RESUMO
Adult oligodendrocyte progenitor cells (OPCs) give rise to myelinating oligodendrocytes through life and play crucial roles in brain homeostasis and plasticity during health and disease. Cannabinoid compounds acting through CB1 receptors promote the proliferation and differentiation of OPCs in vitro and facilitate developmental myelination and myelin repair in vivo. However, CB1 receptor expression in adult OPCs in situ has not been corroborated by anatomical studies and the contribution of this receptor population to the (re)myelination effects of cannabinoids remains a matter of debate. Using electron microscopy methods applied to NG2-EYFP reporter mice we assessed the localization of CB1 receptors in OPCs of the adult mouse hippocampus. To control for the specificity of CB1 receptor immunostaining we generated transgenic mice bearing EYFP expression in NG2 glia and wild-type (NG2-EYFP-CB1 +/+) and knockout (NG2-EYFP-CB1 -/-) for CB1 receptors. Double immunogold and immunoperoxidase labeling for CB1 and EYFP, respectively, revealed that CB1 receptors are present in a low proportion of NG2 positive profiles within hippocampal stratum radiatum of NG2-EYFP-CB1 +/+ mice. Quantitative analysis of immunogold particles in synaptic structures and NG2 profiles showed that CB1 receptors are expressed at lower density in adult OPCs than in glutamatergic cells of the rodent hippocampus. These results highlight the presence of CB1 receptors in adult OPCs thus providing an anatomical substrate for the remyelination promoting effects of cannabinoids and open a novel perspective on the roles of the endocannabinoid system in brain physiology through the modulation of NG2 glia.
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Cannabinoids are known to modulate oligodendrogenesis and developmental CNS myelination. However, the cell-autonomous action of these compounds on oligodendroglial cells in vivo, and the molecular mechanisms underlying these effects have not yet been studied. Here, by using oligodendroglial precursor cell (OPC)-targeted genetic mouse models, we show that cannabinoid CB1 receptors exert an essential role in modulating OPC differentiation at the critical periods of postnatal myelination. We found that selective genetic inactivation of CB1 receptors in OPCs in vivo perturbs oligodendrogenesis and postnatal myelination by altering the RhoA/ROCK signaling pathway, leading to hypomyelination, and motor and cognitive alterations in young adult mice. Conversely, pharmacological CB1 receptor activation, by inducing E3 ubiquitin ligase-dependent RhoA proteasomal degradation, promotes oligodendrocyte development and CNS myelination in OPCs, an effect that was not evident in OPC-specific CB1 receptor-deficient mice. Moreover, pharmacological inactivation of ROCK in vivo overcomes the defects in oligodendrogenesis and CNS myelination, and behavioral alterations found in OPC-specific CB1 receptor-deficient mice. Overall, this study supports a cell-autonomous role for CB1 receptors in modulating oligodendrogenesis in vivo, which may have a profound impact on the scientific knowledge and therapeutic manipulation of CNS myelination by cannabinoids.
Assuntos
Canabinoides , Células Precursoras de Oligodendrócitos , Receptor CB1 de Canabinoide , Animais , Canabinoides/farmacologia , Diferenciação Celular/fisiologia , Inativação Gênica , Camundongos , Bainha de Mielina/metabolismo , Células Precursoras de Oligodendrócitos/metabolismo , Oligodendroglia/metabolismo , Receptor CB1 de Canabinoide/metabolismoRESUMO
BACKGROUND AND PURPOSE: Research on demyelinating disorders aims to find novel molecules that are able to induce oligodendrocyte precursor cell differentiation to promote central nervous system remyelination and functional recovery. Δ9 -Tetrahydrocannabinol (THC), the most prominent active constituent of the hemp plant Cannabis sativa, confers neuroprotection in animal models of demyelination. However, the possible effect of THC on myelin repair has never been studied. EXPERIMENTAL APPROACH: By using oligodendroglia-specific reporter mouse lines in combination with two models of toxin-induced demyelination, we analysed the effect of THC on the processes of oligodendrocyte regeneration and functional remyelination. KEY RESULTS: We show that THC administration enhanced oligodendrocyte regeneration, white matter remyelination and motor function recovery. THC also promoted axonal remyelination in organotypic cerebellar cultures. THC remyelinating action relied on the induction of oligodendrocyte precursor differentiation upon cell cycle exit and via CB1 cannabinoid receptor activation. CONCLUSIONS AND IMPLICATIONS: Overall, our study identifies THC administration as a promising pharmacological strategy aimed to promote functional CNS remyelination in demyelinating disorders.
Assuntos
Doenças Desmielinizantes , Remielinização , Substância Branca , Animais , Diferenciação Celular , Doenças Desmielinizantes/induzido quimicamente , Doenças Desmielinizantes/tratamento farmacológico , Dronabinol/farmacologia , Camundongos , OligodendrogliaRESUMO
Δ9 -Tetrahydrocannabinol (THC), the main bioactive compound found in the plant Cannabis sativa, exerts its effects by activating cannabinoid receptors present in many neural cells. Cannabinoid receptors are also physiologically engaged by endogenous cannabinoid compounds, the so-called endocannabinoids. Specifically, the endocannabinoid 2-arachidonoylglycerol has been highlighted as an important modulator of oligodendrocyte (OL) development at embryonic stages and in animal models of demyelination. However, the potential impact of THC exposure on OL lineage progression during the critical periods of postnatal myelination has never been explored. Here, we show that acute THC administration at early postnatal ages in mice enhanced OL development and CNS myelination in the subcortical white matter by promoting oligodendrocyte precursor cell cycle exit and differentiation. Mechanistically, THC-induced-myelination was mediated by CB1 and CB2 cannabinoid receptors, as demonstrated by the blockade of THC actions by selective receptor antagonists. Moreover, the THC-mediated modulation of oligodendroglial differentiation relied on the activation of the mammalian target of rapamycin complex 1 (mTORC1) signaling pathway, as mTORC1 pharmacological inhibition prevented the THC effects. Our study identifies THC as an effective pharmacological strategy to enhance oligodendrogenesis and CNS myelination in vivo.
Assuntos
Dronabinol , Endocanabinoides , Animais , Dronabinol/farmacologia , Alvo Mecanístico do Complexo 1 de Rapamicina , Camundongos , Oligodendroglia , Receptores de CanabinoidesRESUMO
Recessive dystrophic epidermolysis bullosa (RDEB) is a severe skin disease caused by mutation of the COL7A1 gene. RDEB is associated with high levels of TGF-ß1, which is likely to be involved in the fibrosis that develops in this disease. Endoglin (CD105) is a type III coreceptor for TGF-ß1 and its overexpression in fibroblasts deregulates physiological Smad/Alk1/Alk5 signalling, repressing the synthesis of TGF-ß1 and extracellular matrix (ECM) proteins. Raloxifene is a specific estrogen receptor modulator designated as an orphan drug for hereditary hemorrhagic telangiectasia, a rare vascular disease. Raloxifene stimulates endoglin synthesis, which could attenuate fibrosis. By contrast, the antioxidant N-acetylcysteine may have therapeutic value to rectify inflammation, fibrosis and endothelial dysfunction. Thus, we present here a repurposing strategy based on the molecular and functional screening of fibroblasts from RDEB patients with these drugs, leading us to propose the repositioning of these two well-known drugs currently in clinical use, raloxifene and N-acetylcysteine, to counteract fibrosis and inflammation in RDEB. Both compounds modulate the profibrotic events that may ultimately be responsible for the clinical manifestations in RDEB, suggesting that these findings may also be relevant for other diseases in which fibrosis is an important pathophysiological event.
Assuntos
Acetilcisteína/farmacologia , Reposicionamento de Medicamentos , Epidermólise Bolhosa/genética , Fibroblastos/efeitos dos fármacos , Cloridrato de Raloxifeno/farmacologia , Fator de Crescimento Transformador beta1/genética , Receptores de Activinas Tipo II/genética , Receptores de Activinas Tipo II/metabolismo , Antioxidantes/farmacologia , Estudos de Casos e Controles , Colágeno Tipo VII/genética , Colágeno Tipo VII/metabolismo , Endoglina/genética , Endoglina/metabolismo , Epidermólise Bolhosa/metabolismo , Epidermólise Bolhosa/patologia , Antagonistas de Estrogênios/farmacologia , Proteínas da Matriz Extracelular/genética , Proteínas da Matriz Extracelular/metabolismo , Fibroblastos/metabolismo , Fibroblastos/patologia , Fibrose , Regulação da Expressão Gênica , Humanos , Padrões de Herança , Cultura Primária de Células , Receptor do Fator de Crescimento Transformador beta Tipo I/genética , Receptor do Fator de Crescimento Transformador beta Tipo I/metabolismo , Índice de Gravidade de Doença , Transdução de Sinais , Pele/efeitos dos fármacos , Pele/metabolismo , Pele/patologia , Proteínas Smad/genética , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta1/metabolismoRESUMO
Von Hippel-Lindau (VHL), is a rare autosomal dominant inherited cancer in which the lack of VHL protein triggers the development of multisystemic tumors such us retinal hemangioblastomas (HB), CNS-HB, and clear cell renal cell carcinoma (ccRCC). ccRCC ranks third in terms of incidence and first in cause of death. Standard systemic therapies for VHL-ccRCC have shown limited response, with recurrent surgeries being the only effective treatment. Targeting of ß2-adrenergic receptor (ADRB) has shown therapeutic antitumor benefits on VHL-retinal HB (clinical trial) and VHL-CNS HB (in vitro). Therefore, the in vitro and in vivo antitumor benefits of propranolol (ADRB-1,2 antagonist) and ICI-118,551 (ADRB-2 antagonist) on VHL-/- ccRCC primary cultures and 786-O tumor cell lines have been addressed. Propranolol and ICI-118,551 activated apoptosis inhibited gene and protein expression of HIF-2α, CAIX, and VEGF, and impaired partially the nuclear internalization of HIF-2α and NFĸB/p65. Moreover, propranolol and ICI-118,551 reduced tumor growth on two in vivo xenografts. Finally, ccRCC patients receiving propranolol as off-label treatment have shown a positive therapeutic response for two years on average. In summary, propranolol and ICI-118,551 have shown antitumor benefits in VHL-derived ccRCC, and since ccRCCs comprise 63% of the total RCCs, targeting ADRB2 becomes a promising drug for VHL and other non-VHL tumors.
RESUMO
Phenotypic drug discovery must take advantage of the large amount of clinical data currently available. In this sense, the impact of microRNAs (miRs) on human disease and clinical therapeutic responses is becoming increasingly well documented. Accordingly, it might be possible to use miR-based signatures as phenotypic read-outs of pathological status, for example in cancer. Here, we propose to use the information accumulating regarding the biology of miRs from clinical research in the preclinical arena, adapting it to the use of miR biosensors in the earliest steps of drug screening. Thus, we have used an amperometric dual magnetosensor capable of monitoring a miR-21/miR-205 signature to screen for new drugs that restore these miRs to non-tumorigenic levels in cell models of breast cancer and glioblastoma. In this way we have been able to identify a new chemical entity, 11PS04 ((3aR,7aS)-2-(3-propoxyphenyl)-7,7a-dihydro-3aH-pyrano[3,4-d]oxazol-6(4H)-one), the therapeutic potential of which was suggested in mechanistic assays of disease models, including 3D cell culture (oncospheres) and xenografts. These assays highlighted the potential of this compound to attack cancer stem cells, reducing the growth of breast and glioblastoma tumors in vivo. These data demonstrate the enhanced chain of translatability of this strategy, opening up new perspectives for drug-discovery pipelines and highlighting the potential of miR-based electro-analytical sensors as efficient tools in modern drug discovery.
Assuntos
Técnicas Biossensoriais , MicroRNAs/metabolismo , Células-Tronco Neoplásicas/patologia , Oxazóis/farmacologia , Animais , Antineoplásicos/farmacologia , Carcinogênese/efeitos dos fármacos , Carcinogênese/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glioma/patologia , Fenômenos Magnéticos , Glândulas Mamárias Animais/efeitos dos fármacos , Glândulas Mamárias Animais/patologia , Camundongos , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Oxazóis/química , Reprodutibilidade dos Testes , Esferoides Celulares/efeitos dos fármacos , Esferoides Celulares/metabolismo , Esferoides Celulares/patologia , Temozolomida/farmacologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Controlled delivery of multiple chemotherapeutics can improve the effectiveness of treatments and reduce side effects and relapses. Here in, we used albumin-stabilized gold nanoclusters modified with doxorubicin and SN38 (AuNCs-DS) as combined therapy for cancer. The chemotherapeutics are conjugated to the nanostructures using linkers that release them when exposed to different internal stimuli (Glutathione and pH). This system has shown potent antitumor activity against breast and pancreatic cancer cells. Our studies indicate that the antineoplastic activity observed may be related to the reinforced DNA damage generated by the combination of the drugs. Moreover, this system presented antineoplastic activity against mammospheres, a culturing model for cancer stem cells, leading to an efficient reduction of the number of oncospheres and their size. In summary, the nanostructures reported here are promising carriers for combination therapy against cancer and particularly to cancer stem cells.
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Induced pluripotent stem cells (iPSCs) can be differentiated in vitro and in vivo to all cardiovascular lineages and are therefore a promising cell source for cardiac regenerative therapy. However, iPSC lines do not all differentiate into cardiomyocytes (CMs) with the same efficiency. Here, we show that telomerase-competent iPSCs with relatively long telomeres and high expression of the shelterin-complex protein TRF1 (iPSChighT ) differentiate sooner and more efficiently into CMs than those with relatively short telomeres and low TRF1 expression (iPSClowT ). Ascorbic acid, an enhancer of cardiomyocyte differentiation, further increases the cardiomyocyte yield from iPSChighT but does not rescue the cardiomyogenic potential of iPSClowT . Interestingly, although iPSCslowT differentiate very poorly to the mesoderm and endoderm lineages, they differentiate very efficiently to the ectoderm lineage, indicating that cell fate can be determined by in vitro selection of iPSCs with different telomere content. Our findings highlight the importance of selecting iPSCs with ample telomere reserves in order to generate high numbers of CMs in a fast, reliable, and efficient way. Stem Cells 2017;35:362-373.
Assuntos
Diferenciação Celular , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Miócitos Cardíacos/citologia , Miócitos Cardíacos/metabolismo , Homeostase do Telômero , Animais , Ácido Ascórbico/farmacologia , Diferenciação Celular/efeitos dos fármacos , Linhagem da Célula/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Tamanho Celular/efeitos dos fármacos , Colágeno/metabolismo , Corpos Embrioides/citologia , Corpos Embrioides/metabolismo , Camundongos , Miócitos Cardíacos/efeitos dos fármacos , Homeostase do Telômero/efeitos dos fármacosRESUMO
The molecular mechanisms that drive mammalian cardiomyocytes out of the cell cycle soon after birth remain largely unknown. Here, we identify telomere dysfunction as a critical physiological signal for cardiomyocyte cell-cycle arrest. We show that telomerase activity and cardiomyocyte telomere length decrease sharply in wild-type mouse hearts after birth, resulting in cardiomyocytes with dysfunctional telomeres and anaphase bridges and positive for the cell-cycle arrest protein p21. We further show that premature telomere dysfunction pushes cardiomyocytes out of the cell cycle. Cardiomyocytes from telomerase-deficient mice with dysfunctional telomeres (G3 Terc(-/-)) show precocious development of anaphase-bridge formation, p21 up-regulation, and binucleation. In line with these findings, the cardiomyocyte proliferative response after cardiac injury was lost in G3 Terc(-/-) newborns but rescued in G3 Terc(-/-)/p21(-/-) mice. These results reveal telomere dysfunction as a crucial signal for cardiomyocyte cell-cycle arrest after birth and suggest interventions to augment the regeneration capacity of mammalian hearts.
Assuntos
Pontos de Checagem do Ciclo Celular , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Miócitos Cardíacos/citologia , Miócitos Cardíacos/metabolismo , Telômero/metabolismo , Anáfase , Animais , Animais Recém-Nascidos , Proliferação de Células , Dano ao DNA , Reparo do DNA , Camundongos Endogâmicos C57BL , Modelos Biológicos , Telomerase/metabolismo , Homeostase do TelômeroRESUMO
After myocardial infarction in humans, lost cardiomyocytes are replaced by an irreversible fibrotic scar. In contrast, zebrafish hearts efficiently regenerate after injury. Complete regeneration of the zebrafish heart is driven by the strong proliferation response of its cardiomyocytes to injury. Here we show that, after cardiac injury in zebrafish, telomerase becomes hyperactivated, and telomeres elongate transiently, preceding a peak of cardiomyocyte proliferation and full organ recovery. Using a telomerase-mutant zebrafish model, we found that telomerase loss drastically decreases cardiomyocyte proliferation and fibrotic tissue regression after cryoinjury and that cardiac function does not recover. The impaired cardiomyocyte proliferation response is accompanied by the absence of cardiomyocytes with long telomeres and an increased proportion of cardiomyocytes showing DNA damage and senescence characteristics. These findings demonstrate the importance of telomerase function in heart regeneration and highlight the potential of telomerase therapy as a means of stimulating cell proliferation upon myocardial infarction.
Assuntos
Coração/fisiologia , Regeneração , Telomerase/fisiologia , Proteínas de Peixe-Zebra/fisiologia , Animais , Proliferação de Células , Expressão Gênica , Técnicas de Inativação de Genes , Miocárdio/enzimologia , Miócitos Cardíacos/fisiologia , Técnicas de Cultura de Tecidos , Peixe-ZebraRESUMO
The CB1 cannabinoid receptor regulates cortical progenitor proliferation during embryonic development, but the molecular mechanism of this action remains unknown. Here, we report that CB1-deficient mouse embryos show premature cell cycle exit, decreased Pax6- and Tbr2-positive cell number, and reduced mammalian target of rapamycin complex 1 (mTORC1) activation in the ventricular and subventricular cortical zones. Pharmacological stimulation of the CB1 receptor in cortical slices and progenitor cell cultures activated the mTORC1 pathway and increased the number of Pax6- and Tbr2-expressing cells. Likewise, acute CB1 knockdown in utero reduced mTORC1 activation and cannabinoid-induced Tbr2-positive cell generation. Luciferase reporter and chromatin immunoprecipitation assays revealed that the CB1 receptor drives Tbr2 expression downstream of Pax6 induction in an mTORC1-dependent manner. Altogether, our results demonstrate that the CB1 receptor tunes dorsal telencephalic progenitor proliferation by sustaining the transcriptional activity of the Pax6-Tbr2 axis via the mTORC1 pathway, and suggest that alterations of CB1 receptor signaling, by producing the missexpression of progenitor identity determinants may contribute to neurodevelopmental alterations.
Assuntos
Córtex Cerebral , Regulação da Expressão Gênica no Desenvolvimento/genética , Receptor CB1 de Canabinoide/metabolismo , Transdução de Sinais/genética , Células-Tronco/fisiologia , Proteínas com Domínio T/metabolismo , Animais , Animais Recém-Nascidos , Técnicas de Cultura de Células , Córtex Cerebral/citologia , Córtex Cerebral/embriologia , Córtex Cerebral/crescimento & desenvolvimento , Embrião de Mamíferos , Proteínas do Olho/genética , Proteínas do Olho/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Antígeno Ki-67/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina , Camundongos , Camundongos Transgênicos , Complexos Multiproteicos/genética , Complexos Multiproteicos/metabolismo , Mutação/genética , Proteínas do Tecido Nervoso/metabolismo , Técnicas de Cultura de Órgãos , Fator de Transcrição PAX6 , Fatores de Transcrição Box Pareados/genética , Fatores de Transcrição Box Pareados/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Receptor CB1 de Canabinoide/genética , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Proteínas com Domínio T/genética , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismoRESUMO
The generation and specification of pyramidal neuron subpopulations during development relies on a complex network of transcription factors. The CB(1) cannabinoid receptor is the major molecular target of endocannabinoids and marijuana active compounds. This receptor has been shown to influence neural progenitor proliferation and axonal growth, but its involvement in neuronal differentiation and the functional impact in the adulthood caused by altering its signaling during brain development are not known. Here we show that the CB(1) receptor, by preventing Satb2 (special AT-rich binding protein 2)-mediated repression, increased Ctip2 (COUP-TF interacting protein 2) promoter activity, and Ctip2-positive neuron generation. Unbalanced neurogenic fate determination found in complete CB(1)(-/-) mice and in glutamatergic neuron-specific Nex-CB(1)(-/-) mice induced overt alterations in corticospinal motor neuron generation and subcerebral connectivity, thereby resulting in an impairment of skilled motor function in adult mice. Likewise, genetic deletion of CB(1) receptors in Thy1-YFP-H mice elicited alterations in corticospinal tract development. Altogether, these data demonstrate that the CB(1) receptor contributes to the generation of deep-layer cortical neurons by coupling endocannabinoid signals from the neurogenic niche to the intrinsic proneurogenic Ctip2/Satb2 axis, thus influencing appropriate subcerebral projection neuron specification and corticospinal motor function in the adulthood.
Assuntos
Diferenciação Celular/fisiologia , Proteínas de Ligação à Região de Interação com a Matriz/fisiologia , Neurônios Motores/fisiologia , Células Piramidais/fisiologia , Tratos Piramidais/fisiologia , Receptor CB1 de Canabinoide/fisiologia , Proteínas Repressoras/fisiologia , Fatores de Transcrição/fisiologia , Proteínas Supressoras de Tumor/fisiologia , Animais , Comportamento Animal/fisiologia , Proliferação de Células , Células Cultivadas , Imunofluorescência , Regulação da Expressão Gênica/genética , Regulação da Expressão Gênica/fisiologia , Imuno-Histoquímica , Hibridização In Situ , Camundongos , Camundongos Knockout , Microscopia Confocal , Regiões Promotoras Genéticas/genética , Regiões Promotoras Genéticas/fisiologia , Proteína Quinase C/metabolismo , Tratos Piramidais/citologia , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Endocannabinoids act as neuromodulatory and neuroprotective cues by engaging type 1 cannabinoid receptors. These receptors are highly abundant in the basal ganglia and play a pivotal role in the control of motor behaviour. An early downregulation of type 1 cannabinoid receptors has been documented in the basal ganglia of patients with Huntington's disease and animal models. However, the pathophysiological impact of this loss of receptors in Huntington's disease is as yet unknown. Here, we generated a double-mutant mouse model that expresses human mutant huntingtin exon 1 in a type 1 cannabinoid receptor-null background, and found that receptor deletion aggravates the symptoms, neuropathology and molecular pathology of the disease. Moreover, pharmacological administration of the cannabinoid Δ(9)-tetrahydrocannabinol to mice expressing human mutant huntingtin exon 1 exerted a therapeutic effect and ameliorated those parameters. Experiments conducted in striatal cells show that the mutant huntingtin-dependent downregulation of the receptors involves the control of the type 1 cannabinoid receptor gene promoter by repressor element 1 silencing transcription factor and sensitizes cells to excitotoxic damage. We also provide in vitro and in vivo evidence that supports type 1 cannabinoid receptor control of striatal brain-derived neurotrophic factor expression and the decrease in brain-derived neurotrophic factor levels concomitant with type 1 cannabinoid receptor loss, which may contribute significantly to striatal damage in Huntington's disease. Altogether, these results support the notion that downregulation of type 1 cannabinoid receptors is a key pathogenic event in Huntington's disease, and suggest that activation of these receptors in patients with Huntington's disease may attenuate disease progression.
Assuntos
Corpo Estriado/metabolismo , Doença de Huntington/genética , Neurônios/metabolismo , Receptor CB1 de Canabinoide/genética , Análise de Variância , Animais , Western Blotting , Sobrevivência Celular , Dronabinol/farmacologia , Hormônio Liberador de Hormônio do Crescimento/análogos & derivados , Doença de Huntington/metabolismo , Imageamento por Ressonância Magnética , Masculino , Camundongos , Camundongos Transgênicos , Atividade Motora/efeitos dos fármacos , Receptor CB1 de Canabinoide/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Teste de Desempenho do Rota-RodRESUMO
Cannabinoid-derived drugs are promising agents for the development of novel neuroprotective strategies. Activation of neuronal CB(1) cannabinoid receptors attenuates excitotoxic glutamatergic neurotransmission, triggers prosurvival signalling pathways and palliates motor symptoms in animal models of neurodegenerative disorders. However, in Huntington's disease there is a very early downregulation of CB(1) receptors in striatal neurons that, together with the undesirable psychoactive effects triggered by CB(1) receptor activation, foster the search for alternative pharmacological treatments. Here, we show that CB(2) cannabinoid receptor expression increases in striatal microglia of Huntington's disease transgenic mouse models and patients. Genetic ablation of CB(2) receptors in R6/2 mice, that express human mutant huntingtin exon 1, enhanced microglial activation, aggravated disease symptomatology and reduced mice lifespan. Likewise, induction of striatal excitotoxicity in CB(2) receptor-deficient mice by quinolinic acid administration exacerbated brain oedema, microglial activation, proinflammatory-mediator state and medium-sized spiny neuron degeneration. Moreover, administration of CB(2) receptor-selective agonists to wild-type mice subjected to excitotoxicity reduced neuroinflammation, brain oedema, striatal neuronal loss and motor symptoms. Studies on ganciclovir-induced depletion of astroglial proliferation in transgenic mice expressing thymidine kinase under the control of the glial fibrillary acidic protein promoter excluded the participation of proliferating astroglia in CB(2) receptor-mediated actions. These findings support a pivotal role for CB(2) receptors in attenuating microglial activation and preventing neurodegeneration that may pave the way to new therapeutic strategies for neuroprotection in Huntington's disease as well as in other neurodegenerative disorders with a significant excitotoxic component.
Assuntos
Doença de Huntington , Microglia/metabolismo , Fármacos Neuroprotetores/metabolismo , Receptor CB2 de Canabinoide/metabolismo , Animais , Antibacterianos/farmacologia , Biomarcadores/metabolismo , Corpo Estriado/citologia , Corpo Estriado/efeitos dos fármacos , Corpo Estriado/metabolismo , Corpo Estriado/patologia , Humanos , Proteína Huntingtina , Doença de Huntington/metabolismo , Doença de Huntington/patologia , Doença de Huntington/fisiopatologia , Imageamento por Ressonância Magnética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Minociclina/farmacologia , Degeneração Neural/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Ácido Quinolínico/farmacologia , Receptor CB2 de Canabinoide/genética , Teste de Desempenho do Rota-Rod , Convulsões/fisiopatologiaRESUMO
During brain development, functional neurogenesis is achieved by the concerted action of various steps that include the expansion of progenitor cells, neuronal specification, and establishment of appropriate synapses. Brain patterning and regionalization is regulated by a variety of extracellular signals and morphogens that, together with neuronal activity, orchestrate and regulate progenitor proliferation, differentiation, and neuronal maturation. In the adult brain, CB(1) cannabinoid receptors are expressed at very high levels in selective areas and are engaged by endocannabinoids, which act as retrograde messengers controlling neuronal function and preventing excessive synaptic activity. In addition, the endocannabinoid system is present at early developmental stages of nervous system formation. Recent studies have provided novel information on the role of this endogenous neuromodulatory system in the control of neuronal specification and maturation. Thus, cannabinoid receptors and locally produced endocannabinoids regulate neural progenitor proliferation and pyramidal specification of projecting neurons. CB(1) receptors also control axonal navigation, migration, and positioning of interneurons and excitatory neurons. Loss of function studies by genetic ablation or pharmacological blockade of CB(1) receptors interferes with long-range subcortical projections and, likewise, prenatal cannabinoid exposure induces different functional alterations in the adult brain. Potential implications of these new findings, such as the participation of the endocannabinoid system in the pathogenesis of neurodevelopmental disorders (e.g., schizophrenia) and the regulation of neurogenesis in brain depression, are discussed herein.
Assuntos
Encéfalo , Moduladores de Receptores de Canabinoides/fisiologia , Endocanabinoides , Neurogênese/fisiologia , Transtornos Psicóticos , Animais , Encéfalo/citologia , Encéfalo/crescimento & desenvolvimento , Encéfalo/fisiopatologia , Proliferação de Células , Cognição/fisiologia , Humanos , Transtornos Psicóticos/metabolismo , Transtornos Psicóticos/fisiopatologia , Receptores de Canabinoides/fisiologia , Transdução de Sinais/fisiologia , Células-Tronco/fisiologiaRESUMO
Endocannabinoids act as retrograde messengers that, by inhibiting neurotransmitter release via presynaptic CB(1) cannabinoid receptors, regulate the functionality of many synapses. In addition, the endocannabinoid system participates in the control of neuron survival. Thus, CB(1) receptor activation has been shown to protect neurons from acute brain injury as well as in neuroinflammatory conditions and neurodegenerative diseases. Nonetheless, some studies have reported that cannabinoids can also exert neurotoxic actions. Cannabinoid neuroprotective activity relies on the inhibition of glutamatergic neurotransmission and on other various mechanisms, and is supported by the observation that the brain overproduces endocannabinoids upon damage. Coupling of neuronal CB(1) receptors to cell survival routes such as the phosphatidylinositol 3-kinase/Akt and extracellular signal-regulated kinase pathways may contribute to cannabinoid neuroprotective action. These pro-survival signals occur, at least in part, by the cross-talk between CB(1) receptors and growth factor tyrosine kinase receptors. Besides promoting neuroprotection, a role for the endocannabinoid system in the control of neurogenesis from neural progenitors has been put forward. In addition, activation of CB(2) cannabinoid receptors on glial cells may also participate in neuroprotection by limiting the extent of neuroinflammation. Altogether, these findings support that endocannabinoids constitute a new family of lipid mediators that act as instructive signals in the control of neuron survival.
Assuntos
Moduladores de Receptores de Canabinoides/metabolismo , Endocanabinoides , Neurônios/metabolismo , Receptor CB1 de Canabinoide/metabolismo , Transdução de Sinais , Animais , Sobrevivência Celular/fisiologia , Humanos , Doenças Neurodegenerativas/fisiopatologia , Neuroglia/metabolismo , Receptores Proteína Tirosina Quinases/metabolismoRESUMO
Endocannabinoids (eCBs) have recently been identified as axon guidance cues shaping the connectivity of local GABAergic interneurons in the developing cerebrum. However, eCB functions during pyramidal cell specification and establishment of long-range axonal connections are unknown. Here, we show that eCB signaling is operational in subcortical proliferative zones from embryonic day 12 in the mouse telencephalon and controls the proliferation of pyramidal cell progenitors and radial migration of immature pyramidal cells. When layer patterning is accomplished, developing pyramidal cells rely on eCB signaling to initiate the elongation and fasciculation of their long-range axons. Accordingly, CB(1) cannabinoid receptor (CB(1)R) null and pyramidal cell-specific conditional mutant (CB(1)R(f/f,NEX-Cre)) mice develop deficits in neuronal progenitor proliferation and axon fasciculation. Likewise, axonal pathfinding becomes impaired after in utero pharmacological blockade of CB(1)Rs. Overall, eCBs are fundamental developmental cues controlling pyramidal cell development during corticogenesis.
Assuntos
Axônios/metabolismo , Moduladores de Receptores de Canabinoides/metabolismo , Endocanabinoides , Células Piramidais/metabolismo , Transdução de Sinais , Animais , Antagonistas de Receptores de Canabinoides , Diferenciação Celular , Feminino , Humanos , Camundongos , Camundongos Transgênicos , Gravidez , Células Piramidais/citologia , Receptores de Canabinoides/genética , Receptores de Canabinoides/metabolismoRESUMO
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.
Assuntos
Movimento Celular , Esclerose Múltipla/metabolismo , Esclerose Múltipla/patologia , Células Progenitoras Mieloides/citologia , Células Progenitoras Mieloides/metabolismo , Receptor CB2 de Canabinoide/metabolismo , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Células Cultivadas , Modelos Animais de Doenças , Regulação da Expressão Gênica , Humanos , Camundongos , Camundongos Knockout , Esclerose Múltipla/genética , Receptor CB2 de Canabinoide/deficiência , Receptor CB2 de Canabinoide/genética , Medula Espinal/citologia , Medula Espinal/metabolismoRESUMO
Cannabinoids, the active components of Cannabis sativa L., act in the body by mimicking endogenous substances--the endocannabinoids--that activate specific cell surface receptors. Cannabinoids exert various palliative effects in cancer patients. In addition, cannabinoids inhibit the growth of different types of tumor cells, including glioma cells, in laboratory animals. They do so by modulating key cell signaling pathways, mostly the endoplasmic reticulum stress response, thereby inducing antitumoral actions such as the apoptotic death of tumor cells and the inhibition of tumor angiogenesis. Of interest, cannabinoids seem to be selective antitumoral compounds, as they kill glioma cells, but not their non-transformed astroglial counterparts. On the basis of these preclinical findings, a pilot clinical study of Delta(9)-tetrahydrocannabinol (THC) in patients with recurrent glioblastoma multiforme has been recently run. The good safety profile of THC, together with its possible growth-inhibiting action on tumor cells, justifies the setting up of future trials aimed at evaluating the potential antitumoral activity of cannabinoids.