RESUMEN
Calcium signaling is essential for lymphocyte activation, with genetic disruptions of store-operated calcium (Ca2+) entry resulting in severe immunodeficiency. The inositol 1,4,5-trisphosphate receptor (IP3R), a homo- or heterotetramer of the IP3R1-3 isoforms, amplifies lymphocyte signaling by releasing Ca2+ from endoplasmic reticulum stores following antigen stimulation. Although knockout of all IP3R isoforms in mice causes immunodeficiency, the seeming redundancy of the isoforms is thought to explain the absence of variants in human immunodeficiency. In this study, we identified compound heterozygous variants of ITPR3 (a gene encoding IP3R subtype 3) in two unrelated Caucasian patients presenting with immunodeficiency. To determine whether ITPR3 variants act in a nonredundant manner and disrupt human immune responses, we characterized the Ca2+ signaling capacity, the lymphocyte response, and the clinical phenotype of these patients. We observed disrupted Ca2+ signaling in patient-derived fibroblasts and immune cells, with abnormal proliferation and activation responses following T-cell receptor stimulation. Reconstitution of IP3R3 in IP3R knockout cell lines led to the identification of variants as functional hypomorphs that showed reduced ability to discriminate between homeostatic and induced states, validating a genotype-phenotype link. These results demonstrate a functional link between defective endoplasmic reticulum Ca2+ channels and immunodeficiency and identify IP3Rs as diagnostic targets for patients with specific inborn errors of immunity. These results also extend the known cause of Ca2+-associated immunodeficiency from store-operated entry to impaired Ca2+ mobilization from the endoplasmic reticulum, revealing a broad sensitivity of lymphocytes to genetic defects in Ca2+ signaling.
Asunto(s)
Señalización del Calcio , Calcio , Receptores de Inositol 1,4,5-Trifosfato , Animales , Humanos , Ratones , Calcio/metabolismo , Señalización del Calcio/genética , Señalización del Calcio/inmunología , Homeostasis , Receptores de Inositol 1,4,5-Trifosfato/genética , Receptores de Inositol 1,4,5-Trifosfato/inmunología , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Isoformas de Proteínas/metabolismo , Enfermedades del Sistema Inmune/metabolismoRESUMEN
CRAC, which plays important role in Ca2+-dependent T-lymphocyte activation, is composed of the ER-resident STIM1 and the plasma membrane Orai1 pore-forming subunit. Both accumulate at the immunological synapse (IS) between a T cell and an antigen-presenting cell (APC). We hypothesized that adapter/interacting proteins regulate Orai1 residence in the IS. We could show that mGFP-tagged Orai1-Full channels expressed in Jurkat cells had a biphasic IS-accumulation kinetics peaked at 15 min. To understand the background of Orai1 IS-redistribution we knocked down STIM1 and SAP97 (adaptor protein with a short IS-residency (15 min) and ability to bind Orai1 N-terminus): the mGFP-Orai1-Full channels kept on accumulating in the IS up to the 60th minute in the STIM1- and SAP97-lacking Jurkat cells. Deletion of Orai1 N terminus (mGFP-Orai1-Δ72) resulted in the same time course as described for STIM1/SAP97 knock-down cells. Ca2+-imaging of IS-engaged T-cells revealed that of Orai1 residency modifies the Ca2+-response: cells expressing mGFP-Orai1-Δ72 construct or mGFP-Orai1-Full in SAP-97 knock-down cells showed higher number of Ca2+-oscillation up to the 90th minute after IS formation. Overall, these data suggest that SAP97 may contribute to the short-lived IS-residency of Orai1 and binding of STIM1 to Orai1 N-terminus is necessary for SAP97-Orai1 interaction.
Asunto(s)
Señalización del Calcio/inmunología , Sinapsis Inmunológicas/metabolismo , Proteína ORAI1/metabolismo , Linfocitos T/inmunología , Linfocitos T/metabolismo , Inmunidad Adaptativa , Homólogo 1 de la Proteína Discs Large/antagonistas & inhibidores , Homólogo 1 de la Proteína Discs Large/genética , Homólogo 1 de la Proteína Discs Large/metabolismo , Retículo Endoplásmico/metabolismo , Técnicas de Silenciamiento del Gen , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Células HEK293 , Humanos , Sinapsis Inmunológicas/genética , Sinapsis Inmunológicas/inmunología , Células Jurkat , Cinética , Activación de Linfocitos , Proteínas de Neoplasias/antagonistas & inhibidores , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Proteína ORAI1/química , Proteína ORAI1/genética , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Eliminación de Secuencia , Molécula de Interacción Estromal 1/antagonistas & inhibidores , Molécula de Interacción Estromal 1/genética , Molécula de Interacción Estromal 1/metabolismoRESUMEN
Nicotinic acid adenine dinucleotide 2'-phosphate (NAADP) is a naturally occurring nucleotide that has been shown to be involved in the release of Ca2+ from intracellular stores in a wide variety of cell types, tissues and organisms. Current evidence suggests that NAADP may function as a trigger to initiate a Ca2+ signal that is then amplified by other Ca2+ release mechanisms. A fundamental question that remains unanswered is the identity of the NAADP receptor. Our recent studies have identified HN1L/JPT2 as a high affinity NAADP binding protein that is essential for the modulation of Ca2+ channels.
Asunto(s)
Canales de Calcio/inmunología , Señalización del Calcio/inmunología , Calcio/inmunología , Proteínas Asociadas a Microtúbulos/inmunología , NADP/análogos & derivados , Animales , Humanos , NADP/inmunologíaRESUMEN
The sympathetic nervous system (SNS) controls various physiological functions via the neurotransmitter noradrenaline. Activation of the SNS in response to psychological or physical stress is frequently associated with weakened immunity. Here, we investigated how adrenoceptor signaling influences leukocyte behavior. Intravital two-photon imaging after injection of noradrenaline revealed transient inhibition of CD8+ and CD4+ T cell locomotion in tissues. Expression of ß-adrenergic receptor in hematopoietic cells was not required for NA-mediated inhibition of motility. Rather, chemogenetic activation of the SNS or treatment with adrenergic receptor agonists induced vasoconstriction and decreased local blood flow, resulting in abrupt hypoxia that triggered rapid calcium signaling in leukocytes and halted cell motility. Oxygen supplementation reversed these effects. Treatment with adrenergic receptor agonists impaired T cell responses induced in response to viral and parasitic infections, as well as anti-tumor responses. Thus, stimulation of the SNS impairs leukocyte mobility, providing a mechanistic understanding of the link between adrenergic receptors and compromised immunity.
Asunto(s)
Adrenérgicos/inmunología , Movimiento Celular/inmunología , Inmunidad/inmunología , Leucocitos/inmunología , Sistema Nervioso Simpático/inmunología , Animales , Señalización del Calcio/inmunología , Línea Celular Tumoral , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Receptores Adrenérgicos/inmunología , Transducción de Señal/inmunología , Linfocitos T/inmunologíaRESUMEN
Glycerol monolaurate (GML), a naturally occurring monoglyceride, is widely used commercially for its antimicrobial properties. Interestingly, several studies have shown that GML not only has antimicrobial properties but is also an anti-inflammatory agent. GML inhibits peripheral blood mononuclear cell proliferation and inhibits T cell receptor (TCR)-induced signaling events. In this study, we perform an extensive structure activity relationship analysis to investigate the structural components of GML necessary for its suppression of human T cell activation. Human T cells were treated with analogs of GML, differing in acyl chain length, head group, linkage of acyl chain, and number of laurate groups. Treated cells were then tested for changes in membrane dynamics, LAT clustering, calcium signaling, and cytokine production. We found that an acyl chain with 12-14 carbons, a polar head group, an ester linkage, and a single laurate group at any position are all necessary for GML to inhibit protein clustering, calcium signaling, and cytokine production. Removing the glycerol head group or replacing the ester linkage with a nitrogen prevented derivative-mediated inhibition of protein cluster formation and calcium signaling, while still inhibiting TCR-induced cytokine production. These findings expand our current understanding of the mechanisms of action of GML and the of GML needed to function as a novel immunosuppressant.
Asunto(s)
Señalización del Calcio/efectos de los fármacos , Lauratos/farmacología , Activación de Linfocitos/efectos de los fármacos , Monoglicéridos/farmacología , Receptores de Antígenos de Linfocitos T/inmunología , Linfocitos T/inmunología , Señalización del Calcio/inmunología , HumanosRESUMEN
BACKGROUND: Bronchial remodeling is a key feature of asthma that is already present in preschoolers with wheezing. Moreover, bronchial smooth muscle (BSM) remodeling at preschool age is predictive of asthma at school age. However, the mechanism responsible for BSM remodeling in preschoolers with wheezing remains totally unknown. In contrast, in adult asthma, BSM remodeling has been associated with an increase in BSM cell proliferation related to increased mitochondrial mass and biogenesis triggered by an altered calcium homeostasis. Indeed, BSM cell proliferation was decreased in vitro by the calcium channel blocker gallopamil. OBJECTIVE: Our aim was to investigate the mechanisms involved in BSM cell proliferation in preschoolers with severe wheezing, with special attention to the role of mitochondria and calcium signaling. METHODS: Bronchial tissue samples obtained from 12 preschool controls without wheezing and 10 preschoolers with severe wheezing were used to measure BSM mass and establish primary BSM cell cultures. BSM cell proliferation was assessed by manual counting and flow cytometry, ATP content was assessed by bioluminescence, mitochondrial respiration was assessed by using either the Seahorse or Oroboros technique, mitochondrial mass and biogenesis were assessed by immunoblotting, and calcium response to carbachol was assessed by confocal microscopy. The effect of gallopamil was also evaluated. RESULTS: BSM mass, cell proliferation, ATP content, mitochondrial respiration, mass and biogenesis, and calcium response were all increased in preschoolers with severe wheezing compared with in the controls. Gallopamil significantly decreased BSM mitochondrial biogenesis and mass, as well as cell proliferation. CONCLUSION: Mitochondria are key players in BSM cell proliferation in preschoolers with severe wheezing and could represent a potential target to treat BSM remodeling at an early stage of the disease.
Asunto(s)
Remodelación de las Vías Aéreas (Respiratorias)/inmunología , Bronquios/inmunología , Mitocondrias Musculares/inmunología , Músculo Liso/inmunología , Ruidos Respiratorios/inmunología , Asma/etiología , Asma/inmunología , Asma/patología , Bronquios/patología , Señalización del Calcio/efectos de los fármacos , Señalización del Calcio/inmunología , Células Cultivadas , Preescolar , Femenino , Galopamilo/farmacología , Humanos , Lactante , Masculino , Mitocondrias Musculares/patología , Músculo Liso/patologíaRESUMEN
BACKGROUND: Stromal interaction molecule 1 (STIM1)-mediated store-operated Ca2+ entry (SOCE) is now recognized as the main mechanism of the majority of nonexcitable cell calcium influx. Calcium overload is a primary mechanism of endothelial cell injury during systemic inflammatory response and sepsis. Whether STIM1-mediated SOCE plays a role in calcium overload in vascular endothelial cell injury remains unclear. MATERIALS AND METHODS: To explore the role of STIM1-gated SOCE in vascular endothelial cell calcium overload and inflammation, we established a human septic serum or lipopolysaccharide (LPS)-induced human umbilical vein endothelial cell (HUVEC) experimental system and derived ribonucleic acid interference (RNAi)-mediated STIM1, ORAI1 (orai gene [HGNC: 25896 Entrez Gene: 84876] coding protein, ORAI Calcium Release-Activated Calcium Modulator 1), and transient receptor potential channel 1 (TRPC1) (core components of store-operated Ca2+[SOC]) downregulated HUVECs, as well as STIM1 overinduced HUVECs. RESULTS: Our results show that sepsis serum or LPS stimulation increased STIM1 in HUVECs and increased all cytokines except for VEGF and the inflammatory mediators tumor necrosis factor, intercellular cell adhesion molecule-1, and endothelin-1 in a time-dependent manner. RNAi-mediated knockdown of STIM1 significantly inhibited serum or LPS-induced inflammatory cytokine expression, and STIM1 overexpression in HUVECs promoted LPS-mediated induction of these cytokines. Meanwhile, similar to the blocking effect of the specific SOC inhibitors Gd3+ and La3+ on LPS-induced calcium influx, RNAi-mediated depletion of STIM1 or the SOC proteins TRPC1 and ORAI1 could significantly inhibit serum or LPS-induced extracellular calcium influx, as well as the expression of the inflammatory cytokines tumor necrosis factor, intercellular cell adhesion molecule-1, and endothelin-1. Simultaneous downregulation of the SOCE core units TRPC1 and ORAI1 inhibited LPS-induced calcium influx and cytokine expression, which could not be restored by inducing STIM1. Forced expression of nuclear factor-κB (NF-κB) in HUVECs significantly induced STIM1 expression, whereas RNAi-mediated depletion of NF-κB significantly inhibited STIM1 mRNA levels and significantly reduced the thapsigargin-mediated SOCE calcium influx, which was similar to results with the NF-κB inhibitor wogonin. CONCLUSIONS: Septic serum stimulates the expression of STIM1, cytokines, and inflammatory mediators in HUVECs. STIM1-mediated SOCE is required for Ca2+ influx induced by LPS or septic serum and contributes cytokines and inflammatory mediators in septic serum-stimulated HUVECs. In addition, STIM1-mediated SOCE on Ca2+ influx by septic serum or LPS involves NF-κB signaling.
Asunto(s)
Quemaduras/sangre , Calcio/metabolismo , Endotelio Vascular/patología , Proteínas de Neoplasias/metabolismo , Sepsis/inmunología , Molécula de Interacción Estromal 1/metabolismo , Adulto , Quemaduras/inmunología , Señalización del Calcio/inmunología , Endotelio Vascular/inmunología , Técnicas de Silenciamiento del Gen , Células Endoteliales de la Vena Umbilical Humana , Humanos , Lipopolisacáridos/inmunología , Masculino , Persona de Mediana Edad , Proteínas de Neoplasias/genética , Proteína ORAI1/genética , Proteína ORAI1/metabolismo , Sepsis/sangre , Sepsis/patología , Suero/inmunología , Molécula de Interacción Estromal 1/genéticaRESUMEN
Alveolar macrophages (AM) maintain airway immune balance; however, the regulation of heterogeneity of AMs is incompletely understood. We demonstrate that RGS1 coregulates the immunophenotype of AM subpopulations, including pro- and anti-inflammatory, injury- and repair-associated, and pro- and antifibrotic phenotypes, through the PLC-IP3R signal-dependent intracellular Ca2+ response. Flt3+ AMs and Tie2+ AMs had different immune properties, and RGS1 expression in the cells was targeted by exosomes (EXOs) containing miR-223 and miR-27b-3p that were derived from vascular endothelial cells (EnCs) and type II alveolar epithelial cells (EpCs-II), respectively. Imbalance of AMs was correlated with acute lung injury/acute respiratory distress syndrome (ALI/ARDS) and pulmonary fibrosis (PF) caused a lack of secretion of CD31+ and CD74+ EXOs derived from EnCs and EpCs-II. Timely treatment with EXOs significantly improved endotoxin-induced ALI/ARDS and bleomycin-induced PF in mice. Thus, EnC- and EpC-II-derived EXOs regulate the immune balance of AMs and can be used as potential therapeutic drugs.
Asunto(s)
Lesión Pulmonar Aguda/terapia , Señalización del Calcio/inmunología , Exosomas/trasplante , Macrófagos Alveolares/metabolismo , Proteínas RGS/metabolismo , Síndrome de Dificultad Respiratoria/terapia , Lesión Pulmonar Aguda/metabolismo , Células Epiteliales Alveolares/fisiología , Animales , Células Endoteliales/fisiología , Endotelio/metabolismo , Epitelio/metabolismo , Exosomas/metabolismo , Humanos , Inmunidad , Ratones , Proteínas RGS/genética , Síndrome de Dificultad Respiratoria/metabolismoRESUMEN
Exhausted immune responses to chronic diseases represent a major challenge to global health. We study CD4+ T cells in a mouse model with regulatable antigen presentation. When the cells are driven through the effector phase and are then exposed to different levels of persistent antigen, they lose their T helper 1 (Th1) functions, upregulate exhaustion markers, resemble naturally anergic cells, and modulate their MAPK, mTORC1, and Ca2+/calcineurin signaling pathways with increasing dose and time. They also become unable to help B cells and, at the highest dose, undergo apoptosis. Transcriptomic analyses show the dynamic adjustment of gene expression and the accumulation of T cell receptor (TCR) signals over a period of weeks. Upon antigen removal, the cells recover their functionality while losing exhaustion and anergy markers. Our data suggest an adjustable response of CD4+ T cells to different levels of persisting antigen and contribute to a better understanding of chronic disease.
Asunto(s)
Antígenos/inmunología , Señalización del Calcio/inmunología , Anergia Clonal , Regulación de la Expresión Génica/inmunología , Sistema de Señalización de MAP Quinasas/inmunología , Células TH1/inmunología , Animales , Antígenos/genética , Linfocitos B/inmunología , Señalización del Calcio/genética , Femenino , Perfilación de la Expresión Génica , Sistema de Señalización de MAP Quinasas/genética , Diana Mecanicista del Complejo 1 de la Rapamicina/genética , Diana Mecanicista del Complejo 1 de la Rapamicina/inmunología , Ratones , Ratones Transgénicos , Receptores de Antígenos de Linfocitos T/genética , Receptores de Antígenos de Linfocitos T/inmunologíaRESUMEN
Natural killer (NK) cells are innate cytotoxic lymphocytes that efficiently eliminate malignant and virus-infected cells without prior activation via the directed and focused release of lytic granule contents for target cell lysis. This cytolytic process is tightly regulated at discrete checkpoint stages to ensure the selective killing of diseased target cells and is highly dependent on the coordinated regulation of cytoskeletal components. The actin-binding protein filamin crosslinks cortical actin filaments into orthogonal networks and links actin filament webs to cellular membranes to modulate cell migration, adhesion, and signaling. However, its role in the regulation of NK cell functions remains poorly understood. Here, we show that filamin A (FLNa), a filamin isoform with preferential expression in leukocytes, is recruited to the NK cell lytic synapse and is required for NK cell cytotoxicity through the modulation of conjugate formation with target cells, synaptic filamentous actin (F-actin) accumulation, and cytotoxic degranulation, but not granule polarization. Interestingly, we also find that the loss of FLNa augments the target cell-induced expression of IFN-γ and TNF-α by NK cells, correlating with enhanced activation signals such as Ca2+ mobilization, ERK, and NF-κB, and a delayed down-modulation of the NKG2D receptor. Thus, our results identify FLNa as a new regulator of NK cell effector functions during their decision to kill target cells through a balanced regulation of NK cell cytotoxicity vs cytokine production. Moreover, this study implicates the cross-linking/bundling of F-actin mediated by FLNa as a necessary process coordinating optimal NK effector functions.
Asunto(s)
Actinas/inmunología , Señalización del Calcio/inmunología , Filaminas/inmunología , Inmunidad Celular , Interferón gamma/inmunología , Células Asesinas Naturales/inmunología , Factor de Necrosis Tumoral alfa/inmunología , Línea Celular , HumanosRESUMEN
Antigen recognition by the T-cell receptor induces a cytosolic Ca2+ signal that is crucial for T-cell function. The Ca2+ channel TRPM2 (transient receptor potential cation channel subfamily M member 2) has been shown to facilitate influx of extracellular Ca2+ through the plasma membrane of T cells. Therefore, it was suggested that TRPM2 is involved in T-cell activation and differentiation. However, these results are largely derived from in vitro studies using T-cell lines and non-physiologic means of TRPM2 activation. Thus, the relevance of TRPM2-mediated Ca2+ signaling in T cells remains unclear. Here, we use TRPM2-deficient mice to investigate the function of TRPM2 in T-cell activation and differentiation. In response to TCR stimulation in vitro, Trpm2-/- and WT CD4+ and CD8+ T cells similarly upregulated the early activation markers NUR77, IRF4, and CD69. We also observed regular proliferation of Trpm2-/- CD8+ T cells and unimpaired differentiation of CD4+ T cells into Th1, Th17, and Treg cells under specific polarizing conditions. In vivo, Trpm2-/- and WT CD8+ T cells showed equal specific responses to Listeria monocytogenes after infection of WT and Trpm2-/- mice and after transfer of WT and Trpm2-/- CD8+ T cells into infected recipients. CD4+ T-cell responses were investigated in the model of anti-CD3 mAb-induced intestinal inflammation, which allows analysis of Th1, Th17, Treg, and Tr1-cell differentiation. Here again, we detected similar responses of WT and Trpm2-/- CD4+ T cells. In conclusion, our results argue against a major function of TRPM2 in T-cell activation and differentiation.
Asunto(s)
Diferenciación Celular/inmunología , Activación de Linfocitos/inmunología , Canales Catiónicos TRPM/inmunología , Animales , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Señalización del Calcio/inmunología , Proliferación Celular/fisiología , Listeria monocytogenes/inmunología , Listeriosis/inmunología , Ratones , Ratones Endogámicos C57BL , Linfocitos T Reguladores/inmunología , Células TH1/inmunología , Células Th17/inmunologíaRESUMEN
Intestinal inflammation is a major threat to the health and growth of young animals such as piglets. As a next-generation probiotics, limited studies have shown that Akkermansia muciniphila could alleviate inflammation of intestinal epithelial cells (IECs). In this study, a TNF-α-induced inflammatory model of IPEC-J2 cells, the intestinal porcine enterocytes, was built to evaluate the effects of active or inactive A. muciniphila on the inflammation of IECs. The viability of IPEC-J2 cells was the highest when treated with active (108 copies/mL) or inactive (109 copies/mL) A. muciniphila for 7.5 h (P < 0.01). Treated with 20 ng/mL of TNF-α and followed by a treatment of A. muciniphila, the mRNA level of proinflammatory cytokines (IL-8, IL-1ß, IL-6 and TNF-α) was remarkably reduced (P < 0.05) along with the increased mRNA level of tight junction proteins (ZO-1 and Occludin, P < 0.05). Flow cytometry analysis showed that active or inactive A. muciniphila significantly suppressed the rate of the early and total apoptotic of the inflammatory IPEC-J2 cells (P < 0.05). According to results of transcriptome sequencing, active and inactive A. muciniphila may decline cell apoptosis by down-regulating the expression of key genes in calcium signaling pathway, or up-regulating the expression of key genes in cell cycle signaling pathway. And the bacterium may alleviate the inflammation of IECs by down-regulating the expression of PI3K upstream receptor genes. Our results indicate that A. muciniphila may be a promising NGP targeting intestinal inflammation.
Asunto(s)
Inflamación/dietoterapia , Mucosa Intestinal/inmunología , Probióticos/administración & dosificación , Akkermansia/inmunología , Animales , Señalización del Calcio/inmunología , Línea Celular , Supervivencia Celular/inmunología , Modelos Animales de Enfermedad , Regulación hacia Abajo/inmunología , Células Epiteliales , Humanos , Inflamación/inmunología , Mucosa Intestinal/citología , Fosfatidilinositol 3-Quinasas/metabolismo , Transducción de Señal/inmunología , Porcinos , Factor de Necrosis Tumoral alfa/administración & dosificación , Factor de Necrosis Tumoral alfa/inmunologíaRESUMEN
Most agonists stimulate platelet Ca2+ rises via G-protein coupled receptors (GPCRs) or ITAM-linked receptors (ILRs). Well studied are the GPCRs stimulated by the soluble agonists thrombin (PAR1, PAR4), ADP (P2Y1, P2Y12), and thromboxane A2 (TP), signaling via phospholipase (PLC)ß isoforms. The platelet ILRs glycoprotein VI (GPVI), C-type lectin-like receptor 2 (CLEC2), and FcγRIIa are stimulated by adhesive ligands or antibody complexes and signal via tyrosine protein kinases and PLCγ isoforms. Marked differences exist between the GPCR- and ILR-induced Ca2+ signaling in: (i) dependency of tyrosine phosphorylation; (ii) oscillatory versus continued Ca2+ rises by mobilization from the endoplasmic reticulum; and (iii) smaller or larger role of extracellular Ca2+ entry via STIM1/ORAI1. Co-stimulation of both types of receptors, especially by thrombin (PAR1/4) and collagen (GPVI), leads to a highly enforced Ca2+ rise, involving mitochondrial Ca2+ release, which activates the ion and phospholipid channel, anoctamin-6. This highly Ca2+-dependent process causes swelling, ballooning, and phosphatidylserine expression, establishing a unique platelet population swinging between vital and necrotic (procoagulant 'zombie' platelets). Additionally, the high Ca2+ status of procoagulant platelets induces a set of additional events: (i) Ca2+ dependent cleavage of signaling proteins and receptors via calpain and ADAM isoforms; (ii) microvesiculation; (iii) enhanced coagulation factor binding; and (iv) fibrin-coat formation involving transglutaminases. Given the additive roles of GPCR and ILR in Ca2+ signal generation, high-throughput screening of biomolecules or small molecules based on Ca2+ flux measurements provides a promising way to find new inhibitors interfering with prolonged high Ca2+, phosphatidylserine expression, and hence platelet procoagulant activity.
Asunto(s)
Anoctaminas/metabolismo , Plaquetas/metabolismo , Señalización del Calcio/inmunología , Proteínas de Unión al GTP/metabolismo , HumanosRESUMEN
Aging is an unstoppable process and begins shortly after birth. Each cell of the organism is affected by the irreversible process, not only with equal density but also at varying ages and with different speed. Therefore, aging can also be understood as an adaptation to a continually changing cellular environment. One of these very prominent changes in age affects Ca2+ signaling. Especially immune cells highly rely on Ca2+-dependent processes and a strictly regulated Ca2+ homeostasis. The intricate patterns of impaired immune cell function may represent a deficit or compensatory mechanisms. Besides, altered immune function through Ca2+ signaling can profoundly affect the development of age-related disease. This review attempts to summarize changes in Ca2+ signaling due to channels and receptors in T cells and beyond in the context of aging.
Asunto(s)
Envejecimiento/inmunología , Canales de Calcio/inmunología , Señalización del Calcio/inmunología , Calcio/inmunología , Homeostasis/inmunología , Linfocitos T/inmunología , Animales , HumanosRESUMEN
Alzheimer's disease (AD) is the most common form of dementia, affecting millions of people worldwide. Unfortunately, none of the current treatments are effective at improving cognitive function in AD patients and, therefore, there is an urgent need for the development of new therapies that target the early cause(s) of AD. Intracellular calcium (Ca2+) regulation is critical for proper cellular and neuronal function. It has been suggested that Ca2+ dyshomeostasis is an upstream factor of many neurodegenerative diseases, including AD. For this reason, chemical agents or small molecules aimed at targeting or correcting this Ca2+ dysregulation might serve as therapeutic strategies to prevent the development of AD. Moreover, neurons are not alone in exhibiting Ca2+ dyshomeostasis, since Ca2+ disruption is observed in other cell types in the brain in AD. In this review, we examine the distinct Ca2+ channels and compartments involved in the disease mechanisms that could be potential targets in AD.
Asunto(s)
Enfermedad de Alzheimer/fisiopatología , Enfermedad de Alzheimer/terapia , Señalización del Calcio/inmunología , Calcio/metabolismo , Homeostasis , HumanosRESUMEN
The number and activity of T cell subsets in the atherosclerotic plaques are critical for the prognosis of patients with acute coronary syndrome. ß2 Integrin activation is pivotal for T cell recruitment and correlates with future cardiac events. Despite this knowledge, differential regulation of adhesiveness in T cell subsets has not been explored yet. In this study, we show that in human T cells, SDF-1α-mediated ß2 integrin activation is driven by a, so far, not-described reactive oxidative species (ROS)-regulated calcium influx. Furthermore, we show that CD4+CD28null T cells represent a highly reactive subset showing 25-fold stronger ß2 integrin activation upon SDF-1α stimulation compared with CD28+ T cells. Interestingly, ROS-dependent Ca release was much more prevalent in the pathogenetically pivotal CD28null subset compared with the CD28+ T cells, whereas the established mediators of the classical pathways for ß2 integrin activation (PKC, PI3K, and PLC) were similarly activated in both T cell subsets. Thus, interference with the calcium flux attenuates spontaneous adhesion of CD28null T cells from acute coronary syndrome patients, and calcium ionophores abolished the observed differences in the adhesion properties between CD28+ and CD28null T cells. Likewise, the adhesion of these T cell subsets was indistinguishable in the presence of exogenous ROS/H2O2 Together, these data provide a molecular explanation of the role of ROS in pathogenesis of plaque destabilization.
Asunto(s)
Síndrome Coronario Agudo/inmunología , Antígenos CD18/inmunología , Linfocitos T CD4-Positivos/inmunología , Señalización del Calcio/inmunología , Especies Reactivas de Oxígeno/inmunología , Síndrome Coronario Agudo/patología , Antígenos CD28/inmunología , Linfocitos T CD4-Positivos/patología , Quimiocina CXCL12/inmunología , Femenino , Humanos , MasculinoRESUMEN
An expansion of CD21low B cells has been described in a variety of diseases associated with persistent immune stimulation as in chronic infection, immunodeficiency, or autoimmunity. Different developmental stages of CD21low B cells have been highlighted in specific diseases; however, a systematic comparison of distribution, phenotype, and signaling capacity of these populations has not yet been performed to delineate the pivotal character of this unusual B cell population. Screening of more than 200 patients with autoimmune disease demonstrated that the prevalence of patients with expanded CD21low B cells varies between diseases. The expansion was frequent in patients with systemic lupus erythematosus, in which it correlated to relative B cell lymphopenia and duration of disease. Different proportions of distinct developmental stages of CD21low B cells co-occur in nearly all patients with autoimmune disease. Although in most patients, naive-like and CD27- switched memory B cells were the most prominent CD21low subpopulations, there was no detectable association of the pattern with the underlying disease. Despite their distinct developmental stage, all CD21low B cells share a common core phenotype including the increased expression of inhibitory receptors, associated with an elevated constitutive phosphorylation of proximal signaling molecules downstream of the BCR but impaired Ca2+ mobilization and NF-κB activation after BCR stimulation. Further, this was accompanied by impaired upregulation of CD69, although CD86 upregulation was preserved. Beyond maturation-associated differences, the common core characteristics of all CD21low B cell populations suggests either a common ancestry or a shared sustained imprint by the environment they originated in.
Asunto(s)
Subgrupos de Linfocitos B/inmunología , Señalización del Calcio/inmunología , Lupus Eritematoso Sistémico/inmunología , Memoria a Corto Plazo , Receptores de Complemento 3d/inmunología , Transducción de Señal/inmunología , Adulto , Subgrupos de Linfocitos B/patología , Femenino , Humanos , Masculino , Receptores de Antígenos de Linfocitos B/inmunologíaRESUMEN
While B cells play a significant role in the onset of type-1 diabetes (T1D), little is know about their role in those early stages. Thus, to gain new insights into the role of B cells in T1D, we converted a physiological early pancreas-infiltrating B cell into a novel BCR mouse model using Somatic Cell Nuclear Transfer (SCNT). Strikingly, SCNT-derived B1411 model displayed neither developmental block nor anergy. Instead, B1411 underwent spontaneous germinal center reactions. Without T cell help, B1411-Rag1-/- was capable of forming peri-/intra-pancreatic lymph nodes, and undergoing class-switching. RNA-Seq analysis identified 93 differentially expressed genes in B1411 compared to WT B cells, including Irf7, Usp18, and Mda5 that had been linked to a potential viral etiology of T1D. We also found various members of the oligoadenylate synthase (OAS) family to be enriched in B1411, such as Oas1, which had recently also been linked to T1D. Strikingly, when challenged with glucose B1411-Rag1-/- mice displayed impaired glucose tolerance.
Asunto(s)
Autoinmunidad , Linfocitos B/inmunología , Estado Prediabético/etiología , Estado Prediabético/inmunología , Animales , Basidiomycota/genética , Basidiomycota/metabolismo , Señalización del Calcio/inmunología , Ensamble y Desensamble de Cromatina , Células Clonales/inmunología , Diabetes Mellitus Experimental/etiología , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/inmunología , Femenino , Perfilación de la Expresión Génica , Prueba de Tolerancia a la Glucosa , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Ratones Endogámicos NOD , Ratones Noqueados , Modelos Inmunológicos , Técnicas de Transferencia Nuclear , Estado Prediabético/genética , Receptores de Antígenos de Linfocitos B/genética , Receptores de Antígenos de Linfocitos B/inmunologíaRESUMEN
NK cell-mediated Ab-dependent cellular cytotoxicity (ADCC) is increasingly recognized to play an important role in cancer immunotherapy, transplant rejection, and autoimmunity. However, several aspects of the molecular interactions of IgG subclasses with the Fc-gamma receptor IIIA (FcγRIIIA)/CD16a expressed on NK cells remain unknown. The aim of the current study was to further analyze the role of IgG subclasses and FCGR3A V158F single nucleotide polymorphism (SNP) on Ca2+ signaling and NK cell-mediated ADCC against Daudi target cells in vitro. NK cells were isolated from donors with different FCGR3A SNP. The affinity of rituximab IgG subclasses to CD20 expressed on Daudi cells showed similar dissociation constant as tested by flow cytometry. Induction of Ca2+ signaling, degranulation, intracellular cytokine production, and ADCC was demonstrated for IgG1 and IgG3, to a lesser degree also for IgG4, but not for IgG2. Compared to NK cells carrying the low-affinity (FF) variant for the FCGR3A V158F SNP, binding of IgG1 and IgG3 to NK cells carrying the high-affinity (VV) and VF SNP variants was two- to threefold higher. Variations of FCGR3A SNP among the eight tested donors (1 VV, 3FF, and 4VF) revealed no significant differences of Ca2+ signaling and degranulation; however, ADCC was somewhat weaker in donors with the low-affinity FF variation. In conclusion, this is the first study correlating Ca2+ signaling and NK cell-mediated ADCC triggered by the four IgG subclasses with the FCGR3A V158F SNP. Our findings indicate important differences in the interactions of IgG subclasses with FcγRIIIA/CD16a but no major impact of FCGR3A SNP and may therefore help to better correlate the functional properties of particular engineered therapeutic antibodies in vitro with individual differences of their clinical efficacy.
Asunto(s)
Citotoxicidad Celular Dependiente de Anticuerpos , Antineoplásicos Inmunológicos/farmacología , Señalización del Calcio , Inmunoglobulina G/farmacología , Células Asesinas Naturales/inmunología , Polimorfismo de Nucleótido Simple , Receptores de IgG , Rituximab/farmacología , Antineoplásicos Inmunológicos/farmacocinética , Señalización del Calcio/efectos de los fármacos , Señalización del Calcio/inmunología , Línea Celular Tumoral , Humanos , Neoplasias/inmunología , Neoplasias/patología , Neoplasias/terapia , Receptores de IgG/genética , Receptores de IgG/inmunologíaRESUMEN
TRIM family proteins play integral roles in the innate immune response to virus infection. MG53 (TRIM72) is essential for cell membrane repair and is believed to be a muscle-specific TRIM protein. Here we show human macrophages express MG53, and MG53 protein expression is reduced following virus infection. Knockdown of MG53 in macrophages leads to increases in type I interferon (IFN) upon infection. MG53 knockout mice infected with influenza virus show comparable influenza virus titres to wild type mice, but display increased morbidity accompanied by more accumulation of CD45+ cells and elevation of IFNß in the lung. We find that MG53 knockdown results in activation of NFκB signalling, which is linked to an increase in intracellular calcium oscillation mediated by ryanodine receptor (RyR). MG53 inhibits IFNß induction in an RyR-dependent manner. This study establishes MG53 as a new target for control of virus-induced morbidity and tissue injury.