RESUMO
BACKGROUND: Advanced atrioventricular block (AVB), that is, higher than second-degree Mobitz-1, is an abnormal finding in athletes. Despite intensive investigation, in several cases the pathogenesis remains unknown, but frequently pacemaker implantation is still indicated. Increasing evidence points to circulating anti-Ro/Sjögren syndrome-related antigen A (SSA) antibodies cross-reacting with L-type calcium channel and inhibiting the related current as an epidemiologically relevant and potentially reversible cause of isolated AVB in adults. The aim of the study was to determine the prevalence of anti-Ro/SSA-associated advanced AVBs in a large sample of young athletes. METHODS AND RESULTS: A total of 2536 consecutive athletes aged <40 years without a history of cardiac diseases/interventions were enrolled in a cross-sectional study. Resting and exercise electrocardiography was performed, and those presenting any AVB were further evaluated by 24-hour Holter ECG. Athletes with second-degree AVBs and their mothers underwent anti-Ro/SSA testing. Moreover, purified immunoglobulin G from subjects with anti-Ro/SSA-positive and anti-Ro/SSA-negative advanced AVB were tested on L-type calcium current and L-type-calcium channel expression using tSA201 cells. The global prevalence of advanced AVB in the overall sample was ≈0.1%, but the risk considerably increased (2%) when intensely trained postpubertal male subjects were selectively considered. While none of the athletes with advanced AVB showed heart abnormalities, in 100% of cases anti-Ro/SSA antibodies were detected. Ex vivo experiments showed that immunoglobulin G from anti-Ro/SSA-positive but not -negative subjects with advanced AVB acutely inhibit L-type calcium current and chronically downregulate L-type-calcium channel expression. CONCLUSIONS: Our study provides evidence that advanced AVB occurs in young athletes, in most cases associated with anti-Ro/SSA antibodies blocking L-type calcium channels. These findings may open new avenues for immunomodulating therapies to reduce the risk of life-threatening events in athletes, avoiding or delaying pacemaker implantation.
Assuntos
Anticorpos Antinucleares , Atletas , Bloqueio Atrioventricular , Canais de Cálcio Tipo L , Humanos , Masculino , Feminino , Adulto , Estudos Transversais , Bloqueio Atrioventricular/imunologia , Bloqueio Atrioventricular/epidemiologia , Bloqueio Atrioventricular/diagnóstico , Prevalência , Adulto Jovem , Canais de Cálcio Tipo L/imunologia , Anticorpos Antinucleares/sangue , Anticorpos Antinucleares/imunologia , Adolescente , Eletrocardiografia Ambulatorial , Ribonucleoproteínas/imunologiaRESUMO
BACKGROUND: CACNA1C, as a type of voltage-dependent calcium ion transmembrane channel, played regulatory roles in the development and progress of multiple tumors. This study was aimed to analyze the roles of CACNA1C in ovarian cancer (OC) of overall survival (OS) and to explore its relationships with immunity. METHODS: Single gene mRNA sequencing data and corresponding clinical information were obtained from The Cancer Genome Atlas Database (TCGA) and the International Cancer Genome Consortium (ICGC) datasets. Gene set enrichment analysis (GSEA) was used to identify CACNA1C-related signal pathways. Univariate and multivariate Cox regression analyses were applied to evaluate independent prognostic factors. Besides, associations between CACNA1C and immunity were also explored. RESULTS: CACNA1C had a lower expression in OC tumor tissues than in normal tissues (P < 0.001), with significant OS (P = 0.013) and a low diagnostic efficiency. We further validated the expression levels of CACNA1C in OC by means of the ICGC dataset (P = 0.01), qRT-PCR results (P < 0.001) and the HPA database. Univariate and multivariate Cox hazard regression analyses indicated that CACNA1C could be an independent risk factor of OS for OC patients (both P < 0.001). Five significant CACNA1C-related signaling pathways were identified by means of GSEA. As for genetic alteration analysis, altered CACNA1C groups were significantly associated with OS (P = 0.0169), progression-free survival (P = 0.0404), disease-free survival (P = 0.0417) and disease-specific survival (P = 9.280e-3), compared with unaltered groups in OC. Besides, CACNA1C was dramatically associated with microsatellite instability (MSI) and immunity. CONCLUSIONS: Our results shed light on that CACNA1C could be a prognostic predictor of OS in OC and it was closely related to immunity.
Assuntos
Canais de Cálcio Tipo L/genética , Neoplasias Ovarianas/genética , Antígenos de Neoplasias , Canais de Cálcio Tipo L/imunologia , Canais de Cálcio Tipo L/metabolismo , Feminino , Humanos , Instabilidade de Microssatélites , Mutação , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/metabolismo , Prognóstico , Modelos de Riscos Proporcionais , Mapas de Interação de ProteínasRESUMO
BACKGROUND: Cardiac arrest is a tragic event that causes 1 death roughly every 90 seconds worldwide. Survivors generally undergo a workup to identify the cause of arrest. However, 5% to 10% of cardiac arrests remain unexplained. Because cardiac arrhythmias underlie most cardiac arrests and increasing evidence strongly supports the involvement of autoantibodies in arrhythmogenesis, a large-panel autoantibody screening was performed in patients with cardiac arrest. METHODS: This is an observational, cross-sectional study of patients from the Montreal Heart Institute hospital cohort, a single-center registry of participants. A peptide microarray was designed to screen for immunoglobulin G targeting epitopes from all known cardiac ion channels with extracellular domains. Plasma samples from 23 patients with unexplained cardiac arrest were compared with those from 22 patients with cardiac arrest cases of ischemic origin and a group of 29 age-, sex-, and body mass index-matched healthy subjects. The false discovery rate, least absolute shrinkage and selection operator logistic regression, and random forest methods were carried out jointly to find significant differential immunoglobulin G responses. RESULTS: The autoantibody against the pore domain of the L-type voltage-gated calcium channel was consistently identified as a biomarker of idiopathic cardiac arrest (P=0.002; false discovery rate, 0.007; classification accuracies ≥0.83). Functional studies on human induced pluripotent stem cell-derived cardiomyocytes demonstrated that the anti-L-type voltage-gated calcium channel immunoglobulin G purified from patients with idiopathic cardiac arrest is proarrhythmogenic by reducing the action potential duration through calcium channel inhibition. CONCLUSIONS: The present report addresses the concept of autoimmunity and cardiac arrest. Hitherto unknown autoantibodies targeting extracellular sequences of cardiac ion channels were detected. Moreover, the study identified an autoantibody signature specific to patients with cardiac arrest.
Assuntos
Autoanticorpos/imunologia , Autoantígenos/imunologia , Canais de Cálcio Tipo L/imunologia , Parada Cardíaca/imunologia , Potenciais de Ação , Adulto , Idoso , Sequência de Aminoácidos , Especificidade de Anticorpos , Arritmias Cardíacas/sangue , Arritmias Cardíacas/imunologia , Arritmias Cardíacas/fisiopatologia , Autoanticorpos/sangue , Biomarcadores , Diferenciação Celular , Células Cultivadas , Estudos Transversais , Feminino , Parada Cardíaca/sangue , Parada Cardíaca/epidemiologia , Sistema de Condução Cardíaco/imunologia , Sistema de Condução Cardíaco/fisiopatologia , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Células-Tronco Pluripotentes Induzidas/citologia , Canais Iônicos/imunologia , Masculino , Pessoa de Meia-Idade , Miócitos Cardíacos/imunologia , Técnicas de Patch-Clamp , Biblioteca de Peptídeos , Análise Serial de Proteínas , Quebeque/epidemiologia , Sistema de RegistrosRESUMO
BACKGROUND AND PURPOSE: Hypertension has been the leading preventable cause of premature death worldwide. The aim of this study was to design a more efficient vaccine against novel targets for the treatment of hypertension. EXPERIMENTAL APPROACH: The epitope CE12, derived from the human L-type calcium channel (CaV 1.2), was designed and conjugated with Qß bacteriophage virus-like particles to test the efficacy in hypertensive animals. Further, the hepatitis B core antigen (HBcAg)-CE12-CQ10 vaccine, a bivalent vaccine based on HBcAg virus-like particles and targeting both human angiotensin AT1 receptors and CaV 1.2 channels, was developed and evaluated in hypertensive rodents. KEY RESULTS: The Qß-CE12 vaccine effectively decreased the BP in hypertensive rodents. A monoclonal antibody against CE12 specifically bound to L-type calcium channels and inhibited channel activity. Injection with monoclonal antibody against CE12 effectively reduced the BP in angiotensin II-induced hypertensive mice. The HBcAg-CE12-CQ10 vaccine showed antihypertensive effects in hypertensive mice and relatively superior antihypertensive effects in spontaneously hypertensive rats and ameliorated L-NAME-induced renal injury. In addition, no obvious immune-mediated damage or electrophysiological adverse effects were detected. CONCLUSION AND IMPLICATIONS: Immunotherapy against both AT1 receptors and CaV 1.2 channels decreased the BP in hypertensive rodents effectively and provided protection against hypertensive target organ damage without obvious feedback activation of renin-angiotensin system or induction of dominant antibodies against the carrier protein. Thus, the HBcAg-CE12-CQ10 vaccine may provide a novel and promising therapeutic approach for hypertension.
Assuntos
Pressão Sanguínea/efeitos dos fármacos , Canais de Cálcio Tipo L/imunologia , Hipertensão/prevenção & controle , Receptor Tipo 1 de Angiotensina/imunologia , Vacinas Combinadas/farmacologia , Vacinas de Partículas Semelhantes a Vírus/farmacologia , Angiotensina II , Animais , Canais de Cálcio Tipo L/metabolismo , Modelos Animais de Doenças , Epitopos , Hipertensão/imunologia , Hipertensão/metabolismo , Hipertensão/fisiopatologia , Masculino , Camundongos Endogâmicos BALB C , Ratos Endogâmicos SHR , Receptor Tipo 1 de Angiotensina/metabolismo , VacinaçãoRESUMO
Recent studies have postulated the presence of functional autoantibodies (Abs) against L-type voltage gated calcium channels (VGCCs) in the serum of patients with type 1 diabetes, with various proposed physiological consequences, both islet cell associated and extra-glandular. Arguably, the most potentially damaging effect reported for these Abs is induction of apoptosis in pancreatic beta (ß) cells, yet a convincing pathogenic mechanism remains to be demonstrated. In the current study, we report an assay of reactive oxygen species (ROS) stress induction in the rat insulinoma cell line Rin A12, as determined by 2', 7'-Dichlorofluorescein diacetate (DCF-DA) fluorescence detection by flow cytometry. We demonstrate that incubation of Rin A12 cells with immunoglobulin G (IgG) containing anti-VGCC activity from patients with T1D mediates a significant increase in ROS, with subsequent induction of apoptosis, as determined by positivity for annexin V expression. Neither T1D patient-derived IgG lacking anti-VGCC activity or IgG from healthy donors altered ROS or annexin V expression, indicating the new assay is specific for the detection of functional anti-VGCC Abs. Subsequent screening of IgG samples derived from individual patients indicated a prevalence of approximately 75% in a cohort of 20 patients with T1D. The new cell-based assay provides, for the first time, experimental evidence supporting a plausible pathophysiological mechanism underlying anti-VGCC Ab-mediated apoptosis induction in ß cells. Additionally, the assay is a considerable advance on previously published methods for detecting and characterising the functional activity of anti-VGCC Abs in patient-derived samples.
Assuntos
Bioensaio , Canais de Cálcio Tipo L/imunologia , Diabetes Mellitus Tipo 1 , Imunoglobulina G , Células Secretoras de Insulina , Adolescente , Adulto , Idoso , Animais , Apoptose/imunologia , Autoanticorpos/sangue , Autoanticorpos/imunologia , Canais de Cálcio Tipo L/metabolismo , Linhagem Celular Tumoral , Diabetes Mellitus Tipo 1/sangue , Diabetes Mellitus Tipo 1/imunologia , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Células Secretoras de Insulina/imunologia , Células Secretoras de Insulina/metabolismo , Masculino , Pessoa de Meia-Idade , Ratos , Espécies Reativas de Oxigênio/sangue , Espécies Reativas de Oxigênio/imunologiaRESUMO
BACKGROUND: T lymphocytes express not only cell membrane ORAI calcium release-activated calcium modulator 1 but also voltage-gated calcium channel (Cav) 1 channels. In excitable cells these channels are composed of the ion-forming pore α1 and auxiliary subunits (ß and α2δ) needed for proper trafficking and activation of the channel. Previously, we disclosed the role of Cav1.2 α1 in mouse and human TH2 but not TH1 cell functions and showed that knocking down Cav1 α1 prevents experimental asthma. OBJECTIVE: We investigated the role of ß and α2δ auxiliary subunits on Cav1 α1 function in TH2 lymphocytes and on the development of acute allergic airway inflammation. METHODS: We used Cavß antisense oligonucleotides to knock down Cavß and gabapentin, a drug that binds to and inhibits α2δ1 and α2δ2, to test their effects on TH2 functions and their capacity to reduce allergic airway inflammation. RESULTS: Mouse and human TH2 cells express mainly Cavß1, ß3, and α2δ2 subunits. Cavß antisense reduces T-cell receptor-driven calcium responses and cytokine production by mouse and human TH2 cells with no effect on TH1 cells. Cavß is mainly involved in restraining Cav1.2 α1 degradation through the proteasome because a proteasome inhibitor partially restores the α1 protein level. Gabapentin impairs the T-cell receptor-driven calcium response and cytokine production associated with the loss of α2δ2 protein in TH2 cells. CONCLUSIONS: These results stress the role of Cavß and α2δ2 auxiliary subunits in the stability and activation of Cav1.2 channels in TH2 lymphocytes both in vitro and in vivo, as demonstrated by the beneficial effect of Cavß antisense and gabapentin in allergic airway inflammation.
Assuntos
Canais de Cálcio Tipo L/imunologia , Hipersensibilidade/imunologia , Subunidades Proteicas/imunologia , Linfócitos T/imunologia , Doença Aguda , Alérgenos , Animais , Feminino , Inflamação/imunologia , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , OvalbuminaRESUMO
The obligate intracellular parasite Toxoplasma gondii exploits cells of the immune system to disseminate. Upon T. gondii-infection, γ-aminobutyric acid (GABA)/GABAA receptor signaling triggers a hypermigratory phenotype in dendritic cells (DCs) by unknown signal transduction pathways. Here, we demonstrate that calcium (Ca2+) signaling in DCs is indispensable for T. gondii-induced DC hypermotility and transmigration in vitro. We report that activation of GABAA receptors by GABA induces transient Ca2+ entry in DCs. Murine bone marrow-derived DCs preferentially expressed the L-type voltage-dependent Ca2+ channel (VDCC) subtype Cav1.3. Silencing of Cav1.3 by short hairpin RNA or selective pharmacological antagonism of VDCCs abolished the Toxoplasma-induced hypermigratory phenotype. In a mouse model of toxoplasmosis, VDCC inhibition of adoptively transferred Toxoplasma-infected DCs delayed the appearance of cell-associated parasites in the blood circulation and reduced parasite dissemination to target organs. The present data establish that T. gondii-induced hypermigration of DCs requires signaling via VDCCs and that Ca2+ acts as a second messenger to GABAergic signaling via the VDCC Cav1.3. The findings define a novel motility-related signaling axis in DCs and unveil that interneurons and DCs share common GABAergic motogenic pathways. T. gondii employs GABAergic non-canonical pathways to induce host cell migration and facilitate dissemination.
Assuntos
Canais de Cálcio Tipo L/imunologia , Sinalização do Cálcio , Células Dendríticas/imunologia , Receptores de GABA-A/imunologia , Toxoplasma/imunologia , Toxoplasmose/imunologia , Transferência Adotiva , Animais , Movimento Celular , Células Cultivadas , Células Dendríticas/parasitologia , GABAérgicos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Toxoplasma/fisiologia , Toxoplasmose/parasitologia , Ácido gama-Aminobutírico/imunologiaRESUMO
The prevalence of Mycobacterium tuberculosis (M. tb) strains eliciting drug resistance has necessitated the need for understanding the complexities of host pathogen interactions. The regulation of calcium homeostasis by Voltage Gated Calcium Channel (VGCCs) upon M. tb infection has recently assumed importance in this area. We previously showed a suppressor role of VGCC during M. tb infections and recently reported the mechanisms of its regulation by M. tb. Here in this report, we further characterize the role of VGCC in mediating defence responses of macrophages during mycobacterial infection. We report that activation of VGCC during infection synergistically downmodulates the generation of oxidative burst (ROS) by macrophages. This attenuation of ROS is regulated in a manner which is dependent on Toll like Receptor (TLR) and also on the route of calcium influx, Protein Kinase C (PKC) and by Mitogen Activation Protein Kinase (MAPK) pathways. VGCC activation during infection increases cell survival and downmodulates autophagy. Concomitantly, pro-inflammatory responses such as IL-12 and IFN-γ secretion and the levels of their receptors on cell surface are inhibited. Finally, the ability of phagosomes to fuse with lysosomes in M. bovis BCG and M. tb H37Rv infected macrophages is also compromised when VGCC activation occurs during infection. The results point towards a well-orchestrated strategy adopted by mycobacteria to supress protective responses mounted by the host. This begins with the increase in the surface levels of VGCCs by mycobacteria and their antigens by well-controlled and regulated mechanisms. Subsequent activation of the upregulated VGCC following tweaking of calcium levels by molecular sensors in turn mediates suppressor responses and prepare the macrophages for long term persistent infection.
Assuntos
Canais de Cálcio Tipo L/imunologia , Sinalização do Cálcio/imunologia , Macrófagos/imunologia , Infecções por Mycobacterium/imunologia , Mycobacterium bovis/imunologia , Animais , Linhagem Celular Tumoral , Feminino , Humanos , Interferon gama/imunologia , Interleucina-12/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Espécies Reativas de Oxigênio/imunologiaRESUMO
There is accumulating evidence during sepsis that cardiomyocyte (CM) homeostasis is compromised, resulting in cardiac dysfunction. An important role for complement in these outcomes is now demonstrated. Addition of C5a to electrically paced CMs caused prolonged elevations of intracellular Ca(2+) concentrations during diastole, together with the appearance of spontaneous Ca(2+) transients. In polymicrobial sepsis in mice, we found that three key homeostasis-regulating proteins in CMs were reduced: Na(+)/K(+)-ATPase, which is vital for effective action potentials in CMs, and two intracellular Ca(2+) concentration regulatory proteins, that is, sarcoplasmic/endoplasmic reticulum calcium ATPase 2 and the Na(+)/Ca(2+) exchanger. Sepsis caused reduced mRNA levels and reductions in protein concentrations in CMs for all three proteins. The absence of either C5a receptor mitigated sepsis-induced reductions in the three regulatory proteins. Absence of either C5a receptor (C5aR1 or C5aR2) diminished development of defective systolic and diastolic echocardiographic/Doppler parameters developing in the heart (cardiac output, left ventricular stroke volume, isovolumic relaxation, E' septal annulus, E/E' septal annulus, left ventricular diastolic volume). We also found in CMs from septic mice the presence of defective current densities for Ik1, l-type calcium channel, and Na(+)/Ca(2+) exchanger. These defects were accentuated in the copresence of C5a. These data suggest complement-related mechanisms responsible for development of cardiac dysfunction during sepsis.
Assuntos
Coinfecção/imunologia , Miócitos Cardíacos/imunologia , Miócitos Cardíacos/patologia , Sepse/imunologia , Sepse/fisiopatologia , Animais , Cálcio/metabolismo , Canais de Cálcio Tipo L/imunologia , Coinfecção/microbiologia , Coinfecção/fisiopatologia , Complemento C5a/imunologia , Citoplasma/química , Citoplasma/metabolismo , Coração/fisiopatologia , Camundongos , Miócitos Cardíacos/microbiologia , Receptor da Anafilatoxina C5a/deficiência , Receptor da Anafilatoxina C5a/imunologia , Receptor da Anafilatoxina C5a/fisiologia , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/imunologia , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/metabolismo , Sepse/complicaçõesRESUMO
Cacna1s encodes the α1S subunit (Cav1.1) of voltage-dependent calcium channels, and is required for normal skeletal and cardiac muscle function, where it couples with the ryanodine receptor to regulate muscle contraction. Recently CACNA1S was reported to be expressed on the tips of retinal depolarizing bipolar cells (DBCs) and colocalized with metabotropic glutamate receptor 6 (mGluR6), which is critical to DBC signal transduction. Further, in mGluR6 knockout mice, expression at this location is down regulated. We examined RNAseq data from mouse retina and found expression of a novel isoform of Cacna1s. To determine if CACNA1S was a functional component of the DBC signal transduction cascade, we performed immunohistochemistry to visualize its expression in several mouse lines that lack DBC function. Immunohistochemical staining with antibodies to CACNA1S show punctate labeling at the tips of DBCs in wild type (WT) retinas that are absent in Gpr179 nob5 mutant retinas and decreased in Grm6 -/- mouse retinas. CACNA1S and transient receptor potential cation channel, subfamily M, member 1 (TRPM1) staining also colocalized in WT retinas. Western blot analyses for CACNA1S of either retinal lysates or proteins after immunoprecipitation with the CACNA1S antibody failed to show the presence of bands expected for CACNA1S. Mass spectrometric analysis of CACNA1S immunoprecipitated proteins also failed to detect any peptides matching CACNA1S. Immunohistochemistry and western blotting after expression of GPR179 in HEK293T cells indicate that the CACNA1S antibody used here and in the retinal studies published to date, cross-reacts with GPR179. These data suggest caution should be exercised in conferring a role for CACNA1S in DBC signal transduction based solely on immunohistochemical staining.
Assuntos
Reações Antígeno-Anticorpo , Canais de Cálcio Tipo L/genética , Canais de Cálcio Tipo L/imunologia , Regulação da Expressão Gênica/fisiologia , Receptores Acoplados a Proteínas G/imunologia , Retina/metabolismo , Células Bipolares da Retina/metabolismo , Sequência de Aminoácidos , Animais , Western Blotting , Reações Cruzadas , Feminino , Células HEK293 , Humanos , Imuno-Histoquímica , Síndromes de Imunodeficiência , Masculino , Espectrometria de Massas , Camundongos , Camundongos Knockout , Dados de Sequência Molecular , Doenças da Imunodeficiência Primária , Isoformas de Proteínas , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Transdução de SinaisRESUMO
Trp-His, the anti-atherosclerotic dipeptide, exerted an antiproliferative effect on vascular smooth muscle cells by L-type Ca(2+) channel blocker-like effect. The beneficial potential by the blockade of Ca(2+) channels on chronic intestinal inflammation, including inflammatory bowel disease (IBD), is unclear. Trp-His (100 or 250 mg/kg body weight/day) was administered for 14 days to BALB/c mice, and 5% dextran sodium sulfate (DSS) was administered to induce colitis in the last 7 days. Trp-His reduced DSS-induced typical colitis symptoms and cytokine expression in the colon. Trp-His inhibited interleukin (IL)-8 secretion in tumor necrosis factor (TNF)-α-stimulated HT-29 cells. The inhibitory effect of Trp-His, as well as that of Ca(2+) channel blockers, was impaired by the presence of Ca(2+) channel agonist Bay K 8644. The TNF-α-induced activation of mitogen-activated protein kinases (MAPKs) and IκBα were decreased by Trp-His. These results indicated that the anti-inflammatory effect of Trp-His may be involved in the blockade of L-type Ca(2+) channels.
Assuntos
Anti-Inflamatórios/administração & dosagem , Canais de Cálcio Tipo L/imunologia , Dipeptídeos/administração & dosagem , Doenças Inflamatórias Intestinais/tratamento farmacológico , Intestinos/imunologia , Animais , Canais de Cálcio Tipo L/genética , Feminino , Humanos , Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/imunologia , Interleucina-8/genética , Interleucina-8/imunologia , Intestinos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologiaRESUMO
OBJECTIVE: To explore the effect of interleukin-17A (IL-17A) on the serum level of antiheart autoantibodies in mice with viral myocarditis. METHODS: Male wild-type (WT) and IL-17A-deficient (IL-17A(-/-)) BALB/c mice were intraperitoneally injected with Coxsackie virus B3 (CVB3) for establishing VMC models (VMC-WT group and VMC-IL-17A(-/-) group). Meanwhile, a control group (WT group) of WT mice were established by i.p. administration of phosphate buffered saline (PBS). Paraffin sections of cardiac tissues were made 14 days after CVB3 injection. Myocardial histopathologic changes were evaluated by HE staining. The levels of anti-adenine nucleotide translocator (ANT) autoantibody, anti-ß-myosin heavy chain (ß-MHC) autoantibody and anti-cardiac L-type calcium channel (CACH2) autoantibody in sera were measured by ELISA. RESULTS: Compared with WT group, the levels of anti-ANT-autoantibody and anti-ß-MHC-autoantibody significantly increased in VMC-WT group (P<0.01, P<0.05), while the concentration of anti-CACH2-autoantibody showed no significant difference between WT and VMC-WT groups (P>0.05). Compared with VMC-WT group, the level of anti-ANT-autoantibody was reduced in VMC-IL-17A(-/-) group (P<0.05), while the levels of anti-ß-MHC-autoantibody and anti-CACH2-autoantibody showed no significant difference between them (P>0.05). CONCLUSION: IL-17A contributed to the secretion of anti-ANT-autoantibody of VMC mice, but had no effect on the secretion of anti-ß-MHC-autoantibody and anti-CACH2-autoantibody in VMC mice.
Assuntos
Autoanticorpos/imunologia , Infecções por Coxsackievirus/imunologia , Enterovirus Humano B/imunologia , Interleucina-17/imunologia , Miocardite/imunologia , Animais , Autoanticorpos/sangue , Canais de Cálcio Tipo L/imunologia , Infecções por Coxsackievirus/genética , Infecções por Coxsackievirus/virologia , Enterovirus Humano B/fisiologia , Ensaio de Imunoadsorção Enzimática , Células Hep G2 , Interações Hospedeiro-Patógeno/imunologia , Humanos , Interleucina-17/genética , Interleucina-17/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Translocases Mitocondriais de ADP e ATP/imunologia , Miocardite/genética , Miocardite/virologia , Miocárdio/imunologia , Miocárdio/patologia , Cadeias Pesadas de Miosina/imunologia , Miosina não Muscular Tipo IIB/imunologiaRESUMO
The purpose of this study was to evaluate whether CC-AAbs levels could predict prognosis in CHF patients. A total of 2096 patients with CHF (841 DCM patients and 1255 ICM patients) and 834 control subjects were recruited. CC-AAbs were detected and the relationship between CC-AAbs and patient prognosis was analyzed. During a median follow-up time of 52 months, there were 578 deaths. Of these, sudden cardiac death (SCD) occurred in 102 cases of DCM and 121 cases of ICM. The presence of CC-AAbs in patients was significantly higher than that of controls (both P < 0.001). Multivariate analysis revealed that positive CC-AAbs could predict SCD (HR 3.191, 95% CI 1.598-6.369 for DCM; HR 2.805, 95% CI 1.488-5.288 for ICM) and all-cause mortality (HR 1.733, 95% CI 1.042-2.883 for DCM; HR 2.219, 95% CI 1.461-3.371 for ICM) in CHF patients. A significant association between CC-AAbs and non-SCD (NSCD) was found in ICM patients (HR = 1.887, 95% CI 1.081-3.293). Our results demonstrated that the presence of CC-AAbs was higher in CHF patients versus controls and corresponds to a higher incidence of all-cause death and SCD. Positive CC-AAbs may serve as an independent predictor for SCD and all-cause death in these patients.
Assuntos
Autoanticorpos/sangue , Canais de Cálcio Tipo L/imunologia , Morte Súbita Cardíaca , Insuficiência Cardíaca/sangue , Isquemia Miocárdica/sangue , Adulto , Idoso , Biomarcadores/sangue , Estudos de Casos e Controles , Doença Crônica , Feminino , Insuficiência Cardíaca/imunologia , Insuficiência Cardíaca/mortalidade , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Isquemia Miocárdica/imunologia , Isquemia Miocárdica/mortalidade , PrognósticoRESUMO
The goal of this study is to determine whether patients with paraneoplastic cerebellar degeneration (PCD) and small-cell lung cancer (SCLC) have a specific repertoire of antibodies, if SOX1 antibodies (SOX1-ab) can predict the presence of SCLC, and if antibodies to cell surface antigens occur in this syndrome. Antibody analysis was done using immunohistochemistry on rat brain, immunoblot with recombinant antigens, screening of cDNA expression libraries, and immunolabeling of live neurons in 39 patients with PCD and SCLC. VGCC-ab were measured by RIA, and SOX1-ab, Hu-ab, and ZIC4-ab by immunoblot. Lambert-Eaton myastenic syndrome (LEMS) was present in 10 of 23 patients with electrophysiological studies. At least one antibody was detected in 72% of patients. The individual frequencies were: 49% SOX1-ab, 44% VGCC-ab, 31% Hu-ab, and 13% ZIC4-ab. SOX1-ab occurred in 76% of patients with VGCC-ab and 27% of those without VGCC-ab (pâ=â0.0036). SOX1-ab were not found in 39 patients with sporadic late-onset cerebellar ataxia, 23 with cerebellar ataxia and glutamic acid decarboxylase antibodies, and 73 with PCD and cancer types other than SCLC (31 without onconeural antibodies, 25 with Yo-ab , 17 with Tr-ab). Five patients (13%) had antibodies against unknown neuronal cell surface antigens but none of them improved with immunotherapy. One serum immunoreacted against the axon initial segment of neurons and another serum against ELKS1, a protein highly expressed in the cerebellum that interacts with the beta4-subunit of the VGCC. In conclusion, 72% of patients with PCD and SCLC had one or more antibodies that indicate the presence of this tumor. In these patients, VGCC-ab and SOX1-ab occur tightly associated. SOX1-ab are predictors of SCLC in ataxia patients with a specificity of 100% and sensitivity of 49%. Unlike limbic encephalitis with SCLC, antibodies to cell surface antigens other than VGCC-ab, are infrequent and do not predict response to treatment.
Assuntos
Anticorpos/imunologia , Neoplasias Pulmonares/imunologia , Degeneração Paraneoplásica Cerebelar/imunologia , Carcinoma de Pequenas Células do Pulmão/imunologia , Idoso , Idoso de 80 Anos ou mais , Anticorpos/sangue , Biomarcadores Tumorais/sangue , Canais de Cálcio Tipo L/imunologia , Proteínas ELAV/imunologia , Feminino , Humanos , Neoplasias Pulmonares/complicações , Masculino , Pessoa de Meia-Idade , Proteínas do Tecido Nervoso/imunologia , Degeneração Paraneoplásica Cerebelar/etiologia , Fatores de Transcrição SOXB1/imunologia , Carcinoma de Pequenas Células do Pulmão/complicações , Fatores de Transcrição/imunologiaRESUMO
The human antimicrobial peptide cathelicidin LL-37 has, besides its antimicrobial properties, also been shown to regulate apoptosis in a cell type-specific manner. Mechanisms involved in LL-37-regulated apoptotic signaling are not identified. Here, we show that LL-37 reduces the human osteoblast-like MG63 cell number and cell viability in the micromolar concentration range with an IC50 value of about 5 µM. Treatment with 4 µM LL-37 increased the number of annexin V-positive cells and stimulated activation of caspase 3 showing that LL-37 promotes apoptosis. Treatment with 4 µM LL-37 caused an acute and sustained rise in intracellular Ca(2+) concentration assessed by laser-scanning confocal microscopy of Fluo-4-AM-loaded MG63 cells. LL-37 increased Ca(2+) also in the presence of the respective L- and T-type voltage-sensitive Ca(2+) channel blockers nifedipine and NiCl2. LL-37 had no effect on Ca(2+) in cells incubated with Ca(2+)-free solution. LL-37 (4 and 8 µM) reduced the MG63 cell number both in the presence and absence of Ca(2+) in the medium. In conclusion, LL-37 reduces the osteoblast cell number by promoting apoptosis, and furthermore, LL-37 stimulates Ca(2+) inflow via a mechanism independent of voltage-sensitive Ca(2+) channels. Interestingly, LL-37-induced lowering of the cell number seems to be mediated via a mechanism independent of Ca(2+).
Assuntos
Peptídeos Catiônicos Antimicrobianos/farmacologia , Apoptose/efeitos dos fármacos , Cálcio/imunologia , Osteoblastos/imunologia , Anexina A5/imunologia , Anexina A5/metabolismo , Peptídeos Catiônicos Antimicrobianos/imunologia , Apoptose/imunologia , Cálcio/metabolismo , Canais de Cálcio Tipo L/imunologia , Canais de Cálcio Tipo L/metabolismo , Canais de Cálcio Tipo T/imunologia , Canais de Cálcio Tipo T/metabolismo , Linhagem Celular , Relação Dose-Resposta a Droga , Humanos , Osteoblastos/metabolismo , CatelicidinasRESUMO
AIMS: Calcium channel plays an important role in the autoimmune pathogenesis of idiopathic dilated cardiomyopathy (DCM). Autoantibodies have emerged as a new upstream target of sudden death in DCM. We sought to validate the hypothesis that autoantibodies against l-type calcium channel (CC-AAbs) are arrhythmogenic and lead to sudden death in patients with DCM. METHODS AND RESULTS: We investigated sudden death and ventricular arrhythmias in 80 patients with DCM in a prospective, case follow-up survey. During a follow-up of 32 (SD 8) months, CC-AAbs-positive patients not only had a higher incidence of ventricular tachycardia (VT) but also a higher incidence of sudden death than CC-AAbs-negative patients (for VT: 59.0 vs. 24.4%, P = 0.002 and for sudden death: 20.5 vs. 4.9%, P = 0.045). Further univariate and multivariate analyses showed that the occurrence of CC-AAbs was the strongest independent predictor for sudden death (odds ratio: 10.20, 95% confidence interval: 2.43-36.78, P = 0.0027). Experimental studies in ex vivo systems using affinity-purified CC-AAbs from patients demonstrated that CC-AAbs were able to induce VT by prolongation of action potential duration (APD) and triggered activity by early afterdepolarization (EAD). CONCLUSION: Our results demonstrate for the first time to our knowledge that there is a high incidence of sudden death and VT in CC-AAbs-positive patients with DCM. Furthermore, experimental data from ex vivo systems suggest that CC-AAbs might induce VT by prolongation of APD and triggered activity by EAD.
Assuntos
Autoanticorpos , Canais de Cálcio Tipo L/imunologia , Morte Súbita Cardíaca/etiologia , Potenciais de Ação/fisiologia , Animais , Arritmias Cardíacas/imunologia , Cardiomiopatia Dilatada/complicações , Cardiomiopatia Dilatada/imunologia , Cardiomiopatia Dilatada/mortalidade , Cardiomiopatia Dilatada/fisiopatologia , Morte Súbita Cardíaca/epidemiologia , Modelos Animais de Doenças , Feminino , Seguimentos , Humanos , Modelos Logísticos , Masculino , Estudos Prospectivos , Ratos , Ratos Sprague-Dawley , Taquicardia Ventricular/imunologiaRESUMO
Considering that α-1 repeat region may be involved in the ion binding and translocation of Na(+)-Ca(2+) exchanger (NCX), it is possible that the antibodies against NCX α-1 repeat may have a crucial action on NCX activity. The aim of the present study is to investigate the effect of antibody against α-1 repeat (117-137), designated as α-1(117-137), on NCX activity. The antibody against the synthesized α-1(117-137) was prepared and affinity-purified. Whole-cell patch clamp technique was used to study the change of Na(+)-Ca(2+) exchange current (I(Na/Ca)) in adult rat cardiomyocytes. To evaluate the functional specificity of this antibody, its effects on L-type Ca(2+) current (I(Ca,L)), voltage-gated Na(+) current (I(Na)) and delayed rectifier K(+) current (I(K)) were also observed. The amino acid sequences of α-1(117-137) in NCX and residues 1 076-1 096 within L-type Ca(2+) channel were compared using EMBOSS Pairwise Alignment Algorithms. The results showed that outward and inward I(Na/Ca) were decreased by the antibody against α-1(117-137) dose-dependently in the concentration range from 10 to 160 nmol/L, with IC(50) values of 18.9 nmol/L and 22.4 nmol/L, respectively. Meanwhile, the antibody also decreased I(Ca,L) in a concentration-dependent manner with IC(50) of 22.7 nmol/L. No obvious effects of the antibody on I(Na) and I(K) were observed. Moreover, comparison of the amino acid sequences showed there was 23.8% sequence similarity between NCX α-1(117-137) and residues 1 076-1 096 within L-type Ca(2+) channel. These results suggest that antibody against α-1(117-137) is a blocking antibody to NCX and can also decrease I(Ca,L) in a concentration-dependent manner, while it does not have obvious effects on I(Na) and I(K).
Assuntos
Anticorpos Bloqueadores/farmacologia , Bloqueadores dos Canais de Cálcio/farmacologia , Canais de Cálcio Tipo L , Miócitos Cardíacos/metabolismo , Trocador de Sódio e Cálcio/antagonistas & inibidores , Sequência de Aminoácidos , Animais , Anticorpos Bloqueadores/metabolismo , Canais de Cálcio Tipo L/genética , Canais de Cálcio Tipo L/imunologia , Canais de Cálcio Tipo L/metabolismo , Cobaias , Potenciais da Membrana , Dados de Sequência Molecular , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/fisiologia , Técnicas de Patch-Clamp , Ratos , Ratos Wistar , Trocador de Sódio e Cálcio/genética , Trocador de Sódio e Cálcio/imunologiaRESUMO
Congenital heart block (CHB) is a conduction abnormality that affects hearts of foetuses and/or newborn to mothers with autoantibodies reactive with the intracellular soluble ribonucleoproteins 48-kD La, 52-kD Ro and 60-kD Ro. CHB carries substantial mortality and morbidity, with more than 60% of affected children requiring lifelong pacemakers. Several hypotheses have been proposed to explain the pathogenesis of CHB. These can be grouped under three main hypotheses: Apoptosis, Serotoninergic and Ca channel hypothesis. Here, we discuss these hypotheses and provide recent scientific thinking that will most likely dominate the future of this field of research.
Assuntos
Canais de Cálcio Tipo L/imunologia , Canais de Cálcio Tipo L/metabolismo , Bloqueio Cardíaco/congênito , Bloqueio Cardíaco/etiologia , Bloqueio Cardíaco/imunologia , Doenças do Recém-Nascido/etiologia , Doenças do Recém-Nascido/metabolismo , Modelos Biológicos , Animais , Apoptose/imunologia , Autoanticorpos/imunologia , Feminino , Bloqueio Cardíaco/patologia , Humanos , Recém-Nascido , Doenças do Recém-Nascido/imunologia , Doenças do Recém-Nascido/patologia , Troca Materno-Fetal/imunologia , Gravidez , Receptores 5-HT4 de Serotonina/imunologiaRESUMO
We have recently postulated that functional autoantibodies (Abs) against L-type voltage-gated calcium channels (VGCCs) contribute to autonomic dysfunction in type 1 diabetes (T1D). Previous studies based on whole-organ assays have proven valuable in establishing the mechanism of anti-VGCC Ab activity, but are complex and unsuitable for screening large patient cohorts. In the current study, we used real-time dynamic monitoring of cell impedance to demonstrate that anti-VGCC Abs from patients with T1D inhibit the adherence of Rin A12 cells. The functional effect of the anti-VGCC Abs was mimicked by the dihydropyridine agonist, Bay K8644, and reversed by the antagonist, nicardipine, providing a pharmacological link to the whole-organ studies. IVIg neutralized the effect on cell adhesion of the anti-VGCC Abs, consistent with the presence of anti-idiotypic Abs in IVIg that may prevent the emergence of pathogenic Abs in healthy individuals. The cell impedance assay can be performed in a 96 well plate format, and represents a simple method for detecting the presence of anti-VGCC activity in patient immunoglobulin (IgG). The new cell assay should prove useful for further studies to determine the prevalence of the Ab and its association with symptoms of autonomic dysfunction in patients with T1D.
Assuntos
Autoanticorpos/imunologia , Canais de Cálcio Tipo L/imunologia , Adesão Celular/imunologia , Diabetes Mellitus Tipo 1/imunologia , Impedância Elétrica , Imunoglobulina G/imunologia , Éster Metílico do Ácido 3-Piridinacarboxílico, 1,4-Di-Hidro-2,6-Dimetil-5-Nitro-4-(2-(Trifluormetil)fenil)/farmacologia , Animais , Autoanticorpos/análise , Agonistas dos Canais de Cálcio/farmacologia , Bloqueadores dos Canais de Cálcio/farmacologia , Linhagem Celular , Humanos , Nicardipino/farmacologia , RatosRESUMO
Ca(2+) is a highly versatile intracellular second messenger in many cell types, and regulates many complicated cellular processes, including cell activation, proliferation and apoptosis. Influx of Ca(2+) from the extracellular fluid is required for sustained elevation of the cytosolic Ca(2+) concentration and full activation of Ca(2+)-dependent processes. It is widely accepted that Ca(2+) release-activated Ca(2+) channels are the major routes of Ca(2+) influx in electrically non-excitable cells, including hematopoietic cells, whereas voltage-gated Ca(2+) channels such as L-type Ca(2+) channels (LTCCs) serve as the principal routes of Ca(2+) entry into electrically excitable cells such as neurons and myocytes. However, recent pharmacological and molecular genetic studies have revealed the existence of functional LTCCs and/or LTCC-like channels in a variety of immune cells including mast cells. In this article, we review recent advances in our understanding of Ca(2+) signaling in immune cells with a special interest in mast cells. We highlight roles for LTCCs in antigen receptor-mediated mast cell activation and survival.