RESUMO
T follicular helper (TFH) cells are critical in adaptive immune responses to pathogens and vaccines; however, what drives the initiation of their developmental program remains unclear. Studies suggest that a T cell antigen receptor (TCR)-dependent mechanism may be responsible for the earliest TFH cell-fate decision, but a critical aspect of the TCR has been overlooked: tonic TCR signaling. We hypothesized that tonic signaling influences early TFH cell development. Here, two murine TCR-transgenic CD4+ T cells, LLO56 and LLO118, which recognize the same antigenic peptide presented on major histocompatibility complex molecules but experience disparate strengths of tonic signaling, revealed low tonic signaling promotes TFH cell differentiation. Polyclonal T cells paralleled these findings, with naive Nur77 expression distinguishing TFH cell potential. Two mouse lines were also generated to both increase and decrease tonic signaling strength, directly establishing an inverse relationship between tonic signaling strength and TFH cell development. Our findings elucidate a central role for tonic TCR signaling in early TFH cell-lineage decisions.
Assuntos
Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais , Células T Auxiliares Foliculares/imunologia , Células T Auxiliares Foliculares/metabolismo , Animais , Antígenos/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Diferenciação Celular/imunologia , Antígenos H-2/imunologia , Imunização , Imunofenotipagem , Ativação Linfocitária/imunologia , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Peptídeos/imunologiaRESUMO
In this issue of Immunity, Van Braeckel-Budimmir et al. (2017) reveal that the pathogenic response of mice to a Plasmodium berghei infection is dominated by a Vß8.1 T cell response. Mice lacking Vß8.1 T cells fail to mount a pathogenic response, thus showing that the TCR locus can be an Immune response (Ir) gene.
Assuntos
Malária , Receptores de Antígenos de Linfócitos T alfa-beta , Animais , Camundongos , Receptores de Antígenos de Linfócitos T , Linfócitos TRESUMO
Intricate processes in the thymus and periphery help curb the development and activation of autoreactive T cells. The subtle signals that govern these processes are an area of great interest, but tuning TCR sensitivity for the purpose of affecting T cell behavior remains technically challenging. Previously, our laboratory described the derivation of two TCR-transgenic CD4 T cell mouse lines, LLO56 and LLO118, which recognize the same cognate Listeria epitope with the same affinity. Despite the similarity of the two TCRs, LLO56 cells respond poorly in a primary infection whereas LLO118 cells respond robustly. Phenotypic examination of both lines revealed a substantial difference in their surface of expression of CD5, which serves as a dependable readout of the self-reactivity of a cell. We hypothesized that the increased interaction with self by the CD5-high LLO56 was mediated through TCR signaling, and was involved in the characteristic weak primary response of LLO56 to infection. To explore this issue, we generated an inducible knock-in mouse expressing the self-sensitizing voltage-gated sodium channel Scn5a. Overexpression of Scn5a in peripheral T cells via the CD4-Cre promoter resulted in increased TCR-proximal signaling. Further, Scn5a-expressing LLO118 cells, after transfer into BL6 recipient mice, displayed an impaired response during infection relative to wild-type LLO118 cells. In this way, we were able to demonstrate that tuning of TCR sensitivity to self can be used to alter in vivo immune responses. Overall, these studies highlight the critical relationship between TCR-self-pMHC interaction and an immune response to infection.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Animais , Antígenos CD5/imunologia , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Canal de Sódio Disparado por Voltagem NAV1.5/imunologia , Receptores de Antígenos de Linfócitos T/imunologiaRESUMO
Germline-encoded receptors recognizing common pathogen-associated molecular patterns are a central element of the innate immune system and play an important role in shaping the host response to infection. Many of the innate immune molecules central to these signaling pathways are evolutionarily conserved. LysMD3 is a novel molecule containing a putative peptidoglycan-binding domain that has orthologs in humans, mice, zebrafish, flies, and worms. We found that the lysin motif (LysM) of LysMD3 is likely related to a previously described peptidoglycan-binding LysM found in bacteria. Mouse LysMD3 is a type II integral membrane protein that co-localizes with GM130+ structures, consistent with localization to the Golgi apparatus. We describe here two lines of mLysMD3-deficient mice for in vivo characterization of mLysMD3 function. We found that mLysMD3-deficient mice were born at Mendelian ratios and had no obvious pathological abnormalities. They also exhibited no obvious immune response deficiencies in a number of models of infection and inflammation. mLysMD3-deficient mice exhibited no signs of intestinal dysbiosis by 16S analysis or alterations in intestinal gene expression by RNA sequencing. We conclude that mLysMD3 contains a LysM with cytoplasmic orientation, but we were unable to define a physiological role for the molecule in vivo.
Assuntos
Deleção de Genes , Animais , Autoantígenos/análise , Infecções Bacterianas/genética , Infecções Bacterianas/imunologia , Sistemas CRISPR-Cas , Feminino , Imunidade Inata , Inflamação/genética , Inflamação/imunologia , Masculino , Proteínas de Membrana/análise , Camundongos , Micoses/genética , Micoses/imunologia , Filogenia , Viroses/genética , Viroses/imunologiaRESUMO
Extrafollicular (EF) B-cell responses are increasingly being recognized as an alternative pathway of B-cell activation, particularly in autoimmunity. Critical cellular interactions required for the EF B-cell response are unclear. A key question in autoimmunity, in which Toll-like receptor (TLR) signals are costimulatory and could be sufficient for B-cell activation, is whether T cells are required for the response. This is pivotal, because autoreactive B cells are considered antigen-presenting cells for autoreactive T cells, but where such interactions occur has not been identified. Here, using AM14 site-directed transgenic rheumatoid factor (RF) mice, we report that B cells can be activated, differentiate, and isotype-switch independent of antigen-specific T-cell help, αß T cells, CD40L signaling, and IL-21 signaling to B cells. However, T cells do dramatically enhance the response, and this occurs via CD40L and IL-21 signals. Surprisingly, the response is completely inducible T-cell costimulator ligand independent. These results establish that, although not required, T cells substantially amplify EF autoantibody production and thereby implicate T-independent autoreactive B cells as a potential vector for breaking T-cell tolerance. We suggest that these findings explain why autoreactivity first focuses on self-components for which B cells carry TLR ligands, because these will uniquely be able to activate B cells independently of T cells, with subsequent T-B interactions activating autoreactive T cells, resulting in chronic autoimmunity.
Assuntos
Autoimunidade , Linfócitos B/imunologia , Linfócitos T/imunologia , Receptores Toll-Like/metabolismo , Transferência Adotiva , Animais , Ligantes , Ativação Linfocitária , Cooperação Linfocítica/imunologia , Linfopenia/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Camundongos Transgênicos , Receptores de Antígenos de Linfócitos T alfa-beta/deficiência , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Fator Reumatoide/imunologia , Transdução de Sinais/imunologiaRESUMO
BACKGROUND The presence of anatomical variations of the hepatic artery poses a challenge for normothermic machine perfusion (NMP). Here, we describe our experience of creating a single arterial cannulation for NMP in 3 donor livers with replaced right hepatic artery. CASE REPORT Three donor livers with replaced right hepatic artery were perfused using NMP (OrganOx® metra®) for liver transplantation. To maintain hepatic artery integrity and establish an intact arterial vascular inflow for NMP, a single vasculature was created to allow single arterial cannulation for NMP. A piece of intravenous-line tubing was used as a bridge from the splenic artery to the superior mesenteric artery during the back-table preparation. After 1 h of NMP, the lactate of 2 livers decreased from >10.0 to about 1.0 mmol/L, and the lactate of 1 liver decreased from >4.0 to <0.4 mmol/L. Three livers made >100 mL of bile after 4 h of NMP and were successfully implanted after >10 h of NMP. The recipients spent 2, 3, and 4 days in the Intensive Care Unit and were discharged home at 6, 7, and 9 days, respectively. None of the patients experienced early allograft dysfunction or any early technical complication or non-anastomotic biliary stricture. CONCLUSIONS Creating an intravenous-line tubing bridge from the splenic artery to the superior mesenteric artery prior to NMP of liver grafts associated with replaced right hepatic artery could reduce the cold ischemia time associated with vessel reconstruction and reduce bleeding risk during NMP. This is feasible, safe, and effective.
Assuntos
Transplante de Fígado , Humanos , Artéria Hepática/cirurgia , Artéria Esplênica/cirurgia , Artéria Mesentérica Superior/cirurgia , Preservação de Órgãos , Doadores Vivos , Fígado/irrigação sanguínea , Perfusão , Cateterismo , Lactatos , MesentérioRESUMO
The contribution of self-peptide-MHC signaling in CD4+ T cells to metabolic programming has not been definitively established. In this study, we employed LLO118 and LLO56, two TCRtg CD4+ T cells that recognize the same Listeria epitope. We previously have shown that LLO56 T cells are highly self-reactive and respond poorly in a primary infection, whereas LLO118 cells, which are less self-reactive, respond well during primary infection. We performed metabolic profiling and found that naive LLO118 had a dramatically higher basal respiration rate, a higher maximal respiration rate, and a higher glycolytic rate relative to LLO56. The LLO118 cells also exhibited a greater uptake of 2-NBD-glucose, in vitro and in vivo. We extended the correlation of low self-reactivity (CD5lo) with high basal metabolism using two other CD4+ TCRtg cells with known differences in self-reactivity, AND and Marilyn. We hypothesized that the decreased metabolism resulting from a strong interaction with self was mediated through TCR signaling. We then used an inducible knock-in mouse expressing the Scn5a voltage-gated sodium channel. This channel, when expressed in peripheral T cells, enhanced basal TCR-mediated signaling, resulting in decreased respiration and glycolysis, supporting our hypothesis. Genes and metabolites analysis of LLO118 and LLO56 T cells revealed significant differences in their metabolic pathways, including the glycerol phosphate shuttle. Inhibition of this pathway reverts the metabolic state of the LLO118 cells to be more LLO56 like. Overall, these studies highlight the critical relationship between peripheral TCR-self-pMHC interaction, metabolism, and the immune response to infection.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Animais , Metabolismo Basal , Diferenciação Celular/imunologia , Ativação Linfocitária/imunologia , Camundongos , Camundongos Transgênicos , Receptores de Antígenos de Linfócitos T/genética , Transdução de SinaisRESUMO
CLEC16A is in a locus genetically linked to autoimmune diseases including multiple sclerosis, but the function of this gene in the nervous system is unknown. Here we show that two mouse strains carrying independent Clec16a mutations developed neurodegenerative disease characterized by motor impairments and loss of Purkinje cells. Neurons from Clec16a-mutant mice exhibited increased expression of the autophagy substrate p62, accumulation of abnormal intra-axonal membranous structures bearing the autophagy protein LC3, and abnormal Golgi morphology. Multiple aspects of endocytosis, lysosome and Golgi function were normal in Clec16a-deficient murine embryonic fibroblasts and HeLa cells. However, these cells displayed abnormal bulk autophagy despite unimpaired autophagosome formation. Cultured Clec16a-deficient cells exhibited a striking accumulation of LC3 and LAMP-1 positive autolysosomes containing undigested cytoplasmic contents. Therefore Clec16a, an autophagy protein that is critical for autolysosome function and clearance, is required for Purkinje cell survival.
Assuntos
Lectinas Tipo C/genética , Lisossomos/fisiologia , Proteínas de Transporte de Monossacarídeos/genética , Doença dos Neurônios Motores/patologia , Mutação , Células de Purkinje/citologia , Animais , Autofagia , Sobrevivência Celular , Células Cultivadas , Complexo de Golgi/patologia , Células HeLa , Humanos , Lectinas Tipo C/metabolismo , Camundongos , Proteínas de Transporte de Monossacarídeos/metabolismo , Doença dos Neurônios Motores/genéticaRESUMO
To investigate how CD4(+) T cells function against a bacterial pathogen, we generated a Listeria monocytogenes-specific CD4(+) T cell model. In this system, two TCRtg mouse lines, LLO56 and LLO118, recognize the same immunodominant epitope (LLO190-205) of L. monocytogenes and have identical in vitro responses. However, in vivo LLO56 and LLO118 display vastly different responses during both primary and secondary infection. LLO118 dominates in the primary response and in providing CD8 T cell help. LLO56 predominates in the secondary response. We have also shown that both specific [T cell receptor (TCR)-mediated] and non-specific stimuli (bypassing the TCR) elicit distinct responses from the two transgenics, leading us to conclude that the strength of self-pMHC signaling during development tightly dictates the cell's future response in the periphery. Herein, we review our findings in this transfer system, focusing on the contribution of the immunomodulatory molecule CD5 and the importance of self-interaction in peripheral maintenance of the cell. We also discuss the manner in which individual TCR affinities to foreign and self-pMHC contribute to the outcome of an immune response; our assertion is that there exists a spectrum of possible T cell responses to recognition of cognate antigen during infection, adding immense diversity to the immune system's response to pathogens.
RESUMO
PURPOSE: To investigate the production of monokine induced by gamma-interferon (MIG) during a primary Herpes simplex virus type 1 (HSV-1) infection of the cornea. We hypothesize that multiple CXCR3 ligands are involved in T cell recruitment during HSV-1 corneal infection and that neutrophils have the potential to contribute to their production. MATERIALS AND METHODS: Levels of MIG were evaluated in an in vivo murine model of HSV-1 corneal infection by quantitative ELISA. Cultured murine corneal fibroblast (MCF) cells and purified neutrophils were stimulated in vitro with IFN-γ and IL-1α to determine inducers of MIG. Cellular sources of MIG production in vivo were investigated via cellular depletion studies. Additionally, MIG production resulting from interaction between resident human corneal cells and neutrophils was evaluated in an ex vivo model of human corneal infection. RESULTS: MIG was significantly elevated on days 2-6 and on day 8 following corneal infection. MCF and neutrophils secreted MIG in response to IFN-γ, but not IL-1α stimulation. Co-stimulation with IFN-γ and IL-1α induced a four-fold increase in MIG production by MCF. However, the same combination led to a three-fold decrease in MIG production by neutrophils. In vivo, a 52% reduction in MIG levels was observed in the neutrophil depleted host. In the human ex vivo model, MIG levels were significantly elevated in response to communication between HSV-1 infected corneal tissue and neutrophils. CONCLUSIONS: Here, we report the evidence for the production of MIG, a second CXCR3 ligand, during the primary immune response to HSV-1 corneal infection. Our results support the hypothesis that both neutrophils and resident corneal cells contribute to MIG production in vivo. However, neutrophils produce MIG in response to communication with HSV-1-infected resident corneal cells more efficiently than by direct interaction with virus. In addition, we found that MIG production by neutrophils and resident corneal cells was differentially regulated by IL-1α.
Assuntos
Quimiocina CXCL9/genética , Ceratócitos da Córnea/metabolismo , Modelos Animais de Doenças , Regulação da Expressão Gênica/fisiologia , Herpesvirus Humano 1/fisiologia , Ceratite Herpética/genética , Neutrófilos/metabolismo , Animais , Células Cultivadas , Quimiocina CXCL9/metabolismo , Córnea/metabolismo , Córnea/virologia , Ceratócitos da Córnea/efeitos dos fármacos , Ensaio de Imunoadsorção Enzimática , Feminino , Interferon gama/farmacologia , Interleucina-1alfa/farmacologia , Ceratite Herpética/metabolismo , Ceratite Herpética/virologia , Camundongos , Camundongos Endogâmicos C57BL , Neutrófilos/efeitos dos fármacos , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Many mechanisms of and treatments for type 1 diabetes studied in the NOD mouse model have not been replicated in human disease models. Thus, the field of diabetes research remains hindered by the lack of an in vivo system in which to study the development and onset of autoimmune diabetes. To this end, we characterized a system using human CD4(+) T cells pulsed with autoantigen-derived peptides. Six weeks after injection of as few as 0.5 × 10(6) antigen-pulsed cells into the NOD-Scid Il2rg(-/-) mouse expressing the human HLA-DR4 transgene, infiltration of mouse islets by human T cells was seen. Although islet infiltration occurred with both healthy and diabetic donor antigen-pulsed CD4(+) T cells, diabetic donor injections yielded significantly greater levels of insulitis. Additionally, significantly reduced insulin staining was observed in mice injected with CD4(+) T-cell lines from diabetic donors. Increased levels of demethylated ß-cell-derived DNA in the bloodstream accompanied this loss of insulin staining. Together, these data show that injection of small numbers of autoantigen-reactive CD4(+) T cells can cause a targeted, destructive infiltration of pancreatic ß-cells. This model may be valuable for understanding mechanisms of induction of human diabetes.