RESUMEN
Metabolic dysfunction mutations can impair energy sensing and cause cancer. Loss of function of the mitochondrial tricarboxylic acid (TCA) cycle enzyme subunit succinate dehydrogenase B (SDHB) results in various forms of cancer typified by pheochromocytoma (PC). Here we delineate a signaling cascade where the loss of SDHB induces the Warburg effect, triggers dysregulation of [Ca2+]i, and aberrantly activates calpain and protein kinase Cdk5, through conversion of its cofactor from p35 to p25. Consequently, aberrant Cdk5 initiates a phospho-signaling cascade where GSK3 inhibition inactivates energy sensing by AMP kinase through dephosphorylation of the AMP kinase γ subunit, PRKAG2. Overexpression of p25-GFP in mouse adrenal chromaffin cells also elicits this phosphorylation signaling and causes PC. A potent Cdk5 inhibitor, MRT3-007, reverses this phospho-cascade, invoking a senescence-like phenotype. This therapeutic approach halted tumor progression in vivo. Thus, we reveal an important mechanistic feature of metabolic sensing and demonstrate that its dysregulation underlies tumor progression in PC and likely other cancers.
Asunto(s)
Adenilato Quinasa , Carcinoma Neuroendocrino , Adenilato Quinasa/metabolismo , Animales , Quinasa 5 Dependiente de la Ciclina/metabolismo , Metabolismo Energético , Glucógeno Sintasa Quinasa 3/metabolismo , Ratones , Fosforilación , SuccinatosRESUMEN
Alzheimer's disease (AD) is characterized by the aberrant processing of amyloid precursor protein (APP) and the accumulation of hyperphosphorylated tau, both of which are accompanied by neuroinflammation. Dietary supplementation with spray-dried porcine plasma (SDP) has anti-inflammatory effects in inflammation models. We investigated whether dietary supplementation with SDP prevents the neuropathological features of AD. The experiments were performed in 2- and 6-month-old SAMP8 mice fed a control diet, or a diet supplemented with 8% SDP, for 4 months. AD brain molecular markers were determined by Western blot and real-time PCR. Senescent mice showed reduced levels of p-GSK3ß (Ser9) and an increase in p-CDK5, p-tau (Ser396), sAPPß, and the concentration of Aß40, (all p < 0.05). SDP prevented these effects of aging and reduced Bace1 levels (all p < 0.05). Senescence increased the expression of Mme1 and Ide1 and pro-inflammatory cytokines (Il-17 and Il-18; all p < 0.05); these changes were prevented by SDP supplementation. Moreover, SDP increased Tgf-ß expression (p < 0.05). Furthermore, in aged mice, the gene expression levels of the microglial activation markers Trem2, Ym1, and Arg1 were increased, and SDP prevented these increases (all p < 0.05). Thus, dietary SDP might delay AD onset by reducing its hallmarks in senescent mice.
Asunto(s)
Enfermedad de Alzheimer/prevención & control , Encéfalo/efectos de los fármacos , Suplementos Dietéticos , Plasma , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/patología , Péptidos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Alimentación Animal , Animales , Encéfalo/metabolismo , Encéfalo/patología , Quinasa 5 Dependiente de la Ciclina/metabolismo , Citocinas/genética , Citocinas/metabolismo , Modelos Animales de Enfermedad , Regulación de la Expresión Génica , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Mediadores de Inflamación/metabolismo , Microglía/efectos de los fármacos , Microglía/metabolismo , Microglía/patología , Ovillos Neurofibrilares/efectos de los fármacos , Ovillos Neurofibrilares/metabolismo , Ovillos Neurofibrilares/patología , Fragmentos de Péptidos/metabolismo , Fosforilación , Transducción de Señal , Secado por Pulverización , Sus scrofa , Proteínas tau/metabolismoRESUMEN
Drug addiction is a chronic and recurrent disease associated with learning and memory. Shaped by drug use and cues from the environment, drug memory serves a key role in drugseeking behaviour. Methamphetamine (MA), a globally abused drug, causes cognitive impairment, and endoplasmic reticulum (ER) stress is one of the mechanisms via which this occurs. In the current study, it was hypothesized that ER stress may serve a role in the disturbance of drug memory. The present study demonstrated that 5 mg/kg MA inhibited conditioned place preference behaviour via ER stress, which caused a disruption in longterm potentiation in the hippocampus. When mice were pretreated with the ER stress inhibitors 4phenyl butyric acid or tauroursodeoxycholic acid, drugevoked synaptic plasticity was induced. Western blotting results indicated that treatment with 5 mg/kg MA enhanced the expression of cyclindependent kinase5 and decreased the expression of Ca2+/calmodulindependent protein kinase II α via ER stress. Collectively, the present results suggested that a large dose of MA inhibited drugevoked synaptic plasticity and disrupted drug memory by inducing ER stress.
Asunto(s)
Estrés del Retículo Endoplásmico/efectos de los fármacos , Hipocampo/efectos de los fármacos , Potenciación a Largo Plazo/efectos de los fármacos , Metanfetamina/farmacología , Plasticidad Neuronal/efectos de los fármacos , Animales , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/metabolismo , Estimulantes del Sistema Nervioso Central/farmacología , Quinasa 5 Dependiente de la Ciclina/metabolismo , Hipocampo/metabolismo , Hipocampo/fisiología , Potenciación a Largo Plazo/fisiología , Masculino , Memoria/efectos de los fármacos , Ratones Endogámicos C57BL , Transducción de Señal/efectos de los fármacos , Ácido Tauroquenodesoxicólico/farmacologíaRESUMEN
PURPOSE: Tumor resistance to radiation therapy is a therapeutic challenge in the treatment of patients with non-small cell lung cancer. Cyclin-dependent kinase 5 (CDK5) has been proposed to participate in cell proliferation, migration and invasion, drug resistance, and immune evasion. However, the functions and regulatory mechanisms of CDK5 in lung cancer radioresistance have not been investigated. METHODS AND MATERIALS: DNA damage response and repair were measured by neutral comet assay and γ-H2AX and Rad51 foci staining. The biological functions of CDK5 in lung cancer radioresistance were investigated with clonogenic survival assays and xenograft tumor models. Small interfering RNAs and short hairpin RNAs were used to knock down CDK5 in A549 and H1299 cells. The effects of CDK5 depletion on the tumorigenic behaviors of lung cancer cells were evaluated in vitro and in vivo. Gene expression was examined by RNA-seq and quantitative real-time polymerase chain reaction. RESULTS: We report that CDK5 depletion impairs lung cancer progression and radioresistance in vitro and in vivo. Mechanistically, we identify TAZ, a component of the Hippo pathway, as a critical downstream effector of CDK5. Loss of CDK5 downregulates TAZ expression and attenuates Hippo signaling activation. Importantly, we provide evidence that TAZ is the major effector mediating the biological functions of CDK5 in lung cancer. CONCLUSIONS: These results illustrate that CDK5 activates Hippo signaling via TAZ to participate in tumorigenesis and radioresistance, suggesting that CDK5 may be a promising radiosensitization target for the treatment of lung cancer.
Asunto(s)
Quinasa 5 Dependiente de la Ciclina/metabolismo , Neoplasias Pulmonares/radioterapia , Proteínas Serina-Treonina Quinasas/metabolismo , Tolerancia a Radiación/fisiología , Factores de Transcripción/metabolismo , Células A549 , Aciltransferasas , Animales , Línea Celular Tumoral , Ensayo Cometa , Quinasa 5 Dependiente de la Ciclina/deficiencia , Quinasa 5 Dependiente de la Ciclina/genética , Daño del ADN , Reparación del ADN , Progresión de la Enfermedad , Regulación hacia Abajo , Técnica del Anticuerpo Fluorescente , Técnicas de Silenciamiento del Gen , Silenciador del Gen , Xenoinjertos , Vía de Señalización Hippo , Histonas/análisis , Humanos , Neoplasias Pulmonares/química , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Ratones , Proteínas de Neoplasias/deficiencia , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , ARN Interferente Pequeño , Recombinasa Rad51/análisis , Regulación hacia ArribaRESUMEN
Fragile X syndrome is an X-linked dominant disorder and the most common cause of inherited mental retardation. It is caused by trinucleotide repeat expansion in the fragile X mental retardation 1 gene (FMR1) at the Xq27.3. The expansion blocks expression of the gene product, Fragile X Mental Retardation Protein (FMRP). The syndrome includes mild to moderate mental retardation and behavioral manifestations such as tactile defensiveness, gaze avoidance, repetitive motor mannerisms, perseverative (repetitive) speech, hyperarousal and it frequently includes seizures. This behavioral phenotype overlaps significantly with autism spectrum disorder. The knockout mice lack normal Fmr1 protein and show macro-orchidism, learning deficits, and hyperactivity. Consequently, this knockout mouse may serve as a valuable tool in the elucidation of the physiological role of FMR1 and the mechanisms involved in macroorchidism, abnormal behavior, abnormalities comparable to those of human fragile X patients. In this study we evaluated the effects of taurine on the testicular physiology to better understand the cellular mechanisms underlying macro-orchidism. We found that there was a significant decrease in the number of Leydig cells in the testis of fragile X mouse. Furthermore, the expression of somatostatin was drastically decreased and differential expression pattern of CDK5 in fragile X mouse testis. In the control testis, CDK is expressed in primary and secondary spermatids whereas in the Fmr1 ko mice CDK 5 is expressed mainly in spermatogonia. Taurine supplementation led to an increase in CDK5 expression in both controls and Ko mice. CDKs (Cyclin-dependent kinases) are a group of serine/threonine protein kinases activated by binding to a regulatory subunit cyclin. Over 20 functionally diverse proteins involved in cytoskeleton dynamics, cell adhesion, transport, and membrane trafficking act as CDK5 substrates elucidating the molecular mechanisms of CDK5 function. CDK5 phosphorylates a diverse list of substrates, implicating it in the regulation of a range of cellular processes. CDK5 is expressed in Leydig cells, Sertoli cells, spermatogonia and peritubular cells indicating a role in spermatogenesis. In this study we examined the expression levels of CDK5 and how it is affected by taurine supplementation in the testes and found that taurine plays an important role in testicular physiology and corrected some of the pathophysiology observed in the fragile x mouse testis.
Asunto(s)
Quinasa 5 Dependiente de la Ciclina/metabolismo , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Síndrome del Cromosoma X Frágil/genética , Taurina/farmacología , Testículo/fisiopatología , Animales , Suplementos Dietéticos , Modelos Animales de Enfermedad , Síndrome del Cromosoma X Frágil/fisiopatología , Masculino , Ratones , Ratones Noqueados , Expansión de Repetición de TrinucleótidoRESUMEN
Protein-protein interactions (PPIs) are at the core of virtually all biological processes in cells. Consequently, targeting PPIs is emerging at the forefront of drug discovery. Cellular assays which closely recapitulate native conditions in vivo are instrumental to understand how small molecule drugs can modulate such interactions. We have integrated MultiBacMam, a baculovirus-based mammalian gene delivery tool we developed, with bimolecular fluorescence complementation (BiFC), giving rise to a highly efficient system for assay development, identification and characterization of PPI modulators. We used our system to analyze compounds impacting on CDK5-p25 PPI, which is implicated in numerous diseases including Alzheimer's. We evaluated our tool-kit with the known inhibitor p5T, and we established a mini-screen to identify compounds that modulate this PPI in dose-response experiments. Finally, we discovered several compounds disrupting CDK5-p25 PPI, which had not been identified by other screening or structure-based methods before.
Asunto(s)
Quinasa 5 Dependiente de la Ciclina/metabolismo , Evaluación Preclínica de Medicamentos/métodos , Proteínas del Tejido Nervioso/metabolismo , Mapas de Interacción de Proteínas/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas/farmacología , Animales , Células COS , Línea Celular , Chlorocebus aethiops , Quinasa 5 Dependiente de la Ciclina/antagonistas & inhibidores , Descubrimiento de Drogas/métodos , Fluorescencia , Humanos , Simulación del Acoplamiento Molecular , Proteínas del Tejido Nervioso/antagonistas & inhibidores , Bibliotecas de Moléculas Pequeñas/químicaRESUMEN
Alzheimer's disease (AD) is a major and devastating neurodegenerative disease, and the amyloid-ß (Aß) hypothesis is still the central theory for AD pathogenesis. Meanwhile, another major mental illness, depression, is one of the risk factors for AD. From a high-throughput screening (HTS), amoxapine, a typical secondary amine tricyclic antidepressant (TCA), was identified to reduce Aß production. A follow-up investigation on antidepressants showed that most of the TCAs harbour similar activity. Previous studies have indicated that TCAs improve cognitive function in AD mouse models as well as in preliminary clinical data; however, the underlying mechanism is controversial, and the effect on Aß is elusive. Thus, we developed a secondary screening to determine the molecular target of amoxapine, and serotonin receptor 6 (HTR6) was identified. Knockdown of HTR6 reduced the amoxapine's effect, while the HTR6 antagonist SB258585 mimicked the activity of amoxapine. Further mechanistic study showed that amoxapine and SB258585 reduced Aß generation through multiple HTR6-mediated targets, including ß-arrestin2 and CDK5. Taken together, our study suggests that amoxapine, though no longer a first-line drug for the treatment of depression, may be beneficial for AD and further structural modification of TCAs may lead to desirable therapeutic agents to treat both AD and depression.
Asunto(s)
Amoxapina/farmacología , Péptidos beta-Amiloides/metabolismo , Antidepresivos Tricíclicos/farmacología , Neuronas/metabolismo , Receptores de Serotonina/genética , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/efectos de los fármacos , Línea Celular , Quinasa 5 Dependiente de la Ciclina/metabolismo , Evaluación Preclínica de Medicamentos , Técnicas de Silenciamiento del Gen , Células HEK293 , Humanos , Neuronas/citología , Neuronas/efectos de los fármacos , Piperazinas/farmacología , Receptores de Serotonina/metabolismo , Sulfonamidas/farmacología , Arrestina beta 2/metabolismoRESUMEN
OBJECTIVE: To investigate the effect of GAPT, an extract mixture from Radix Ginseng, Rhizoma Acor tatarinowii, Radix Polygalae and Radix Curcuma (containing ingredient of turmeric), etc. on expression of tau protein and its phosphorylation related enzyme in hippocampal neurons of APPV717I transgenic mice. METHODS: Sixty three-month-old APPV717I transgenic mice were randomly divided into model group, donepezil group [0.92 mg/(kgâ¢d)], the low, medium and high dosage of GAPT groups [0.075, 0.15, 0.30 g/(kgâ¢d), 12 in each group], and 12 three-month-old C57BL/6J mice were set as a normal control group, treatments were administered orally once a day respectively, and both the normal group and model group were given 0.5% sodium carboxymethyl cellulose solution. Immunohistochemistry (IHC) and Western blot analysis were used to detect the expression of total tau protein (Tau-5), cyclin-dependent kinase 5 (CDK5) and protein phosphatase 2A (PP2A) in hippocampal neurons of experimental mice after 8-month drug administration (11 months old). RESULTS: In the model group, the expression of Tau-5 and CDK5 were increased, whereas the expression of PP2A was decreased in hippocampal neurons, which were signifificantly different compared with that in the normal group (all P<0.01). IHC test indicated the number and area of either Tau-5 or CDK5 positive cells were decreased with a dose-depended way in GAPT groups, and an increase of PP2A. Compared with the model group, the changes were signifificant in GAPT groups (P<0.05 or P<0.01). Similar results were shown by Western blot. CONCLUSION: GAPT could attenuate abnormal hyperphosphorylation of tau protein in hippocampal neurons of APPV717I transgenic mice via inhibiting the expression of CDK5 and activating the expression of PP2A.
Asunto(s)
Medicamentos Herbarios Chinos/farmacología , Hipocampo/patología , Neuronas/enzimología , Proteínas tau/metabolismo , Animales , Región CA1 Hipocampal/patología , Quinasa 5 Dependiente de la Ciclina/metabolismo , Femenino , Inmunohistoquímica , Masculino , Ratones Endogámicos C57BL , Ratones Transgénicos , Neuronas/efectos de los fármacos , Fosforilación/efectos de los fármacos , Proteína Fosfatasa 2/metabolismoRESUMEN
Developing selective inhibitors for a particular kinase remains a major challenge in kinase-targeted drug discovery. Here we performed a multi-step virtual screening for dual-specificity tyrosine-phosphorylation-regulated kinase 1A (DYRK1A) inhibitors by focusing on the selectivity for DYRK1A over cyclin-dependent kinase 5 (CDK5). To examine the key factors contributing to the selectivity, we constructed logistic regression models to discriminate between actives and inactives for DYRK1A and CDK5, respectively, using residue-based binding free energies. The residue-based parameters were calculated by molecular mechanics-generalized Born surface area (MM-GBSA) decomposition methods for kinase-ligand complexes modeled by computer ligand docking. Based on the findings from the logistic regression models, we built a three-dimensional (3D) pharmacophore model and chose filter criteria for the multi-step virtual screening. The virtual hit compounds obtained from the screening were assessed for their inhibitory activities against DYRK1A and CDK5 by in vitro assay. Our screening identified two novel selective DYRK1A inhibitors with IC50 values of several µM for DYRK1A and >100µM for CDK5, which can be further optimized to develop more potent selective DYRK1A inhibitors.
Asunto(s)
Evaluación Preclínica de Medicamentos , Inhibidores de Proteínas Quinasas/análisis , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Adenosina Trifosfato/metabolismo , Secuencia de Aminoácidos , Sitios de Unión , Bioensayo , Quinasa 5 Dependiente de la Ciclina/metabolismo , Humanos , Ligandos , Modelos Logísticos , Simulación del Acoplamiento Molecular , Inhibidores de Proteínas Quinasas/química , Proteínas Serina-Treonina Quinasas/química , Proteínas Tirosina Quinasas/química , Máquina de Vectores de Soporte , Quinasas DyrKRESUMEN
OBJECTIVES: To explore the protective effects of Tongmai Yizhi Decoction (, TYD), a Chinese herb complex prescription against the impairment of cognitive functions and memory loss in amyloid beta 1-40 (Aß1-40) peptide and ibotenic (IBO)-induced Alzheimer's disease (AD) model rats. METHODS: The in vivo model was established by injecting Aß1-40 and IBO into left hippocampal CA1 area of Sprague-Dawley (SD) rat to mimic AD. Totally 32 SD rats were divided into 4 groups, including sham operation group, AD model group, TYD group [AD rats treated with TYD at the dosage of 19.44 g/(kgâ¢d) for 4 weeks] and huperzine A group [AD rats treated with huperzine A at the dosage of 40.5 µg/(kgâ¢d) for 4 weeks]. Spatial learning and memory level was detected by Morris Water Maze test. Histological morphology in the hippocampus was tested by hematoxylin-eosin (HE) staining. Cyclin-dependent kinase-5 (Cdk5) protein and gene expression level were investigated by Western blot analysis and real-time quantitative polymerase chain reaction (RT-qPCR), respectively. RESULTS: Aß1-40 and IBO treatment induced longer escape latency of rats, compared with sham operation group from day 25 (P<0.01). However, TYD and huperzine A obviously shortened the escape latency from day 26 (P<0.01). Moreover, the effect of TYD was similar to huperzine A (P>0.05). Furthermore, HE staining also showed that TYD and huperzine A reversed the neuropathological changes in the hippocampus triggered by Aß1-40 and IBO. TYD and huperzine A effectively reduced the expression levels of Cdk5 protein and gene located in rat hippocampus, compared with the AD model group (P<0.01). CONCLUSION: TYD could be a promising neuroprotective agent for protecting neuron from AD injury through inhibiting Cdk5 expression.
Asunto(s)
Enfermedad de Alzheimer/tratamiento farmacológico , Medicamentos Herbarios Chinos/uso terapéutico , Fármacos Neuroprotectores/uso terapéutico , Alpinia , Enfermedad de Alzheimer/patología , Animales , Cognición/efectos de los fármacos , Quinasa 5 Dependiente de la Ciclina/metabolismo , Modelos Animales de Enfermedad , Regulación hacia Abajo/efectos de los fármacos , Femenino , Hipocampo/efectos de los fármacos , Masculino , Aprendizaje por Laberinto/efectos de los fármacos , Memoria/efectos de los fármacos , Extractos Vegetales , Ratas , Ratas Sprague-DawleyRESUMEN
The pathological features of Alzheimer's disease (AD) include extracellular neuritic plaques containing ß-amyloid (Aß) peptide, a cleaved fragment of amyloid precursor protein (APP) via ß-site amyloid precursor protein-cleaving enzyme 1 (BACE1) and intracellular neurofibrillary tangles (NFTs) composed of hyperphosphorylated tau. Cyclin-dependent kinase 5 (Cdk5) is increasingly thought to play a pivotal role in the pathogenesis of AD, both as a regulator of the production of Aß and through its well-established role as a tau kinase. Fuzhisan (FZS), a Chinese herbal complex prescription, has been used for the treatment AD for over 20 years, and is known to enhance the cognitive ability in AD patients as well as in AD model rats. To investigate mechanisms of AD and the potential therapy of FZS in AD, we treated senescence-accelerated mouse SAMP8 mice, a useful model of AD-related memory impairment, with FZS by intragastrical administration for 8 weeks and Donepizel was used as a positive control. The results showed that FZS (0.3, 0.6, and 1.2 g/kg/day) improved impaired cognitive ability of aged SAMP8 mice in a dose-dependent manner. FZS robustly decreased Aß level and phosphorylation of tau. This was accompanied by a significant decrease in the BACE1 level and phosphorylated APP (Thr668). Futhermore, The p25/Cdk5 pathway was markedly down-regulated by FZS treatment. These results indicated that the memory ameliorating effect of FZS may be, in part, by regulation the p25/Cdk5 pathway which may contribute to down-regulation of Aß and tau hyperphosphorylation.
Asunto(s)
Enfermedad de Alzheimer/tratamiento farmacológico , Péptidos beta-Amiloides/biosíntesis , Medicamentos Herbarios Chinos/farmacología , Hipocampo/efectos de los fármacos , Ovillos Neurofibrilares/efectos de los fármacos , Proteínas tau/metabolismo , Envejecimiento , Enfermedad de Alzheimer/metabolismo , Animales , Quinasa 5 Dependiente de la Ciclina/metabolismo , Modelos Animales de Enfermedad , Regulación hacia Abajo , Hipocampo/metabolismo , Masculino , Ratones , Ovillos Neurofibrilares/metabolismo , Fosforilación/efectos de los fármacosRESUMEN
Flavonoids have been shown to improve cognitive function and delay the dementia progression. However, the underlying mechanisms remain elusive. In the present study, we examined the effect of Scutellaria baicalensis stem-leaf total flavonoids (SSTFs) extracted from S. baicalensis Georgi on spatial learning and memory in a vascular dementia (VaD) rat model and explored its molecular mechanisms. The VaD rats were developed by permanent bilateral occlusion of the common carotid artery. Seven days after recovery, the VaD rats were treated with either 50 or 100 mg/kg of SSTF for 60 days. The spatial learning and memory was evaluated in the Morris water maze (MWM) test. The tau hyperphosphorylation and the levels of the related protein kinases or phosphatases were examined by western blot analysis. In VaD rats, SSTF treatment at 100 mg/kg significantly reduced the escape latency in training trial in MWM test. In the probe trial, SSTF treatment increased the searching time and travel distance in the target quadrant. SSTF treatment inhibited the tau phosphorylation in both cortex and hippocampus in VaD rats. Meanwhile, SSTF reduced the activity of glycogen synthase kinase 3ß and cyclin-dependent kinase 5 in VaD rats. In contrast, SSTF treatment increased the level of the protein phosphatase 2A subunit B in VaD rats. SSTF treatment significantly improved the spatial cognition in VaD rats. Our results suggest that SSTF may alleviate tau-hyperphosphorylation-induced neurotoxicity through coordinating the activity of kinases and phosphatase after a stroke. SSTF may be developed into promising novel therapeutics for VaD.
Asunto(s)
Demencia Vascular/tratamiento farmacológico , Flavonoides/farmacología , Memoria/efectos de los fármacos , Scutellaria baicalensis , Aprendizaje Espacial/efectos de los fármacos , Animales , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Isquemia Encefálica/complicaciones , Quinasa 5 Dependiente de la Ciclina/metabolismo , Demencia Vascular/etiología , Demencia Vascular/psicología , Medicamentos Herbarios Chinos/farmacología , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Masculino , Aprendizaje por Laberinto/efectos de los fármacos , Fosforilación/efectos de los fármacos , Proteína Fosfatasa 2/metabolismo , Ratas , Ratas Sprague-Dawley , Proteínas tau/metabolismoRESUMEN
Although numerous studies have reported the influence of extremely low frequency magnetic field (ELF-MF) exposure on human health, its effects on cognitive deficits in Alzheimer's disease (AD) have remained under debate. Moreover, the influence of ELF-MF on hyperphosphorylated tau, which is one of the most common pathological hallmarks of AD, has not been reported to date. Therefore, transgenic mice (3xTg) were used in the present study. 3xTg mice, which express an APP/PS1 mutation combined with a tau (P301L) mutation and that develop cognitive deficits at 6 months of age, were subjected to ELF-MF (50Hz, 500µT) exposure or sham exposure daily for 3 months. We discovered that ELF-MF exposure ameliorated cognitive deficits and increased synaptic proteins in 3xTg mice. The protective effects of ELF-MF exposure may have also been caused by the inhibition of apoptosis and/or decreased oxidative stress levels that were observed in the hippocampus tissues of treated mice. Furthermore, tau hyperphosphorylation was decreased in vivo because of ELF-MF exposure, and this decrease was induced by the inhibition of GSK3ß and CDK5 activities and activation of PP2Ac. We are the first to report that exposure to ELF-MF can attenuate tau phosphorylation. These findings suggest that ELF-MF exposure could act as a valid therapeutic strategy for ameliorating cognitive deficits and attenuating tau hyperphosphorylation in AD.
Asunto(s)
Enfermedad de Alzheimer/complicaciones , Trastornos del Conocimiento/etiología , Trastornos del Conocimiento/terapia , Magnetoterapia/métodos , Proteínas tau/metabolismo , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/radioterapia , Precursor de Proteína beta-Amiloide/genética , Animales , Apoptosis/genética , Apoptosis/efectos de la radiación , Condicionamiento Psicológico/fisiología , Condicionamiento Psicológico/efectos de la radiación , Quinasa 5 Dependiente de la Ciclina/genética , Quinasa 5 Dependiente de la Ciclina/metabolismo , Modelos Animales de Enfermedad , Relación Dosis-Respuesta en la Radiación , Regulación de la Expresión Génica/genética , Regulación de la Expresión Génica/efectos de la radiación , Glucógeno Sintasa Quinasa 3 beta/genética , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Humanos , Aprendizaje por Laberinto/fisiología , Aprendizaje por Laberinto/efectos de la radiación , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Mutación/genética , Fosforilación/efectos de la radiación , Presenilina-1/genética , Especies Reactivas de Oxígeno/metabolismo , Proteínas tau/genéticaRESUMEN
Adult neurogenesis, a process that consists in the generation of new neurons from adult neural stem cells, represents a remarkable illustration of the brain structural plasticity abilities. The hypothalamus, a brain region that plays a key role in the neuroendocrine regulations including reproduction, metabolism or food intake, houses neural stem cells located within a hypothalamic neurogenic niche. In adult sheep, a seasonal mammalian species, previous recent studies have revealed photoperiod-dependent changes in the hypothalamic cell proliferation rate. In addition, doublecortin (DCX), a microtubule-associated protein expressed in immature migrating neurons, is highly present in the vicinity of the hypothalamic neurogenic niche. With the aim to further explore the mechanism underlying adult sheep hypothalamic neurogenesis, we first show that new neuron production is also seasonally regulated since the density of DCX-positive cells changes according to the photoperiodic conditions at various time points of the year. We then demonstrate that cyclin-dependant kinase-5 (Cdk5) and p35, two proteins involved in DCX phosphorylation and known to be critically involved in migration processes, are co-expressed with DCX in young hypothalamic neurons and are capable of in vivo interaction. Finally, to examine the migratory potential of these adult-born neurons, we reveal the rostro-caudal extent of DCX labeling on hypothalamic sagittal planes. DCX-positive cells are found in the most rostral nuclei of the hypothalamus, including the preoptic area many of which co-expressed estrogen receptor-α. Thus, beyond the confirmation of the high level of neuron production during short photoperiod in sheep, our results bring new and compelling elements in support of the existence of a hypothalamic migratory path that is responsive to seasonal stimuli.
Asunto(s)
Movimiento Celular , Hipotálamo/fisiología , Células-Madre Neurales/fisiología , Neurogénesis , Fotoperiodo , Animales , Quinasa 5 Dependiente de la Ciclina/metabolismo , Proteínas de Dominio Doblecortina , Hipotálamo/citología , Hipotálamo/metabolismo , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Células-Madre Neurales/citología , Células-Madre Neurales/metabolismo , Neuropéptidos/metabolismo , Estaciones del Año , OvinosRESUMEN
Cyclin-dependent kinase 5 (Cdk5) is an important serine/threonine kinase that plays critical roles in many physiological processes. Recently, Cdk5 has been reported to phosphorylate TRPV1 at threonine 407 (Thr-407) in humans (Thr-406 in rats), which enhances the function of TRPV1 channel and promotes thermal hyperalgesia in the complete Freund's adjuvant (CFA)-induced inflammatory pain rats. However, the underlying mechanisms are still unknown. Here, we demonstrate that Cdk5 phosphorylates TRPV1 at Threonine 406 and promotes the surface localization of TRPV1, leading to inflammatory thermal hyperalgesia. The mutation of Thr-406 of TRPV1 to alanine reduced the interaction of TRPV1 with the cytoskeletal elements and decreased the binding of TRPV1 with the motor protein KIF13B, which led to reduced surface distribution of TRPV1. Disrupting the phosphorylation of TRPV1 at Thr-406 dramatically reduced the surface level of TRPV1 in HEK 293 cells after transient expression and the channel function in cultured dorsal root ganglion (DRG) neurons. Notably, intrathecal administration of the interfering peptide against the phosphorylation of Thr-406 alleviated heat hyperalgesia and reduced the surface level of TRPV1 in inflammatory pain rats. Together, these results demonstrate that Cdk5-mediated phosphorylation of TRPV1 at Thr-406 increases the surface level and the function of TRPV1, while the TAT-T406 peptide can effectively attenuate thermal hyperalgesia. Our studies provide a potential therapy for inflammatory pain.
Asunto(s)
Quinasa 5 Dependiente de la Ciclina/metabolismo , Hiperalgesia/etiología , Inflamación/complicaciones , Canales Catiónicos TRPV/metabolismo , Alanina/genética , Animales , Calcio/metabolismo , Células Cultivadas , Quinasa 5 Dependiente de la Ciclina/genética , Modelos Animales de Enfermedad , Adyuvante de Freund/toxicidad , Ganglios Espinales/citología , Humanos , Inflamación/inducido químicamente , Cinesinas , Masculino , Mutación/genética , Neuronas/metabolismo , Péptidos/uso terapéutico , Fosforilación , Fosfotransferasas/metabolismo , Ratas , Ratas Sprague-Dawley , Canales Catiónicos TRPV/genética , Treonina/genética , Treonina/metabolismoRESUMEN
OBJECTIVE: To observe the effects of polydatin on learning and memory and cyclin-dependent kinase 5 (Cdk5) kinase activity in the hippocampus of rats with chronic alcoholism. METHODS: Forty rats were randomly divided into 4 groups: control group, chronic alcoholism group, low and high polydatin group. The rat chronic alcoholism model was established by ethanol 3.0 g/(kg · d) (intragastric administration). The abstinence scoring was used to evaluate the rats withdrawal symptoms; cognitive function was measured by Morris water maze experiment; Cdk5 protein expression in the hippocampus was detected by immunofluorescence; Cdk5 kinase activity in the hippocampus was detected by liquid scintillation counting method. RESULTS: The abstinence score, escape latency, Cdk5 kinase activity in chronic alcoholism group rats were significantly higher than those of control group (P < 0.05). The abstinence score, escape latency in high polydatin group rats were significantly lower than those of chronic alcoholism group (P < 0.05); Cdk5 kinase activity in high and low polydatin group rats was significantly lower than that of chronic alcoholism group( P < 0.05); immunofluorescence showed that the Cdk5 positive cells of chronic alcoholism group were significantly increased compared with control group (P < 0.05), and the Cdk5 positive cells of polydatin groups were significantly decreased compared with chronic alcoholism group ( P < 0.05). CONCLUSION: Polydatin-reduced the chronic alcoholism damage may interrelate with regulation of Cdk5 kinase activity.
Asunto(s)
Alcoholismo/fisiopatología , Quinasa 5 Dependiente de la Ciclina/metabolismo , Glucósidos/farmacología , Hipocampo/efectos de los fármacos , Aprendizaje/efectos de los fármacos , Memoria/efectos de los fármacos , Estilbenos/farmacología , Animales , Medicamentos Herbarios Chinos/farmacología , Hipocampo/enzimología , RatasRESUMEN
Calpain mediated cleavage of CDK5 natural precursor p35 causes a stable complex formation of CDK5/p25, which leads to hyperphosphorylation of tau. Thus inhibition of this complex is a viable target for numerous acute and chronic neurodegenerative diseases involving tau protein, including Alzheimer's disease. Since CDK5 has the highest sequence homology with its mitotic counterpart CDK2, our primary goal was to design selective CDK5/p25 inhibitors targeting neurodegeneration. A novel structure-based virtual screening protocol comprised of e-pharmacophore models and virtual screening workflow was used to identify nine compounds from a commercial database containing 2.84 million compounds. An ATP non-competitive and selective thieno[3,2-c]quinolin-4(5H)-one inhibitor (10) with ligand efficiency (LE) of 0.3 was identified as the lead molecule. Further SAR optimization led to the discovery of several low micromolar inhibitors with good selectivity. The research represents a new class of potent ATP non-competitive CDK5/p25 inhibitors with good CDK2/E selectivity.
Asunto(s)
Adenosina Trifosfato/química , Quinasa 5 Dependiente de la Ciclina/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/química , Quinolonas/química , Adenosina Trifosfato/metabolismo , Sitios de Unión , Análisis por Conglomerados , Quinasa 5 Dependiente de la Ciclina/metabolismo , Evaluación Preclínica de Medicamentos , Humanos , Enlace de Hidrógeno , Cinética , Simulación del Acoplamiento Molecular , Unión Proteica , Inhibidores de Proteínas Quinasas/metabolismo , Estructura Terciaria de Proteína , Quinolonas/metabolismo , Relación Estructura-Actividad , Proteínas tau/metabolismoRESUMEN
Since Alzheimer disease (AD) is a multifactorial disease, recent therapeutical approaches concentrate on the development of a multitargeting drug. Various protein kinases are known to be involved in the progression of AD. A first series of 3-ethoxycarbonyl-1-aza-9-oxafluorenes has been synthesized and biologically evaluated as AD-relevant protein kinase inhibitors. A concentration-dependent inhibition of important AD-relevant kinases has been characterized after the selectivity of kinase inhibition had been demonstrated. Structure-activity relationships of protein kinase inhibition are discussed and first multitargeting inhibitors have been identified.
Asunto(s)
Enfermedad de Alzheimer/enzimología , Proteína Quinasa CDC2/metabolismo , Quinasa 5 Dependiente de la Ciclina/metabolismo , Diseño de Fármacos , Fluorenos/química , Fluorenos/farmacología , Glucógeno Sintasa Quinasa 3/metabolismo , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Enfermedad de Alzheimer/tratamiento farmacológico , Animales , Compuestos Aza/síntesis química , Compuestos Aza/química , Compuestos Aza/farmacología , Bioensayo , Sistemas de Liberación de Medicamentos , Evaluación Preclínica de Medicamentos , Activación Enzimática/efectos de los fármacos , Fluorenos/síntesis química , Glucógeno Sintasa Quinasa 3 beta , Humanos , Estructura Molecular , Inhibidores de Proteínas Quinasas/síntesis química , Células Sf9 , Relación Estructura-ActividadRESUMEN
Although neurons are not productively infected with HIV-1, neuronal injury and death are frequently seen in the brains of AIDS patients with neurological and neurocognitive disorders. Evidently, viral proteins including Tat and cellular inflammatory factors released by activated and/or infected microglia, macrophages, and astrocytes contribute to neuronal cell death. Several studies have demonstrated that HIV-1 associated neuronal cell injury is mediated by dysregulation of signaling pathways that are controlled, in part, by a class of serine/threonine kinases. In this study, we demonstrate that pDING, a novel plant-derived phosphate binding protein has the capacity to reduce the severity of injury and death caused by HIV-1 and its neurotoxic Tat protein. We demonstrate that pDING, also called p27SJ/p38SJ, protects cells from the loss of neuronal processes induced by Tat and promotes neuronal outgrowth after Tat-mediated injury. Further, expression of pDING prevents Tat-induced oxidative stress and mitochondrial permeability. With its profound phosphatase activity, pDING controls the activity of several kinases including MAPK, Cdk5, and their downstream target protein, MEF2, which is implicated in neuronal cell protection. Our results show that expression of pDING in neuronal cells diminishes the level of hyperphosphorylated forms of Cdk5 and MEF2 caused by Tat and the other neurotoxic agents that are secreted by the HIV-1 infected cells. These observations suggest that pDING, through its phosphatase activity, has the ability to manipulate the state of phosphorylation and activity of several factors involved in neuronal cell health in response to HIV-1.
Asunto(s)
Neuronas/efectos de los fármacos , Proteínas de Unión a Fosfato/farmacología , Proteínas de Plantas/farmacología , Productos del Gen tat del Virus de la Inmunodeficiencia Humana/toxicidad , Muerte Celular , Células Cultivadas , Quinasa 5 Dependiente de la Ciclina/genética , Quinasa 5 Dependiente de la Ciclina/metabolismo , Humanos , Hypericum/química , Mitocondrias/efectos de los fármacos , Mitocondrias/fisiología , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Estrés Oxidativo/efectos de los fármacos , Proteínas de Unión a Fosfato/metabolismo , Monoéster Fosfórico Hidrolasas/genética , Monoéster Fosfórico Hidrolasas/metabolismoRESUMEN
BACKGROUND: Buyang Huanwu Decoction (BYHWD), a traditional Chinese medicine formula, has been shown to exert a variety of pharmacological effects including neuroprotective properties. However, the mechanism of neuroprotection is not fully understood. This study was designed to explore the mechanism of BYHWD in the treatment of spinal ischemia-reperfusion injury in rats. METHODS: Twenty-eight male Sprague-Dawley rats, weighting 250-280 g, were used, and were randomly divided into four groups with 7 animals in each: sham operation group (Control), spinal ischemia with saline (SI + Saline), spinal ischemia with BYHWD (SI + BYHWD), and spinal ischemia with roscovitine (SI + R). After 60 minutes of spinal ischemia followed by 72 hours of reperfusion, motor function of hind limbs, spinal ischemic infarction volume, the number of apoptotic cells, and cyclin-dependent kinase 5 (Cdk5) were examined. RESULT: Ischemia-reperfusion resulted in injury of the spines, while BYHWD significantly improved spinal function. The spinal infarction volume, number of apoptotic cells, and Cdk5 were decreased by administration of BYHWD. The similar improvements were seen with the pre-treatment of roscovitine. CONCLUSIONS: BYHWD prevented the ischemia-reperfusion-induced spinal injury in rats. The protective function of BYHWD was, in part, linked with inhibition of Cdk5.