RESUMEN
Diversity of neural crest derivatives has been studied with a variety of approaches during embryonic development. In mammals Cre-LoxP lineage tracing is a robust means to fate map neural crest relying on cre driven from regulatory elements of early neural crest genes. Sox10 is an essential transcription factor for normal neural crest development. A variety of efforts have been made to label neural crest derivatives using partial Sox10 regulatory elements to drive cre expression. To date published Sox10-cre lines have focused primarily on lineage tracing in specific tissues or during early fetal development. We describe two new Sox10-cre BAC transgenes, constitutive (cre) and inducible (cre/ERT2), that contain the complete repertoire of Sox10 regulatory elements. We present a thorough expression profile of each, identifying a few novel sites of Sox10 expression not captured by other neural crest cre drivers. Comparative mapping of expression patterns between the Sox10-cre and Sox10-cre/ERT2 transgenes identified a narrow temporal window in which Sox10 expression is present in mesenchymal derivatives prior to becoming restricted to neural elements during embryogenesis. In more caudal structures, such as the intestine and lower urinary tract, our Sox10-cre BAC transgene appears to be more efficient in labeling neural crest-derived cell types than Wnt1-cre. The analysis reveals consistent expression of Sox10 in non-neural crest derived glandular epithelium, including salivary, mammary, and urethral glands of adult mice. These Sox10-cre and Sox10-cre/ERT2 transgenic lines are verified tools that will enable refined temporal and cell-type specific lineage analysis of neural crest derivatives as well as glandular tissues that rely on Sox10 for proper development and function.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Mesodermo/embriología , Cresta Neural/embriología , Factores de Transcripción SOXE/biosíntesis , Cráneo/embriología , Transgenes , Animales , Mesodermo/citología , Ratones , Ratones Transgénicos , Cresta Neural/citología , Factores de Transcripción SOXE/genética , Cráneo/citologíaRESUMEN
The migration and fate of cranial and vagal neural crest-derived progenitor cells (NCPCs) have been extensively studied; however, much less is known about sacral NCPCs particularly in regard to their distribution in the urogenital system. To construct a spatiotemporal map of NCPC migration pathways into the developing lower urinary tract, we utilized the Sox10-H2BVenus transgene to visualize NCPCs expressing Sox10. Our aim was to define the relationship of Sox10-expressing NCPCs relative to bladder innervation, smooth muscle differentiation, and vascularization through fetal development into adulthood. Sacral NCPC migration is a highly regimented, specifically timed process, with several potential regulatory mileposts. Neuronal differentiation occurs concomitantly with sacral NCPC migration, and neuronal cell bodies are present even before the pelvic ganglia coalesce. Sacral NCPCs reside within the pelvic ganglia anlagen through 13.5 days post coitum (dpc), after which they begin streaming into the bladder body in progressive waves. Smooth muscle differentiation and vascularization of the bladder initiate prior to innervation and appear to be independent processes. In adult bladder, the majority of Sox10+ cells express the glial marker S100ß, consistent with Sox10 being a glial marker in other tissues. However, rare Sox10+ NCPCs are seen in close proximity to blood vessels and not all are S100ß+, suggesting either glial heterogeneity or a potential nonglial role for Sox10+ cells along vasculature. Taken together, the developmental atlas of Sox10+ NCPC migration and distribution profile of these cells in adult bladder provided here will serve as a roadmap for future investigation in mouse models of lower urinary tract dysfunction.
Asunto(s)
Movimiento Celular , Cresta Neural/citología , Sacro/citología , Sistema Urogenital/inervación , Animales , Diferenciación Celular , Embrión de Mamíferos/citología , Embrión de Mamíferos/metabolismo , Endotelio Vascular/metabolismo , Ganglios/metabolismo , Mesodermo/metabolismo , Ratones Transgénicos , Miocitos del Músculo Liso/citología , Cresta Neural/metabolismo , Neuroglía/citología , Neuroglía/metabolismo , Factores de Transcripción SOXE/metabolismo , Células Madre/citología , Células Madre/metabolismo , Factores de Tiempo , Sistema Urogenital/irrigación sanguíneaRESUMEN
CD40 interacts with CD40L and plays an essential role in immune regulation and homeostasis. Recent research findings, however, support a pathogenic role of CD40 in a number of autoimmune diseases. We previously showed that memory B cells from relapsing-remitting multiple sclerosis (RRMS) patients exhibited enhanced proliferation with CD40 stimulation compared with healthy donors. In this study, we used a multiparameter phosflow approach to analyze the phosphorylation status of NF-κB and three major MAPKs (P38, ERK, and JNK), the essential components of signaling pathways downstream of CD40 engagement in B cells from MS patients. We found that memory and naive B cells from RRMS and secondary progressive MS patients exhibited a significantly elevated level of phosphorylated NF-κB (p-P65) following CD40 stimulation compared with healthy donor controls. Combination therapy with IFN-ß-1a (Avonex) and mycophenolate mofetil (Cellcept) modulated the hyperphosphorylation of P65 in B cells of RRMS patients at levels similar to healthy donor controls. Lower disease activity after the combination therapy correlated with the reduced phosphorylation of P65 following CD40 stimulation in treated patients. Additionally, glatiramer acetate treatment also significantly reduced CD40-mediated P65 phosphorylation in RRMS patients, suggesting that reducing CD40-mediated p-P65 induction may be a general mechanism by which some current therapies modulate MS disease.
Asunto(s)
Linfocitos B/inmunología , Antígenos CD40/inmunología , Acetato de Glatiramer/administración & dosificación , Interferón beta-1a/administración & dosificación , Activación de Linfocitos/efectos de los fármacos , Sistema de Señalización de MAP Quinasas , Esclerosis Múltiple , Ácido Micofenólico/administración & dosificación , Factor de Transcripción ReIA/inmunología , Anciano , Linfocitos B/patología , Quimioterapia Combinada , Quinasas MAP Reguladas por Señal Extracelular/inmunología , Femenino , Humanos , Memoria Inmunológica/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/inmunología , Masculino , Persona de Mediana Edad , Esclerosis Múltiple/tratamiento farmacológico , Esclerosis Múltiple/inmunología , Esclerosis Múltiple/patología , Fosforilación/efectos de los fármacos , Fosforilación/inmunologíaRESUMEN
The contribution of autoantibody-producing plasma cells in multiple sclerosis (MS) remains unclear. Anti-CD20 B cell depletion effectively reduces disease activity in MS patients, but it has a minimal effect on circulating autoantibodies and oligoclonal bands in the cerebrospinal fluid. Recently we reported that MEDI551, an anti-CD19 mAb, therapeutically ameliorates experimental autoimmune encephalomyelitis (EAE), the mouse model of MS. MEDI551 potently inhibits pathogenic adaptive immune responses, including depleting autoantibody-producing plasma cells. In the present study, we demonstrated that CD19 mAb treatment ameliorates EAE more effectively than does CD20 mAb. Myelin oligodendrocyte glycoprotein-specific Abs and short-lived and long-lived autoantibody-secreting cells were nearly undetectable in the CD19 mAb-treated mice, but they remained detectable in the CD20 mAb-treated mice. Interestingly, residual disease severity in the CD20 mAb-treated animals positively correlated with the frequency of treatment-resistant plasma cells in the bone marrow. Of note, treatment-resistant plasma cells contained a substantial proportion of CD19(+)CD20(-) plasma cells, which would have otherwise been targeted by CD19 mAb. These data suggested that CD19(+)CD20(-) plasma cells spared by anti-CD20 therapy likely contribute to residual EAE severity by producing autoreactive Abs. In patients with MS, we also identified a population of CD19(+)CD20(-) B cells in the cerebrospinal fluid that would be resistant to CD20 mAb treatment.
Asunto(s)
Anticuerpos Monoclonales/uso terapéutico , Antígenos CD19/inmunología , Antígenos CD20/inmunología , Linfocitos B/inmunología , Encefalomielitis Autoinmune Experimental/inmunología , Células Plasmáticas/inmunología , Adulto , Animales , Anticuerpos Monoclonales/inmunología , Autoanticuerpos/análisis , Líquido Cefalorraquídeo/química , Líquido Cefalorraquídeo/citología , Líquido Cefalorraquídeo/inmunología , Modelos Animales de Enfermedad , Encefalomielitis Autoinmune Experimental/fisiopatología , Encefalomielitis Autoinmune Experimental/terapia , Femenino , Humanos , Masculino , Ratones , Persona de Mediana Edad , Esclerosis Múltiple/inmunología , Glicoproteína Mielina-Oligodendrócito/inmunologíaRESUMEN
BACKGROUND: We previously found that subjects with amnestic mild cognitive impairment exhibit a pro-inflammatory immune profile in the cerebrospinal fluid similar to multiple sclerosis, a central nervous system autoimmune disease. We therefore hypothesized that early neuroinflammation would reflect increases in brain amyloid burden during amnestic mild cognitive impairment. METHODS: Cerebrospinal fluid and blood samples were collected from 24 participants with amnestic mild cognitive impairment (12 men, 12 women; 66 ± 6 years; 0.5 Clinical Dementia Rating) enrolled in the AETMCI study. Analyses of cerebrospinal fluid and blood included immune profiling by multi-parameter flow cytometry, genotyping for apolipoprotein (APO)ε, and quantification of cytokine and immunoglobin levels. Amyloid (A)ß deposition was determined by 18F-florbetapir positron emission tomography. Spearman rank order correlations were performed to assess simple linear correlation for parameters including amyloid imaging, central and peripheral immune cell populations, and protein cytokine levels. RESULTS: Soluble Aß42 in the cerebrospinal fluid declined as Aß deposition increased overall and in the precuneous and posterior cingulate cortices. Lymphocyte profiling revealed a significant decline in T cell populations in the cerebrospinal fluid, specifically CD4+ T cells, as Aß deposition in the posterior cingulate cortex increased. In contrast, increased Aß burden correlated positively with increased memory B cells in the cerebrospinal fluid, which was exacerbated in APOε4 carriers. For peripheral circulating lymphocytes, only B cell populations decreased with Aß deposition in the precuneous cortex, as peripheral T cell populations did not correlate with changes in brain amyloid burden. CONCLUSIONS: Elevations in brain Aß burden associate with a shift from T cells to memory B cells in the cerebrospinal fluid of subjects with amnestic mild cognitive impairment in this exploratory cohort. These data suggest the presence of cellular adaptive immune responses during Aß accumulation, but further study needs to determine whether lymphocyte populations contribute to, or result from, Aß dysregulation during memory decline on a larger cohort collected at multiple centers. TRIAL REGISTRATION: AETMCI NCT01146717.
Asunto(s)
Inmunidad Adaptativa/fisiología , Péptidos beta-Amiloides/metabolismo , Encéfalo/metabolismo , Disfunción Cognitiva , Citocinas/metabolismo , Linfocitos/patología , Anciano , Compuestos de Anilina/metabolismo , Apolipoproteínas E/genética , Encéfalo/diagnóstico por imagen , Disfunción Cognitiva/sangre , Disfunción Cognitiva/líquido cefalorraquídeo , Disfunción Cognitiva/patología , Glicoles de Etileno/metabolismo , Femenino , Citometría de Flujo , Humanos , Masculino , Persona de Mediana Edad , Tomografía de Emisión de PositronesRESUMEN
Plasmablasts are a highly differentiated, antibody secreting B cell subset whose prevalence correlates with disease activity in Multiple Sclerosis (MS). For most patients experiencing partial transverse myelitis (PTM), plasmablasts are elevated in the blood at the first clinical presentation of disease (known as a clinically isolated syndrome or CIS). In this study we found that many of these peripheral plasmablasts are autoreactive and recognize primarily gray matter targets in brain tissue. These plasmablasts express antibodies that over-utilize immunoglobulin heavy chain V-region subgroup 4 (VH4) genes, and the highly mutated VH4+ plasmablast antibodies recognize intracellular antigens of neurons and astrocytes. Most of the autoreactive, highly mutated VH4+ plasmablast antibodies recognize only a portion of cortical neurons, indicating that the response may be specific to neuronal subgroups or layers. Furthermore, CIS-PTM patients with this plasmablast response also exhibit modest reactivity toward neuroantigens in the plasma IgG antibody pool. Taken together, these data indicate that expanded VH4+ peripheral plasmablasts in early MS patients recognize brain gray matter antigens. Peripheral plasmablasts may be participating in the autoimmune response associated with MS, and provide an interesting avenue for investigating the expansion of autoreactive B cells at the time of the first documented clinical event.
Asunto(s)
Autoanticuerpos/metabolismo , Linfocitos B/inmunología , Encéfalo/inmunología , Esclerosis Múltiple/inmunología , Células Plasmáticas/inmunología , Adulto , Anciano , Astrocitos/inmunología , Astrocitos/patología , Linfocitos B/patología , Encéfalo/patología , Encefalomielitis Autoinmune Experimental/inmunología , Encefalomielitis Autoinmune Experimental/patología , Femenino , Sustancia Gris/inmunología , Sustancia Gris/patología , Humanos , Inmunoglobulina G/metabolismo , Masculino , Persona de Mediana Edad , Esclerosis Múltiple/patología , Neuromielitis Óptica/inmunología , Neuromielitis Óptica/patología , Neuronas/inmunología , Neuronas/patología , Células Plasmáticas/fisiología , Accidente Cerebrovascular/inmunología , Accidente Cerebrovascular/patología , Adulto JovenRESUMEN
Plasma cells and the autoreactive Abs they produce are suspected to contribute to the pathogenesis of multiple sclerosis, but recent attempts to target these components of humoral immunity have failed. MEDI551, an anti-CD19 Ab that depletes mature B cells including plasma cells may offer a compelling alternative that reduces pathogenic adaptive immune responses while sparing regulatory mechanisms. Indeed, our data demonstrate that a single dose of MEDI551, given before or during ongoing experimental autoimmune encephalomyelitis, disrupts development of the disease. Leukocyte infiltration into the spinal cord is significantly reduced, as well as short-lived and long-lived autoreactive CD138(+) plasma cells in the spleen and bone marrow, respectively. In addition, potentially protective CD1d(hi)CD5(+) regulatory B cells show resistance to depletion, and myelin-specific Foxp3(+) regulatory T cells are expanded. Taken together, these results demonstrate that MEDI551 disrupts experimental autoimmune encephalomyelitis by inhibiting multiple proinflammatory components whereas preserving regulatory populations.
Asunto(s)
Inmunidad Adaptativa/efectos de los fármacos , Anticuerpos/farmacología , Antígenos CD19/inmunología , Médula Ósea/inmunología , Encefalomielitis Autoinmune Experimental/prevención & control , Médula Espinal/inmunología , Animales , Antígenos CD19/genética , Antígenos CD1d/genética , Antígenos CD1d/inmunología , Linfocitos B Reguladores/inmunología , Linfocitos B Reguladores/patología , Médula Ósea/patología , Supervivencia Celular , Encefalomielitis Autoinmune Experimental/inducido químicamente , Encefalomielitis Autoinmune Experimental/genética , Encefalomielitis Autoinmune Experimental/inmunología , Factores de Transcripción Forkhead/genética , Factores de Transcripción Forkhead/inmunología , Expresión Génica , Depleción Linfocítica , Masculino , Ratones , Ratones Transgénicos , Glicoproteína Mielina-Oligodendrócito , Células Plasmáticas/inmunología , Células Plasmáticas/patología , Ingeniería de Proteínas , Transducción de Señal , Médula Espinal/patología , Bazo/inmunología , Bazo/patología , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/patologíaRESUMEN
The cytokines IL-6 and IL-10 are produced by cells of the adaptive and innate arms of the immune system and they appear to play key roles in genetically diverse autoimmune diseases such as relapsing remitting multiple sclerosis (MS), rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). Whereas previous intense investigations focused on the generation of autoantibodies and their contribution to immune-mediated pathogenesis in these diseases; more recent attention has focused on the roles of cytokines such as IL-6 and IL-10. In response to pathogens, antigen presenting cells (APC), including B cells, produce IL-6 and IL-10 in order to up-or down-regulate immune cell activation and effector responses. Evidence of elevated levels of the proinflammatory cytokine IL-6 has been routinely observed during inflammatory responses and in a number of autoimmune diseases. Our recent studies suggest that MS peripheral blood B cells secrete higher quantities of IL-6 and less IL-10 than B cells from healthy controls. Persistent production of IL-6, in turn, contributes to T cell expansion and the functional hyperactivity of APC such as MS B cells. Altered B cell activity can have a profound impact on resultant T cell effector functions. Enhanced signaling through the IL-6 receptor can effectively inhibit cytolytic activity, induce T cell resistance to IL-10-mediated immunosuppression and increase skewing of autoreactive T cells to a pathogenic Th17 phenotype. Our recent findings and studies by others support a role for the indirect attenuation of B cell responses by Glatiramer acetate (GA) therapy. Our studies suggest that GA therapy temporarily permits homeostatic regulatory mechanisms to be reinstated. Future studies of mechanisms underlying dysregulated B cell cytokine production could lead to the identification of novel targets for improved immunoregulatory therapies for autoimmune diseases.
Asunto(s)
Autoinmunidad , Interleucina-10/metabolismo , Interleucina-6/metabolismo , Esclerosis Múltiple/inmunología , Animales , Modelos Animales de Enfermedad , Humanos , Inmunomodulación , Esclerosis Múltiple/genética , Esclerosis Múltiple/terapiaRESUMEN
BACKGROUND: Repetitive hypoxic preconditioning (RHP) creates an anti-inflammatory phenotype that protects from stroke-induced injury for months after a 2-week treatment. The mechanisms underlying long-term tolerance are unknown, though one exposure to hypoxia significantly increased peripheral B cell representation. For this study, we sought to determine if RHP specifically recruited B cells into the protected ischemic hemisphere, and whether RHP could phenotypically alter B cells prior to stroke onset. METHODS: Adult, male SW/ND4 mice received RHP (nine exposures over 2 weeks; 8 to 11 % O2; 2 to 4 hours) or identical exposures to 21 % O2 as control. Two weeks following RHP, a 60-minute transient middle cerebral artery occlusion was induced. Standard techniques quantified CXCL13 mRNA and protein expression. Two days after stroke, leukocytes were isolated from brain tissue (70:30 discontinuous Percoll gradient) and profiled on a BD-FACS Aria flow cytometer. In a separate cohort without stroke, sorted splenic CD19+ B cells were isolated 2 weeks after RHP and analyzed on an Illumina MouseWG-6 V2 Bead Chip. Final gene pathways were determined using Ingenuity Pathway Analysis. Student's t-test or one-way analysis of variance determined significance (P < 0.05). RESULTS: CXCL13, a B cell-specific chemokine, was upregulated in post-stroke cortical vessels of both groups. In the ischemic hemisphere, RHP increased B cell representation by attenuating the diapedesis of monocyte, macrophage, neutrophil and T cells, to quantities indistinguishable from the uninjured, contralateral hemisphere. Pre-stroke splenic B cells isolated from RHP-treated mice had >1,900 genes differentially expressed by microarray analysis. Genes related to B-T cell interactions, including antigen presentation, B cell differentiation and antibody production, were profoundly downregulated. Maturation and activation were arrested in a cohort of B cells from pre-stroke RHP-treated mice while regulatory B cells, a subset implicated in neurovascular protection from stroke, were upregulated. CONCLUSIONS: Collectively, our data characterize an endogenous neuroprotective phenotype that utilizes adaptive immune mechanisms pre-stroke to protect the brain from injury post-stroke. Future studies to validate the role of B cells in minimizing injury and promoting central nervous system recovery, and to determine whether B cells mediate an adaptive immunity to systemic hypoxia that protects from subsequent stroke, are needed.
Asunto(s)
Linfocitos B/metabolismo , Terapia de Inmunosupresión , Infarto de la Arteria Cerebral Media/complicaciones , Precondicionamiento Isquémico , Animales , Antígenos CD/metabolismo , Linfocitos B/patología , Proliferación Celular , Quimiocina CXCL1/metabolismo , Modelos Animales de Enfermedad , Endotelio/metabolismo , Endotelio/patología , Citometría de Flujo , Lateralidad Funcional , Regulación de la Expresión Génica/fisiología , Masculino , Ratones , Análisis por Micromatrices , Fosfopiruvato Hidratasa/metabolismo , Factores de TiempoRESUMEN
Progressive multifocal leukoencephalopathy (PML) induced by JC virus (JCV) is a risk for natalizumab-treated multiple sclerosis (MS) patients. Here we characterize the JCV-specific T cell responses in healthy donors and natalizumab-treated MS patients to reveal functional differences that may account for the development of natalizumab-associated PML. CD4 and CD8 T cell responses specific for all JCV proteins were readily identified in MS patients and healthy volunteers. The magnitude and quality of responses to JCV and cytomegalovirus (CMV) did not change from baseline through several months of natalizumab therapy. However, the frequency of T cells producing IL-10 upon mitogenic stimulation transiently increased after the first dose. In addition, MS patients with natalizumab-associated PML were distinguished from all other subjects in that they either had no detectable JCV-specific T cell response or had JCV-specific CD4 T cell responses uniquely dominated by IL-10 production. Additionally, IL-10 levels were higher in the CSF of individuals with recently diagnosed PML. Thus, natalizumab-treated MS patients with PML have absent or aberrant JCV-specific T cell responses compared with non-PML patients, and changes in T cell-mediated control of JCV replication may be a risk factor for developing PML. Our data suggest further approaches to improved monitoring, treatment and prevention of PML in natalizumab-treated patients.
Asunto(s)
Anticuerpos Monoclonales Humanizados/administración & dosificación , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Inmunidad Celular/efectos de los fármacos , Virus JC/inmunología , Leucoencefalopatía Multifocal Progresiva/inmunología , Esclerosis Múltiple/inmunología , Anticuerpos Monoclonales Humanizados/efectos adversos , Linfocitos T CD4-Positivos/patología , Linfocitos T CD8-positivos/patología , Citomegalovirus/inmunología , Infecciones por Citomegalovirus/tratamiento farmacológico , Infecciones por Citomegalovirus/inmunología , Infecciones por Citomegalovirus/patología , Femenino , Humanos , Interleucina-10/inmunología , Leucoencefalopatía Multifocal Progresiva/tratamiento farmacológico , Leucoencefalopatía Multifocal Progresiva/patología , Leucoencefalopatía Multifocal Progresiva/virología , Masculino , Esclerosis Múltiple/tratamiento farmacológico , Esclerosis Múltiple/patología , Esclerosis Múltiple/virología , Natalizumab , Factores de RiesgoRESUMEN
BACKGROUND AND OBJECTIVES: Patients with Alzheimer dementia display evidence of amyloid-related neurodegeneration. Our focus was to determine whether such patients also display evidence of a disease-targeting adaptive immune response mediated by CD4+ T cells. To test this hypothesis, we evaluated the CSF immune profiles of patients with Alzheimer clinical syndrome (ACS), who display clinically defined dementia. METHODS: Innate and adaptive immune profiles of patients with ACS were measured using multicolor flow cytometry. CSF-derived CD4+ and CD8+ T-cell receptor repertoire genetics were measured using next-generation sequencing. Brain-specific autoantibody signatures of CSF-derived antibody pools were measured using array technology or ELISA. CSF from similar-age healthy controls (HCs) was used as a comparator cohort. RESULTS: Innate cells were expanded in the CSF of patients with ACS in comparison to HCs, and innate cell expansion increased with age in the patients with ACS, but not HCs. Despite innate cell expansion in the CSF, the frequency of total CD4+ T cells reduced with age in the patients with ACS. T-cell receptor repertoire genetics indicated that T-cell clonal expansion is enhanced, and diversity is reduced in the patients with ACS compared with similar-age HCs. DISCUSSION: Examination of CSF indicates that CD4+ T cell-mediated adaptive immune responses are altered in patients with ACS. Understanding the underlying mechanisms affecting adaptive immunity will help move us toward the goal of slowing cognitive decline.
Asunto(s)
Inmunidad Adaptativa/inmunología , Enfermedad de Alzheimer/líquido cefalorraquídeo , Enfermedad de Alzheimer/inmunología , Autoanticuerpos/líquido cefalorraquídeo , Linfocitos T CD4-Positivos/metabolismo , Inmunidad Innata/inmunología , Anciano , Linfocitos T CD8-positivos/metabolismo , Estudios de Cohortes , Femenino , Humanos , Masculino , Persona de Mediana Edad , SíndromeRESUMEN
Recent evidence suggests that B- and T-cell interactions may be paramount in relapsing-remitting MS (RRMS) disease pathogenesis. We hypothesized that memory B-cell pools from RRMS patients may specifically harbor a subset of potent neuro-APC that support neuro-Ag reactive T-cell proliferation and cytokine secretion. To test this hypothesis, we compared CD80 and HLA-DR expression, IL-10 and lymphotoxin-α secretion, neuro-Ag binding capacity, and neuro-Ag presentation by memory B cells from RRMS patients to naïve B cells from RRMS patients and to memory and naïve B cells from healthy donors (HD). We identified memory B cells from some RRMS patients that elicited CD4(+) T-cell proliferation and IFN-γ secretion in response to myelin basic protein and myelin oligodendrocyte glycoprotein. Notwithstanding the fact that the phenotypic parameters that promote efficient Ag presentation were observed to be similar between RRMS and HD memory B cells, a corresponding capability to elicit CD4(+) T-cell proliferation in response to myelin basic protein and myelin oligodendrocyte glycoprotein was not observed in HD memory B cells. Our results demonstrate for the first time that the memory B-cell pool in RRMS harbors neuro-Ag specific B cells that can activate T cells.
Asunto(s)
Linfocitos B/inmunología , Linfocitos T CD4-Positivos/inmunología , Memoria Inmunológica/inmunología , Interferón gamma/biosíntesis , Proteína Básica de Mielina/inmunología , Glicoproteína Asociada a Mielina/inmunología , Adulto , Estudios de Cohortes , Femenino , Citometría de Flujo , Humanos , Inmunofenotipificación , Interferón gamma/sangre , Interferón gamma/inmunología , Activación de Linfocitos , Linfotoxina-alfa/inmunología , Masculino , Persona de Mediana Edad , Esclerosis Múltiple Recurrente-Remitente/inmunología , Proteínas de la Mielina , Glicoproteína Mielina-Oligodendrócito , Adulto JovenRESUMEN
Mechanically ventilated surgical patients have a variety of bacterial flora that are often undetectable by traditional culture methods. The source of infection in many of these patients remains unclear. To address this clinical problem, the microbiome profile and host inflammatory response in bronchoalveolar lavage samples from the surgical intensive care unit were examined relative to clinical pathology diagnoses. The hypothesis was tested that clinical diagnosis of respiratory tract flora were similar to culture positive lavage samples in both microbiome and inflammatory profile. Bronchoalveolar lavage samples were collected in the surgical intensive care unit as standard of care for intubated individuals with a clinical pulmonary infection score of >6 or who were expected to be intubated for >48 hours. Cytokine analysis was conducted with the Bioplex Pro Human Th17 cytokine panel. The microbiome of the samples was sequenced for the 16S rRNA region using the Ion Torrent. Microbiome diversity analysis showed the culture-positive samples had the lowest levels of diversity and culture negative with the highest based upon the Shannon-Wiener index (culture positive: 0.77 ± 0.36, respiratory tract flora: 2.06 ± 0.73, culture negative: 3.97 ± 0.65). Culture-negative samples were not dominated by a single bacterial genera. Lavages classified as respiratory tract flora were more similar to the culture-positive in the microbiome profile. A comparison of cytokine expression between groups showed increased levels of cytokines (IFN-g, IL-17F, IL-1B, IL-31, TNF-a) in culture-positive and respiratory tract flora groups. Culture-positive samples exhibited a more robust immune response and reduced diversity of bacterial genera. Lower cytokine levels in culture-negative samples, despite a greater number of bacterial species, suggest a resident nonpathogenic bacterial community may be indicative of a normal pulmonary environment. Respiratory tract flora samples were most similar to the culture-positive samples and may warrant classification as culture-positive when considering clinical treatment.
Asunto(s)
Bacterias/inmunología , Pulmón/microbiología , Microbiota/inmunología , Neumonía Asociada al Ventilador/inmunología , Respiración Artificial/efectos adversos , Adulto , Anciano , Bacterias/genética , Bacterias/aislamiento & purificación , Líquido del Lavado Bronquioalveolar/microbiología , Citocinas/inmunología , Citocinas/metabolismo , ADN Bacteriano/aislamiento & purificación , Femenino , Humanos , Unidades de Cuidados Intensivos , Pulmón/inmunología , Masculino , Persona de Mediana Edad , Neumonía Asociada al Ventilador/microbiología , ARN Ribosómico 16S/genética , Respiración Artificial/métodosRESUMEN
B cells are highly potent antigen presenting cells of their cognate antigens. However, it remains unknown whether B cells can orchestrate Th17-mediated responses against neuro-antigens. We report that MS patients and healthy donors had a similar frequency of antigen-specific Th1 and Th17 cells, and distribution of T effector and T central memory cells. Notwithstanding these similarities, the application of an in vitro assay demonstrated that the B cells derived from a subset of MS patients exhibited the capability of coordinating Th17 responses directed toward neuro-antigens. These observations underscore the B cell's contribution to the putative underpinnings of multiple sclerosis.
Asunto(s)
Linfocitos B/fisiología , Esclerosis Múltiple Recurrente-Remitente/patología , Proteína Básica de Mielina/farmacología , Glicoproteína Mielina-Oligodendrócito/inmunología , Células Th17/metabolismo , Antígenos CD , Linfocitos B/efectos de los fármacos , Linfocitos B/inmunología , Proliferación Celular/efectos de los fármacos , Proliferación Celular/fisiología , Células Cultivadas , Técnicas de Cocultivo , Citocinas/metabolismo , Femenino , Citometría de Flujo , Humanos , Activación de Linfocitos , Masculino , Proteína Básica de Mielina/inmunología , Glicoproteína Mielina-Oligodendrócito/farmacología , Células Th17/efectos de los fármacosRESUMEN
Alzheimer's disease (AD) is a progressive, neurodegenerative disease that may involve inflammatory responses in the central nervous system (CNS). Our objective was to determine whether patients with amnestic mild cognitive impairment (aMCI), a preclinical stage of AD, have inflammatory characteristics similar to patients with multiple sclerosis (MS), a known CNS inflammatory disease. The frequency of lymphocytes and levels of pro-inflammatory cytokines in the cerebrospinal fluid of aMCI patients was comparable to MS patients or patients at high risk to develop MS. Thus, brain inflammation occurs early at the preclinical stage of AD and may have an important role in pathology.
Asunto(s)
Enfermedad de Alzheimer/inmunología , Disfunción Cognitiva/inmunología , Citocinas/líquido cefalorraquídeo , Enfermedades Desmielinizantes/inmunología , Esclerosis Múltiple/inmunología , Anciano , Enfermedad de Alzheimer/líquido cefalorraquídeo , Estudios de Casos y Controles , Linaje de la Célula , Disfunción Cognitiva/líquido cefalorraquídeo , Citocinas/inmunología , Enfermedades Desmielinizantes/líquido cefalorraquídeo , Humanos , Linfocitos/citología , Linfocitos/inmunología , Persona de Mediana Edad , Esclerosis Múltiple/líquido cefalorraquídeoRESUMEN
IMPORTANCE: This study describes what is, to our knowledge, the previously unknown effect of glatiramer acetate therapy on B cells in patients with relapsing-remitting multiple sclerosis (MS). OBJECTIVE: To determine whether glatiramer acetate therapy normalizes dysregulated B-cell proliferation and cytokine production in patients with MS. DESIGN, SETTING, AND PARTICIPANTS: Twenty-two patients with MS who were receiving glatiramer acetate therapy and 22 treatment-naive patients with MS were recruited at The University of Texas Southwestern Medical Center MS clinic. Cell samples from healthy donors were obtained from HemaCare (Van Nuys, California) or Carter Blood Bank (Dallas, Texas). Treatment-naive patients with MS had not received any disease-modifying therapies for at least 3 months before the study. EXPOSURES: Glatiramer acetate therapy for at least 3 months at the time of the study. MAIN OUTCOMES AND MEASURES: B-cell phenotype and proliferation and immunoglobulin and cytokine secretion. RESULTS: A restoration of interleukin 10 production by peripheral B cells was observed in patients undergoing glatiramer acetate therapy as well as a significant reduction of interleukin 6 production in a subset of patients who received therapy for less than 32 months. Furthermore, proliferation in response to high-dose CD40L was altered and immunoglobulin production was elevated in in vitro-activated B cells obtained from patients who received glatiramer acetate. CONCLUSIONS AND RELEVANCE: Glatiramer acetate therapy remodels the composition of the B-cell compartment and influences cytokine secretion and immunoglobulin production. These data suggest that glatiramer acetate therapy affects several aspects of dysregulated B-cell function in MS that may contribute to the therapeutic mechanisms of glatiramer acetate.
Asunto(s)
Linfocitos B/efectos de los fármacos , Inmunosupresores/uso terapéutico , Esclerosis Múltiple Recurrente-Remitente/tratamiento farmacológico , Péptidos/uso terapéutico , Adulto , Citocinas/efectos de los fármacos , Femenino , Acetato de Glatiramer , Humanos , Masculino , Persona de Mediana Edad , Resultado del Tratamiento , Adulto JovenRESUMEN
The pathogenic role for B cells in the context of relapsing remitting multiple sclerosis (MS) is incompletely defined. Although classically considered a T cell-mediated disease, B cell-depleting therapies showed efficacy in treating the clinical symptoms of RRMS without decreasing plasma cells or total immunoglobulin (Ig) levels. Here, we discuss the potential implications of antibody-independent B cell effector functions that could contribute to autoimmunity with particular focus on antigen presentation, cytokine secretion, and stimulation of T cell subsets. We highlight differences between memory and naïve B cells from MS patients such as our recent findings of hyper-proliferation from MS memory B cells in response to CD40 engagement. We discuss the implications of IL6 overproduction in contrast to limited IL10 production by B cells from MS patients and comment on the impact of these functions on yet unexplored aspects of B cells in autoimmune disease. Finally, we contextualize B cell effector functions with respect to current immunomodulatory therapies for MS and show that glatiramer acetate (GA) does not directly modulate B cell proliferation or cytokine secretion.
Asunto(s)
Anticuerpos/inmunología , Subgrupos de Linfocitos B/inmunología , Esclerosis Múltiple Recurrente-Remitente/inmunología , Linfocitos B Reguladores/inmunología , Citocinas/biosíntesis , Citocinas/inmunología , Humanos , Memoria Inmunológica , Inflamación/inmunología , Esclerosis Múltiple Recurrente-Remitente/metabolismoRESUMEN
Relative positions of neurons within mature murine pelvic ganglia based on expression of neurotransmitters have been described. However the spatial organization of developing innervation in the murine urogenital tract (UGT) and the gene networks that regulate specification and maturation of neurons within the pelvic ganglia of the lower urinary tract (LUT) are unknown. We used whole-mount immunohistochemistry and histochemical stains to localize neural elements in 15.5 days post coitus (dpc) fetal mice. To identify potential regulatory factors expressed in pelvic ganglia, we surveyed expression patterns for known or probable transcription factors (TF) annotated in the mouse genome by screening a whole-mount in situ hybridization library of fetal UGTs. Of the 155 genes detected in pelvic ganglia, 88 encode TFs based on the presence of predicted DNA-binding domains. Neural crest (NC)-derived progenitors within the LUT were labeled by Sox10, a well-known regulator of NC development. Genes identified were categorized based on patterns of restricted expression in pelvic ganglia, pelvic ganglia and urethral epithelium, or pelvic ganglia and urethral mesenchyme. Gene expression patterns and the distribution of Sox10+, Phox2b+, Hu+, and PGP9.5+ cells within developing ganglia suggest previously unrecognized regional segregation of Sox10+ progenitors and differentiating neurons in early development of pelvic ganglia. Reverse transcription-PCR of pelvic ganglia RNA from fetal and post-natal stages demonstrated that multiple TFs maintain post-natal expression, although Pax3 is extinguished before weaning. Our analysis identifies multiple potential regulatory genes including TFs that may participate in segregation of discrete lineages within pelvic ganglia. The genes identified here are attractive candidate disease genes that may now be further investigated for their roles in malformation syndromes or in LUT dysfunction.
RESUMEN
Multiple sclerosis is a chronic debilitating autoimmune disease of the central nervous system. The contribution of B cells in the pathoetiology of MS has recently been highlighted by the emergence of rituximab, an anti-CD20 monoclonal antibody that specifically depletes B cells, as a potent immunomodulatory therapy for the treatment of MS. However, a clearer understanding of the impact B cells have on the neuro-inflammatory component of MS pathogenesis is needed in order to develop novel therapeutics whose affects on B cells would be beneficial and not harmful. Since T cells are known mediators of the pathology of MS, the goal of this review is to summarize what is known about the interactions between B cells and T cells, and how current and emerging immunotherapies may impact B-T cell interactions in MS.