenotype we traced to defective actin cytoskeletal dynamics. These activated B cells mediated TLR3-induced diabetes protection. CONCLUSIONS: Immunotherapies must account for both B cell location and activation, and these properties may differ in autoimmune and healthy settings.
Asunto(s)
Antivirales/farmacología , Linfocitos B/inmunología , Diabetes Mellitus/prevención & control , Poli I-C/farmacología , Receptor Toll-Like 3/metabolismo , Proteínas Adaptadoras del Transporte Vesicular/metabolismo , Animales , Diabetes Mellitus/metabolismo , Femenino , Citometría de Flujo , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Ratones SCID , Transducción de Señal/efectos de los fármacosRESUMEN
A series of trivalent lanthanide hydroxysulfates, Ln(OH)SO(4), (Ln = Pr through Yb, except radioactive Pm) has been synthesized via hydrothermal methods from Ln(2)(SO(4))(3)·8H(2)O by reaction with aqueous NaOH at 170 °C in Teflon lined Parr steel autoclaves, and were characterized by single crystal X-ray diffraction and FT-IR spectroscopy. Two types of arrangements were found in the solid state. The lighter (Ln = Pr-Nd, Sm-Gd) and heavier lanthanide(III) hydroxysulfates (Tb-Yb) are each isostructural. Both structure types exhibit the monoclinic space group P2(1)/n, but the unit cell content is doubled with two crystallographically distinct LnO(8) polyhedra for the heavier lanthanide compounds. The lighter complexes maintain the coordination number 9, forming a three-dimensional extended lattice. The heavier counterparts exhibit the coordination number 8, and arrange as infinite columns of two crystallographically different LnO(8) polyhedra, while extending along the "c" axis. These columns of LnO(8) polyhedra are surrounded and separated by six columns of sulfate ions, also elongating in the "c" direction. The rigid sulfate entities seem to obstruct the closing in of the lighter LnO(9) polyhedra, and show an inclining degree of torsion into the "ac" layers. The crystal lattice of the lighter 4f complexes can sufficiently withstand the tension buildup, caused by the decreasing Ln(3+) radius, up to Gd(OH)SO(4). The energy profile of this structural arrangement then seems to exceed levels at which this structure type is favorable. The lattice arrangement of the heavier Ln-analogues seems to offer a lower energy profile. This appears to be the preferred arrangement for the heavier lanthanide hydroxysulfates, whose crystal lattice exhibits more flexibility, as the coordination sphere of these analogues is less crowded. The IR absorbance frequencies of the hydroxide ligands correlate as a function of the Ln(3+) ionic radius. This corresponds well with the X-ray single crystal analysis data.