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1.
Mol Oncol ; 13(7): 1589-1604, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31162799

RESUMO

Activation of the cyclic adenosine monophosphate/protein kinase A (cAMP/PKA) pathway induces glial differentiation of glioblastoma (GBM) cells, but the mechanism by which microRNA (miRNA) regulate this process remains poorly understood. In this study, by performing miRNA genomics and loss- and gain-of-function assays in dibutyryl-cAMP-treated GBM cells, we identified a critical negative regulator, hsa-miR-1275, that modulates a set of genes involved in cancer progression, stem cell maintenance, and cell maturation and differentiation. Additionally, we confirmed that miR-1275 directly and negatively regulates the protein expression of glial fibrillary acidic protein (GFAP), a marker of mature astrocytes. Of note, tri-methyl-histone H3 (Lys27) (H3K27me3), downstream of the PKA/polycomb repressive complex 2 (PRC2) pathway, accounts for the downregulation of miR-1275. Furthermore, decreased miR-1275 expression and induction of GFAP expression were also observed in dibutyryl-cAMP-treated primary cultured GBM cells. In a patient-derived glioma stem cell tumor model, a cAMP elevator and an inhibitor of H3K27me3 methyltransferase inhibited tumor growth, induced differentiation, and reduced expression of miR-1275. In summary, our study shows that epigenetic inhibition of miR-1275 by the cAMP/PKA/PRC2/H3K27me3 pathway mediates glial induction of GBM cells, providing a new mechanism and novel targets for differentiation-inducing therapy.

2.
Mar Drugs ; 17(5)2019 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-31060323

RESUMO

High intraocular pressure (IOP)-induced retinal ischemia leads to acute glaucoma, which is one of the leading causes of irreversible visual-field loss, characterized by loss of retinal ganglion cells (RGCs) and axonal injury in optic nerves (ONs). Oxidative stress and the inflammatory response play an important role in the ischemic injury of retinal and optic nerves. We focus on 5α-androst-3ß, 5α, 6ß-triol (TRIOL), a synthetic neuroactive derivative of natural marine steroids 24-methylene-cholest-3ß, 5α, 6ß, 19-tetrol and cholestane-3ß, 5α, 6ß-triol, which are two neuroactive polyhydroxysterols isolated from the soft coral Nephthea brassica and the gorgonian Menella kanisa, respectively. We previously demonstrated that TRIOL was a neuroprotective steroid with anti-inflammatory and antioxidative activities. However, the potential role of TRIOL on acute glaucoma and its underlying mechanisms remains unclear. Here, we report TRIOL as a promising neuroprotectant that can protect RGCs and their axons/dendrites from ischemic-reperfusion (I/R) injury in an acute intraocular hypertension (AIH) model. Intravitreal injection of TRIOL significantly alleviated the loss of RGCs and the damage of axons and dendrites in rats and mice with acute glaucoma. As NF-E2-related factor 2 (Nrf2) is one of the most critical regulators in oxidative and inflammatory injury, we further evaluated the effect of TRIOL on Nrf2 knockout mice, and the neuroprotective role of TRIOL on retinal ischemia was not observed in Nrf2 knockout mice, indicating that activation of Nrf2 is responsible for the neuroprotection of TRIOL. Further experiments demonstrated that TRIOL can activate and upregulate Nrf2, along with its downstream hemeoxygenase-1 (HO-1), by negative regulation of Kelch-like ECH (Enoyl-CoA Hydratase) associated Protein-1 (Keap1). In conclusion, our study shed new light on the neuroprotective therapy of retinal ischemia and proposed a promising marine drug candidate, TRIOL, for the therapeutics of acute glaucoma.


Assuntos
Androstanóis/farmacologia , Fator 2 Relacionado a NF-E2/deficiência , Fármacos Neuroprotetores/farmacologia , Traumatismo por Reperfusão/tratamento farmacológico , Células Ganglionares da Retina/efeitos dos fármacos , Esteroides/farmacologia , Animais , Técnicas de Cultura de Células , Hipóxia Celular/efeitos dos fármacos , Modelos Animais de Doenças , Glaucoma , Heme Oxigenase-1/metabolismo , Inflamação/tratamento farmacológico , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microglia/efeitos dos fármacos , Microglia/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Hipertensão Ocular/tratamento farmacológico , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley
3.
Cell Death Dis ; 10(5): 358, 2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-31043589

RESUMO

Given that glioma stem cells (GSCs) play a critical role in the initiation and chemoresistance in glioblastoma multiforme (GBM), targeting GSCs is an attractive strategy to treat GBM. Utilizing an anti-cancer compound library, we identified R406, the active metabolite of a FDA-approved Syk inhibitor for immune thrombocytopenia (ITP), with remarkable cytotoxicity against GSCs but not normal neural stem cells. R406 significantly inhibited neurosphere formation and triggered apoptosis in GSCs. R406 induced a metabolic shift from glycolysis to oxidative phosphorylation (OXPHOS) and subsequently production of excess ROS in GSCs. R406 also diminished tumor growth and efficiently sensitized gliomas to temozolomide in GSC-initiating xenograft mouse models. Mechanistically, the anti-GSC effect of R406 was due to the disruption of Syk/PI3K signaling in Syk-positive GSCs and PI3K/Akt pathway in Syk-negative GSCs respectively. Overall, these findings not only identify R406 as a promising GSC-targeting agent but also reveal the important role of Syk and PI3K pathways in the regulation of energy metabolism in GSCs.

4.
Molecules ; 24(2)2019 Jan 11.
Artigo em Inglês | MEDLINE | ID: mdl-30641903

RESUMO

Neuroinflammation has been well recognized as a key pathological event in acute glaucoma. The medical therapy of acute glaucoma mainly focuses on lowering intraocular pressure (IOP), while there are still scarce anti-inflammatory agents in the clinical treatment of acute glaucoma. Here we reported that ß,3α,5α-trihydroxy-androst-6-one (sterone), a novel synthetic polyhydric steroid, blocked neuroinflammation mediated by microglia/macrophages and alleviated the loss of retinal ganglion cells (RGCs) caused by acute intraocular hypertension (AIH). The results showed that sterone significantly inhibited the morphological changes, the up-regulation of inflammatory biomarker ionized calcium-binding adapter molecule 1 (Iba-1), and the mRNA increase of proinflammatory tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and interleukin-6 (IL-6) induced by lipopolysaccharide (LPS) in BV2 microglia and RAW264.7 macrophages. Moreover, immunofluorescence and western blotting analysis revealed that sterone markedly abrogated the nuclear translocation and phosphorylation of nuclear factor-κB (NF-κB) p65 subunit. Furthermore, sterone significantly suppressed the inflammatory microglial activation and RGCs' reduction caused by retinal ischemia/reperfusion (I/R) injury in a rat AIH model. These results suggest sterone may be a potential candidate in the treatment of acute glaucoma caused by microglial activation-mediated neuroinflammatory injury.


Assuntos
Microglia/efeitos dos fármacos , Microglia/metabolismo , Fármacos Neuroprotetores/farmacologia , Hipertensão Ocular/metabolismo , Hipertensão Ocular/fisiopatologia , Células Ganglionares da Retina/efeitos dos fármacos , Células Ganglionares da Retina/metabolismo , Esteroides/farmacologia , Doença Aguda , Animais , Anti-Inflamatórios/química , Anti-Inflamatórios/farmacologia , Contagem de Células , Sobrevivência Celular/efeitos dos fármacos , Modelos Animais de Doenças , Glaucoma/tratamento farmacológico , Glaucoma/etiologia , Glaucoma/metabolismo , Glaucoma/fisiopatologia , Lipopolissacarídeos/efeitos adversos , Camundongos , Estrutura Molecular , NF-kappa B/metabolismo , Fármacos Neuroprotetores/síntese química , Hipertensão Ocular/tratamento farmacológico , Hipertensão Ocular/etiologia , Células RAW 264.7 , Ratos , Traumatismo por Reperfusão/etiologia , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/patologia , Esteroides/síntese química
6.
J Virol ; 92(6)2018 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-29263275

RESUMO

Oncolytic virotherapy is an emerging treatment modality that uses replication-competent viruses to destroy cancer cells. M1 is a naturally occurring alphavirus (Togaviridae) which shows potent oncolytic activities against many cancers. Accumulation of unfolded proteins during virus replication leads to a transcriptional/translational response known as the unfolded protein response (UPR), which might counteract the antitumor effect of the oncolytic virus. In this report, we show that either pharmacological or biological inhibition of IRE1α or PERK, but not ATF6, substantially increases the oncolytic effects of the M1 virus. Moreover, inhibition of IRE1α blocks M1 virus-induced autophagy, which restricts the antitumor effects of the M1 virus through degradation of viral protein, in glioma cells. In addition, IRE1α suppression significantly increases the oncolytic effect of M1 virus in an orthotopic glioma model. From a molecular pathology study, we found that IRE1α is expressed at lower levels in higher-grade gliomas, suggesting greater antitumor efficacy of the oncolytic virus M1. Taken together, these findings illustrate a defensive mechanism of glioma cells against the oncolytic virus M1 and identify possible approaches to enhance the oncolytic viral protein accumulation and the subsequent lysis of tumor cells.IMPORTANCE Although oncolytic virotherapy is showing great promise in clinical applications, not all patients are benefiting. Identifying inhibitory signals in refractory cancer cells for each oncolytic virus would provide a good chance to increase the therapeutic effect. Here we describe that infection with the oncolytic virus M1 triggers the unfolded protein response (UPR) and subsequent autophagy, while blocking the UPR-autophagy axis significantly potentiates the antitumor efficacy of M1 in vitro and in vivo A survey of cancer tissue banks revealed that IRE1α, a key element in the UPR pathway, is commonly downregulated in higher-grade human gliomas, suggesting favorable prospects for the application of M1. Our work provides a potential predictor and target for enhancement of the therapeutic effectiveness of the M1 virus. We predict that the mechanism-based combination therapy will promote cancer virotherapy in the future.


Assuntos
Autofagia/imunologia , Endorribonucleases/deficiência , Glioma/terapia , Proteínas de Neoplasias/deficiência , Terapia Viral Oncolítica , Vírus Oncolíticos , Proteínas Serina-Treonina Quinases/deficiência , Togaviridae , Animais , Autofagia/genética , Linhagem Celular Tumoral , Cercopithecus aethiops , Cricetinae , Endorribonucleases/imunologia , Feminino , Glioma/genética , Glioma/imunologia , Humanos , Camundongos Endogâmicos BALB C , Camundongos Nus , Proteínas de Neoplasias/imunologia , Proteínas Serina-Treonina Quinases/imunologia , Resposta a Proteínas não Dobradas/genética , Resposta a Proteínas não Dobradas/imunologia , Células Vero , Ensaios Antitumorais Modelo de Xenoenxerto
7.
Hum Gene Ther ; 29(8): 950-961, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28750564

RESUMO

Oncolytic virotherapy is a novel and intriguing treatment strategy for cancer therapy. However, the clinical potential of oncolytic virus as single agent is limited. M1 virus is a promising oncolytic virus that has been tested in preclinical studies. In this study, we investigated the effect of the combination use of M1 virus and Bcl-2 family inhibitors. A chemical compounds screening including ten Bcl-2 family inhibitors demonstrated that pan-Bcl-2 inhibitors selectively augmented M1 virus oncolysis in cancer cells at very low doses. The mechanism of the enhanced antitumor effect of pan-Bcl-2 inhibitors with M1 virus is mainly due to the inhibition of Bcl-xL, which synergizes with M1-induced upregulation of Bak to trigger apoptosis. In xenograft mouse models and patient-derived tumor tissues, the combination of M1 and pan-Bcl-2 inhibitors significantly inhibited tumor growth and prolonged survival, suggesting the potential therapeutic value of this strategy. These findings offer insights into the synergy between Bcl-xL inhibition and oncolytic virus M1 as a combination anticancer treatment modality.

8.
J Neurochem ; 144(2): 186-200, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-29205357

RESUMO

Hyperglycolysis, observed within the penumbra zone during brain ischemia, was shown to be detrimental for tissue survival because of lactate accumulation and reactive oxygen species overproduction in clinical and experimental settings. Recently, mounting evidence suggests that glycolytic reprogramming and induced metabolic enzymes can fuel the activation of peripheral immune cells. However, the possible roles and details regarding hyperglycolysis in neuroinflammation during ischemia are relatively poorly understood. Here, we investigated whether overactivated glycolysis could activate microglia and identified the crucial regulators of neuroinflammatory responses in vitro and in vivo. Using BV 2 and primary microglial cultures, we found hyperglycolysis and induction of the key glycolytic enzyme hexokinase 2 (HK2) were essential for microglia-mediated neuroinflammation under hypoxia. Mechanistically, HK2 up-regulation led to accumulated acetyl-coenzyme A, which accounted for the subsequent histone acetylation and transcriptional activation of interleukin (IL)-1ß. The inhibition and selective knockdown of HK2 in vivo significantly protected against ischemic brain injury by suppressing microglial activation and IL-1ß production in male Sprague-Dawley rats subjected to transient middle cerebral artery occlusion (MCAo) surgery. We provide novel insights for HK2 specifically serving as a neuroinflammatory determinant, thus explaining the neurotoxic effect of hyperglycolysis and indicating the possibility of selectively targeting HK2 as a therapeutic strategy in acute ischemic stroke.

9.
PLoS One ; 12(10): e0184578, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28976984

RESUMO

Mycoplasma infection has been reported to be associated with cancer migration, invasion, epithelial-mesenchymal transition as well as the resistance to nucleoside analogues chemotherapeutic drugs. In this study, we found that the sensitivity of hepatocarcinoma cells to Cisplatin, Gemcitabine and Mitoxantrone was increased by mycoplasma elimination. Similar to the effect of anti-mycoplasma agent, interrupting the interaction between Mycoplasma hyorhinis membrane protein P37 and Annexin A2 of host cells using the N-terminal of ANXA2 polypeptide enhanced the sensitivity of HCC97L cells to Gemcitabine and Mitoxantrone. Meanwhile, we did not observe any changes in expression or distribution of multidrug resistance associated transporters, ATP-Binding Cassette protein B1, C1 and G2, on the removal of mycoplasma. These results suggest that mycoplasma induces a resistance to multiple drugs in hepatocarcinoma cells which required the interaction of P37 and Annexin A2. The pathway downstream this interaction needs to be explored.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Anexina A2/metabolismo , Carcinoma Hepatocelular/patologia , Neoplasias Hepáticas/patologia , Mycoplasma hyorhinis/fisiologia , Antibacterianos/farmacologia , Antineoplásicos/farmacologia , Azitromicina/farmacologia , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/microbiologia , Linhagem Celular Tumoral , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacologia , Resistência a Múltiplos Medicamentos , Resistencia a Medicamentos Antineoplásicos , Fluoroquinolonas/farmacologia , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/microbiologia , Mitoxantrona/farmacologia , Moxifloxacina , Mycoplasma hyorhinis/efeitos dos fármacos , Mycoplasma hyorhinis/genética , Mycoplasma hyorhinis/isolamento & purificação , Ligação Proteica , Reação em Cadeia da Polimerase em Tempo Real
10.
Sci Transl Med ; 9(404)2017 Aug 23.
Artigo em Inglês | MEDLINE | ID: mdl-28835517

RESUMO

Oncolytic virotherapy is rapidly progressing through clinical evaluation. However, the therapeutic efficacy of oncolytic viruses in humans has been less than expected from preclinical studies. We describe an anticancer drug screen for compounds that enhance M1 oncolytic virus activity in hepatocellular carcinoma (HCC). An inhibitor of the valosin-containing protein (VCP) was identified as the top sensitizer, selectively increasing potency of the oncolytic virus up to 3600-fold. Further investigation revealed that VCP inhibitors cooperated with M1 virus-suppressed inositol-requiring enzyme 1α (IRE1α)-X-box binding protein 1 (XBP1) pathway and triggered irresolvable endoplasmic reticulum (ER) stress, subsequently promoting robust apoptosis in HCC. We show that VCP inhibitor improved the oncolytic efficacy of M1 virus in several mouse models of HCC and primary HCC tissues. Finally, this combinatorial therapeutic strategy was well tolerated in nonhuman primates. Our study identifies combined VCP inhibition and oncolytic virus as a potential treatment for HCC and demonstrates promising therapeutic potential.


Assuntos
Antineoplásicos/metabolismo , Carcinoma Hepatocelular/terapia , Carcinoma Hepatocelular/virologia , Neoplasias Hepáticas/terapia , Neoplasias Hepáticas/virologia , Vírus Oncolíticos/metabolismo , Proteína com Valosina/antagonistas & inibidores , Animais , Apoptose , Efeito Espectador , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Terapia Combinada , Estresse do Retículo Endoplasmático , Endorribonucleases/metabolismo , Ensaios de Triagem em Larga Escala , Humanos , Neoplasias Hepáticas/patologia , Vírus Oncolíticos/patogenicidade , Primatas , Proteínas Serina-Treonina Quinases/metabolismo , Proteína com Valosina/metabolismo , Proteína 1 de Ligação a X-Box/metabolismo
11.
Biochem Biophys Res Commun ; 483(2): 892-896, 2017 02 05.
Artigo em Inglês | MEDLINE | ID: mdl-28082198

RESUMO

Neuroinflammation is one of key pathologic element in neurological diseases including stroke, traumatic brain injury, Alzheimer' s Disease, Parkinson's Disease, and multiple sclerosis as well. Up-regulation of endothelial adhesion molecules, which facilitate leukocyte adhesion to the endothelium, is the vital process of endothelial cells mediated neuroinflammation. Androst-3ß, 5α, 6ß-triol (Triol) is a synthetic steroid which has been reported to have neuroprotective effects in hypoxia/re-oxygenation-induced neuronal injury model. In the present study, we firstly investigated whether Triol inhibited the TNF-α-induced inflammatory response in rat brain microvascular endothelial cells (RBMECs). Our data showed that Triol decreased TNF-α-induced expression of vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) and the adhesion of neutrophil to RBMECs. We also found that Triol inhibited TNF-α-induced degradation of IκBα and phosphorylation of NF-κBp65 that are required for NF-κB activation. Furthermore, Triol significantly reversed TNF-α-induced down-expression of CYLD, which is a deubiquitinase that negatively regulates activation of NF-κB. These results suggest that Triol displays an anti-inflammatory effect on TNF-α-induced RBMECs via downregulating of CYLD-NF-κB signaling pathways and might have a potential benefit in therapeutic neuroinflammation related diseases.


Assuntos
Androstanóis/farmacologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Fármacos Neuroprotetores/farmacologia , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Animais , Adesão Celular/efeitos dos fármacos , Células Cultivadas , Regulação para Baixo/efeitos dos fármacos , Células Endoteliais/citologia , Humanos , Molécula 1 de Adesão Intercelular/metabolismo , Inibidor de NF-kappaB alfa/metabolismo , Neutrófilos/citologia , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Fosforilação , Ratos , Transdução de Sinais/efeitos dos fármacos , Fator de Transcrição RelA/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Ubiquitina Tiolesterase/metabolismo , Molécula 1 de Adesão de Célula Vascular/metabolismo
12.
Cell Rep ; 18(2): 468-481, 2017 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-28076790

RESUMO

Glioblastoma multiforme (GBM) is among the most aggressive of human cancers. Although differentiation therapy has been proposed as a potential approach to treat GBM, the mechanisms of induced differentiation remain poorly defined. Here, we established an induced differentiation model of GBM using cAMP activators that specifically directed GBM differentiation into astroglia. Transcriptomic and proteomic analyses revealed that oxidative phosphorylation and mitochondrial biogenesis are involved in induced differentiation of GBM. Dibutyryl cyclic AMP (dbcAMP) reverses the Warburg effect, as evidenced by increased oxygen consumption and reduced lactate production. Mitochondrial biogenesis induced by activation of the CREB-PGC1α pathway triggers metabolic shift and differentiation. Blocking mitochondrial biogenesis using mdivi1 or by silencing PGC1α abrogates differentiation; conversely, overexpression of PGC1α elicits differentiation. In GBM xenograft models and patient-derived GBM samples, cAMP activators also induce tumor growth inhibition and differentiation. Our data show that mitochondrial biogenesis and metabolic switch to oxidative phosphorylation drive the differentiation of tumor cells.


Assuntos
Astrócitos/patologia , Neoplasias Encefálicas/patologia , Diferenciação Celular , AMP Cíclico/metabolismo , Glioblastoma/metabolismo , Glioblastoma/patologia , Glicólise , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , 8-Bromo Monofosfato de Adenosina Cíclica/análogos & derivados , 8-Bromo Monofosfato de Adenosina Cíclica/farmacologia , Astrócitos/metabolismo , Astrócitos/ultraestrutura , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/ultraestrutura , Morte Celular/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Perfilação da Expressão Gênica , Proteína Glial Fibrilar Ácida/metabolismo , Glioblastoma/genética , Glioblastoma/ultraestrutura , Glicólise/efeitos dos fármacos , Humanos , Biogênese de Organelas , Fosforilação Oxidativa/efeitos dos fármacos , Proteômica , Transdução de Sinais , Ensaios Antitumorais Modelo de Xenoenxerto
13.
Nat Commun ; 7: 13107, 2016 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-27708285

RESUMO

Snakes have numerous features distinctive from other tetrapods and a rich history of genome evolution that is still obscure. Here, we report the high-quality genome of the five-pacer viper, Deinagkistrodon acutus, and comparative analyses with other representative snake and lizard genomes. We map the evolutionary trajectories of transposable elements (TEs), developmental genes and sex chromosomes onto the snake phylogeny. TEs exhibit dynamic lineage-specific expansion, and many viper TEs show brain-specific gene expression along with their nearby genes. We detect signatures of adaptive evolution in olfactory, venom and thermal-sensing genes and also functional degeneration of genes associated with vision and hearing. Lineage-specific relaxation of functional constraints on respective Hox and Tbx limb-patterning genes supports fossil evidence for a successive loss of forelimbs then hindlimbs during snake evolution. Finally, we infer that the ZW sex chromosome pair had undergone at least three recombination suppression events in the ancestor of advanced snakes. These results altogether forge a framework for our deep understanding into snakes' history of molecular evolution.


Assuntos
Evolução Biológica , Elementos de DNA Transponíveis , Serpentes/anatomia & histologia , Serpentes/genética , Animais , Linhagem da Célula , Evolução Molecular , Feminino , Membro Anterior , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Genoma , Membro Posterior , Lagartos/genética , Masculino , Filogenia , Recombinação Genética , Cromossomos Sexuais , Transcriptoma
14.
Tumour Biol ; 37(11): 14721-14731, 2016 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-27629139

RESUMO

The melastatin-like transient receptor potential 7 (TRPM7) has been implicated in proliferation or apoptosis of some cancers, indicating the potential of TRPM7 as an anti-anaplastic target. Here, we identified the characteristic TRPM7 channel currents in human malignant glioma MGR2 cells, which could be blocked by a pharmacologic inhibitor Gd3+. We mined the clinical sample data from Oncomine Database and found that human malignant glioma tissues expressed higher TRPM7 mRNA than normal brain ones. Importantly, we identified a widely used clinical anesthetic midazolam as a TRPM7 inhibitor. Midazolam treatment for seconds suppressed the TRPM7 currents and calcium influx, and treatment for 48 h inhibited the TRPM7 expression. The inhibitory effect on TRPM7 accounts for the proliferation loss and G0/G1 phase cell cycle arrest induced by midazolam. Our data demonstrates that midazolam represses proliferation of human malignant glioma cells through inhibiting TRPM7 currents, which may be further potentiated by suppressing the expression of TRPM7. Our result indicates midazolam as a pharmacologic lead compound with brain-blood barrier permeability for targeting TRPM7 in the glioma.


Assuntos
Ansiolíticos/farmacologia , Cálcio/metabolismo , Proliferação de Células/efeitos dos fármacos , Glioma/tratamento farmacológico , Midazolam/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Canais de Cátion TRPM/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Western Blotting , Ciclo Celular/efeitos dos fármacos , Mineração de Dados , Bases de Dados Factuais , Imunofluorescência , Glioma/metabolismo , Glioma/patologia , Humanos , Processamento de Imagem Assistida por Computador/métodos , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Canais de Cátion TRPM/genética , Canais de Cátion TRPM/metabolismo , Células Tumorais Cultivadas
15.
Hum Gene Ther ; 27(9): 700-11, 2016 09.
Artigo em Inglês | MEDLINE | ID: mdl-27296553

RESUMO

Cancers figure among the leading causes of morbidity and mortality worldwide. The number of new cases is expected to rise by about 70% over the next 2 decades. Development of novel therapeutic agents is urgently needed for clinical cancer therapy. Alphavirus M1 is a Getah-like virus isolated from China with a genome of positive single-strand RNA. We have previously identified that alphavirus M1 is a naturally existing oncolytic virus with significant anticancer activity against different kinds of cancer (e.g., liver cancer, bladder cancer, and colon cancer). To support the incoming clinical trial of intravenous administration of alphavirus M1 to cancer patients, we assessed the safety of M1 in adult nonhuman primates. We previously presented the genome sequencing data of the cynomolgus macaques (Macaca fascicularis), which was demonstrated as an ideal animal species for virus infection study. Therefore, we chose cynomolgus macaques of either sex for the present safety study of oncolytic virus M1. In the first round of administration, five experimental macaques were intravenously injected with six times of oncolytic virus M1 (1 × 10(9) pfu/dose) in 1 week, compared with five vehicle-injected control animals. The last two rounds of injections were further completed in the following months in the same way as the first round. Body weight, temperature, complete blood count, clinical biochemistries, cytokine profiles, lymphocytes subsets, neutralizing antibody, and clinical symptoms were closely monitored at different time points. Magnetic resonance imaging was also performed to assess the possibility of encephalitis or arthritis. As a result, no clinical, biochemical, immunological, or medical imaging or other pathological evidence of toxicity was found during the whole process of the study. Our results in cynomolgus macaques suggested the safety of intravenous administration of oncolytic virus M1 in cancer patients in the future.


Assuntos
Alphavirus/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Vetores Genéticos/administração & dosagem , Vírus Oncolíticos/imunologia , Alphavirus/genética , Animais , Modelos Animais de Doenças , Feminino , Humanos , Injeções Intravenosas , Macaca fascicularis , Masculino , Vírus Oncolíticos/genética
16.
Mol Med Rep ; 13(3): 2499-505, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26821268

RESUMO

Glutamate is the principal neurotransmitter in the central nervous system. Glutamate-mediated excitotoxicity is the predominant cause of cerebral damage. Recent studies have shown that lysosomal membrane permeabilization (LMP) is involved in ischemia­associated neuronal death in non­human primates. This study was designed to investigate the effect of glutamate on lysosomal stability in primary cultured cortical neurons. Glutamate treatment for 30 min induced the permeabilization of lysosomal membranes as assessed by acridine orange redistribution and immunofluorescence of cathepsin B in the cytoplasm. Inhibition of glutamate excitotoxicity by the NMDA receptor antagonist MK­801 and the calcium chelator ethylene glycol­bis (2­aminoethylether)­N, N, N', N'­tetraacetic acid, rescued lysosomes from permeabilization. The role of calpain and reactive oxygen species (ROS) in inducing LMP was also investigated. Ca2+ overload following glutamate treatment induced the activation of calpain and the production of ROS, which are two major contributors to neuronal death. It has been reported that lysosomal­associated membrane protein 2 (LAMP2) and heat shock protein (HSP)70 are two calpain substrates that promote LMP in cancer cells; however, it was found that calpains were activated by glutamate, but only LAMP2 was subsequently degraded. Furthermore, LMP was not alleviated by treatment with the calpain inhibitors calpeptin and SJA6017, which blocked the cleavage of the calpain substrate α­fodrin. It was demonstrated that LMP was significantly alleviated by treatment with the antioxidant N­Acetyl­L­cysteine, indicating that LMP involvement in early glutamate excitotoxicity may be mediated partly by ROS rather than calpain activation. Overall, these data shed light on the role of ROS-mediated LMP in early glutamate excitotoxicity.


Assuntos
Ácido Glutâmico/farmacologia , Lisossomos/metabolismo , Neurônios/metabolismo , Animais , Calpaína/metabolismo , Permeabilidade da Membrana Celular/efeitos dos fármacos , Células Cultivadas , Córtex Cerebral/citologia , Lisossomos/efeitos dos fármacos , Cultura Primária de Células , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/metabolismo
17.
Mol Ther ; 24(1): 156-65, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26373347

RESUMO

Oncolytic virotherapy is a novel and emerging treatment modality that uses replication-competent viruses to destroy cancer cells. Although diverse cancer cell types are sensitive to oncolytic viruses, one of the major challenges of oncolytic virotherapy is that the sensitivity to oncolysis ranges among different cancer cell types. Furthermore, the underlying mechanism of action is not fully understood. Here, we report that activation of cyclic adenosine monophosphate (cAMP) signaling significantly sensitizes refractory cancer cells to alphavirus M1 in vitro, in vivo, and ex vivo. We find that activation of the cAMP signaling pathway inhibits M1-induced expression of antiviral factors in refractory cancer cells, leading to prolonged and severe endoplasmic reticulum (ER) stress, and cell apoptosis. We also demonstrate that M1-mediated oncolysis, which is enhanced by cAMP signaling, involves the factor, exchange protein directly activated by cAMP 1 (Epac1), but not the classical cAMP-dependent protein kinase A (PKA). Taken together, cAMP/Epac1 signaling pathway activation inhibits antiviral factors and improves responsiveness of refractory cancer cells to M1-mediated virotherapy.


Assuntos
Alphavirus/genética , Colforsina/administração & dosagem , AMP Cíclico/metabolismo , Neoplasias/terapia , Transdução de Sinais/efeitos dos fármacos , Animais , Apoptose , Linhagem Celular Tumoral , Colforsina/farmacologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Fatores de Troca do Nucleotídeo Guanina/genética , Células HCT116 , Humanos , Camundongos , Neoplasias/genética , Terapia Viral Oncolítica , Vírus Oncolíticos/genética
18.
Neuro Oncol ; 17(12): 1578-88, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26014048

RESUMO

BACKGROUND: Malignant glioma is the most lethal primary tumor of the central nervous system, with notable cell invasion causing significant recurrence. Suppression of glioma invasion is very important for improving clinical outcomes. Drugs that directly disrupt the cytoskeleton have been developed for this purpose; however, drug resistance and unsatisfactory selectivity have limited their clinical use. Previously, we reported that protein kinase A (PKA, also known as cyclic-AMP dependent protein kinase) activation induced the differentiation of glioma cells. METHODS: We used several small molecular inhibitors and RNA interference, combined with wound healing assays, Matrigel transwell assay, and microscopic observation, to determine whether activation of the PKA pathway could inhibit the invasion of human glioma cells. RESULTS: Activation of PKA decreased the invasion of glioma cells. The mechanism operated via transcriptional upregulation of microtubule-associated protein 2 (MAP2), which was activated by the PKA pathway and led to ossification of microtubule dynamics via polymerization of tubulin. This resulted in morphological changes and a reduction in glioma cell invasion. Furthermore, chromosome immunoprecipitation and quantitative real-time polymerase chain reaction showed that signal transducer and activator of transcription 3 (STAT3) is involved in the transcriptional upregulation of MAP2. CONCLUSION: Our findings suggested that PKA may represent a potential target for anti-invasion glioma therapy and that the downstream modulators (eg, STAT3/MAP2) partially mediate the effects of PKA.


Assuntos
Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Glioma/metabolismo , Glioma/patologia , Proteínas Associadas aos Microtúbulos/metabolismo , Transcrição Genética , Animais , Linhagem Celular Tumoral , Humanos , Invasividade Neoplásica , Ratos , Transdução de Sinais , Regulação para Cima
19.
Steroids ; 98: 166-72, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25578735

RESUMO

Imbalance of excitation and inhibition in neurons is implicated in the pathogenesis of epilepsy. Voltage-gated sodium channels, which play a vital role in regulating neuronal excitability, are one of the major targets for developing anti-epileptic drugs. Here we provide evidence that cholestane-3ß,5α,6ß-triol (triol), a major metabolic oxysterol of cholesterol, is an effective state-dependent negative sodium channels modulator. Triol reduced Na(+) current density in a concentration-dependent manner. 10 µM triol shifted steady-state/fast/slow inactivation curves of sodium channels toward the hyperpolarizing direction. Additionally, triol reduced voltage-gated sodium currents in a voltage- and frequency-dependent manner. In a kainic acid-induced seizures mouse model, triol (25 mg/kg) significantly increased the latency of seizure onset and attenuated seizure severity. Our findings provide novel insights for understanding the modulatory role of a small molecular oxysterol on voltage-gated sodium channels and suggest triol may represent a novel and promising candidate for epilepsy intervention.


Assuntos
Anticonvulsivantes/farmacologia , Colestanóis/farmacologia , Epilepsia/tratamento farmacológico , Bloqueadores do Canal de Sódio Disparado por Voltagem/farmacologia , Canais de Sódio Disparados por Voltagem/metabolismo , Animais , Anticonvulsivantes/química , Colestanóis/química , Colesterol/química , Colesterol/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Epilepsia/induzido quimicamente , Epilepsia/metabolismo , Agonistas de Aminoácidos Excitatórios/efeitos adversos , Agonistas de Aminoácidos Excitatórios/farmacologia , Ácido Caínico/efeitos adversos , Ácido Caínico/farmacologia , Camundongos , Bloqueadores do Canal de Sódio Disparado por Voltagem/química
20.
Acta Pharm Sin B ; 4(6): 447-53, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26579416

RESUMO

In this work, retinal penetration of fluorescein was achieved in vitro by covalent attachment of taurine to fluorescein, yielding the F-Tau conjugate. Nuclear magnetic resonance (NMR) and high resolution mass spectrometry (HRMS) were used to confirm the successful synthesis of F-Tau. The cellular uptake of F-Tau in adult retinal pigment epithelial cells (ARPE-19) and human retinal microvascular endothelial cells (hRMECs) was visualized via confocal scanning microscopy. The results indicated an improvement of solubility and a reduction of logP of F-Tau compared with fluorescein. As compared with fluorescein, F-Tau showed little toxicity, and was retained longer by cells in uptake experiments. F-Tau also displayed higher transepithelial permeabilities than fluorescein in ARPE-19 and hRMECs monolayer cells (P<0.05). These results showed that taurine may be a useful ligand for targeting small-molecule hydrophobic pharmaceuticals into the retina.

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