RESUMEN
Brain stroke is one of the leading causes of death worldwide. We explored a potential stroke-related role for a newly found microRNA, miR-1247-3p, and one of its target genes, caspase-2, predicted by TargetScanVert. In the present study, we found that miR-1247-3p was downregulated during ischemia/reperfusion (I/R) and that LV-miR-1247-3p overexpression attenuated brain impairment induced by I/R. Similar results were observed in neuro2a (N2a) cells treated with oxygen-glucose deprivation/reoxygenation (OGD/R). Caspase-2 was upregulated in the I/R and OGD/R model, while Z-VDVAD-FMK - the inhibitor of caspase-2-inhibited apoptosis of N2a cells induced by OGD/R. An miR-1247-3p mimic inhibited caspase-2 expression and attenuated apoptosis of N2a cells induced by OGD/R. Myocardin-related transcription factor-A (MRTF-A) overexpression upregulated miR-1247 and mature miR-1247-3p levels and attenuated apoptosis induced by OGD/R, whereas its anti-apoptotic function could be blocked by a miR-1247-3p inhibitor. Hence, we conclude that miR-1247-3p may protect cells during brain stroke. This study offers insights for the development of effective therapeutics for promoting the survival of cerebral neurons during brain I/R injury.
Asunto(s)
Caspasa 2/metabolismo , MicroARNs/genética , Accidente Cerebrovascular/genética , Animales , Apoptosis/genética , Caspasa 2/genética , Línea Celular , Regulación hacia Abajo , Glucosa/metabolismo , Infarto de la Arteria Cerebral Media/genética , Infarto de la Arteria Cerebral Media/metabolismo , Infarto de la Arteria Cerebral Media/patología , Masculino , Ratones , Ratones Endogámicos C57BL , MicroARNs/metabolismo , Neuronas/metabolismo , Proteínas Nucleares/metabolismo , Oxígeno/metabolismo , Daño por Reperfusión/genética , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Transducción de Señal , Accidente Cerebrovascular/metabolismo , Accidente Cerebrovascular/patología , Transactivadores/genética , Transactivadores/metabolismoRESUMEN
A series of new Olaparib derivatives was designed and synthesized, and their inhibitory activities against poly (ADP-ribose) polymerases-1 (PARP-1) enzyme and cancer cell line MDA-MB-436 in vitro were evaluated. The results showed that compound 5l exhibited the most potent inhibitory effects on PARP-1 enzyme (16.10 ± 1.25 nM) and MDA-MB-436 cancer cell (11.62 ± 2.15 µM), which was close to that of Olaparib. As a PARP-1 inhibitor had been reported to be viable to neuroprotection, in order to search for new multitarget-directed ligands (MTDLs) for the treatment of Alzheimer's disease (AD), the inhibitory activities of the synthesized compounds against the enzymes AChE (from electric eel) and BChE (from equine serum) were also tested. Compound 5l displayed moderate BChE inhibitory activity (9.16 ± 0.91 µM) which was stronger than neostigmine (12.01 ± 0.45 µM) and exhibited selectivity for BChE over AChE to some degree. Molecular docking studies indicated that 5l could bind simultaneously to the catalytic active of PARP-1, but it could not interact well with huBChE. For pursuit of PARP-1 and BChE dual-targeted inhibitors against AD, small and flexible non-polar groups introduced to the compound seemed to be conducive to improving its inhibitory potency on huBChE, while keeping phthalazine-1-one moiety unchanged which was mainly responsible for PARP-1 inhibitory activity. Our research gave a clue to search for new agents based on AChE and PARP-1 dual-inhibited activities to treat Alzheimer's disease.
Asunto(s)
Acetilcolinesterasa/metabolismo , Antineoplásicos/farmacología , Inhibidores de la Colinesterasa/farmacología , Simulación del Acoplamiento Molecular , Ftalazinas/química , Ftalazinas/farmacología , Piperazinas/química , Piperazinas/farmacología , Poli(ADP-Ribosa) Polimerasa-1/antagonistas & inhibidores , Animales , Antineoplásicos/síntesis química , Antineoplásicos/química , Butirilcolinesterasa/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Inhibidores de la Colinesterasa/síntesis química , Inhibidores de la Colinesterasa/química , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Electrophorus , Caballos , Humanos , Estructura Molecular , Ftalazinas/síntesis química , Piperazinas/síntesis química , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Relación Estructura-ActividadRESUMEN
Alzheimer's disease (AD) is a chronic neurodegenerative disease of the elderly that seriously affects the quality of life and the life expectancy of those affected. There is, as yet, no effective drug treatment of AD, although several acetylcholinesterase (AChE) inhibitors and a glutamate antagonist can provide relief from its symptoms. Recent studies have indicated that the overactivation of poly(ADP-ribose) polymerase-1 (PARP-1) may promote nerve cell death in the brains of AD patients, implying that PARP-1 inhibition may have therapeutic value for the treatment of AD. Therefore, it is important to investigate novel agents with both AChE- and PARP-1-inhibitory bioactivities. In this study, the structure-based virtual screening of PARP-1 inhibitors was performed to search for potential agents with high affinities for AChE. The dynamic stability of the selected AChE-ligand complexes was investigated by molecular dynamics (MD) simulation. Two compounds, CID57390505 and CID71605390, showed high affinities for and stability in complex with AChE in docking and MD simulations. Thus, our in silico research identified two compounds with AChE and PARP-1 dual-targeted activities, indicating that this technique could aid attempts to develop more potent agents against AD.
Asunto(s)
Acetilcolinesterasa/química , Inhibidores de la Colinesterasa/química , Inhibidores Enzimáticos/química , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Poli(ADP-Ribosa) Polimerasa-1/química , Enfermedad de Alzheimer/tratamiento farmacológico , Inhibidores de la Colinesterasa/farmacología , Descubrimiento de Drogas , Inhibidores Enzimáticos/farmacología , Humanos , Concentración 50 Inhibidora , Ligandos , Estructura Molecular , Poli(ADP-Ribosa) Polimerasa-1/antagonistas & inhibidores , Relación Estructura-Actividad CuantitativaRESUMEN
Escin, as an internally applied anti-inflammatory agent, has been widely used in the treatment of inflammation and edema resulting from trauma or operation in the clinic. However, the effect of its external use on cutaneous inflammation and edema remains unexplored. In the present study, the anti-inflammatory and anti-edematous effects of external use of escin were studied in carrageenan-induced paw edema and histamine-induced capillary permeability in rats, paraxylene-induced ear swelling in mice, and cotton pellet-induced granuloma in rats. Effects of external use of escin gel on prostaglandin E2 (PGE2), tumor necrosis factor-α (TNF-α), and interleukin-1ß (IL-1ß) were determined by ELISA. The anti-inflammatory mechanism was explored by detecting the expression of glucocorticoid receptor (GR) with Western blotting and Real-time PCR analyses, with further exploration of nuclear factor-κB (NF-κB), p38 mitogen-activated protein kinase (P38MAPK) and activator protein-1 (AP-1) expressions. We demonstrated that external use of escin showed significant anti-inflammatory effects on acute and chronic inflammation in different animal models and its anti-inflammatory effects might be related to down-regulation of PGE2, TNF-α, and IL-1ß. The results also showed that escin exerted its anti-inflammatory effects by promoting the expression of GR, with the possible mechanism being inhibition of the expressions of GR-related signaling molecules such as NF-κB and AP-1.
Asunto(s)
Antiinflamatorios/administración & dosificación , Edema/tratamiento farmacológico , Escina/administración & dosificación , Extractos Vegetales/administración & dosificación , Receptores de Glucocorticoides/inmunología , Aesculus/química , Animales , Dinoprostona/inmunología , Edema/genética , Edema/inmunología , Femenino , Humanos , Interleucina-1beta/genética , Interleucina-1beta/inmunología , Masculino , Ratones , Ratas , Ratas Sprague-Dawley , Receptores de Glucocorticoides/genética , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/inmunologíaRESUMEN
Eco-pharmacovigilance (EPV) is a practical and powerful approach to minimize the potential risks posed by pharmaceutical residues in environment. However, it is impracticable to practise rigorous and unitary EPV process for all the existing and new pharmaceuticals. Here, we focused on non-steroidal anti-inflammatory drugs (NSAIDs), and discussed the necessity and potential opportunities of practising EPV of NSAIDs. We found that the consumption of NSAIDs is huge and ubiquitous across the globe. NSAIDs were worldwidely reported as one of the most dominant and frequently detected groups in environmental matrices including wastewater, surface water, suspended solids, sediments, groundwater, even drinking water. Besides, there is definitive evidence for the adverse impacts of NSAID residues on scavenging birds and aquatic species. These data suggested the necessity of implementing EPV of NSAIDs. From the perspective of drug administration, we identified some things that can be done as management practice options for EPV implementation on NSAIDs.
Asunto(s)
Antiinflamatorios no Esteroideos/análisis , Contaminación Ambiental/análisis , Farmacovigilancia , Animales , Antiinflamatorios no Esteroideos/efectos adversos , Contaminación Ambiental/efectos adversos , Contaminación Ambiental/prevención & control , Humanos , Agua/análisisRESUMEN
Our recent study has revealed that the myocardin-related transcription factor-A (MRTF-A) is involved in the apoptosis of cortical neurons induced by ischemia/reperfusion (I/R). Histone deacetylase 5 (HDAC5) and histone acetyltransferase p300 (P300) are two well-known regulators for transcription factors; however, their roles in MRTF-A-related effect on neuronal injuries during I/R are still unclear. In this study, in a model rat cerebral I/R injury via middle cerebral artery occlusion and reperfusion, we found that the expression and activity of HDAC5 was upregulated, whereas p300 and MRTF-A were downregulated both in expression and activity during I/R. Their expression changes and the interaction of the MRTF-A with HDAC5 or p300 were further verified by double immunofluorescence and co-immunoprecipitation. In cultured neuronal apoptosis model induced by H2O2, MRTF-A exhibited an anti-apoptotic effect by enhancing the transcription of Bcl-2 and Mcl-1 via CArG box binding. MRTF-A-induced anti-apoptotic effect was effectively inhibited by HDAC5, but was significantly enhanced by p300. The results suggest that both HDAC5 and p300 are involved in MRTF-A-mediated effect on neuronal apoptosis during ischemia/reperfusion injury, but with opposite effects.
Asunto(s)
Apoptosis/fisiología , Proteína p300 Asociada a E1A/metabolismo , Histona Desacetilasas/metabolismo , Neuronas/metabolismo , Daño por Reperfusión/metabolismo , Factores de Transcripción/metabolismo , Animales , Masculino , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Ratas , Ratas Sprague-Dawley , Daño por Reperfusión/patología , Transcripción Genética/fisiologíaRESUMEN
SIRT2 is involved in the development of a variety of cancers. Shikonin is a natural compound that is known to have antitumor effects. This study aims to assess the effects of shikonin on the development and metastatic progression of colorectal cancer (CRC) through regulation of SIRT2 expression and whether this effect is related to the phosphorylation of extracellular signal-regulated kinases (ERKs). The results demonstrated that SIRT2 is downregulated in CRC biopsy samples (n=31) compared with the adjacent non-cancerous tissues (ANCT, n=26). Furthermore, CRC metastases were positive for SIRT2 despite a lack of expression in the primary tumor. In addition, data from an in vitro assay revealed that overexpression of SIRT2 inhibited the proliferation and metastatic progression of SW480 cells while blocking of SIRT2 expression induced the proliferation and metastatic progression of HT29 cells. Shikonin inhibited the viability, migration and invasion of SW480 cells and it also inhibited the tumor growth in the nude mice model; while AGK2 (a specific inhibitor of SIRT2) reversed these effects. Epidermal growth factor (EGF, an activator of ERK) and ERK-overexpression inhibited the effects of shikonin on SIRT2 expression, proliferation and metastasis in SW480 cells. However, this proliferative effect of EGF was reversed by SIRT2 overexpression. In conclusion, these results suggest that SIRT2 is a new therapeutic target for the treatment of CRC. The antitumor effects of shikonin on CRC seem to be mediated by SIRT2 upregulation via phospho-ERK inhibition.
Asunto(s)
Antineoplásicos/farmacología , Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/patología , Naftoquinonas/farmacología , Sirtuina 2/metabolismo , Animales , Carcinogénesis , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Transformación Celular Neoplásica , Regulación hacia Abajo/efectos de los fármacos , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Masculino , Ratones , Persona de Mediana Edad , Invasividad Neoplásica , Metástasis de la NeoplasiaRESUMEN
Myocardin-related transcription factor-A (MRTF-A) highly expressed in brain has been demonstrated to promote neuronal survival via regulating the transcription of related target genes as a powerful co-activator of serum response factor (SRF). However, the role of MRTF-A in Alzheimer's disease (AD) is still unclear. Here, we showed that MRTF-A was significantly downregulated in cortex of the Aß25-35-induced AD rats, which played a key role in Aß25-35 induced cerebral neuronal degeneration in vitro. Bilateral intracerebroventricular injection of Aß25-35 caused significantly MRTF-A expression decline in cortex of rats, along with significant neuron apoptosis and plasticity damage. In vitro, transfection of MRTF-A into primary cultured cortical neurons prevented Aß25-35 induced neuronal apoptosis and synapses injury. And luciferase reporter assay determined that MRTF-A could bind to and enhance the transactivity of the Mcl-1 (Myeloid cell leukemia-1) and Arc (activity-regulated cytoskeletal-associated protein) promoters by activating the key CArG box element. These data demonstrated that the decreasing of endogenous MRTF-A expression might contribute to the development of AD, whereas the upregulation MRTF-A in neurons could effectively reduce Aß25-35 induced synapse injury and cell apoptosis. And the underlying mechanism might be partially due to MRTF-A-mediated the transcription and expression of Mcl-1 and Arc by triggering the CArG box.
Asunto(s)
Péptidos beta-Amiloides/metabolismo , Fragmentos de Péptidos/metabolismo , Factores de Transcripción/metabolismo , Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/genética , Animales , Apoptosis/fisiología , Supervivencia Celular , Corteza Cerebral/metabolismo , Modelos Animales de Enfermedad , Regulación hacia Abajo , Masculino , Degeneración Nerviosa/metabolismo , Neuronas/metabolismo , Proteínas Nucleares , Fragmentos de Péptidos/genética , Regiones Promotoras Genéticas , Ratas , Ratas Sprague-Dawley , Factor de Respuesta Sérica/metabolismo , Sinapsis/fisiología , Transactivadores , Factores de Transcripción/genética , Transcripción Genética , Activación Transcripcional , Transfección , Regulación hacia ArribaRESUMEN
Multidrug resistance (MDR) limits the anticancer effects of chemotherapy in patients with metastatic colorectal cancer (CRC). Oxaliplatin is a common component of combinational therapeutic regimens administered to patients with metastatic CRC; however, it is also used as a constituent of adjuvant therapy for patients at a risk of recurrent disease. In the present study, we investigated the role of stanniocalcin 2 (STC2) in chemoresistance. STC2 knockdown sensitized chemoresistant CRC cells to oxaliplatin. Moreover, the expression of exogenous STC2 in chemonaïve CRC cells induced oxaliplatin resistance. We confirmed that STC2 upregulated P-glycoprotein (P-gp) expression in CRC cells. Furthermore, shRNA against phosphoinositide 3-kinase (PI3K) or Akt inhibited the action of STC2 on P-gp upregulation and MDR in CRC. To our knowledge, this is the first report to demonstrate the induction of oxaliplatin resistance in CRC cells in response to STC2 stimulation of P-gp via the PI3K/Akt signaling pathway.
Asunto(s)
Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Neoplasias Colorrectales/metabolismo , Resistencia a Antineoplásicos/efectos de los fármacos , Glicoproteínas/metabolismo , Glicoproteínas/farmacología , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Péptidos y Proteínas de Señalización Intercelular/farmacología , Compuestos Organoplatinos/farmacología , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Técnicas de Silenciamiento del Gen , Glicoproteínas/antagonistas & inhibidores , Humanos , Oxaliplatino , Inhibidores de las Quinasa Fosfoinosítidos-3 , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , ARN Interferente Pequeño/farmacología , Regulación hacia ArribaRESUMEN
We recently demonstrated that activation of tyrosine receptor kinase B (TrkB) by 7, 8-dihydroxyflavone (7, 8-DHF), the selective TrkB agonist, increased surface alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) receptors (AMPARs) AMPA receptor subunit GluR1 (GluA1) subunit expression at the synapses of Fragile X Syndrome mutant mice. This present study investigated the effects of 7, 8-DHF on both memory function and synapse structure in relation to the synapse protein level of AMPARs in the Tg2576 Alzheimer's disease (AD) mouse model. The study found that chronic oral administration of 7, 8-DHF significantly improved spatial memory and minimized dendrite loss in the hippocampus of Tg2576 mice. A key feature of 7, 8-DHF action was the increased expression of both GluA1 and GluA2 at synapses. Interestingly, 7, 8-DHF had no effect on the attenuation of amyloid precursor protein or Aß exhibiting in the Tg2576 AD brains, yet it activated the phosphorylation of TrkB receptors and its downstream signals including CaMKII, Akt, Erk1/2, and cAMP-response element-binding protein. Importantly, cyclotraxin B (a TrkB inhibitor), U0126 (a Ras-ERK pathway inhibitor), Wortmannin (an Akt phosphorylation inhibitor), and KN-93 (a CaMKII inhibitor) counteracted the enhanced expression and phosphorylation of AMPAR subunits induced by 7, 8-DHF. Collectively, our results demonstrated that 7, 8-DHF acted on TrkB and resolved learning and memory impairments in the absence of reduced amyloid in amyloid precursor protein transgenic mice partially through improved synaptic structure and enhanced synaptic AMPARs. The findings suggest that the application of 7, 8-DHF may be a promising new approach to improve cognitive abilities in AD. We provided extensive data demonstrating that 7, 8-dihydroflavone, the TrkB agonist, improved Tg2576 mice spatial memory. This improvement is correlated with a reversion to normal values of GluA1 and GluA2 AMPA receptor subunits and dendritic spines in CA1. This work suggests that 7, 8-DHF is a suitable drug to potentiate in vivo Tropomyosin receptor kinase B (TrkB) signaling in the Alzheimer's disease mice model.
Asunto(s)
Enfermedad de Alzheimer/complicaciones , Flavanonas/uso terapéutico , Trastornos de la Memoria , Receptor trkB/metabolismo , Receptores AMPA/metabolismo , Sinapsis/metabolismo , Enfermedad de Alzheimer/genética , Precursor de Proteína beta-Amiloide/genética , Animales , Modelos Animales de Enfermedad , Conducta Exploratoria/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/genética , Humanos , Aprendizaje por Laberinto/efectos de los fármacos , Trastornos de la Memoria/tratamiento farmacológico , Trastornos de la Memoria/etiología , Trastornos de la Memoria/patología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Mutación/genética , Subunidades de Proteína/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Sinapsis/efectos de los fármacos , Sinapsis/ultraestructura , Sinaptosomas/efectos de los fármacos , Sinaptosomas/metabolismoRESUMEN
Baicalin has been shown to provide the neuroprotective effect by alleviating cerebral ischemia injury. However, little's known about the underlying mechanism. Here, a cerebral artery occlusion (MACO)/reperfusion rat model and rat primary cortical neuron culture exposed to hydrogen peroxide (H2O2) were established to evaluate the effect of baicalin on ischemia-induced neuronal apoptosis. We found baicalin can significantly less neurological deficit and reduced infarct volume in vivo. And it efficiently inhibited neuronal apoptosis in vivo and vitro, which was especially characterized by the enhancing of transcription and expression of myeloid cell leukemia-1 (MCL-1) and B-cell lymphoma-2 (BCL-2) in a dose-dependent manner. Furthermore, Baicalin markedly increased myocardin-related transcription factor-A (MRTF-A) level either in ischemic hemisphere or in primary cortical neuron cultures, whiles the anti-apoptosis effect of baicalin was significantly inhibited by transfected with the small interfering RNA of MRTF-A (MRTF-A siRNA) in primary cortical neuron cultures. The luciferase assays also indicated baicalin enhanced the transactivity of MCL-1 and BCL-2 promoter by activating the key CArG box (CC [A/T] 6GG) element, which was reduced by MRTF-A siRNA, suggesting MRTF-A may participate the anti-apoptosis effect of baicalin, and MRTF-A was involved in the transcriptional activity of MCL-1 and BCL-2 that was induced by baicalin. LY294002 (phosphatidylinositol-3 kinase (PI3K) inhibitor) and PD98059 (extracellular signal regulates kinase-1/2 (ERK1/2) inhibitor) obviously reduced baicalin-induced MRTF-A expression and transactivity and expression of MCL-1 and BCL-2, which further abolished the anti-apoptotic effect of baicalin on neuronal apoptosis. Taken together, our data provided the evidence demonstrating the neuroprotective effect of baicalin partially due to MRTF-A-mediated transactivity and expression of MCL-1 and BCL-2 by triggering the CArG box, which might be controlled by the activation of PI3K and ERK1/2.
Asunto(s)
Apoptosis/efectos de los fármacos , Flavonoides/farmacología , Hipoxia-Isquemia Encefálica/prevención & control , Neuronas/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Factores de Transcripción/metabolismo , Activación Transcripcional/efectos de los fármacos , Animales , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/metabolismo , Cromonas/farmacología , Relación Dosis-Respuesta a Droga , Hipoxia-Isquemia Encefálica/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Morfolinas/farmacología , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Inhibidores de las Quinasa Fosfoinosítidos-3 , Cultivo Primario de Células , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , ARN Interferente Pequeño/farmacología , Ratas , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Recent studies showed that hyperglycemia enhanced brain damage when subjected to transient cerebral ischemic stroke. However, the etiologic link between them has been less known. In the present study, based on an experimental rat's model of hyperlipidemia combined with cerebral ischemia-reperfusion injury (I/R), we herein showed that hyperlipidemia induced by high-fat diet (HFD) resulted in considerable increase in serum triglycerides, cholesterol and low-density lipoprotein cholesterol, and remarkable decrease in serum high-density lipoprotein cholesterol, which associated with an exacerbation on neurological deficit, cerebral infarct and terminal deoxynucleotidyl transferase-mediated nick end labeling-positive cells in the ischemic hemisphere of cerebral I/R rats treated with HFD diet. The data showed that serum superoxide dismutase activity and glutathione peroxides content were significantly decreased, while malondialdehyde level was obviously increased by hyperlipidemia or cerebral I/R alone, especially by coexistence of hyperlipidemia and cerebral I/R; meantime, hyperlipidemia also enhanced cerebral I/R-induced protein expression of cytochrome P450 2E1 (CYP2E1) and the levels of pro-inflammatory factors tumor necrosis factor-α and IL-6 in the ischemic hemispheres. Furthermore, the combined action of hyperlipidemia and cerebral I/R resulted in a protein increase expression of intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 compared to hyperlipidemia or cerebral I/R alone. Meanwhile, this study also showed that hyperlipidemia significantly enhanced cerebral I/R-induced transfer of cytochrome c from mitochondria to cytosolic and the protein expressions of Apaf-1 and caspase-3, but also decreased cerebral I/R-induced bcl-2 protein expression. The results reveal that hyperlipidemia exacerbates cerebral I/R-induced injury through the synergistic effect on CYP2E1 induction, which further induces reactive oxygen species formation, oxidative stress, inflammation and neuronal apoptosis by coexistence of hyperlipidemia and cerebral I/R.
Asunto(s)
Apoptosis/fisiología , Hiperlipidemias/metabolismo , Inflamación/metabolismo , Estrés Oxidativo/fisiología , Daño por Reperfusión/metabolismo , Animales , Dieta Alta en Grasa/efectos adversos , Modelos Animales de Enfermedad , Infarto de la Arteria Cerebral Media/metabolismo , Masculino , Ratas , Ratas Sprague-DawleyRESUMEN
Stable microtubules (MTs) is involved the mechanism of diabetic cardiomyopathy (DCM), which is induced by acetylation of α-tubulin. The present study investigated whether SIRT2, a deacetylase, regulates MT stability through α-tubulin deacetylation in DCM and whether the receptor of advanced glycation end products (AGEs) signaling pathway is involved in this effect. Type 1 diabetic mellitus (T1DM) rats model was established by a single intraperitoneal injection of streptozotocin (STZ, 65 mg/kg), and neonatal rat cardiomyocytes were also cultured. Heart function was detected by Doppler. MT stability was elevated by ß-tubulin expression density. The protein expression of SIRT2, acetylated α-tubulin and AGEs receptor were detected by immunohistochemistry or Western blots. The interaction of SIRT2 and acetylated α-tubulin was detected by Co-immunoprecipitation. In an animal model of T1DM, Western blots and immunohistochemistry revealed downregulation of SIRT2 but upregulation of the acetylated α-tubulin protein. These effects were reduced by treatment of aminoguanidine, an inhibitor of AGEs production. HDAC6 expression did not regulated in heart. In primary cultures of neonatal rat cardiomyocytes, the AGEs treatment impaired the SIRT2/acetylated α-tubulin signaling pathway, and SIRT2-overexpression reversed the function of AGEs on cardiomyocytes. In addition, gene silencing of AGEs receptor alleviated the impairment effect of AGEs on cardiomyocytes. In conclusion, these data demonstrate that AGEs/AGEs receptor promote MT stabilization via the suppression of the SIRT2/acetylated α-tubulin signaling pathway in DCM development.
Asunto(s)
Diabetes Mellitus Experimental/enzimología , Diabetes Mellitus Tipo 1/enzimología , Cardiomiopatías Diabéticas/enzimología , Microtúbulos/enzimología , Miocitos Cardíacos/enzimología , Sirtuina 2/metabolismo , Acetilación , Animales , Células Cultivadas , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/fisiopatología , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/fisiopatología , Cardiomiopatías Diabéticas/genética , Cardiomiopatías Diabéticas/fisiopatología , Ecocardiografía Doppler , Regulación Enzimológica de la Expresión Génica , Productos Finales de Glicación Avanzada/metabolismo , Masculino , Unión Proteica , Estabilidad Proteica , Interferencia de ARN , Ratas Sprague-Dawley , Receptor para Productos Finales de Glicación Avanzada/genética , Receptor para Productos Finales de Glicación Avanzada/metabolismo , Transducción de Señal , Sirtuina 2/genética , Transfección , Tubulina (Proteína)/metabolismo , Función Ventricular Izquierda , Presión VentricularRESUMEN
PRIMARY OBJECTIVE: To examine the neuroprotection of baicalin, a flavonoid compound derived from the dried root of Scutellaria baicalensis Georgi, on neurons. RESEARCH DESIGN: A rat PC12 cell line was used to study the neuroprotection and possible mechanisms of baicalin on H2O2-induced neuron damage. METHODS: Three anti- and one pro-apoptosis genes in PC12 cells were examined. Cell apoptosis was induced by H2O2 and apoptotic rate was obtained by flow cytometry. MTT for cell viability, immunofluorescence microscopy for promoter activity and western blot for gene expression were also employed. RESULTS: Data of MTT reduction assay and flow cytometry revealed that viability loss and apoptotic rate were reduced by pre-treatment of PC12 cells with baicalin for 24 hours. Baicalin was also found to increase SOD, GSH-Px activities and to decrease MDA level. Results from Western blot and immunofluorescence microscopy showed baicalin increased the expressions of survivin, Bcl-2 and p-STAT3 and decreased caspase-3 expression which were attenuated by AG-490. CONCLUSIONS: The results point to the possibility of the neuroprotective effects of baicalin on neuronal apoptosis induced by oxidative stress and indicate that activation of the JAK/STAT signalling pathway might involve the anti-apoptotic effect of baicalin.
Asunto(s)
Antioxidantes/farmacología , Apoptosis/efectos de los fármacos , Isquemia Encefálica/patología , Flavonoides/farmacología , Fármacos Neuroprotectores/farmacología , Estrés Oxidativo/efectos de los fármacos , Animales , Western Blotting , Isquemia Encefálica/tratamiento farmacológico , Caspasa 3/metabolismo , Supervivencia Celular/efectos de los fármacos , Técnica del Anticuerpo Fluorescente , Peróxido de Hidrógeno , Ratones , RatasRESUMEN
AIM: To investigate the correlation of hyperlipemia (HL) and acute cerebral ischemia/reperfusion (I/R) injury on liver damage and its mechanism. METHODS: Rats were divided into 4 groups: control, HL, I/R and HL+I/R. After the induction of HL via a high-fat diet for 18 wk, middle cerebral artery occlusion was followed by 24 h of reperfusion to capture I/R. Serum alanine transaminase (ALT) and aspartate aminotransferase (AST) were analyzed as part of liver function tests and liver damage was further assessed by histological examination. Hepatocyte apoptosis was evaluated by terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay. The expression of genes related to apoptosis (caspase-3, bcl-2) was assayed by immunohistochemistry and Western blotting. Serum tumor necrosis factor-α (TNF-α), interleukin-1 (IL-1) and liver mitochondrial superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), malondialdehyde (MDA) and Ca(2+) levels were measured to determine inflammatory and oxidative/antioxidative status respectively. Microsomal hydroxylase activity of the cytochrome P450 2E1 (CYP2E1)-containing enzyme was measured with aniline as the substrate, and CYP2E1 expression in the liver tissue and microsome was determined by immunohistochemistry and Western blotting respectively. RESULTS: HL alone induced by high-fat diet for 18 wk resulted in liver damage, indicated by histopathological analysis, and a considerable increase in serum ALT (25.13 ± 16.90 vs 9.56 ± 1.99, P < 0.01) and AST levels (18.01 ± 10.00 vs 11.33 ± 4.17, P < 0.05) compared with control. Moreover, HL alone induced hepatocyte apoptosis, which was determined by increased TUNEL-positive cells (4.47 ± 0.45 vs 1.5 ± 0.22, P < 0.01), higher caspase-3 and lower bcl-2 expression. Interestingly, compared with those in control, HL or I/R groups, massive increases of serum ALT (93.62 ± 24.00 vs 9.56 ± 1.99, 25.13 ± 16.90 or 12.93 ± 6.14, P < 0.01) and AST (82.32 ± 26.92 vs 11.33 ± 4.17, 18.01 ± 10.00 or 14.00 ± 6.19, P < 0.01) levels in HL+I/R group were observed suggesting severe liver damage, which was confirmed by liver histology. In addition, HL combined with I/R also caused significantly increased hepatocyte apoptosis, as evidenced by increased TUNEL-positive cells (6.20 ± 0.29 vs 1.5 ± 0.22, 4.47 ± 0.45 or 1.97 ± 0.47, P < 0.01), elevated expression of caspase-3 and lower expression of bcl-2. Furthermore, when compared to HL or I/R alone, HL plus I/R enhanced serum TNF-α, IL-1, liver mitochondrial MDA and Ca(2+) levels, suppressed SOD and GSH-Px in liver mitochondria, and markedly up-regulated the activity (11.76 ± 2.36 vs 4.77 ± 2.31 or 3.11 ± 1.35, P < 0.01) and expression (3.24 ± 0.38 vs 1.98 ± 0.88 or 1.72 ± 0.58, P < 0.01) of CYP2E1 in liver. CONCLUSION: The coexistence of HL and acute cerebral I/R induces severe liver damage, suggesting that cerebral ischemic stroke would exaggerate the damage of liver caused by HL. This effect is possibly due to enhanced CYP2E1 induction which further promotes oxidative damage, inflammation and hepatocyte apoptosis.
Asunto(s)
Encéfalo/irrigación sanguínea , Hiperlipidemias/complicaciones , Hepatopatías/etiología , Daño por Reperfusión/complicaciones , Enfermedad Aguda , Alanina Transaminasa/sangre , Animales , Apoptosis , Aspartato Aminotransferasas/sangre , Biomarcadores/sangre , Western Blotting , Calcio/metabolismo , Caspasa 3/metabolismo , Citocromo P-450 CYP2E1/metabolismo , Dieta Alta en Grasa , Modelos Animales de Enfermedad , Glutatión Peroxidasa/metabolismo , Hiperlipidemias/etiología , Hiperlipidemias/metabolismo , Inmunohistoquímica , Etiquetado Corte-Fin in Situ , Infarto de la Arteria Cerebral Media/complicaciones , Interleucina-1/sangre , Hígado/metabolismo , Hígado/patología , Hepatopatías/sangre , Hepatopatías/patología , Masculino , Malondialdehído/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Ratas , Ratas Sprague-Dawley , Daño por Reperfusión/etiología , Daño por Reperfusión/metabolismo , Superóxido Dismutasa/metabolismo , Factores de Tiempo , Factor de Necrosis Tumoral alfa/sangreRESUMEN
MRTF-A, known as one of the myocardin-related transcription factors, is widely found in newborn rat cortical or hippocampus neurons as well as in adult rat forebrain. Recent studies indicate that MRTF-A elevates SRF-driven transcription and enhances its stimulation by neurotrophic factor (BDNF). However, the mechanism underlying the contribution of the MRTF-A to neuronal survival is not completely understood. In this study, we investigated the effect of MRTF-A on neuronal apoptosis and its underlying mechanism. First of all, our study demonstrated that MRTF-A expression decreased obviously in rats' brains during the early period of cerebral ischemia-reperfusion. In order to estimate the effect of MRTF-A on neuronal apoptosis in vitro, we used an established experimental paradigm in which MRTF-A protected cortical neurons against both hypoxia-trophic deprivation and hydrogen peroxide-induced apoptosis. Obviously, over-expression of wild-type MRTF-A in cortical neurons inhibited apoptosis rate and enhanced anti-apoptotic gene-MCL-1. In contrast, co-expression of MRTF-A and the small interfering RNA of MRTF-A (siRNA) reversed the effect of neuroprotection and the upregulation on MCL-1 expression afforded by MRTF-A. Our study also determined whether the effect of MRTF-A up-regulating on MCL-1 expression is correlated to MRTF-A-enhancing CArG box transcription. The result showed that over-expression of wild-type MRTF-A upregulated the transcription activity of MCL reporter gene via driving the binding-domain CArG box in MCL-1 promoter, which was also reversed by co-expression of MRTF-A siRNA. In addition, we also found that BDNF neuroprotection on apoptosis induced by hypoxia-trophic withdrawal was not only inhibited by LY29004 and PD98059 but also partially blocked by transfection of dominant-negative MRTF-A in cortical neurons via enhancing the expression of anti-apoptotic gene-MCL-1, suggesting a downstream neuroprotective mechanism to BDNF neuroprotection on apoptosis.
Asunto(s)
Apoptosis/fisiología , Hipoxia-Isquemia Encefálica/genética , Neuronas/metabolismo , Factores de Transcripción/biosíntesis , Animales , Apoptosis/genética , Células COS , Supervivencia Celular/genética , Chlorocebus aethiops , Regulación de la Expresión Génica , Hipoxia-Isquemia Encefálica/patología , Hipoxia-Isquemia Encefálica/prevención & control , Masculino , Neuronas/patología , Ratas , Ratas Sprague-Dawley , Factores de Transcripción/genéticaRESUMEN
In this study, we investigated the anti-tumor effects and possible mechanisms of fucoxanthin, which has been reported to inhibit tumor proliferation and induce apoptosis in vitro or in vivo. Human gastric adenocarcinoma MGC-803 cells were treated with fucoxanthin (25µM, 50µM or 75µM). Data of flow cytometry revealed that fucoxanthin (50µM or 75µM) increased the ratio of cell in G2/M phase and apoptotic MGC-803 cells varying on a dose-dependent manner. Results from reverse transcriptase-polymerase chain reaction and Western blot showed that treatment with fucoxanthin (50µM or 75µM) significantly decreased the expressions of CyclinB1, survivin and STAT3 in MGC-803 cells in a dose-dependent manner both at the time of 24h and 48h. In addition, immunofluorescence microscopy analysis also revealed the suppressed expressions of CyclinB1 and survivin by fucoxanthin. After pretreatment with AG490 (the inhibitor for JAK/STAT signal pathway), the expressions of p-STAT3 and survivin remained also slightly lower than the vehicle control group. Co-treated with fucoxanthin (75µM) and AG490, the reduction on the expressions of STAT3, p-STAT3 and CyclinB1 by fucoxanthin were attenuated while that of survivin was enhanced. Taken together, fucoxanthin can down-regulate the expressions of CyclinB1 and survivin, inducing cell cycle arrest in G2/M phase, and apoptosis in MGC-803 cells. The reduction of CyclinB1 by fucoxanthin was associated with JAK/STAT signal pathway.
Asunto(s)
Adenocarcinoma/patología , Apoptosis/efectos de los fármacos , Quinasas Janus/metabolismo , Factores de Transcripción STAT/metabolismo , Transducción de Señal/efectos de los fármacos , Neoplasias Gástricas/patología , Xantófilas/farmacología , Adenocarcinoma/genética , División Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Ciclina B1/genética , Fase G2/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Proteínas Inhibidoras de la Apoptosis/genética , Factor de Transcripción STAT3/genética , Neoplasias Gástricas/genética , SurvivinRESUMEN
Scutellarin (Scu) is the major active principle (flavonoid) extracted from Erigeron breviscapus (Vant.) Hand-Mazz, a Chinese herbal medicine. In this paper, we investigated the effects of Scu on brain injury through the inhibition of AIF-mediated apoptosis induced by transient focal brain ischemia in rats. Rats were treated with Scu for 7 d and then subjected to cerebral ischemia/reperfusion (I/R) injury induced by a middle cerebral artery occlusion (MCAO). After 2 h of ischemia and 22 h of reperfusion, the infarct volume and the neurological deficit were determined by TTC staining and Longa's score. IN SITU end-labeling of nuclear DNA fragments (TUNEL) was employed to determine the degree of DNA fragmentation. NAD content and PARP activity in brain homogenate were determined. The expression of AIF in the nucleus was analyzed by Western blot. The present study showed that Scu significantly reduced the infarct volume and ameliorated the neurological deficit. An increase in the number of TUNEL-positive cells and a decrease in the NAD level were also observed after 2 h of ischemia and 22 h of reperfusion. At the same time, Scu (50 and 75 mg kg (-1), i. g.) treatment reversed brain NAD depletion and reduced DNA fragmentation. Scu also inhibited PARP overactivation and AIF translocation from the mitochondria to the nucleus following cerebral I/R. These findings suggested that the neuroprotective effects of Scu on brain ischemic injury-induced apoptosis might be associated with inhibition of PARP-dependent mitochondrial dysfunction and subsequent translocation of AIF.
Asunto(s)
Apigenina/uso terapéutico , Factor Inductor de la Apoptosis/metabolismo , Isquemia Encefálica/tratamiento farmacológico , Erigeron , Glucuronatos/uso terapéutico , Mitocondrias/efectos de los fármacos , Fármacos Neuroprotectores/uso terapéutico , Extractos Vegetales/uso terapéutico , Animales , Apigenina/farmacología , Apoptosis/efectos de los fármacos , Factor Inductor de la Apoptosis/genética , Transporte Biológico , Encéfalo/efectos de los fármacos , Núcleo Celular/metabolismo , Fragmentación del ADN/efectos de los fármacos , Glucuronatos/farmacología , Infarto de la Arteria Cerebral Media/tratamiento farmacológico , Masculino , Mitocondrias/metabolismo , NAD/metabolismo , Fármacos Neuroprotectores/farmacología , Fitoterapia , Extractos Vegetales/farmacología , Poli(ADP-Ribosa) Polimerasas/metabolismo , Ratas , Ratas Sprague-DawleyRESUMEN
OBJECTIVE: To investigate the relationship between trans, trans-muconic acid (ttMA) as benzene metabolite of occupational workers and benzene concentration in air. METHODS: A rapid and sensitive high-performance liquid chromatography was developed to determine the level of urinary ttMA. ttMA was extrated from urinary samples in liquid-liquid phase a ODS (2) (5u) column (phi 4.6 mm x 150 mm) and detected at wavelength 264 nm in a UV detector using vanillic acid as an internal standard. The mobile phase was acetaticacid/tetrahydrofuran/methanol/water (v/v, 1:2:10:87). The method was validated with 56 urine samples collected from occupationally benzene-exposed individuals. RESULTS: A correlation coefficient (r = 0.9963) was found for ttMA ranging 0.10-10.00 microg/mL. The limit of detection was 0.10 microg/mL. The recovery and reproducibility were generally over 90%. There was a positive correlation between ttMA and benzene level in air. The equation was Y = 0.859 + 0.108C (before work, r = 0.6200) or Y = 1.980 + 0.179C (after work, r = 0.7930). CONCLUSION: This method can be used to determine and control the level of urinary ttMA in those who are occupationally exposed to benzene.