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1.
Int J Mol Sci ; 22(5)2021 Mar 08.
Artigo em Inglês | MEDLINE | ID: mdl-33800170

RESUMO

IL-34 has been recently identified as a ligand for CSF1R that regulates various cellular processes including cell proliferation, survival, and differentiation. Although the binding of IL-34 to CSF1R modulates several cancer-driving signaling pathways, little is known about the role of IL-34/CSF1R signaling in breast cancer. Herein, we report that IL-34 induces epithelial cell transformation and breast tumorigenesis through activation of MEK/ERK and JNK/c-Jun pathways. IL-34 increased the phosphorylation of MEK1/2, ERK1/2, JNK1/2, and c-Jun through CSF1R in mouse skin epidermal JB6 C141 cells and human breast cancer MCF7 cells. IL-34 enhanced c-Fos and c-Jun promoter activity, resulting in increased AP-1 transactivation activity in JB6 Cl41 and MCF7 cells. Moreover, PIN1 enhanced IL-34-induced phosphorylation of MEK1/2, ERK1/2, JNK1/2, and c-Jun in JB6 Cl41 and MCF7 cells. Inhibition of PIN1 using juglone prevented the IL-34-induced transformation of JB6 C141 cells. Similarly, silencing of PIN1 reduced the IL-34-induced tumorigenicity of MCF7 cells. Consistent with these results, the synergistic model showed that treatment with juglone suppressed the IL-34-induced growth of tumors formed by 4T1 cells in BALB/c mice. Our study demonstrates the role of IL-34-induced MEK/ERK and JNK/c-Jun cascades in breast cancer and highlights the regulatory role of PIN1 in IL-34-induced breast tumorigenesis.


Assuntos
Neoplasias da Mama/metabolismo , Transformação Celular Neoplásica/metabolismo , Células Epiteliais/metabolismo , Interleucinas/metabolismo , Sistema de Sinalização das MAP Quinases , Proteínas de Neoplasias/metabolismo , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Animais , Neoplasias da Mama/patologia , Transformação Celular Neoplásica/patologia , Células Epiteliais/patologia , Feminino , Humanos , Células MCF-7 , Camundongos , Camundongos Endogâmicos BALB C
2.
Eur J Med Chem ; 216: 113298, 2021 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-33689933

RESUMO

Tumor-associated macrophages (TAMs) are predominantly associated with tumor growth. Colony-stimulating factor 1 receptor (CSF1R) acts as a key regulator of TAM survival and differentiation and is a molecular target for cancer therapies. Herein, novel CSF1R inhibitors were identified through a replacement strategy for the hinge-binding moiety. The introduction of imidazo[1,2-a]pyridine (49) or pyrazolo[1,5-a]pyridine (50) as hinge binders led to 87% and 82% inhibition at 10 nM for CSF1R in the enzymatic assay, with IC50 values of 25 nM and 27 nM in MNFS60 cells, respectively. These derivatives significantly inhibited CSF1R phosphorylation in cells. Our approach could be utilized as a strategy to discover novel kinase inhibitors.


Assuntos
Desenho de Fármacos , Inibidores de Proteínas Quinases/química , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/antagonistas & inibidores , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Humanos , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Fosforilação/efeitos dos fármacos , Inibidores de Proteínas Quinases/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Pirazóis/química , Pirazóis/metabolismo , Pirazóis/farmacologia , Piridinas/química , Piridinas/metabolismo , Piridinas/farmacologia , Células RAW 264.7 , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Relação Estrutura-Atividade
3.
Int J Mol Sci ; 22(5)2021 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-33652870

RESUMO

In this review, we focus on the emerging roles of microglia in the brain, with particular attention to synaptic plasticity in health and disease. We present evidence that ramified microglia, classically believed to be "resting" (i.e., inactive), are instead strongly implicated in dynamic and plastic processes. Indeed, there is an intimate relationship between microglia and neurons at synapses which modulates activity-dependent functional and structural plasticity through the release of cytokines and growth factors. These roles are indispensable to brain development and cognitive function. Therefore, approaches aimed at maintaining the ramified state of microglia might be critical to ensure normal synaptic plasticity and cognition. On the other hand, inflammatory signals associated with Alzheimer's disease are able to modify the ramified morphology of microglia, thus leading to synapse loss and dysfunction, as well as cognitive impairment. In this context, we highlight microglial TREM2 and CSF1R as emerging targets for disease-modifying therapy in Alzheimer's disease (AD) and other neurodegenerative disorders.


Assuntos
Doença de Alzheimer/patologia , Microglia/patologia , Sinapses/patologia , Doença de Alzheimer/metabolismo , Doença de Alzheimer/fisiopatologia , Animais , Humanos , Inflamação/metabolismo , Inflamação/patologia , Inflamação/fisiopatologia , Glicoproteínas de Membrana/análise , Glicoproteínas de Membrana/metabolismo , Microglia/metabolismo , Plasticidade Neuronal , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/análise , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Receptores Imunológicos/análise , Receptores Imunológicos/metabolismo , Sinapses/metabolismo
4.
Int J Mol Sci ; 22(5)2021 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-33652607

RESUMO

Spermatogenesis is a complex process, in which spermatogonial cells proliferate and differentiate in the seminiferous tubules of the testis to generate sperm. This process is under the regulation of endocrine and testicular paracrine/autocrine factors. In the present study, we demonstrated that colony stimulating factor-1 (CSF-1) is produced by mouse testicular macrophages, Leydig, Sertoli, peritubular cells and spermatogonial cells (such as CDH1-positively stained cells; a marker of spermatogonial cells). In addition, we demonstrated the presence of CSF-1 and its receptor (CSF-1R) in testicular macrophages, Leydig, Sertoli, peritubular cells and spermatogonial cells of human testis. We also show that the protein levels of CSF-1 were the highest in testis of 1-week-old mice and significantly decreased with age (2-12-week-old). However, the transcriptome levels of CSF-1 significantly increased in 2-3-week-old compared to 1-week-old, and thereafter significantly decreased with age. On the other hand, the transcriptome levels of CSF-1R was significantly higher in mouse testicular tissue of all examined ages (2-12-week-old) compared to 1-week-old. Our results demonstrate the involvement of CSF-1 in the induction the proliferation and differentiation of spermatogonial cells to meiotic and postmeiotic stages (BOULE- and ACROSIN-positive cells) under in vitro culture conditions, using methylcellulose culture system (MCS). Thus, it is possible to suggest that CSF-1 system, as a testicular paracrine/autocrine system, is involved in the development of different stages of spermatogenesis and may be used in the development of future therapeutic strategies for treatment of male infertility.


Assuntos
Fator Estimulador de Colônias de Macrófagos/metabolismo , Espermatogênese , Testículo/metabolismo , Animais , Humanos , Masculino , Camundongos , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Testículo/citologia
5.
J Neurosci ; 41(6): 1274-1287, 2021 02 10.
Artigo em Inglês | MEDLINE | ID: mdl-33380470

RESUMO

Microglia have crucial roles in sculpting synapses and maintaining neural circuits during development. To test the hypothesis that microglia continue to regulate neural circuit connectivity in adult brain, we have investigated the effects of chronic microglial depletion, via CSF1R inhibition, on synaptic connectivity in the visual cortex in adult mice of both sexes. We find that the absence of microglia dramatically increases both excitatory and inhibitory synaptic connections to excitatory cortical neurons assessed with functional circuit mapping experiments in acutely prepared adult brain slices. Microglia depletion leads to increased densities and intensities of perineuronal nets. Furthermore, in vivo calcium imaging across large populations of visual cortical neurons reveals enhanced neural activities of both excitatory neurons and parvalbumin-expressing interneurons in the visual cortex following microglia depletion. These changes recover following adult microglia repopulation. In summary, our new results demonstrate a prominent role of microglia in sculpting neuronal circuit connectivity and regulating subsequent functional activity in adult cortex.SIGNIFICANCE STATEMENT Microglia are the primary immune cell of the brain, but recent evidence supports that microglia play an important role in synaptic sculpting during development. However, it remains unknown whether and how microglia regulate synaptic connectivity in adult brain. Our present work shows chronic microglia depletion in adult visual cortex induces robust increases in perineuronal nets, and enhances local excitatory and inhibitory circuit connectivity to excitatory neurons. Microglia depletion increases in vivo neural activities of both excitatory neurons and parvalbumin inhibitory neurons. Our new results reveal new potential avenues to modulate adult neural plasticity by microglia manipulation to better treat brain disorders, such as Alzheimer's disease.


Assuntos
Microglia/metabolismo , Rede Nervosa/metabolismo , Estimulação Luminosa/métodos , Córtex Visual/metabolismo , Aminopiridinas/farmacologia , Animais , Feminino , Masculino , Camundongos , Microglia/química , Microglia/efeitos dos fármacos , Rede Nervosa/química , Rede Nervosa/efeitos dos fármacos , Pirróis/farmacologia , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/antagonistas & inibidores , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Córtex Visual/química , Córtex Visual/efeitos dos fármacos
6.
Nat Commun ; 11(1): 1833, 2020 04 14.
Artigo em Inglês | MEDLINE | ID: mdl-32286255

RESUMO

Small molecule inhibitor of the bromodomain and extraterminal domain (BET) family proteins is a promising option for cancer treatment. However, current BET inhibitors are limited by their potency or oral bioavailability. Here we report the discovery and characterization of NHWD-870, a BET inhibitor that is more potent than three major clinical stage BET inhibitors BMS-986158, OTX-015, and GSK-525762. NHWD-870 causes tumor shrinkage or significantly suppresses tumor growth in nine xenograft or syngeneic models. In addition to its ability to downregulate c-MYC and directly inhibit tumor cell proliferation, NHWD-870 blocks the proliferation of tumor associated macrophages (TAMs) through multiple mechanisms, partly by reducing the expression and secretion of macrophage colony-stimulating factor CSF1 by tumor cells. NHWD-870 inhibits CSF1 expression through suppressing BRD4 and its target HIF1α. Taken together, these results reveal a mechanism by which BRD4 inhibition suppresses tumor growth, and support further development of NHWD-870 to treat solid tumors.


Assuntos
Comunicação Celular , Proteínas de Ciclo Celular/antagonistas & inibidores , Macrófagos/patologia , Neoplasias/patologia , Fatores de Transcrição/antagonistas & inibidores , Administração Oral , Animais , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Modelos Animais de Doenças , Regulação para Baixo , Desenho de Fármacos , Feminino , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Fator Estimulador de Colônias de Macrófagos/metabolismo , Camundongos Endogâmicos BALB C , Camundongos Nus , Fosforilação , Proteínas Proto-Oncogênicas c-myc/metabolismo , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Transdução de Sinais , Fatores de Transcrição/metabolismo , Resultado do Tratamento
7.
Biol Pharm Bull ; 43(2): 325-333, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32009119

RESUMO

Colony stimulating factor 1 (CSF1) receptor (CSF1R) is a receptor protein-tyrosine kinase specifically expressed in monocyte-lineage cells, such as monocytes and macrophages. In this study, we characterized the pharmacological properties of an azetidine compound, JTE-952 ((2S)-3-{[2-({3-[4-(4-cyclopropylbenzyloxy)-3-methoxyphenyl]azetidine-1-yl}carbonyl)pyridin-4-yl]methoxy}propane-1,2-diol), which is a novel CSF1R tyrosine kinase inhibitor. JTE-952 potently inhibited human CSF1R kinase activity, with a half maximal inhibitory concentration of 11.1 nmol/L, and inhibited the phosphorylation of CSF1R in human macrophages and the CSF1-induced proliferation of human macrophages. It also inhibited human tropomyosin-related kinase A activity, but only at concentrations 200-fold higher than that required to inhibit the activity of CSF1R in inducing the proliferation of human macrophages. JTE-952 displayed no marked inhibitory activity against other kinases. JTE-952 potently inhibited lipopolysaccharide-induced proinflammatory cytokine production by human macrophages and in whole blood. JTE-952 (≥3 mg/kg given orally) also significantly attenuated the CSF1-induced priming of lipopolysaccharide-induced tumor necrosis factor-alpha production in mice and arthritis severity in a mouse model of collagen-induced arthritis. Taken together, these results indicate that JTE-952 is an orally available compound with potent and specific inhibitory activity against CSF1R, both in vitro and in vivo. JTE-952 is a potentially clinically useful agent for various human inflammatory diseases, including rheumatoid arthritis.


Assuntos
Azetidinas/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Animais , Artrite Experimental/tratamento farmacológico , Azetidinas/farmacocinética , Células Cultivadas , Citocinas/sangue , Citocinas/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos DBA , Ratos Endogâmicos Lew , Receptor trkA/metabolismo
8.
FASEB J ; 34(1): 1679-1694, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31914683

RESUMO

Increased pro-inflammatory cytokine levels and proliferation of activated microglia have been found in Parkinson's disease (PD) patients and animal models of PD, suggesting that targeting of the microglial inflammatory response may result in neuroprotection in PD. Microglial proliferation is regulated by many factors, but colony stimulating factor-1 receptor (CSF1R) has emerged as a primary factor. Using data mining techniques on existing microarray data, we found that mRNA expression of the CSF1R ligand, CSF-1, is increased in the brain of PD patients compared to controls. In two different neurotoxic mouse models of PD, acute MPTP and sub-chronic LPS treatment, mRNA and protein levels of CSF1R and CSF-1 were significantly increased. Treatment with the CSF1R inhibitor GW2580 significantly attenuated MPTP-induced CSF1R activation and Iba1-positive cell proliferation, without a reduction of the basal Iba1-positive population in the substantia nigra. GW2580 treatment also significantly decreased mRNA levels of pro-inflammatory factors, without alteration of anti-inflammatory mediators, and significantly attenuated the MPTP-induced loss of dopamine neurons and motor behavioral deficits. Importantly, these effects were observed in the absence of overt microglial depletion, suggesting that targeting CSF1R signaling may be a viable neuroprotective strategy in PD that disrupts pro-inflammatory signaling, but maintains the beneficial effects of microglia.


Assuntos
Anisóis/farmacologia , Anti-Inflamatórios/farmacologia , Proliferação de Células/efeitos dos fármacos , Neurônios Dopaminérgicos/efeitos dos fármacos , Microglia/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Pirimidinas/farmacologia , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Animais , Citocinas/metabolismo , Modelos Animais de Doenças , Dopamina/metabolismo , Neurônios Dopaminérgicos/metabolismo , Neurônios Dopaminérgicos/patologia , Humanos , Lipopolissacarídeos/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Macrófagos/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microglia/metabolismo , Microglia/patologia , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/metabolismo , Doença de Parkinson/patologia , Transdução de Sinais/efeitos dos fármacos , Substância Negra/efeitos dos fármacos , Substância Negra/metabolismo , Substância Negra/patologia
9.
Arterioscler Thromb Vasc Biol ; 40(3): 714-732, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31996022

RESUMO

OBJECTIVE: Calcification of atherosclerotic plaque is traditionally associated with increased cardiovascular event risk; however, recent studies have found increased calcium density to be associated with more stable disease. 3-hydroxy-3-methylglutaryl coenzymeA reductase inhibitors or statins reduce cardiovascular events. Invasive clinical studies have found that statins alter both the lipid and calcium composition of plaque but the molecular mechanisms of statin-mediated effects on plaque calcium composition remain unclear. We recently defined a macrophage Rac (Ras-related C3 botulinum toxin substrate)-IL-1ß (interleukin-1 beta) signaling axis to be a key mechanism in promoting atherosclerotic calcification and sought to define the impact of statin therapy on this pathway. Approach and Results: Here, we demonstrate that statin therapy is independently associated with elevated coronary calcification in a high-risk patient population and that statins disrupt the complex between Rac1 and its inhibitor RhoGDI (Rho GDP-dissociation inhibitor), leading to increased active (GTP bound) Rac1 in primary monocytes/macrophages. Rac1 activation is prevented by rescue with the isoprenyl precursor geranylgeranyl diphosphate. Statin-treated macrophages exhibit increased activation of NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells), increased IL-1ß mRNA, and increased Rac1-dependent IL-1ß protein secretion in response to inflammasome stimulation. Using an animal model of calcific atherosclerosis, inclusion of statin in the atherogenic diet led to a myeloid Rac1-dependent increase in atherosclerotic calcification, which was associated with increased serum IL-1ß expression, increased plaque Rac1 activation, and increased plaque expression of the osteogenic markers, alkaline phosphatase and RUNX2 (Runt-related transcription factor 2). CONCLUSIONS: Statins are capable of increasing atherosclerotic calcification through disinhibition of a macrophage Rac1-IL-1ß signaling axis.


Assuntos
Aterosclerose/tratamento farmacológico , Atorvastatina/uso terapêutico , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Macrófagos/efeitos dos fármacos , Neuropeptídeos/metabolismo , Placa Aterosclerótica , Calcificação Vascular/enzimologia , Proteínas rac1 de Ligação ao GTP/metabolismo , Idoso , Animais , Aterosclerose/enzimologia , Aterosclerose/genética , Aterosclerose/patologia , Células Cultivadas , Modelos Animais de Doenças , Humanos , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Macrófagos/enzimologia , Macrófagos/patologia , Masculino , Camundongos Knockout para ApoE , Neuropeptídeos/deficiência , Neuropeptídeos/genética , Prenilação , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/genética , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Estudos Retrospectivos , Transdução de Sinais , Calcificação Vascular/genética , Calcificação Vascular/patologia , Proteínas rac1 de Ligação ao GTP/deficiência , Proteínas rac1 de Ligação ao GTP/genética , Inibidor alfa de Dissociação do Nucleotídeo Guanina rho/metabolismo
10.
J Enzyme Inhib Med Chem ; 35(1): 311-324, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31809612

RESUMO

Hybridization of reported weakly active antiproliferative hit 5-amino-4-pyrimidinol derivative with 2-anilino-4-phenoxypyrimidines suggests a series of 2,5-diamino-4-pyrimidinol derivatives as potential antiproliferative agents. Few compounds belonging to the proposed series were reported as CSF1R/DAPK1 inhibitors as anti-tauopathies. However, the correlation between CSF1R/DAPK1 signalling pathways and cancer progression provides motives to reprofile them against cancer therapy. The compounds were synthesised, characterized, and evaluated against M-NFS-60 cells and a kinase panel which bolstered predictions of their antiproliferative activity and suggested the involvement of diverse molecular targets. Compound 6e, the most potent in the series, showed prominent broad-spectrum antiproliferative activity inhibiting the growth of hematological, NSCLC, colon, CNS, melanoma, ovarian, renal, prostate and breast cancers by 84.1, 52.79, 72.15, 66.34, 66.48, 51.55, 55.95, 61.85, and 60.87%, respectively. Additionally, it elicited an IC50 value of 1.97 µM against M-NFS-60 cells and good GIT absorption with Pe value of 19.0 ± 1.1 × 10-6 cm/s (PAMPA-GIT). Molecular docking study for 6e with CSF1R and DAPK1 was done to help to understand the binding mode with both kinases. Collectively, compound 6e could be a potential lead compound for further development of anticancer therapies.


Assuntos
Antineoplásicos/farmacologia , Proteínas Quinases Associadas com Morte Celular/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Pirimidinas/farmacologia , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/antagonistas & inibidores , Antineoplásicos/síntese química , Antineoplásicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proteínas Quinases Associadas com Morte Celular/metabolismo , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Simulação de Acoplamento Molecular , Estrutura Molecular , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Pirimidinas/síntese química , Pirimidinas/química , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Relação Estrutura-Atividade
11.
Acta Neuropathol Commun ; 7(1): 186, 2019 11 21.
Artigo em Inglês | MEDLINE | ID: mdl-31753024

RESUMO

Numerous clinical studies have established the debilitating neurocognitive side effects of chemotherapy in the treatment of breast cancer, often referred as chemobrain. We hypothesize that cognitive impairments are associated with elevated microglial inflammation in the brain. Thus, either elimination of microglia or restoration of microglial function could ameliorate cognitive dysfunction. Using a rodent model of chronic Adriamycin (ADR) treatment, a commonly used breast cancer chemotherapy, we evaluated two strategies to ameliorate chemobrain: 1) microglia depletion using the colony stimulating factor-1 receptor (CSF1R) inhibitor PLX5622 and 2) human induced pluripotent stem cell-derived microglia (iMG)-derived extracellular vesicle (EV) treatment. In strategy 1 mice received ADR once weekly for 4 weeks and were then administered CSF1R inhibitor (PLX5622) starting 72 h post-ADR treatment. ADR-treated animals given a normal diet exhibited significant behavioral deficits and increased microglial activation 4-6 weeks later. PLX5622-treated mice exhibited no ADR-related cognitive deficits and near complete depletion of IBA-1 and CD68+ microglia in the brain. Cytokine and RNA sequencing analysis for inflammation pathways validated these findings. In strategy 2, 1 week after the last ADR treatment, mice received retro-orbital vein injections of iMG-EV (once weekly for 4 weeks) and 1 week later, mice underwent behavior testing. ADR-treated mice receiving EV showed nearly complete restoration of cognitive function and significant reductions in microglial activation as compared to untreated ADR mice. Our data demonstrate that ADR treatment elevates CNS inflammation that is linked to cognitive dysfunction and that attenuation of neuroinflammation reverses the adverse neurocognitive effects of chemotherapy.


Assuntos
Antibióticos Antineoplásicos/toxicidade , Disfunção Cognitiva/metabolismo , Doxorrubicina/toxicidade , Células-Tronco Pluripotentes Induzidas/transplante , Mediadores da Inflamação/metabolismo , Compostos Orgânicos/uso terapêutico , Animais , Disfunção Cognitiva/induzido quimicamente , Disfunção Cognitiva/terapia , Humanos , Inflamação/induzido quimicamente , Inflamação/metabolismo , Inflamação/terapia , Mediadores da Inflamação/antagonistas & inibidores , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Compostos Orgânicos/farmacologia , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/antagonistas & inibidores , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo
12.
Int J Mol Sci ; 20(23)2019 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-31757032

RESUMO

Vascular targeting with pro-thrombotic antibody-conjugates is a promising biological treatment for brain arteriovenous malformations (bAVMs). However, targeted drug delivery relies on the identification of unique or overexpressed markers on the surface of a target cell. In the absence of inherent biological markers, stereotactic radiosurgery may be used to prime induction of site-specific and targetable molecular changes on the endothelial surface. To investigate lumen-accessible, endothelial targets induced by radiation, we combined Gamma knife surgery in an AVM animal model with in vivo biotin-labeling and comparative proteomics. Two proteins, αB-crystallin (CRYAB)-a small heat shock protein that normally acts as an intracellular chaperone to misfolded proteins-and activated leukocyte cell adhesion molecule CD166, were further validated for endothelial surface expression after irradiation. Immunostaining of endothelial cells in vitro and rat AVM tissue ex vivo confirmed de novo induction of CRYAB following irradiation (20 Gy). Western analysis demonstrated that CRYAB accumulated intracellularly as a 20 kDa monomer, but, at the cell surface, a novel 65 kDa protein was observed, suggesting radiation stimulates translocation of an atypical CRYAB isoform. In contrast, CD166 had relatively high expression in non-irradiated cells, localized predominantly to the lateral surfaces. Radiation increased CD166 surface exposure by inducing translocation from intercellular junctions to the apical surface without significantly altering total protein levels. These findings reinforce the dynamic molecular changes induced by radiation exposure, particularly at the cell surface, and support further investigation of radiation as a priming mechanism and these molecules as putative targets for focused drug delivery in irradiated tissue.


Assuntos
Cristalinas/metabolismo , Células Endoteliais/efeitos da radiação , Malformações Arteriovenosas Intracranianas/radioterapia , Proteínas Associadas aos Microtúbulos/metabolismo , Radiocirurgia/efeitos adversos , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Animais , Membrana Celular/metabolismo , Células Cultivadas , Células Endoteliais/metabolismo , Raios gama/efeitos adversos , Malformações Arteriovenosas Intracranianas/metabolismo , Camundongos , Transporte Proteico , Ratos , Ratos Sprague-Dawley
13.
Cell Rep ; 29(6): 1539-1554.e7, 2019 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-31693894

RESUMO

Age-related loss of skeletal muscle innervation by motor neurons leads to impaired neuromuscular function and is a well-established clinical phenomenon. However, the underlying pathogenesis remains unclear. Studying mice, we find that the number of motor units (MUs) can be maintained by counteracting neurotoxic microglia in the aged spinal cord. We observe that marked innervation changes, detected by motor unit number estimation (MUNE), occur prior to loss of muscle function in aged mice. This coincides with gene expression changes indicative of neuronal remodeling and microglial activation in aged spinal cord. Voluntary exercise prevents loss of MUs and reverses microglia activation. Depleting microglia by CSF1R inhibition also prevents the age-related decline in MUNE and neuromuscular junction disruption, implying a causal link. Our results suggest that age-related changes in spinal cord microglia contribute to neuromuscular decline in aged mice and demonstrate that removal of aged neurotoxic microglia can prevent or reverse MU loss.


Assuntos
Envelhecimento/metabolismo , Microglia/metabolismo , Neurônios Motores/metabolismo , Condicionamento Físico Animal/fisiologia , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/antagonistas & inibidores , Envelhecimento/patologia , Animais , Linhagem Celular , Bases de Dados Genéticas , Humanos , Células-Tronco Pluripotentes Induzidas , Macrófagos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microglia/enzimologia , Microglia/fisiologia , Neurônios Motores/citologia , Neurônios Motores/patologia , Músculo Esquelético/metabolismo , Músculo Esquelético/fisiopatologia , Junção Neuromuscular/metabolismo , Plasticidade Neuronal/genética , RNA-Seq , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/genética , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Medula Espinal/enzimologia , Medula Espinal/metabolismo , Medula Espinal/fisiopatologia
14.
Proc Natl Acad Sci U S A ; 116(51): 26029-26037, 2019 12 17.
Artigo em Inglês | MEDLINE | ID: mdl-31772011

RESUMO

Hypoxic preconditioning reduces disease severity in a mouse model of multiple sclerosis (MS), in part by enhancing the barrier properties of spinal cord blood vessels. Because other studies have shown that similar levels of hypoxia transiently increase permeability of central nervous system (CNS) blood vessels, the goal of this study was to define the impact of chronic mild hypoxia (CMH, 8% O2) on the integrity of spinal cord blood vessels and the responses of neighboring glial cells. Using extravascular fibrinogen as a marker of vascular disruption, we found that CMH triggered transient vascular leak in spinal cord blood vessels, particularly in white matter, which was associated with clustering and activation of Mac-1-positive microglia around disrupted vessels. Microglial depletion with the colony stimulating factor-1 receptor (CSF-1R) inhibitor PLX5622, while having no effect under normoxic conditions, profoundly increased vascular leak in both white and gray matter during CMH, and this was associated with disruption of astrocyte-vascular coupling and enhanced loss of tight junction proteins. Microglial repair of leaky blood vessels was blocked by a peptide that inhibits the interaction between fibrinogen and its Mac-1 integrin receptor. These findings highlight an important role for microglia in maintaining vascular integrity in the hypoxic spinal cord and suggest that a fibrinogen-Mac-1 interaction underpins this response. As relative hypoxia is experienced in many situations including high altitude, lung disease, obstructive sleep apnea, and age-related CNS ischemia/hypoxia, our findings have important implications regarding the critical role of microglia in maintaining vascular integrity in the CNS.


Assuntos
Hipóxia/metabolismo , Microglia/metabolismo , Esclerose Múltipla/metabolismo , Medula Espinal/irrigação sanguínea , Medula Espinal/metabolismo , Animais , Antígenos de Superfície/metabolismo , Astrócitos/metabolismo , Sistema Nervoso Central/irrigação sanguínea , Modelos Animais de Doenças , Endotélio/irrigação sanguínea , Endotélio/metabolismo , Feminino , Fibrinogênio , Substância Cinzenta/metabolismo , Isquemia/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Esclerose Múltipla/complicações , Neuroglia , Compostos Orgânicos/antagonistas & inibidores , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/efeitos dos fármacos , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Substância Branca/metabolismo
15.
Sci Adv ; 5(10): eaax7031, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31633029

RESUMO

Smoking is the largest preventable cause of death and disease in the United States. However, <5% of quit attempts are successful, underscoring the urgent need for novel therapeutics. Microglia are one untapped therapeutic target. While previous studies have shown that microglia mediate both inflammatory responses in the brain and brain plasticity, little is known regarding their role in nicotine dependence and withdrawal phenotypes. Here, we examined microglial changes in the striatum-a mesolimbic region implicated in the rewarding effects of drugs and the affective disruptions occurring during withdrawal. We show that both nicotine and withdrawal induce microglial morphological changes; however, proinflammatory effects and anxiogenic behaviors were observed only during nicotine withdrawal. Pharmacological microglial depletion during withdrawal prevented these effects. These results define differential effects of nicotine and withdrawal on inflammatory signaling in the brain, laying the groundwork for development of future smoking cessation therapeutics.


Assuntos
Microglia/patologia , Núcleo Accumbens/metabolismo , Síndrome de Abstinência a Substâncias/patologia , Animais , Ansiedade/etiologia , Modelos Animais de Doenças , Locomoção , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microglia/metabolismo , NADPH Oxidase 2/metabolismo , Nicotina/administração & dosagem , Compostos Orgânicos/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/antagonistas & inibidores , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Síndrome de Abstinência a Substâncias/complicações , Síndrome de Abstinência a Substâncias/metabolismo
16.
Technol Cancer Res Treat ; 18: 1533033819883626, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31635541

RESUMO

OBJECTIVE: The aim of this study was to evaluate the relationship between cytokine production, GM-CSF receptor (CSF2RA), and IL-1 receptor (IL1R2) expression in mammary adenocarcinoma and their association with it histopathological parameters and lymph node metastasis. METHODS: We analyzed tumor biopsy samples (cultured in vitro) from 50 women (aged 43-75) with invasive ductal mammary adenocarcinomas. Enzyme-linked immunosorbent assay method the concentrations of interleukin 2, interleukin 6, interleukin 8, interleukin 10, interleukin 17, interleukin 18, interleukin 1ß, interleukin 1Ra, tumor necrosis factor α, interferon γ, granulocyte colony-stimulating factor, granulocyte macrophage colony-stimulating factor, and vascular endothelial growth factor A were determined in culture supernatants. The expression of CSF2RA and IL1R2 in tumor biopsy was evaluated by immunohistochemical method. RESULTS: We showed that the "cytokine profile" of a tumor (the ability of tumor cells and its microenvironment to produce different cytokines) is very individual. It has been shown that the features of the cytokine profile of the mammary adenocarcinoma are important for the formation and realization of the metastatic potential of the mammary adenocarcinoma. We found correlations between some histopathological parameters of mammary adenocarcinoma and coefficients KGM-CSF/CSF2RA and KIL-1ß/IL1R2, which are the ratios of concentrations of granulocyte macrophage colony-stimulating factor and interleukin -1ß to expression of CSF2RA and IL1R2, respectively. KGM-CSF/CSF2RA positively correlated with highly differentiated cells, and KIL-1ß/IL1R2 positively correlated with the number of mitoses, poorly differentiated cells, and a number of lymph nodes with metastases. KGM-CSF/CSF2RA positively correlated with the concentrations of interleukin 6, interleukin 8, interleukin 1Ra, and granulocyte colony-stimulating factor. KIL-1ß/IL1R2 positively correlated with concentrations of interleukin 1ß and interferon γ and negative correlated with the concentrations of vascular endothelial growth factor A and tumor necrosis factor α. It is shown that KIL-1ß/IL1R2 can be considered as a prognostic indicator predicting the probability of mammary adenocarcinoma metastasis to regional lymph nodes. CONCLUSIONS: The ratios of granulocyte macrophage colony-stimulating factor and interleukin 1ß cytokines, produced in tumor, to the expression of CSF2RA and IL1R2 depend on levels of interleukin 6, interleukin 8, tumor necrosis factor α, interferon γ, granulocyte colony-stimulating factor, and vascular endothelial growth factor A and are important factors affecting the progression and metastasis of the breast cancer.


Assuntos
Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Citocinas/biossíntese , Linfonodos/patologia , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Receptores Tipo II de Interleucina-1/metabolismo , Adulto , Idoso , Biópsia , Neoplasias da Mama/genética , Análise por Conglomerados , Feminino , Expressão Gênica , Humanos , Imuno-Histoquímica , Metástase Linfática , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Curva ROC , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/genética , Receptores Tipo II de Interleucina-1/genética
17.
Front Immunol ; 10: 2055, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31552022

RESUMO

Pre-clinical models and clinical trials demonstrate that targeting the action of the cytokine, granulocyte macrophage-colony stimulating factor (GM-CSF), can be efficacious in inflammation/autoimmunity reinforcing the importance of understanding how GM-CSF functions; a significant GM-CSF-responding cell in this context is likely to be the monocyte. This article summarizes critically the literature on the downstream cellular pathways regulating GM-CSF interaction with monocytes (and macrophages), highlighting some contentious issues, and conclusions surrounding this biology. It also suggests future directions which could be undertaken so as to more fully understand this aspect of GM-CSF biology. Given the focus of this collection of articles on monocytes, the following discussion in general will be limited to this population or to its more mature progeny, the macrophage, even though GM-CSF biology is broader than this.


Assuntos
Citocinas/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/imunologia , Inflamação/imunologia , Macrófagos/imunologia , Monócitos/imunologia , Transdução de Sinais/imunologia , Autoimunidade/imunologia , Citocinas/metabolismo , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Humanos , Inflamação/metabolismo , Macrófagos/metabolismo , Monócitos/metabolismo , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/imunologia , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo
18.
J Labelled Comp Radiopharm ; 62(13): 903-908, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31524293

RESUMO

In this practitioner protocol, the radiochemical synthesis of [11 C]CPPC is described in detail, and a quality control summary of three validation productions is presented. The results indicate that the radiotracer product can be produced in good radiochemical yield (> 60 mCi (2.22 GBq) at end-of-synthesis (EOS)), at high specific activity (molar activity > 11,435 mCi/µmole (423 GBq/µmole) at EOS) and high chemical and radiochemical purity. The entire production conforms to current Good Manufacturing Practice (cGMP) requirements. The final product is formulated as a sterile, pyrogen-free solution suitable for human injection.


Assuntos
Furanos/química , Furanos/síntese química , Microglia/metabolismo , Tomografia por Emissão de Pósitrons/métodos , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Biomarcadores/metabolismo , Técnicas de Química Sintética , Controle de Qualidade , Radioquímica
19.
Brain ; 142(10): 3243-3264, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31504240

RESUMO

Neuroinflammation and microglial activation are significant processes in Alzheimer's disease pathology. Recent genome-wide association studies have highlighted multiple immune-related genes in association with Alzheimer's disease, and experimental data have demonstrated microglial proliferation as a significant component of the neuropathology. In this study, we tested the efficacy of the selective CSF1R inhibitor JNJ-40346527 (JNJ-527) in the P301S mouse tauopathy model. We first demonstrated the anti-proliferative effects of JNJ-527 on microglia in the ME7 prion model, and its impact on the inflammatory profile, and provided potential CNS biomarkers for clinical investigation with the compound, including pharmacokinetic/pharmacodynamics and efficacy assessment by TSPO autoradiography and CSF proteomics. Then, we showed for the first time that blockade of microglial proliferation and modification of microglial phenotype leads to an attenuation of tau-induced neurodegeneration and results in functional improvement in P301S mice. Overall, this work strongly supports the potential for inhibition of CSF1R as a target for the treatment of Alzheimer's disease and other tau-mediated neurodegenerative diseases.


Assuntos
Imidazóis/farmacologia , Microglia/efeitos dos fármacos , Piridinas/farmacologia , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Doença de Alzheimer/patologia , Animais , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Estudo de Associação Genômica Ampla , Humanos , Imidazóis/metabolismo , Camundongos , Camundongos Transgênicos , Microglia/fisiologia , Doenças Neurodegenerativas/tratamento farmacológico , Neurogênese , Neuroimunomodulação/efeitos dos fármacos , Neuroimunomodulação/fisiologia , Piridinas/metabolismo , Receptores de GABA/genética , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/antagonistas & inibidores , Tauopatias/tratamento farmacológico , Proteínas tau/genética
20.
Radiat Res ; 192(6): 612-620, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31560640

RESUMO

Radiation-induced acute myeloid leukemia (rAML) in C3H mice is commonly developed through inactivation of PU.1 transcription factor encoded in Sfpi1 on chromosome 2. PU.1 inactivation involves two steps: hemizygous deletion of the Sfpi1 gene (DSG) and point mutation of the allele Sfpi1 gene (PMASG). In this study, we investigated the dose-rate dependence of the frequency of both DSG and PMASG in hematopoietic stem cells (HSCs) of C3H mice that received a total of 3 Gy gamma-ray exposure at dose rates of 20 mGy/day, 200 mGy/day or 1,000 mGy/min. All mice were followed for 250 days from start of irradiation. Fluorescent in situ hybridization of the Sfpi1 gene site indicated that frequency of HSCs with DSG was proportional to dose rate. In cell surface profiles, PU.1-inactivated HSCs by both DSG and PMASG were still positive for PU.1, but negative for GM-CSF receptor-α (GMCSFRα), which is transcriptionally regulated by PU.1. Immunofluorescent staining analysis of both PU.1 and GM-CSFRα also showed dose-rate-dependent levels of PU.1-inactivated HSCs. This study provides evidence that both DSG and PMASG are dose-rate dependent; these experimental data offer new insights into the dose-rate effects in HSCs that can lead to radiation-induced leukemogenesis.


Assuntos
Células-Tronco Hematopoéticas/efeitos da radiação , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Induzida por Radiação/tratamento farmacológico , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/fisiologia , Transativadores/genética , Transativadores/fisiologia , Alelos , Animais , Carcinogênese , Membrana Celular/metabolismo , Proliferação de Células , Relação Dose-Resposta a Droga , Raios gama , Deleção de Genes , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Hibridização In Situ , Leucemia Mieloide Aguda/genética , Leucemia Induzida por Radiação/genética , Masculino , Camundongos , Camundongos Endogâmicos C3H , Mutação Puntual , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo
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