RESUMEN
Mesothelin (MSLN) is a cell surface glycoprotein overexpressed in several solid malignancies, including gastric, lung, mesothelioma, pancreatic and ovarian cancers. While several MSLN-targeting therapeutic approaches are in development, only limited efficacy has been achieved in patients. A potential shortcoming of several described antibody-based approaches is that they target the membrane distal region of MSLN and, additionally, are known to be handicapped by the high levels of circulating soluble MSLN in patients. We show here, using monoclonal antibodies (mAbs) targeting different MSLN-spanning epitopes, that the membrane-proximal region resulted in more efficient killing of MSLN-positive tumor cells in antibody-dependent cell-mediated cytotoxicity (ADCC) assays. Surprisingly, no augmented killing was observed in antibody-dependent cellular phagocytosis (ADCP) by mAbs targeting this membrane-proximal region. To further increase the ADCP potential, we, therefore, generated bispecific antibodies (bsAbs) coupling a high-affinity MSLN binding arm to a blocking CD47 arm. Here, targeting the membrane-proximal domain of MSLN demonstrated enhanced ADCP activity compared to membrane-distal domains when the bsAbs were used in in vitro phagocytosis killing assays. Importantly, the superior anti-tumor activity was also translated in xenograft tumor models. Furthermore, we show that the bsAb approach targeting the membrane-proximal epitope of MSLN optimized ADCC activity by augmenting FcγR-IIIA activation and enhanced ADCP via a more efficient blockade of the CD47/SIRPα axis.
Asunto(s)
Anticuerpos Biespecíficos/inmunología , Anticuerpos Monoclonales/inmunología , Antineoplásicos/inmunología , Antígeno CD47/inmunología , Proteínas Ligadas a GPI/inmunología , Animales , Anticuerpos Biespecíficos/farmacología , Anticuerpos Monoclonales/farmacología , Citotoxicidad Celular Dependiente de Anticuerpos/inmunología , Antineoplásicos/farmacología , Epítopos/inmunología , Humanos , Inmunoterapia/métodos , Mesotelina , Ratones , Fagocitosis/inmunología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
CD19 is a B cell-specific receptor that regulates the threshold of B cell receptor (BCR)-mediated cell proliferation. A CD47xCD19 bispecific antibody (biAb) was generated to target and deplete B cells via multiple antibody-mediated mechanisms. Interestingly, the biAb, constructed of a CD19 binding arm and a CD47 binding arm, inhibited BCR-mediated B-cell proliferation with an effect even more potent than a CD19 monoclonal antibody (mAb). The inhibitory effect of the biAb was not attributable to CD47 binding because a monovalent or bivalent anti-CD47 mAb had no effect on B cell proliferation. Fluorescence resonance energy transfer analysis demonstrated that co-engaging CD19 and CD47 prevented CD19 clustering and its migration to BCR clusters, while only engaging CD19 (with a mAb) showed no impact on either CD19 clustering or migration. The lack of association between CD19 and the BCR resulted in decreased phosphorylation of CD19 upon BCR activation. Furthermore, the biAb differentially modulated BCR-induced gene expression compared to a CD19 mAb. Taken together, this unexpected role of CD47xCD19 co-ligation in inhibiting B cell proliferation illuminates a novel approach in which two B cell surface molecules can be tethered, to one another in order, which may provide a therapeutic benefit in settings of autoimmunity and B cell malignancies.
Asunto(s)
Anticuerpos Biespecíficos/farmacología , Antígenos CD19/metabolismo , Linfocitos B/metabolismo , Antígeno CD47/metabolismo , Receptores de Antígenos de Linfocitos B/metabolismo , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Humanos , Activación de Linfocitos/efectos de los fármacosRESUMEN
CD47, an ubiquitously expressed innate immune checkpoint receptor that serves as a universal "don't eat me" signal of phagocytosis, is often upregulated by hematologic and solid cancers to evade immune surveillance. Development of CD47-targeted modalities is hindered by the ubiquitous expression of the target, often leading to rapid drug elimination and hemotoxicity including anemia. To overcome such liabilities, we have developed a fully human bispecific antibody, NI-1701, designed to coengage CD47 and CD19 selectively on B cells. NI-1701 demonstrates favorable elimination kinetics with no deleterious effects seen on hematologic parameters following single or multiple administrations to nonhuman primates. Potent in vitro and in vivo activity is induced by NI-1701 to kill cancer cells across a plethora of B-cell malignancies and control tumor growth in xenograft mouse models. The mechanism affording maximal tumor growth inhibition by NI-1701 is dependent on the coengagement of CD47/CD19 on B cells inducing potent antibody-dependent cellular phagocytosis of the targeted cells. NI-1701-induced control of tumor growth in immunodeficient NOD/SCID mice was more effective than that achieved with the anti-CD20 targeted antibody, rituximab. Interestingly, a synergistic effect was seen when tumor-implanted mice were coadministered NI-1701 and rituximab leading to significantly improved tumor growth inhibition and regression in some animals. We describe herein, a novel bispecific antibody approach aimed at sensitizing B cells to become more readily phagocytosed and eliminated thus offering an alternative or adjunct therapeutic option to patients with B-cell malignancies refractory/resistant to anti-CD20-targeted therapy. Mol Cancer Ther; 17(8); 1739-51. ©2018 AACR.
Asunto(s)
Anticuerpos Biespecíficos/genética , Leucemia/genética , Leucemia/terapia , Linfoma de Células B/genética , Linfoma de Células B/terapia , Animales , Antígenos CD19 , Antígeno CD47 , Humanos , Leucemia/patología , Linfoma de Células B/patología , Ratones , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: Increased expression of toll-like receptor 4 (TLR4) and its endogenous ligands, is characteristic of rheumatoid arthritis (RA) synovitis. In this study, we evaluated how these TLR4 ligands may drive pathogenic processes and whether the fine profiling of anti-citrullinated protein antibodies (ACPA) based on their target specificity might provide a simple means to predict therapeutic benefit when neutralizing TLR4 in this disease. METHODS: The capacity of RA synovial fluids (RASF) to stimulate cytokine production in monocytes from patients with RA was analyzed by ELISA. The presence of TLR4 activators in RASF was determined by measuring the levels of ACPA, ACPA subtypes with reactivity to specific citrullinated peptides and other TLR4 ligands. Neutralization of TLR4 signaling was investigated using NI-0101, a therapeutic antibody that targets TLR4. RESULTS: RASF exhibited a heterogeneous capacity to induce production of proinflammatory cytokines by monocytes isolated from patients with RA. Such cytokine responses were significantly modified by TLR4 blockade achieved using NI-0101. The analysis of the content of RASF and matched sera demonstrated that ACPA fine specificities in patient samples predict cellular response to anti-TLR4 exposure in vitro. CONCLUSION: TLR4 represents a possible therapeutic target in RA. Our study demonstrates that TLR4 inhibition in an ex vivo model of RA pathogenesis can significantly modulate cytokine release and does so in specific subgroups of RA patient-derived samples. It also suggests that ACPA fine profiling has the potential to identify RA patients with a predominantly TLR4-driven pathotype that could be used to predict preferential response to TLR4 antagonism.
Asunto(s)
Artritis Reumatoide/inmunología , Autoanticuerpos/inmunología , Líquido Sinovial/inmunología , Receptor Toll-Like 4/inmunología , Anciano , Anticuerpos Monoclonales Humanizados/farmacología , Autoantígenos/inmunología , Citocinas/inmunología , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Masculino , Persona de Mediana Edad , Péptidos Cíclicos/inmunología , Receptor Toll-Like 4/antagonistas & inhibidoresRESUMEN
Increased expression of endogenous Toll-like receptor 4 (TLR4) ligands (e.g., Tenascin-C, S100A8/A9, citrullinated fibrinogen (cFb) immune complexes) has been observed in patients with rheumatoid arthritis (RA). However, their roles in RA pathogenesis are not well understood. Here, we investigated the expression kinetics and role of endogenous TLR4 ligands in the murine model of collagen-induced arthritis (CIA). Tenascin-C was upregulated in blood early in CIA, and correlated positively with the clinical score at day 56. Levels of S100A8/A9 increased starting from day 28, peaking at day 42, and correlated positively with joint inflammation. Levels of anti-cFb antibodies increased during the late phase of CIA and correlated positively with both joint inflammation and cartilage damage. Blockade of TLR4 activation at the time of the first TLR4 ligand upregulation prevented clinical and histological signs of arthritis. A TLR4-dependent role was also observed for Tenascin-C and cFb immune complexes in osteoclast differentiation in vitro. Taken together, our data suggests that the pathogenic contribution of TLR4 in promoting joint inflammation and bone erosion during CIA occurs via various TLR4 ligands arising at different stages of disease. The data also suggests that Blockade of TLR4 with monoclonal antibodies is a promising strategy in RA treatment.
Asunto(s)
Artritis Reumatoide/inmunología , Huesos/patología , Calgranulina A/sangre , Tenascina/sangre , Receptor Toll-Like 4/metabolismo , Animales , Complejo Antígeno-Anticuerpo/sangre , Artritis Experimental/inmunología , Artritis Reumatoide/patología , Artritis Reumatoide/fisiopatología , Calgranulina A/genética , Diferenciación Celular , Colágeno , Modelos Animales de Enfermedad , Fibrinógeno/inmunología , Articulaciones/inmunología , Ligandos , Ratones , Ratones Endogámicos DBA , Osteoclastos/metabolismo , Análisis Espacio-Temporal , Tenascina/genética , Receptor Toll-Like 4/genética , Receptor Toll-Like 4/inmunologíaRESUMEN
B cells play a major role in the pathogenesis of many autoimmune disorders, including rheumatoid arthritis, systemic lupus erythematosus, multiple sclerosis, and type I diabetes mellitus, as indicated by the efficacy of B cell-targeted therapies in these diseases. Therapeutic effects of the most commonly used B cell-targeted therapy, anti-CD20 mAb, are contingent upon long-term depletion of peripheral B cells. In this article, we describe an alternative approach involving the targeting of CD79, the transducer subunit of the B cell AgR. Unlike anti-CD20 mAbs, the protective effects of CD79-targeted mAbs do not require cell depletion; rather, they act by inducing an anergic-like state. Thus, we describe a novel B cell-targeted approach predicated on the induction of B cell anergy.
Asunto(s)
Enfermedades Autoinmunes/prevención & control , Linfocitos B/inmunología , Antígenos CD79/inmunología , Anergia Clonal/inmunología , Animales , Anticuerpos Monoclonales/inmunología , Autoinmunidad/inmunología , Femenino , Activación de Linfocitos/inmunología , Recuento de Linfocitos , Depleción Linfocítica , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Ratones NoqueadosRESUMEN
Fc-modified anti-human CD3ε monoclonal antibodies (mAbs) are in clinical development for the treatment of autoimmune diseases. These next generation mAbs have completed clinical trials in patients with type-1 diabetes and inflammatory bowel disease demonstrating a narrow therapeutic window. Lowered doses are ineffective, yet higher pharmacologically-active doses cause an undesirable level of adverse events. Thus, there is a critical need for a return to bench research to explore ways of improving clinical outcomes. Indeed, we recently reported that a short course of treatment affords synergy, providing long-term disease amelioration when combining anti-mouse CD3 and anti-mouse tumor necrosis factor mAbs in experimental arthritis. Such strategies may widen the window between risk and benefit; however, to more accurately assess experimentally the biology and pharmacology, reagents that mimic the current development candidates were required. Consequently, we engineered an Fc-modified anti-mouse CD3ε mAb, 2C11-Novi. Here, we report the functional characterization of 2C11-Novi demonstrating that it does not bind FcγR in vitro and elicits little cytokine release in vivo, while maintaining classical pharmacodynamic effects (CD3-TCR downregulation and T cell killing). Furthermore, we observed that oral administration of 2C11-Novi ameliorated progression of remitting-relapsing experimental autoimmune encephalitis in mice, significantly reducing the primary acute and subsequent relapse phase of the disease. With innovative approaches validated in two experimental models of human disease, 2C11-Novi represents a meaningful tool to conduct further mechanistic studies aiming at exploiting the immunoregulatory properties of Fc-modified anti-CD3 therapies via combination therapy using parenteral or oral routes of administration.
Asunto(s)
Anticuerpos Monoclonales de Origen Murino , Artritis Experimental , Complejo CD3/inmunología , Encefalomielitis Autoinmune Experimental , Ingeniería de Proteínas , Animales , Anticuerpos Monoclonales de Origen Murino/genética , Anticuerpos Monoclonales de Origen Murino/inmunología , Anticuerpos Monoclonales de Origen Murino/farmacología , Artritis Experimental/tratamiento farmacológico , Artritis Experimental/inmunología , Encefalomielitis Autoinmune Experimental/tratamiento farmacológico , Encefalomielitis Autoinmune Experimental/inmunología , Femenino , Humanos , Masculino , Ratones , Receptores de IgG/inmunologíaRESUMEN
OBJECTIVE: The goal of rheumatoid arthritis (RA) treatment is to achieve clinical remission in order to limit structural damage and physical disability. To this end, recent emphasis has been placed on aggressive treatment early in the course of disease with drugs such as anti-tumor necrosis factor (anti-TNF) agents. As T cells are also thought to play an important role in the initiation of RA, we hypothesized that targeting both TNF and T cells would result in better outcomes. The aim of this study was to examine the efficacy of combined therapy with anti-CD3 and anti-TNF in experimental RA. METHODS: Two anti-mouse antibodies were developed as surrogate reagents for anti-TNF and anti-CD3 therapies. Collagen-induced arthritis (CIA) was induced in DBA/1 mice, and antibodies were injected intraperitoneally, either alone on in combination, at predetermined subtherapeutic doses. The frequency and number of pathogenic and regulatory CD4+ T cell subsets in the draining lymph nodes were determined in order to investigate the mechanisms of action. RESULTS: Strikingly, the combination of the two antibodies demonstrated a potent synergy in established CIA, with long-term inhibition of disease progression and protection from joint destruction. The results did not demonstrate any enhancement of CD25+FoxP3+ regulatory T cells, but a profound depletion of pathogenic T cells from the draining lymph nodes was associated with reduced numbers of T cells in the joints. CONCLUSION: A short course of combination therapy with anti-CD3 and anti-TNF efficiently depletes pathogenic T cells from the draining lymph nodes, reducing the numbers of T cells in the joints and affording long-lasting inhibition of established CIA.
Asunto(s)
Anticuerpos Monoclonales/uso terapéutico , Artritis Experimental/tratamiento farmacológico , Complejo CD3/inmunología , Linfocitos T CD4-Positivos/efectos de los fármacos , Factor de Necrosis Tumoral alfa/inmunología , Animales , Anticuerpos Monoclonales/farmacología , Artritis Experimental/inmunología , Artritis Experimental/patología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/patología , Articulaciones/efectos de los fármacos , Articulaciones/patología , Ratones , Ratones Endogámicos DBA , Resultado del TratamientoRESUMEN
An increasing number of data involve immunoreceptors in brain development, synaptic plasticity and behavior. However it has yet to be determined whether these proteins in fact transmit an immunoreceptor-like signal in non-hematopoietic neuronal cells. The recruitment and activation of the Syk family tyrosine kinases, Syk and ZAP-70, being a critical step in this process, we conducted a thorough analysis of Syk/ZAP-70 expression pattern in nervous tissues. Syk/ZAP-70 is present in neurons of different structures including the cerebellum, the hippocampus, the visual system and the olfactory system. During the olfactory system ontogeny the protein is detected from the 16th embryonic day and persists in adulthood. Importantly, Syk was phosphorylated on tyrosine residues representative of an active form of the kinase in specialized neuronal subpopulations comprising rostral migratory stream neuronal progenitor cells, hippocampal pyramidal cells, retinal ganglion cells and cerebellar granular cells. Phospho-Syk staining was also observed in synapse-rich regions such as the olfactory bulb glomeruli and the retina inner plexiform layer. Furthermore, our work on cultured primary hippoccampal neurons indicates that as for hematopoietic cells, Syk phosphorylation is readily induced upon pervanadate treatment. Therefore, Syk appears to be a serious candidate in connecting immunoreceptors to downstream adaptor/effector molecules in neurons.
Asunto(s)
Encéfalo/enzimología , Encéfalo/crecimiento & desarrollo , Regulación del Desarrollo de la Expresión Génica/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Neuronas/enzimología , Proteínas Tirosina Quinasas/metabolismo , Transducción de Señal/fisiología , Animales , Animales Recién Nacidos , Células Cultivadas , Cerebelo/citología , Cerebelo/embriología , Cerebelo/enzimología , Hipocampo/citología , Hipocampo/embriología , Hipocampo/enzimología , Péptidos y Proteínas de Señalización Intracelular/genética , Neuronas/citología , Neuronas/metabolismo , Bulbo Olfatorio/citología , Bulbo Olfatorio/embriología , Bulbo Olfatorio/enzimología , Fosforilación/fisiología , Proteínas Tirosina Quinasas/biosíntesis , Proteínas Tirosina Quinasas/genética , Ratas , Ratas Wistar , Retina/citología , Retina/embriología , Retina/enzimología , Organismos Libres de Patógenos Específicos , Células Madre/citología , Células Madre/enzimología , Células Madre/metabolismo , Quinasa Syk , Proteína Tirosina Quinasa ZAP-70/biosíntesis , Proteína Tirosina Quinasa ZAP-70/genética , Proteína Tirosina Quinasa ZAP-70/metabolismoRESUMEN
IL-6-mediated T cell-driven immune responses are associated with signaling occurring through the membrane-bound cognate receptor α-chain (mIL-6Rα). Once formed, IL-6-mIL-6Rα complexes induce the homodimerization and subsequent phosphorylation of the ubiquitously expressed signal-transducing protein, gp130. This signaling event is defined as classical IL-6 signaling. However, many inflammatory processes assigned to IL-6 may be mediated via binding a naturally occurring soluble IL-6Rα, which forms an agonistic complex (IL-6/soluble IL-6Rα) capable of evoking responses on a wide range of cell types that lack mIL-6Rα (IL-6 trans-signaling). To dissect the differential contribution of the two IL-6 signaling pathways in cell-mediated inflammatory processes, we pharmaceutically targeted each using two murine models of human arthritis. Whereas intra-articular neutralization of trans-signaling attenuated local inflammatory responses, the classical pathway was found to be obligate and sufficient to induce pathogenic T cells and humoral responses, leading to systemic disease. Our data illustrate that mechanisms occurring in the secondary lymphoid organs underlying arthropathies are mediated via the classical pathway of IL-6 signaling, whereas trans-signaling contributes only at the local site, that is, in the affected tissues.
Asunto(s)
Artritis Experimental/inmunología , Autoinmunidad/inmunología , Interleucina-6/inmunología , Transducción de Señal/inmunología , Linfocitos T/inmunología , Animales , Artritis Experimental/metabolismo , Separación Celular , Citocinas/biosíntesis , Ensayo de Inmunoadsorción Enzimática , Citometría de Flujo , Expresión Génica , Interleucina-6/metabolismo , Activación de Linfocitos/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Receptores de Interleucina-6/inmunología , Receptores de Interleucina-6/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Resonancia por Plasmón de Superficie , TransfecciónRESUMEN
BACKGROUND: In many neuroinflammatory diseases, dendritic cells (DCs) accumulate in several compartments of the central nervous system (CNS), including the cerebrospinal fluid (CSF). Myeloid DCs invading the inflamed CNS are thus thought to play a major role in the initiation and perpetuation of CNS-targeted autoimmune responses. We previously reported that, in normal rats, DCs injected intra-CSF migrated outside the CNS and reached the B-cell zone of cervical lymph nodes. However, there is yet no information on the migratory behavior of CSF-circulating DCs under neuroinflammatory conditions. METHODOLOGY/PRINCIPAL FINDINGS: To address this issue, we performed in vivo transfer experiments in rats suffering from experimental autoimmune encephalomyelitis (EAE), a model of multiple sclerosis. EAE or control rats were injected intra-CSF with bone marrow-derived myeloid DCs labeled with the fluorescent marker carboxyfluorescein diacetate succinimidyl ester (CFSE). In parallel experiments, fluorescent microspheres were injected intra-CSF to EAE rats in order to track endogenous antigen-presenting cells (APCs). Animals were then sacrificed on day 1 or 8 post-injection and their brain and peripheral lymph nodes were assessed for the presence of microspheres(+) APCs or CFSE(+) DCs by immunohistology and/or FACS analysis. Data showed that in EAE rats, DCs injected intra-CSF substantially infiltrated several compartments of the inflamed CNS, including the periventricular demyelinating lesions. We also found that in EAE rats, as compared to controls, a larger number of intra-CSF injected DCs reached the cervical lymph nodes. This migratory behavior was accompanied by an accentuation of EAE clinical signs and an increased systemic antibody response against myelin oligodendrocyte glycoprotein, a major immunogenic myelin antigen. CONCLUSIONS/SIGNIFICANCE: Altogether, these results indicate that CSF-circulating DCs are able to both survey the inflamed brain and to reach the cervical lymph nodes. In EAE and maybe multiple sclerosis, CSF-circulating DCs may thus support the immune responses that develop within and outside the inflamed CNS.
Asunto(s)
Encéfalo/patología , Líquido Cefalorraquídeo/citología , Encefalomielitis Autoinmune Experimental/líquido cefalorraquídeo , Ganglios Linfáticos/patología , Animales , Western Blotting , Encefalomielitis Autoinmune Experimental/patología , Ensayo de Inmunoadsorción Enzimática , Femenino , CuelloRESUMEN
Cannabinoids are potential agents for the development of therapeutic strategies against multiple sclerosis. Here we analyzed the role of the peripheral CB(2) cannabinoid receptor in the control of myeloid progenitor cell trafficking toward the inflamed spinal cord and their contribution to microglial activation in an animal model of multiple sclerosis (experimental autoimmune encephalomyelitis, EAE). CB(2) receptor knock-out mice showed an exacerbated clinical score of the disease when compared with their wild-type littermates, and this occurred in concert with extended axonal loss, T-lymphocyte (CD4(+)) infiltration, and microglial (CD11b(+)) activation. Immature bone marrow-derived CD34(+) myeloid progenitor cells, which play a role in neuroinflammatory pathologies, were shown to express CB(2) receptors and to be abundantly recruited toward the spinal cords of CB(2) knock-out EAE mice. Bone marrow-derived cell transfer experiments further evidenced the increased contribution of these cells to microglial replenishment in the spinal cords of CB(2)-deficient animals. In line with these observations, selective pharmacological CB(2) activation markedly reduced EAE symptoms, axonal loss, and microglial activation. CB(2) receptor manipulation altered the expression pattern of different chemokines (CCL2, CCL3, CCL5) and their receptors (CCR1, CCR2), thus providing a mechanistic explanation for its role in myeloid progenitor recruitment during neuroinflammation. These findings demonstrate the protective role of CB(2) receptors in EAE pathology; provide evidence for a new site of CB(2) receptor action, namely the targeting of myeloid progenitor trafficking and its contribution to microglial activation; and support the potential use of non-psychoactive CB(2) agonists in therapeutic strategies for multiple sclerosis and other neuroinflammatory disorders.
Asunto(s)
Movimiento Celular , Esclerosis Múltiple/metabolismo , Esclerosis Múltiple/patología , Células Progenitoras Mieloides/citología , Células Progenitoras Mieloides/metabolismo , Receptor Cannabinoide CB2/metabolismo , Animales , Células de la Médula Ósea/citología , Células de la Médula Ósea/metabolismo , Células Cultivadas , Modelos Animales de Enfermedad , Regulación de la Expresión Génica , Humanos , Ratones , Ratones Noqueados , Esclerosis Múltiple/genética , Receptor Cannabinoide CB2/deficiencia , Receptor Cannabinoide CB2/genética , Médula Espinal/citología , Médula Espinal/metabolismoRESUMEN
The interface between the blood and the cerebrospinal fluid (CSF) is formed by the choroid plexuses (CPs), which are specialized structures located within the brain ventricles. They are composed of a vascularized stroma surrounded by a tight epithelium that controls molecular and cellular traffic between the blood and the CSF. Cells expressing myeloid markers are present within the choroidal stroma. However, the exact identity, maturation state, and functions of these CP-associated myeloid cells are not fully clarified. We show here that this cell population contains immature myeloid progenitors displaying a high proliferative potential. Thus, in neonate rats and, to a lesser extent, in adult rats, cultured CP stroma cells form large colonies of macrophages, in response to M-CSF or GM-CSF, while, under the same conditions, peripheral blood monocytes do not. In addition, under GM-CSF treatment, free-floating colonies of CD11c(+) monocytic cells are generated which, when restimulated with GM-CSF and IL-4, differentiate into OX62(+)/MHC class II(+) dendritic cells. Interestingly, in CP stroma cultures, myeloid cells are found in close association with fibroblastic-like cells expressing the neural stem-cell marker nestin. Similarly, in the developing brain, macrophages and nestin(+) fibroblastic cells accumulate in vivo within the choroidal stroma. Taken together, these results suggest that the CP stroma represents a niche for myeloid progenitors and may serve as a reservoir for brain macrophages.
Asunto(s)
Plexo Coroideo/citología , Células Dendríticas/citología , Macrófagos/citología , Células Madre/citología , Animales , Animales Recién Nacidos , Células de la Médula Ósea/citología , Técnicas de Cultivo de Célula , Diferenciación Celular/efectos de los fármacos , División Celular/efectos de los fármacos , Citometría de Flujo , Factor Estimulante de Colonias de Granulocitos y Macrófagos/farmacología , Antígenos de Histocompatibilidad Clase II/fisiología , Humanos , Interleucina-4/farmacología , Macrófagos/efectos de los fármacos , Ratones , Ratas , Células Madre/efectos de los fármacos , Células del Estroma/citologíaRESUMEN
The lack of draining lymphatic vessels in the central nervous system (CNS) contributes to the so-called "CNS immune privilege." However, despite such a unique anatomic feature, dendritic cells (DCs) are able to migrate from the CNS to cervical lymph nodes through a yet unknown pathway. In this report, labeled bone marrow-derived myeloid DCs were injected stereotaxically into the cerebrospinal fluid (CSF) or brain parenchyma of normal rats. We found that DCs injected within brain parenchyma migrate little from their site of injection and do not reach cervical lymph nodes. In contrast, intra-CSF-injected DCs either reach cervical lymph nodes or, for a minority of them, infiltrate the subventricular zone, where neural stem cells reside. Surprisingly, DCs that reach cervical lymph nodes preferentially target B-cell follicles rather than T-cell-rich areas. This report sheds a new light on the specific role exerted by CSF-infiltrating DCs in the control of CNS-targeted immune responses.