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BACKGROUND: Choline kinase alpha (CHKA), an essential gene in phospholipid metabolism, is among the modulated MALAT1-targeted transcripts in advanced and metastatic prostate cancer (PCa). METHODS: We analyzed CHKA mRNA by qPCR upon MALAT1 targeting in PCa cells, which is characterized by high dose-responsiveness to the androgen receptor (AR) and its variants. Metabolome analysis of MALAT1-depleted cells was performed by quantitative High-resolution 1 H-Nuclear Magnetic Resonance (NMR) spectroscopy. In addition, CHKA genomic regions were evaluated by chromatin immunoprecipitation (ChIP) in order to assess MALAT1-dependent histone-tail modifications and AR recruitment. RESULTS: In MALAT1-depleted cells, the decrease of CHKA gene expression was associated with reduced total choline-containing metabolites compared to controls, particularly phosphocholine (PCho). Upon MALAT1 targeting a significant increase in repressive histone modifications was observed at the CHKA intron-2, encompassing relevant AR binding sites. Combining of MALAT1 targeting with androgen treatment prevented MALAT1-dependent CHKA silencing in androgen-responsive (LNCaP) cells, while it did not in hormone-refractory cells (22RV1 cells). Moreover, AR nuclear translocation and its activation were detected by confocal microscopy analysis and ChIP upon MALAT1 targeting or androgen treatment. CONCLUSIONS: These findings support the role of MALAT1 as a CHKA activator through putative association with the liganded or unliganded AR, unveiling its targeting as a therapeutic option from a metabolic rewiring perspective.
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BACKGROUND: More effective strategies are needed to promote poststroke functional recovery. Here, we evaluated the impact of bihemispheric transcranial direct current stimulation (tDCS) on forelimb motor function recovery and the underlying mechanisms in mice subjected to focal ischemia of the motor cortex. METHODS: Photothrombotic stroke was induced in the forelimb brain motor area, and tDCS was applied once per day for 3 consecutive days, starting 72 hours after stroke. Grid-walking, single pellet reaching, and grip strength tests were conducted to assess motor function. Local field potentials were recorded to evaluate brain connectivity. Western immunoblotting, ELISA, quantitative real-time polymerase chain reaction, and Golgi-Cox staining were used to uncover tDCS-mediated stroke recovery mechanisms. RESULTS: Among our results, tDCS increased the rate of motor recovery, anticipating it at the early subacute stage. In this window, tDCS enhanced BDNF (brain-derived neurotrophic factor) expression and dendritic spine density in the peri-infarct motor cortex, along with increasing functional connectivity between motor and somatosensory cortices. Treatment with the BDNF TrkB (tropomyosin-related tyrosine kinase B) receptor inhibitor, ANA-12, prevented tDCS effects on motor recovery and connectivity as well as the increase of spine density, pERK (phosphorylated extracellular signal-regulated kinase), pCaMKII (phosphorylated calcium/calmodulin-dependent protein kinase II), pMEF (phosphorylated myocyte-enhancer factor), and PSD (postsynaptic density)-95. The tDCS-promoted rescue was paralleled by enhanced plasma BDNF level, suggesting its potential role as circulating prognostic biomarker. CONCLUSIONS: The rate of motor recovery is accelerated by tDCS applied in the subacute phase of stroke. Anticipation of motor recovery via vicariate pathways or neural reserve recruitment would potentially enhance the efficacy of standard treatments, such as physical therapy, which is often delayed to a later stage when plastic responses are progressively lower.
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Corteza Motora , Accidente Cerebrovascular , Estimulación Transcraneal de Corriente Directa , Animales , Factor Neurotrófico Derivado del Encéfalo , Modelos Animales de Enfermedad , Humanos , Ratones , Plasticidad Neuronal , Accidente Cerebrovascular/terapia , Estimulación Transcraneal de Corriente Directa/métodosRESUMEN
Theranostics based on two-photon excitation of therapeutics in the NIR region is an emerging and powerful tool in cancer therapy since this radiation deeply penetrates healthy biological tissues and produces selective cell death. Aggregates of gold nanoparticles coated with glutathione corona functionalized with the dansyl chromophore (a-DG-AuNPs) were synthesized and found efficient nanodevice for applications in photothermal therapy (PTT). Actually the nanoparticle aggregation enhances the quenching of radiative excitation and the consequent conversion into heat. The a-DG-AuNPs are readily internalized in Hep G2 where the chromophore acts as both antenna and transducer of the NIR radiation under two-photons excitation, determining efficient cell ablation via photothermal effect.
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Terapia por Luz de Baja Intensidad/métodos , Nanopartículas del Metal/administración & dosificación , Neoplasias/terapia , Terapia Fototérmica/métodos , Nanomedicina Teranóstica/métodos , Animales , Línea Celular Tumoral , Terapia Combinada/instrumentación , Terapia Combinada/métodos , Fluorescencia , Glutatión/química , Oro/química , Humanos , Rayos Infrarrojos/uso terapéutico , Rayos Láser , Terapia por Luz de Baja Intensidad/instrumentación , Nanopartículas del Metal/química , Ratones , Neoplasias/patología , Fosfatidilcolinas/química , Fotones/uso terapéutico , Terapia Fototérmica/instrumentación , Nanomedicina Teranóstica/instrumentaciónRESUMEN
Consistent body of evidence shows that transcranial direct-current stimulation (tDCS) over the primary motor cortex (M1) facilitates motor learning and promotes recovery after stroke. However, the knowledge of molecular mechanisms behind tDCS effects needs to be deepened for a more rational use of this technique in clinical settings. Here we characterized the effects of anodal tDCS of M1, focusing on its impact on glutamatergic synaptic transmission and plasticity. Mice subjected to tDCS displayed increased long-term potentiation (LTP) and enhanced basal synaptic transmission at layer II/III horizontal connections. They performed better than sham-stimulated mice in the single-pellet reaching task and exhibited increased forelimb strength. Dendritic spine density of layer II/III pyramidal neurons was also increased by tDCS. At molecular level, tDCS enhanced: 1) BDNF expression, 2) phosphorylation of CREB, CaMKII, and GluA1, and 3) S-nitrosylation of GluA1 and HDAC2. Blockade of nitric oxide synthesis by L-NAME prevented the tDCS-induced enhancement of GluA1 phosphorylation at Ser831 and BDNF levels, as well as of miniature excitatory postsynaptic current (mEPSC) frequency, LTP and reaching performance. Collectively, these findings demonstrate that anodal tDCS engages plasticity mechanisms in the M1 and highlight a role for nitric oxide (NO) as a novel mediator of tDCS effects.
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Corteza Motora/fisiología , Plasticidad Neuronal/fisiología , Óxido Nítrico/fisiología , Transducción de Señal/fisiología , Estimulación Transcraneal de Corriente Directa/métodos , Animales , Electrodos , Masculino , Ratones , Ratones Endogámicos C57BL , Corteza Motora/efectos de los fármacos , NG-Nitroarginina Metil Éster/farmacología , Plasticidad Neuronal/efectos de los fármacos , Óxido Nítrico/antagonistas & inhibidores , Técnicas de Cultivo de Órganos , Transducción de Señal/efectos de los fármacosRESUMEN
The unconventional gaseous transmitter nitric oxide (NO) markedly influences most of mechanisms involved in the regulation of intracellular Ca2+ homeostasis. In excitable cells, Ca2+ signaling mainly depends on the activity of voltage-gated Ca2+ channels (VGCCs). In the present paper, we will review data from our laboratory and others characterizing NO-induced modulation of Ca(v)1 (L-type) and Ca(v)2.2 (N-type) channels. In particular, we will explore experimental evidence indicating that NO's inhibition of channel gating is produced via cGMP-dependent protein kinase and examine some of the numerous cell functions that are potentially influenced by the action of NO on Ca2+ channels.