RESUMEN
Targeting myeloid cells, especially microglia, for the treatment of neuroinflammatory diseases such as multiple sclerosis (MS), is underappreciated. Our in silico drug screening reveals topoisomerase 1 (TOP1) inhibitors as promising drug candidates for microglial modulation. We show that TOP1 is highly expressed in neuroinflammatory conditions, and TOP1 inhibition using camptothecin (CPT) and its FDA-approved analog topotecan (TPT) reduces inflammatory responses in microglia/macrophages and ameliorates neuroinflammation in vivo. Transcriptomic analyses of sorted microglia from LPS-challenged mice reveal an altered transcriptional phenotype following TPT treatment. To target myeloid cells, we design a nanosystem using ß-glucan-coated DNA origami (MyloGami) loaded with TPT (TopoGami). MyloGami shows enhanced specificity to myeloid cells while preventing the degradation of the DNA origami scaffold. Myeloid-specific TOP1 inhibition using TopoGami significantly suppresses the inflammatory response in microglia and mitigates MS-like disease progression. Our findings suggest that TOP1 inhibition in myeloid cells represents a therapeutic strategy for neuroinflammatory diseases and that the myeloid-specific nanosystems we designed may also benefit the treatment of other diseases with dysfunctional myeloid cells.
Asunto(s)
Enfermedades Neuroinflamatorias , Inhibidores de Topoisomerasa I , Animales , ADN , Macrófagos , Ratones , Inhibidores de Topoisomerasa I/farmacología , Topotecan/farmacologíaRESUMEN
BACKGROUND: Fluorescent reporter labeling and promoter-driven Cre-recombinant technologies have facilitated cellular investigations of physiological and pathological processes, including the widespread use of the Cx3cr1CreER-Eyfp/wt mouse strain for studies of microglia. METHODS: Immunohistochemistry, Flow Cytometry, RNA sequencing and whole-genome sequencing were used to identify the subpopulation of microglia in Cx3cr1CreER-Eyfp/wt mouse brains. Genetically mediated microglia depletion using Cx3cr1CreER-Eyfp/wtRosa26DTA/wt mice and CSF1 receptor inhibitor PLX3397 were used to deplete microglia. Primary microglia proliferation and migration assay were used for in vitro studies. RESULTS: We unexpectedly identified a subpopulation of microglia devoid of genetic modification, exhibiting higher Cx3cr1 and CX3CR1 expression than Cx3cr1CreER-Eyfp/wtCre+Eyfp+ microglia in Cx3cr1CreER-Eyfp/wt mouse brains, thus termed Cx3cr1highCre-Eyfp- microglia. This subpopulation constituted less than 1% of all microglia under homeostatic conditions, but after Cre-driven DTA-mediated microglial depletion, Cx3cr1highCre-Eyfp- microglia escaped depletion and proliferated extensively, eventually occupying one-third of the total microglial pool. We further demonstrated that the Cx3cr1highCre-Eyfp- microglia had lost their genetic heterozygosity and become homozygous for wild-type Cx3cr1. Therefore, Cx3cr1highCre-Eyfp- microglia are Cx3cr1wt/wtCre-Eyfp-. Finally, we demonstrated that CX3CL1-CX3CR1 signaling regulates microglial repopulation both in vivo and in vitro. CONCLUSIONS: Our results raise a cautionary note regarding the use of Cx3cr1CreER-Eyfp/wt mouse strains, particularly when interpreting the results of fate mapping, and microglial depletion and repopulation studies.
Asunto(s)
Microglía , Transducción de Señal , Animales , Receptor 1 de Quimiocinas CX3C/genética , Receptor 1 de Quimiocinas CX3C/metabolismo , Ratones , Ratones Transgénicos , Microglía/metabolismoRESUMEN
T cell-driven diseases account for considerable morbidity and disability globally and there is an urgent need for new targeted therapies. Both cancer cells and activated T cells have an altered redox balance, and up-regulate the DNA repair protein MTH1 that sanitizes the oxidized nucleotide pool to avoid DNA damage and cell death. Herein we suggest that the up-regulation of MTH1 in activated T cells correlates with their redox status, but occurs before the ROS levels increase, challenging the established conception of MTH1 increasing as a direct response to an increased ROS status. We also propose a heterogeneity in MTH1 levels among activated T cells, where a smaller subset of activated T cells does not up-regulate MTH1 despite activation and proliferation. The study suggests that the vast majority of activated T cells have high MTH1 levels and are sensitive to the MTH1 inhibitor TH1579 (Karonudib) via induction of DNA damage and cell cycle arrest. TH1579 further drives the surviving cells to the MTH1low phenotype with altered redox status. TH1579 does not affect resting T cells, as opposed to the established immunosuppressor Azathioprine, and no sensitivity among other major immune cell types regarding their function can be observed. Finally, we demonstrate a therapeutic effect in a murine model of experimental autoimmune encephalomyelitis. In conclusion, we show proof of concept of the existence of MTH1high and MTH1low activated T cells, and that MTH1 inhibition by TH1579 selectively suppresses pro-inflammatory activated T cells. Thus, MTH1 inhibition by TH1579 may serve as a novel treatment option against autoreactive T cells in autoimmune diseases, such as multiple sclerosis.
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Enzimas Reparadoras del ADN , Monoéster Fosfórico Hidrolasas , Animales , Daño del ADN , Enzimas Reparadoras del ADN/metabolismo , Recuento de Linfocitos , Ratones , Monoéster Fosfórico Hidrolasas/genética , Linfocitos T/metabolismoRESUMEN
Macrophage plasticity enables cells to obtain different functions over a broad proinflammatory and repairing spectrum. In different conditions, macrophages can be induced by high-mobility group box 1 (HMGB1), a nuclear DNA-binding protein that activates innate immunity, to polarize towards a pro- (M1) or anti-inflammatory (M2) phenotype. In this study, we investigated the phenotypes of murine bone-marrow-derived macrophages (BMDMs) induced by different HMGB1 redox isoforms in depth. Our results demonstrate that disulfide HMGB1 (dsHMGB1) induces a unique macrophage phenotype that secretes pro-inflammatory cytokines, rather than inducing metabolic changes leading to nitric oxide production. Fully reduced HMGB1 (frHMGB1) did not induce macrophage polarization. The migrating function of BMDMs was measured by scratch assay after the stimulation with dsHMGB1 and frHMGB1. Both dsHMGB1 and frHMGB1 induced cell migration. We found that dsHMGB1 mediates cytokine secretion and cellular motility, mainly through toll-like receptor 4 (TLR4). Importantly, our data shows that dsHMGB1 and frHMGB1 induce distinct BMDM polarization phenotypes, and that dsHMGB1 induces a unique phenotype differing from the classical proinflammatory macrophage phenotype.
Asunto(s)
Movimiento Celular/efectos de los fármacos , Disulfuros/química , Proteína HMGB1 , Macrófagos/metabolismo , Animales , Femenino , Proteína HMGB1/química , Proteína HMGB1/farmacología , Ratones , Oxidación-Reducción , Receptor Toll-Like 4/metabolismoRESUMEN
Intratumoral heterogeneity is a characteristic of glioblastomas that contain an intermixture of cell populations displaying different glioblastoma subtype gene expression signatures. Proportions of these populations change during tumor evolution, but the occurrence and regulation of glioblastoma subtype transition is not well described. To identify regulators of glioblastoma subtypes we utilized a combination of in vitro experiments and in silico analyses, using experimentally generated as well as publicly available data. Through this combined approach SOX2 was identified to confer a proneural glioblastoma subtype gene expression signature. SFRP2 was subsequently identified as a SOX2-antagonist, able to induce a mesenchymal glioblastoma subtype signature. A subset of patient glioblastoma samples with high SFRP2 and low SOX2 expression was particularly enriched with mesenchymal subtype samples. Phenotypically, SFRP2 decreased tumor sphere formation, stemness as assessed by limiting dilution assay, and overall cell proliferation but increased cell motility, whereas SOX2 induced the opposite effects. Furthermore, an SFRP2/non-canonical-WNT/KLF4/PDGFR/phospho-AKT/SOX2 signaling axis was found to be involved in the mesenchymal transition. Analysis of human tumor tissue spatial gene expression patterns showed distinct expression of SFRP2- and SOX2-correlated genes in vascular and cellular areas, respectively. Finally, conditioned media from SFRP2 overexpressing cells increased CD206 on macrophages. Together, these findings present SFRP2 as a SOX2-antagonist with the capacity to induce a mesenchymal subtype transition in glioma cells located in vascular tumor areas, highlighting its role in glioblastoma tumor evolution and intratumoral heterogeneity.
Asunto(s)
Transición Epitelial-Mesenquimal/genética , Regulación Neoplásica de la Expresión Génica , Glioblastoma/etiología , Glioblastoma/metabolismo , Proteínas de la Membrana/genética , Factores de Transcripción SOXB1/genética , Proteínas Portadoras , Línea Celular Tumoral , Perfilación de la Expresión Génica , Glioblastoma/patología , Humanos , Factor 4 Similar a Kruppel/metabolismo , Macrófagos/inmunología , Macrófagos/metabolismo , Macrófagos/patología , Proteínas de la Membrana/metabolismo , Especificidad de Órganos , Unión Proteica , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptores del Factor de Crecimiento Derivado de Plaquetas/metabolismo , Factores de Transcripción SOXB1/metabolismo , Transducción de SeñalRESUMEN
Microglia, predominant parenchymal resident macrophages in the central nervous system (CNS), are crucial players in neurodevelopment and CNS homeostasis. In disease conditions, pro-inflammatory microglia predominate over their regulatory counterparts, and are thus a potential immunotherapeutic target. It has been well documented that microglia can be effectively depleted using both conditional genetic Cx3cr1Cre-diphtheria toxin receptor (DTR)/diphtheria toxin subunit A (DTA) animal models and pharmacological colony-stimulating factor 1 receptor (CSF1R) inhibitors. Recent advances using these approaches have expanded our knowledge of the multitude of tasks conducted by microglia in both homeostasis and diseases. Importantly, experimental microglial depletion has been proven to exert neuroprotective effects in an increasing number of disease models, mostly explained by reduced neuroinflammation. However, the comprehensive effects of additional targets such as circulating monocytes and peripheral tissue macrophages during microglial depletion periods have not been investigated widely, and for those studies addressing the issue the conclusions are mixed. In this study, we demonstrate that experimental microglial depletion using both Cx3cr1CreER/+Rosa26DTA/+ mice and different doses of CSF1R inhibitor PLX3397 exert crucial influences on circulating monocytes and peripheral tissue macrophages. Our results suggest that effects on peripheral immunity should be considered both in interpretation of microglial depletion studies, and especially in the potential translation of microglial depletion and replacement therapies.
Asunto(s)
Macrófagos/metabolismo , Microglía/metabolismo , Fármacos Neuroprotectores/farmacología , Receptores de Factor Estimulante de Colonias de Granulocitos y Macrófagos/metabolismo , Aminopiridinas/farmacología , Animales , Femenino , Factor de Crecimiento Similar a EGF de Unión a Heparina/genética , Factor de Crecimiento Similar a EGF de Unión a Heparina/metabolismo , Ratones , Ratones Transgénicos , Pirroles/farmacología , Receptores de Factor Estimulante de Colonias de Granulocitos y Macrófagos/genéticaRESUMEN
Multiple sclerosis (MS) is a chronic neuroinflammatory disorder of the central nervous system (CNS) that usually presents in young adults and predominantly in females. Microglia, a major resident immune cell in the CNS, are critical players in both CNS homeostasis and disease. We have previously demonstrated that microglia can be efficiently depleted by the administration of tamoxifen in Cx3cr1CreER/+Rosa26DTA/+ mice, with ensuing repopulation deriving from both the proliferation of residual CNS resident microglia and the engraftment of peripheral monocyte-derived microglia-like cells. In this study, tamoxifen was administered to Cx3cr1CreER/+Rosa26DTA/+ and Cx3cr1CreER/+ female and male mice. Experimental autoimmune encephalomyelitis (EAE), a widely used animal model of MS, was induced by active immunization with myelin oligodendrocyte glycoprotein (MOG) one month after tamoxifen injections in Cx3cr1CreER/+Rosa26DTA/+ mice and Cx3cr1CreER/+ mice, a time point when the CNS niche was colonized by microglia derived from both CNS microglia and peripherally-derived macrophages. We demonstrate that engraftment of microglia-like cells following microglial depletion exacerbated EAE in Cx3cr1CreER/+Rosa26DTA/+ female mice as assessed by clinical symptoms and the expression of CNS inflammatory factors, but these findings were not evident in male mice. Higher major histocompatibility complex class II expression and cytokine production in the female CNS contributed to the sex-dependent EAE severity in mice following engraftment of microglia-like cells. An underestimated yet marked sex-dependent microglial activation pattern may exist in the injured CNS during EAE.
Asunto(s)
Sistema Nervioso Central/citología , Encefalomielitis Autoinmune Experimental/metabolismo , Antígenos de Histocompatibilidad Clase II/metabolismo , Macrófagos/metabolismo , Microglía/citología , Monocitos/metabolismo , Esclerosis Múltiple/metabolismo , Glicoproteína Mielina-Oligodendrócito/inmunología , Animales , Sistema Nervioso Central/inmunología , Sistema Nervioso Central/metabolismo , Citocinas/metabolismo , Modelos Animales de Enfermedad , Femenino , Activación de Macrófagos/inmunología , Macrófagos/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Microglía/inmunología , Microglía/metabolismo , Monocitos/inmunología , Tamoxifeno/administración & dosificaciónRESUMEN
Microglia are implicated in the pathophysiology of several neurodegenerative disorders, including Alzheimer's disease. While the role of microglia and peripheral macrophages in regulating amyloid beta pathology has been well characterized, the impact of these distinct cell subsets on tau pathology remains poorly understood. We and others have recently demonstrated that monocytes can engraft the brain and give rise to long-lived parenchymal macrophages, even under nonpathological conditions. We undertook the current study to investigate the regulation of tau pathology by microglia and peripheral macrophages using hTau transgenic mice, which do not exhibit microglial activation/pathology or macrophage engraftment. To assess the direct impact of microglia on tau pathology we developed a protocol for long-term microglial depletion in Cx3cr1CreER R26DTA mice and crossed them with hTau mice. We then depleted microglia up to 3 months in both young and old mice, but no net change in forebrain soluble oligomeric tau or total or phosphorylated levels of aggregated tau was recorded. To investigate the consequence of peripherally-derived parenchymal macrophages on tau aggregation we partially repopulated the hTau microglial pool with peripheral macrophages, but this also did not affect levels of tau oligomers or insoluble aggregates. Our study questions the direct involvement of microglia or peripheral macrophages in the development of tau pathology in the hTau model.
Asunto(s)
Enfermedad de Alzheimer/patología , Macrófagos/metabolismo , Microglía/metabolismo , Tauopatías/patología , Proteínas tau/metabolismo , Enfermedad de Alzheimer/metabolismo , Animales , Encéfalo/patología , Modelos Animales de Enfermedad , Ratones Transgénicos , Microglía/patología , Monocitos/metabolismoRESUMEN
OBJECTIVES: In a pilot study we aimed to identify biomarkers in repeated muscle biopsies and paired blood samples, taken before and after conventional immunosuppressive therapy, in order to predict long-term therapeutic response in patients with idiopathic inflammatory myopathies (IIM). METHODS: Muscle biopsies were selected from 13 new onset patients, six responders and seven non-responders. Repeated muscle biopsies after a median of 11 months follow-up were available from 9 patients and paired peripheral blood mononuclear cells (PBMCs) from 5 patients. Treatment response after 3 years was defined by MMT-8 measuring muscle strength and the ACR/EULAR 2016 improvement criteria. Frozen biopsy sections were immunohistochemically stained for expression of CD3, CD66b, IL-15, CD68, CD163 and myosin heavy chain neonatal (MHCn). PBMCs were analysed by flow cytometry for monocyte phenotypes (CD14, CD16, CD68, CX3CR1, and CCR2). RESULTS: Before treatment there were no significant differences in any clinical or muscle biopsy variables or monocyte subsets between responders and non-responders. MMT-8 was significantly higher compared to baseline in the responders at 3-year follow-up. In responders the expression of CD68 in the repeated biopsies was significantly lower compared to non-responders (p<0.05). CONCLUSIONS: Baseline biopsy, monocyte profile or clinical data did not predict long-term treatment response, but in the repeated biopsy within 1 year of immunosuppressive treatment, the lower number of macrophages (CD68+) seemed to predict a more favourable long-term clinical response with regard to improved muscle strength.
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Monocitos/citología , Músculo Esquelético/patología , Miositis/terapia , Biomarcadores/análisis , Biopsia , Estudios de Seguimiento , Humanos , Leucocitos Mononucleares/citología , Monocitos/clasificación , Fenotipo , Proyectos PilotoRESUMEN
Amyloid-ß (Aß) plaque deposits in the brain are considered to be one of the main pathological markers of Alzheimer's disease (AD). The sequential proteolytic cleavage of amyloid precursor protein (APP) by the aspartyl proteases ß-site APP-cleaving enzyme 1 (BACE1) and γ-secretase produces Aß. Therefore, BACE1 inhibition is a very attractive target for the treatment of AD. Our previous work identified a DNA aptamer named A1 that can bind to BACE1 with high affinity and specificity and exhibits a distinct inhibitory effect on BACE1 activity in an AD cell model. The purpose of this research was to test the effect of aptamer A1 in Tg6799 mice. Four-month-old Tg6799 mice were randomly divided into two groups and treated with aptamer A1 and ineffective aptamer A1scr, respectively, by intracerebroventricular injection. Subsequent behavioral experiments showed that treatment with the aptamer A1 improved the cognitive abilities of the AD mice. Western blot indicated that BACE1 and soluble amyloid precursor protein ß (sAPPß) expression significantly decreased in the A1-treated mice. Moreover, aptamer A1 reduced the content of Aß42 and the number and density of senile plaques in AD mice. Therefore, our results indicate that aptamer A1 is a novel specific and potent BACE1 inhibitor and is a promising potential target for the treatment of AD.
Asunto(s)
Enfermedad de Alzheimer/terapia , Secretasas de la Proteína Precursora del Amiloide/genética , Péptidos beta-Amiloides/genética , Ácido Aspártico Endopeptidasas/genética , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/patología , Secretasas de la Proteína Precursora del Amiloide/antagonistas & inhibidores , Péptidos beta-Amiloides/farmacología , Animales , Aptámeros de Nucleótidos/genética , Aptámeros de Nucleótidos/farmacología , Ácido Aspártico Endopeptidasas/antagonistas & inhibidores , Encéfalo/efectos de los fármacos , Encéfalo/patología , Modelos Animales de Enfermedad , Terapia Genética/métodos , Humanos , Infusiones Intraventriculares , Ratones , Ratones TransgénicosRESUMEN
Alzheimer's disease (AD) is an irreversible, progressive neurodegenerative disorder of the central nervous system that causes severe cognitive impairment. One of the most significant pathological features of AD is the accumulation of ßamyloid (Aß) peptide in the brain. Resveratrol (Res) is a polyphenol derived from peanuts, red grapes and other plants, which has received increasing attention due to its neuroprotective features. Tg6799 mice are transgenic mice with five familial AD (FAD) mutations that are also known as 5XFAD mice. The present study aimed to investigate the effects of Res on Tg6799 mice. The transgenic mice were randomly divided into the Res treatment group and the vehicle control group, and were treated with 0.5% Res solution (60 mg/kg) or volumematched normal saline, respectively. Treatment was administered by oral gavage daily for 60 consecutive days. Res reduced amyloid plaque formation and the levels of Aß42, and ßsecretase 1 levels were also significantly decreased. Furthermore, Res was able to reduce the expression of amyloid precursor protein and its cleavage products. The administration of Res to Tg6799 mice also improved their spatial working memory, as measured by the Ymaze test, and rescued spatial memory deficits, as measured using the Morris water maze test; however, Res did not affect their motor function. In conclusion, this study suggested that Res may reduce Aßinduced neuronal damage, thus preventing memory loss.
Asunto(s)
Enfermedad de Alzheimer/tratamiento farmacológico , Precursor de Proteína beta-Amiloide/fisiología , Trastornos del Conocimiento/prevención & control , Modelos Animales de Enfermedad , Placa Amiloide/prevención & control , Presenilina-1/fisiología , Resveratrol/farmacología , Animales , Antiinflamatorios no Esteroideos/farmacología , Conducta Animal/efectos de los fármacos , Trastornos del Conocimiento/metabolismo , Trastornos del Conocimiento/patología , Masculino , Trastornos de la Memoria/metabolismo , Trastornos de la Memoria/patología , Trastornos de la Memoria/prevención & control , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Placa Amiloide/metabolismo , Placa Amiloide/patologíaRESUMEN
Microglia are prominent immune cells in the central nervous system (CNS) and are critical players in both neurological development and homeostasis, and in neurological diseases when dysfunctional. Our previous understanding of the phenotypes and functions of microglia has been greatly extended by a dearth of recent investigations. Distinct genetically defined subsets of microglia are now recognized to perform their own independent functions in specific conditions. The molecular profiling of single microglial cells indicates extensively heterogeneous reactions in different neurological disorders, resulting in multiple potentials for crosstalk with other kinds of CNS cells such as astrocytes and neurons. In settings of neurological diseases it could thus be prudent to establish effective cell-based therapies by targeting entire microglial networks. Notably, activated microglial depletion through genetic targeting or pharmacological therapies within a suitable time window can stimulate replenishment of the CNS niche with new microglia. Additionally, enforced repopulation through provision of replacement cells also represents a potential means of exchanging dysfunctional with functional microglia. In each setting the newly repopulated microglia might have the potential to resolve ongoing neuroinflammation. In this review, we aim to summarize the most recent knowledge of microglia and to highlight microglial depletion and subsequent repopulation as a promising cell replacement therapy. Although glial cell replacement therapy is still in its infancy and future translational studies are still required, the approach is scientifically sound and provides new optimism for managing the neurotoxicity and neuroinflammation induced by activated microglia.
Asunto(s)
Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Microglía/fisiología , Enfermedades del Sistema Nervioso/terapia , Animales , Microglía/patologíaRESUMEN
High-grade gliomas (HGGs) are the most aggressive and invasive primary brain tumors. The platelet-derived growth factor (PDGF) signaling pathway drives HGG progression, and enhanced expression of PDGF receptors (PDGFRs) is a well-established aberration in a subset of glioblastomas (GBMs). PDGFRA is expressed in glioma cells, whereas PDGFRB is mostly restricted to the glioma-associated stroma. Here we show that the spatial location of TAMMs correlates with the expansion of a subset of tumor cells that have acquired expression of PDGFRB in both mouse and human low-grade glioma and HCGs. Furthermore, M2-polarized microglia but not bone marrow (BM)-derived macrophages (BMDMs) induced PDGFRB expression in glioma cells and stimulated their migratory capacity. These findings illustrate a heterotypic cross-talk between microglia and glioma cells that may enhance the migratory and invasive capacity of the latter by inducing PDGFRB.
RESUMEN
Circulating monocytes can compete for virtually any tissue macrophage niche and become long-lived replacements that are phenotypically indistinguishable from their embryonic counterparts. As the factors regulating this process are incompletely understood, we studied niche competition in the brain by depleting microglia with >95% efficiency using Cx3cr1CreER/+R26DTA/+ mice and monitored long-term repopulation. Here we show that the microglial niche is repopulated within weeks by a combination of local proliferation of CX3CR1+F4/80lowClec12a- microglia and infiltration of CX3CR1+F4/80hiClec12a+ macrophages that arise directly from Ly6Chi monocytes. This colonization is independent of blood brain barrier breakdown, paralleled by vascular activation, and regulated by type I interferon. Ly6Chi monocytes upregulate microglia gene expression and adopt microglia DNA methylation signatures, but retain a distinct gene signature from proliferating microglia, displaying altered surface marker expression, phagocytic capacity and cytokine production. Our results demonstrate that monocytes are imprinted by the CNS microenvironment but remain transcriptionally, epigenetically and functionally distinct.
Asunto(s)
Encéfalo/inmunología , Linaje de la Célula/inmunología , Regulación de la Expresión Génica/inmunología , Microglía/inmunología , Monocitos/inmunología , Traslado Adoptivo , Animales , Antígenos de Diferenciación/genética , Antígenos de Diferenciación/inmunología , Antígenos Ly/genética , Antígenos Ly/inmunología , Proteínas Bacterianas/inmunología , Células de la Médula Ósea/citología , Células de la Médula Ósea/inmunología , Encéfalo/citología , Encéfalo/efectos de la radiación , Receptor 1 de Quimiocinas CX3C/genética , Receptor 1 de Quimiocinas CX3C/inmunología , Linaje de la Célula/efectos de la radiación , Proliferación Celular , Metilación de ADN , Genes Reporteros , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/inmunología , Interferón Tipo I/genética , Interferón Tipo I/inmunología , Lectinas Tipo C/genética , Lectinas Tipo C/inmunología , Proteínas Luminiscentes/inmunología , Macrófagos/citología , Macrófagos/inmunología , Macrófagos/efectos de la radiación , Ratones , Ratones Transgénicos , Microglía/citología , Microglía/efectos de la radiación , Monocitos/citología , Monocitos/efectos de la radiación , Monocitos/trasplante , Fagocitosis , Receptores Mitogénicos/genética , Receptores Mitogénicos/inmunología , Transducción de Señal , Quimera por Trasplante , Irradiación Corporal TotalRESUMEN
The cytokine transforming growth factor-ß (TGF-ß) regulates the development and homeostasis of several tissue-resident macrophage populations, including microglia. TGF-ß is not critical for microglia survival but is required for the maintenance of the microglia-specific homeostatic gene signature1,2. Under defined host conditions, circulating monocytes can compete for the microglial niche and give rise to long-lived monocyte-derived macrophages residing in the central nervous system (CNS)3-5. Whether monocytes require TGF-ß for colonization of the microglial niche and maintenance of CNS integrity is unknown. We found that abrogation of TGF-ß signaling in CX3CR1+ monocyte-derived macrophages led to rapid onset of a progressive and fatal demyelinating motor disease characterized by myelin-laden giant macrophages throughout the spinal cord. Tgfbr2-deficient macrophages were characterized by high expression of genes encoding proteins involved in antigen presentation, inflammation and phagocytosis. TGF-ß is thus crucial for the functional integration of monocytes into the CNS microenvironment.
Asunto(s)
Encéfalo/inmunología , Enfermedades Desmielinizantes/inmunología , Macrófagos/patología , Médula Espinal/inmunología , Factor de Crecimiento Transformador beta/inmunología , Animales , Encéfalo/metabolismo , Encéfalo/patología , Enfermedades Desmielinizantes/metabolismo , Enfermedades Desmielinizantes/patología , Macrófagos/inmunología , Macrófagos/metabolismo , Ratones , Transducción de Señal , Médula Espinal/metabolismo , Médula Espinal/patología , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
Synthetic hormones in wastewater effluents released into the aquatic environments may interfere with the normal endocrine systems of fish in receiving streams. Norgestrel (NGT) is a synthetic progestin widely used in oral contraceptives and frequently detected in wastewater effluents. In this study, adult female mosquitofish (Gambusia affinis) were exposed to three environmentally relevant concentrations of norgestrel (NGT) (i.e., 3.6, 35.8, and 368.0â¯ngâ¯L-1) for 42â¯d, fin morphology, histology of the ovary, and reproductive behaviors were evaluated. The results showed that NGT at all three concentrations caused an increased frequency of atretic follicular cells in ovaries and impaired mating behaviors exhibited by males toward the NGT-exposed females. In mosquitofish exposed to NGT at 35.8 and 368â¯ngâ¯L-1, the anal fin of females had an increased length ratio of ray4/ray 6, an increased width of ray 3, and increased number of segments in ray 3. The histopathological analysis showed that exposure to NGT increased the incidence of spermatogenesis in ovaries. Mating behavior was impaired 58.4%, 65.7%, and 76.4% (Pâ¯<â¯0.01 in all cases) when mosquitofish were exposed to NGT at 3.6, 35.6 and 368.0â¯ngâ¯L-1, respectively. The rapid masculinization, the increased frequency of atretic follicles, the incidence of spermatogenesis in the ovary of female fish, and the altered reproductive behaviors suggest that wild populations of mosquitofish could be similarly affected inhabiting in NGT contaminated environments.
Asunto(s)
Conducta Animal/efectos de los fármacos , Ciprinodontiformes/fisiología , Norgestrel/toxicidad , Reproducción/efectos de los fármacos , Caracteres Sexuales , Aletas de Animales/anatomía & histología , Aletas de Animales/efectos de los fármacos , Animales , Exposición a Riesgos Ambientales/análisis , Femenino , Masculino , Oocitos/citología , Oocitos/efectos de los fármacos , Ovario/química , Ovario/efectos de los fármacos , Contaminantes Químicos del Agua/toxicidadRESUMEN
Based on the discordance of human epidermal growth factor receptor-2 (HER2) expression between primary and metastatic/recurrent breast cancer, HER2 molecular imaging, which had potential to systemically assess and dynamically monitor HER2 expression, might improve the selection of patients for anti-HER2 therapy. In this study, designed ankyrin repeat protein (DARPin) G3, a novel binding protein with picomolar affinity for HER2, was used and multifunctional superparamagnetic nanoparticles modified with fluorescein-5-maleimide-labeled DARPin G3 (SPIO-G3-5MF) were developed for HER2 imaging. Our results showed that SPIO-G3-5MF nanoparticles, which possessed uniform size of about 100 nm, favorable dispersity and low cytotoxicity, could selectively bind to HER2-positive breast cancer cells even in the presence of trastuzumab. Biodistribution assay demonstrated that abundant accumulation and long retention of SPIO-G3-5MF were observed in HER2-positive transplantation breast tumors although a portion of SPIO-G3-5MF nanoparticles were unavoidably captured by liver and spleen. Further MR imaging revealed that SPIO-G3-5MF could selectively image HER2-positive transplantation breast tumors, yielding remarkable T2 signal reduction (50.33 ± 2.90% at 6 h and 47.29 ± 9.36% at 24 h). Our study suggested that SPIO-G3-5MF might be a promising MR molecular probe for diagnosing and monitoring HER2 expression state of breast cancer in the future.
Asunto(s)
Repetición de Anquirina , Neoplasias de la Mama/diagnóstico por imagen , Fluoresceína/química , Nanopartículas de Magnetita/química , Receptor ErbB-2/metabolismo , Proteínas Recombinantes/química , Animales , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Supervivencia Celular , Femenino , Compuestos Férricos/química , Xenoinjertos , Humanos , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Sondas Moleculares/química , Recurrencia Local de Neoplasia/diagnóstico por imagen , Trasplante de Neoplasias , Tamaño de la Partícula , Propiedades de Superficie , Trastuzumab/farmacologíaRESUMEN
Ischemia-reperfusion (IR) injury involves complex pathological processes in which reduction of nitric oxide (NO) bioavailability is suggested as a key factor. Inorganic nitrate can form NO in vivo via NO synthase-independent pathways and may thus provide beneficial effects during IR. Herein we evaluated the effects of dietary nitrate supplementation in a renal IR model. Male mice (C57BL/6J) were fed nitrate-supplemented chow (1.0mmol/kg/day) or standard chow for two weeks prior to 30min ischemia and during the reperfusion period. Unilateral renal IR caused profound tubular and glomerular damage in the ischemic kidney. Renal function, assessed by plasma creatinine levels, glomerular filtration rate and renal plasma flow, was also impaired after IR. All these pathologies were significantly improved by nitrate. Mechanistically, nitrate treatment reduced renal superoxide generation, pro-inflammatory cytokines (IL-1ß, IL-6 and IL-12 p70) and macrophage infiltration in the kidney. Moreover, nitrate reduced mRNA expression of pro-inflammatory cytokines and chemo attractors, while increasing anti-inflammatory cytokines in the injured kidney. In another cohort of mice, two weeks of nitrate supplementation lowered superoxide generation and IL-6 expression in bone marrow-derived macrophages. Our study demonstrates protective effect of dietary nitrate in renal IR injury that may be mediated via modulation of oxidative stress and inflammatory responses. These novel findings suggest that nitrate supplementation deserve further exploration as a potential treatment in patients at high risk of renal IR injury.
Asunto(s)
Lesión Renal Aguda/tratamiento farmacológico , Interleucina-6/metabolismo , Macrófagos/efectos de los fármacos , Nitratos/uso terapéutico , Estrés Oxidativo , Daño por Reperfusión/tratamiento farmacológico , Animales , Células Cultivadas , Suplementos Dietéticos , Interleucina-6/genética , Riñón/irrigación sanguínea , Activación de Macrófagos , Macrófagos/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Nitratos/administración & dosificación , Nitratos/farmacología , Superóxidos/metabolismoRESUMEN
Microglia are the principal resident immune cells in the central nervous system and are believed to be versatile players in both inflammatory and physiological contexts. On the one hand, in order to safeguard the microenvironment microglia can be rapidly activated by contact with microbial products or cell debris, thereby exerting the functions of innate immunity via phagocytosis and secretion of cytokines and chemokines. Conversely, microglia can also assist in brain development, synaptic plasticity and neural repair through the production of neurotrophic factors and clearance of myelin debris. It is now well accepted that the dysfunction of microglia and microglia-induced neuroinflammation are implicated in the occurrence and progression of many neurological diseases. Although the past decade has witnessed major progress in understanding of multi-tasking microglia, what remains largely enigmatic is the relative importance of microglia at different disease stages and how microglia should be targeted for optimal therapeutic efficacy. Notably, microglia depletion through genetic targeting or pharmacological therapies can be viewed as effective tools to stimulate new microglia to repopulate the central nervous system. Microglia depletion and subsequent repopulation at defined stages in various experimental animal model disorders allow us to extend our knowledge of molecular mechanisms, thus holding promise for designing strategies to resolve neuroinflammation and promote recovery. Herein we highlight the highly plastic and diverse phenotypes of microglia and outline the lessons learned from microglia depletion approaches.