RESUMEN
La infección causada por el SARS-CoV-2 se convirtió en pandemia y causó millones de muertes e incalculables pérdidas económicas en todo el mundo. En ese contexto se introdujeron terapias para contener la infección hasta que se pudiera vacunar a la población. Las propuestas terapéuticas surgieron mayormente de medicamentos ya establecidos cuyos mecanismos de acción podían interferir con el ciclo de infección del virus o con algunos de los mecanismos fisiopatológicos dependientes del hospedero. Los fármacos monoligandos actuaban sobre un único blanco farmacológico para tratar una enfermedad compleja y multifactorial como la COVID-19, lo cual resultó, quizás, la principal causa del fracaso con las intervenciones no-vacunales. En esta comunicación breve se realizó una reseña de los mecanismos farmacológicos del azul de metileno en la terapia de la COVID-19. La multiplicidad de mecanismos documentados relacionados con el bloqueo de la patogenia viral, su bajo costo, el bajo perfil toxicológico, la farmacocinética privilegiada y la capacidad nacional de producirlo convirtieron a este antiguo medicamento en una opción terapéutica en los protocolos de tratamiento a pacientes graves y críticos.
Infection caused by SARS-CoV-2 became pandemic and caused millions of deaths and incalculable economic losses worldwide. In this context, therapies were introduced to contain the infection until the population could be vaccinated. Therapeutic proposals emerged from already established drugs whose mechanisms of action could interfere with the virus infection cycle or with some of the host-dependent pathophysiological mechanisms. Monoligand drugs acted on a single drug target to treat a complex and multifactorial disease like COVID-19, which was the main cause of failure with non-vaccine interventions. In this brief communication, a review of the pharmacological mechanisms of methylene blue in the therapy of COVID-19 was performed. The multiplicity of documented mechanisms related to the blockade of viral pathogenesis, its low cost, low toxicological profile, privileged pharmacokinetics and national capacity to produce it made this drug a therapeutic option in the treatment protocols of severe and critical patients.
RESUMEN
Ferroptosis is an iron-dependent cell death-driven by excessive peroxidation of polyunsaturated fatty acids (PUFAs) of membranes. A growing body of evidence suggests the induction of ferroptosis as a cutting-edge strategy in cancer treatment research. Despite the essential role of mitochondria in cellular metabolism, bioenergetics, and cell death, their function in ferroptosis is still poorly understood. Recently, mitochondria were elucidated as an important component in cysteine-deprivation-induced (CDI) ferroptosis, which provides novel targets in the search for new ferroptosis-inducing compounds (FINs). Here, we identified the natural mitochondrial uncoupler nemorosone as a ferroptosis inducer in cancer cells. Interestingly, nemorosone triggers ferroptosis by a double-edged mechanism. In addition to decreasing the glutathione (GSH) levels by blocking the System xc cystine/glutamate antiporter (SLC7A11), nemorosone increases the intracellular labile Fe2+ pool via heme oxygenase-1 (HMOX1) induction. Interestingly, a structural variant of nemorosone (O-methylated nemorosone), having lost the capacity to uncouple mitochondrial respiration, does not trigger cell death anymore, suggesting that the mitochondrial bioenergetic disruption via mitochondrial uncoupling is necessary for nemorosone-induced ferroptosis. Our results open novel opportunities for cancer cell killing by mitochondrial uncoupling-induced ferroptosis.
Asunto(s)
Ferroptosis , Neoplasias , Humanos , Muerte Celular , Benzofenonas/farmacología , Neoplasias/metabolismo , Glutatión/metabolismoRESUMEN
The use of nanomaterials rationally engineered to treat cancer is a burgeoning field that has reported great medical achievements. Iron-based polymeric nano-formulations with precisely tuned physicochemical properties are an expanding and versatile therapeutic strategy for tumor treatment. Recently, a peculiar type of regulated necrosis named ferroptosis has gained increased attention as a target for cancer therapy. Here, we show for the first time that novel iron oxide nanoparticles coated with gallic acid and polyacrylic acid (IONP-GA/PAA) possess intrinsic cytotoxic activity on various cancer cell lines. Indeed, IONP-GA/PAA treatment efficiently induces ferroptosis in glioblastoma, neuroblastoma, and fibrosarcoma cells. IONP-GA/PAA-induced ferroptosis was blocked by the canonical ferroptosis inhibitors, including deferoxamine and ciclopirox olamine (iron chelators), and ferrostatin-1, the lipophilic radical trap. These ferroptosis inhibitors also prevented the lipid hydroperoxide generation promoted by the nanoparticles. Altogether, we report on novel ferroptosis-inducing iron encapsulated nanoparticles with potent anti-cancer properties, which has promising potential for further in vivo validation.
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Ferroptosis , Nanopartículas , Neoplasias , Apoptosis , Línea Celular Tumoral , Hierro/metabolismo , Nanopartículas Magnéticas de Óxido de HierroRESUMEN
BACKGROUND: Ischemic stroke produces a large health impact worldwide, with scarce therapeutic options. OBJECTIVE: This study aimed to reveal the role of NADPH oxidase and neuroinflammatory genes in the cerebral anti-ischemic effects of C-Phycocyanin (C-PC), the chief biliprotein of Spirulina platensis. METHODS: Rats with either focal cerebral ischemia/reperfusion (I/R) or acute brain hypoperfusion, received C-PC at different doses, or a vehicle, for up to 6 h post-stroke. Neurological, behavioral and histochemical parameters were assessed in I/R rats at 24 h. Cerebral gene expression and hippocampal neuron viability were evaluated in hypoperfused rats at acute (24 h) or chronic phases (30 days), respectively. A molecular docking analysis of NOX2 and C-PC-derived Phycocyanobilin (PCB) was also performed. RESULTS: C-PC, obtained with a purity of 4.342, significantly reduced the infarct volume and neurological deficit in a dose-dependent manner, and improved the exploratory activity of I/R rats. This biliprotein inhibited NOX2 expression, a crucial NADPH oxidase isoform in the brain, and the superoxide increase produced by the ischemic event. Moreover, C-PC-derived PCB showed a high binding affinity in silico with NOX2. C-PC downregulated the expression of pro-inflammatory genes (IFN-γ, IL-6, IL-17A, CD74, CCL12) and upregulated immune suppressive genes (Foxp3, IL-4, TGF-ß) in hypoperfused brain areas. This compound also decreased chronic neuronal death in the hippocampus of hypoperfused rats. CONCLUSION: These results suggest that the inhibition of cerebral NADPH oxidase and the improvement of neuroinflammation are key mechanisms mediating the neuroprotective actions of C-PC against brain ischemia.
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Isquemia Encefálica , Fármacos Neuroprotectores , Daño por Reperfusión , Animales , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/metabolismo , Modelos Animales de Enfermedad , Simulación del Acoplamiento Molecular , NADPH Oxidasas/metabolismo , Fármacos Neuroprotectores/farmacología , Fármacos Neuroprotectores/uso terapéutico , Ficocianina/farmacología , Ficocianina/uso terapéutico , Ratas , Daño por Reperfusión/tratamiento farmacológicoRESUMEN
The number of people with dementia worldwide is estimated at 50 million by 2018 and continues to rise mainly due to increasing aging and population growth. Clinical impact of current interventions remains modest and all efforts aimed at the identification of new therapeutic approaches are therefore critical. Previously, we showed that JM-20, a dihydropyridine-benzodiazepine hybrid molecule, protected memory processes against scopolamine-induced cholinergic dysfunction. In order to gain further insight into the therapeutic potential of JM-20 on cognitive decline and Alzheimer's disease (AD) pathology, here we evaluated its neuroprotective effects after chronic aluminum chloride (AlCl3) administration to rats and assessed possible alterations in several types of episodic memory and associated pathological mechanisms. Oral administration of aluminum to rodents recapitulates several neuropathological alterations and cognitive impairment, being considered a convenient tool for testing the efficacy of new therapies for dementia. We used behavioral tasks to test spatial, emotional- associative and novel object recognition memory, as well as molecular, enzymatic and histological assays to evaluate selected biochemical parameters. Our study revealed that JM-20 prevented memory decline alongside the inhibition of AlCl3 -induced oxidative stress, increased AChE activity, TNF-α and pro-apoptotic proteins (like Bax, caspase-3, and 8) levels. JM-20 also protected against neuronal damage in the hippocampus and prefrontal cortex. Our findings expanded our understanding of the ability of JM-20 to preserve memory in rats under neurotoxic conditions and confirm its potential capacity to counteract cognitive impairment and etiological factors of AD by breaking the progression of key steps associated with neurodegeneration.
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Cloruro de Aluminio/toxicidad , Benzodiazepinas/farmacología , Trastornos de la Memoria/inducido químicamente , Memoria/efectos de los fármacos , Neuronas/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Niacina/análogos & derivados , Cloruro de Aluminio/antagonistas & inhibidores , Animales , Hipocampo/efectos de los fármacos , Masculino , Aprendizaje por Laberinto/efectos de los fármacos , Trastornos de la Memoria/tratamiento farmacológico , Mitocondrias/efectos de los fármacos , Prueba del Laberinto Acuático de Morris/efectos de los fármacos , Niacina/farmacología , Prueba de Campo Abierto/efectos de los fármacos , Corteza Prefrontal/efectos de los fármacos , Ratas , Ratas Wistar , Prueba de Desempeño de Rotación con Aceleración ConstanteRESUMEN
The disruption of iron homeostasis is an important factor in the loss of mitochondrial function in neural cells, leading to neurodegeneration. Here, we assessed the protective action of gossypitrin (Gos), a naturally occurring flavonoid, on iron-induced neuronal cell damage using mouse hippocampal HT-22 cells and mitochondria isolated from rat brains. Gos was able to rescue HT22 cells from the damage induced by 100 µM Fe(II)-citrate (EC50 8.6 µM). This protection was linked to the prevention of both iron-induced mitochondrial membrane potential dissipation and ATP depletion. In isolated mitochondria, Gos (50 µM) elicited an almost complete protection against iron-induced mitochondrial swelling, the loss of mitochondrial transmembrane potential and ATP depletion. Gos also prevented Fe(II)-citrate-induced mitochondrial lipid peroxidation with an IC50 value (12.45 µM) that was about nine time lower than that for the tert-butylhydroperoxide-induced oxidation. Furthermore, the flavonoid was effective in inhibiting the degradation of both 15 and 1.5 mM 2-deoxyribose. It also decreased Fe(II) concentration with time, while increasing O2 consumption rate, and impairing the reduction of Fe(III) by ascorbate. Gos-Fe(II) complexes were detected by UV-VIS and IR spectroscopies, with an apparent Gos-iron stoichiometry of 2:1. Results suggest that Gos does not generally act as a classical antioxidant, but it directly affects iron, by maintaining it in its ferric form after stimulating Fe(II) oxidation. Metal ions would therefore be unable to participate in a Fenton-type reaction and the lipid peroxidation propagation phase. Hence, Gos could be used to treat neuronal diseases associated with iron-induced oxidative stress and mitochondrial damage.
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Flavonoides/farmacología , Hierro/efectos adversos , Mitocondrias/metabolismo , Neuronas/citología , Adenosina Trifosfato/metabolismo , Animales , Línea Celular , Supervivencia Celular/efectos de los fármacos , Ácido Cítrico/efectos adversos , Compuestos Ferrosos/efectos adversos , Peroxidación de Lípido/efectos de los fármacos , Masculino , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Ratones , Mitocondrias/efectos de los fármacos , Neuronas/efectos de los fármacos , Neuronas/metabolismo , RatasRESUMEN
OBJECTIVE: JM-20, a novel hybrid synthetic molecule, has been reported to have antioxidant, mitoprotective, anti-excitotoxic, anti-apoptotic and anti-inflammatory properties. However, the neuroprotective effect of JM-20 against memory impairment in preclinical AD-like models has not been analyzed. The aim of this study was to evaluate the potential neuroprotection of JM-20 that preserves essential memory process from cholinergic dysfunction and other molecular damages. METHODS: The effects of JM-20 on scopolamine (1 mg/kg)-induced cognitive disorders were studied. Male Wistar rats (220-230 g) were treated with JM-20 and/or scopolamine, and behavioral tasks were performed. The AChE activity, superoxide dismutase activity, catalase activity, MDA and T-SH level on brain tissue were determined by spectrophotometric methods. Mitochondrial functionality parameters were measured after behavioral tests. Histological analyses on hippocampus and prefrontal cortex were processed with hematoxylin and eosin, and neuronal and axonal damage were determined. RESULTS: The behavioral, biochemical and histopathological studies revealed that oral pre-treatment with JM-20 (8 mg/kg) significantly attenuated the scopolamine-induced memory deficits, mitochondrial malfunction, oxidative stress, and prevented AChE hyperactivity probably due to specific inhibition of AChE enzyme. It was also observed marked histological protection on hippocampal and prefrontal-cortex regions. CONCLUSIONS: The multimodal action of this molecule could mediate the memory protection here observed and suggest that it may modulate different pathological aspects of memory deï¬cits associated with AD in humans.
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Benzodiazepinas/farmacología , Inhibidores de la Colinesterasa/farmacología , Disfunción Cognitiva/tratamiento farmacológico , Memoria/efectos de los fármacos , Niacina/análogos & derivados , Nootrópicos/farmacología , Acetilcolinesterasa/metabolismo , Animales , Antioxidantes/metabolismo , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Encéfalo/patología , Disfunción Cognitiva/metabolismo , Disfunción Cognitiva/patología , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Peroxidación de Lípido/efectos de los fármacos , Peroxidación de Lípido/fisiología , Masculino , Memoria/fisiología , Trastornos de la Memoria/tratamiento farmacológico , Trastornos de la Memoria/metabolismo , Trastornos de la Memoria/patología , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Niacina/farmacología , Distribución Aleatoria , Ratas Wistar , EscopolaminaRESUMEN
Stroke is frequently associated with severe neurological decline and mortality, and its incidence is expected to increase due to aging population. The only available pharmacological treatment for cerebral ischemia is thrombolysis, with narrow therapeutic windows. Efforts aimed to identify new therapeutics are crucial. In this study, we look into plausible molecular and cellular targets for JM-20, a new hybrid molecule, against ischemic stroke in vivo. Male Wistar rats were subjected to 90 min middle cerebral artery occlusion (MCAO) following 23 h of reperfusion. Animals treated with 8 mg/kg JM-20 (p.o., 1 h after reperfusion) showed minimal neurological impairment and lower GABA and IL-1ß levels in CSF when compared to damaged rats that received vehicle. Immunocontent of pro-survival, phosphorylated Akt protein decreased in the cortex after 24 h as result of the ischemic insult, accompanied by decreased number of NeuN+ cells in the peri-infarct cortex, cornu ammonis 1 (CA1) and dentate gyrus (DG) areas. Widespread reactive astrogliosis in both cortex and hippocampus (CA1, CA3, and DG areas) was observed 24 h post-ischemia. JM-20 prevented the activated Akt reduction, neuronal death, and astrocytes reactivity throughout the brain. Overall, the results reinforce the pharmacological potential of JM-20 as neuroprotective agent and provide important evidences about its molecular and cellular targets in this model of cerebral ischemia.
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Astrocitos/patología , Benzodiazepinas/uso terapéutico , Infarto Encefálico/tratamiento farmacológico , Encéfalo/patología , Infarto de la Arteria Cerebral Media/tratamiento farmacológico , Infarto de la Arteria Cerebral Media/patología , Neuronas/patología , Niacina/análogos & derivados , Animales , Astrocitos/efectos de los fármacos , Astrocitos/metabolismo , Benzodiazepinas/farmacología , Infarto Encefálico/líquido cefalorraquídeo , Infarto Encefálico/patología , Región CA3 Hipocampal/efectos de los fármacos , Región CA3 Hipocampal/metabolismo , Región CA3 Hipocampal/patología , Muerte Celular/efectos de los fármacos , Giro Dentado/efectos de los fármacos , Giro Dentado/metabolismo , Giro Dentado/patología , Proteína Ácida Fibrilar de la Glía/metabolismo , Gliosis/metabolismo , Gliosis/patología , Infarto de la Arteria Cerebral Media/líquido cefalorraquídeo , Interleucina-10/líquido cefalorraquídeo , Interleucina-1beta/líquido cefalorraquídeo , Masculino , Neuronas/efectos de los fármacos , Niacina/farmacología , Niacina/uso terapéutico , Fosforilación/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Ratas Wistar , Resultado del Tratamiento , Ácido gamma-Aminobutírico/líquido cefalorraquídeoRESUMEN
Oxidative stress and mitochondrial dysfunction are two pathophysiological factors often associated with the neurodegenerative process involved in Parkinson's disease (PD). The aim of this study was to investigate the effects of a novel hybrid molecule, named JM-20, in different in vitro and in vivo models of PD induced by rotenone. To perform in vitro studies, SHSY-5Y cells were exposed to rotenone and/or treated with JM-20. To perform in vivo studies male Wistar rats were intoxicated with rotenone (2.5 mg/kg) via intraperitoneal injection and/or treated with JM-20 (40 mg/kg) administered via oral (for 25 days, both treatment). Rats were evaluated for global motor activity by measurement of locomotor activity. In addition, the effects on mortality, general behavior and redox parameters were also investigated. JM-20 protected SHSY-5Y cells against rotenone-induced cytotoxicity, evidenced by a significant diminution of cell death. In in vivo studies, JM-20 prevented rotenone-induced vertical exploration and locomotion frequency reductions, moreover prevented body weight loss and mortality induced by rotenone. It also improved the redox state of rotenone-exposured animals by increasing superoxide dismutase and catalase activities, total tissue-SH levels and decreasing malondialdehyde concentrations. Finally, JM-20 inhibited spontaneous mitochondrial swelling and membrane potential dissipation in isolated rats brain mitochondria. These results demonstrate that JM-20 is a potential neuroprotective agent against rotenone-induced damage in both in vitro and in vivo models, resulting in reduced neuronal oxidative injury and protection of mitochondria from impairment.
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Benzodiazepinas/farmacología , Fármacos Neuroprotectores/farmacología , Síndromes de Neurotoxicidad/prevención & control , Niacina/análogos & derivados , Rotenona/toxicidad , Animales , Peso Corporal/efectos de los fármacos , Encéfalo/metabolismo , Catalasa/metabolismo , Muerte Celular/efectos de los fármacos , Células Cultivadas , Humanos , Masculino , Mitocondrias/metabolismo , Actividad Motora/efectos de los fármacos , Niacina/farmacología , Estrés Oxidativo/efectos de los fármacos , Ratas , Superóxido Dismutasa/metabolismoRESUMEN
Introducción: Croton wagneri Müll. Arg. es una planta endémica del Ecuador, que dentro de la cosmovisión andina ha sido utilizada por los ancestros como planta medicinal. Objetivo: Evaluar la toxicidad aguda por vía oral del extracto hidroalcohólico de C. wagneri y su efecto irritante sobre la mucosa bucal. Método: Se evaluó la toxicidad aguda por vía oral del extracto hidroalcohólico de C. wagneri en 24 ratas hembras albinas Wistar procedentes de Centro para la Producción de Animales de Laboratorio (CENPALAB) de acuerdo con las normas de la Organización para la Cooperación Económica y el Desarrollo (OECD/OCDE 423). Los animales se dividieron en 4 grupos para recibir los extractos de las hojas, los tallos, las inflorescencias y de la planta completa en dosis de 5, 50, 300 y 2 000 mg/kg por vía oral, respectivamente. Después de 14 días de observación se sacrificaron los animales para su evaluación histopatológica. También se evaluó la acción irritante sobre la mucosa bucal en 30 hámster sirios dorados. A los siete días se sacrificaron los animales para su evaluación. Se utilizó el programa estadístico GraphPad Prism 5 para las evaluaciones biológicas. Resultados: Los extractos de las distintas partes de la planta no manifiestan ningún signo clínico de lesión en los pulmones, los riñones, el corazón, el bazo o el estómago, ni con la administración por vía oral de la dosis máxima probada de 2 000 mg/kg, tampoco irritación de la mucosa. Conclusiones: No se observaron cambios en el comportamiento de las ratas albinas Wistar ni en el de los hámsteres sirios dorados, por lo que se concluye que no hubo cambios en los signos clínicos de los animales en estudio.
Introduction: Croton wagneri Müll. Arg. is a plant species endemic to Ecuador used for medicinal purposes in ancestral Andean tradition. Objective: Evaluate the acute toxicity of orally administered C. wagneri hydroalcoholic extract and its irritating effect on the oral mucosa. Method: Acute toxicity of orally administered C. wagneri hydroalcoholic extract was evaluated in 24 female albino Wistar rats obtained from the Center for Laboratory Animal Breeding (CENPALAB) in compliance with standards issued by the Organization for Economic Co-operation and Development (OECD/OCDE 423). The animals were divided into four groups to receive oral administration of extracts from leaves, stems, inflorescences and the whole plant at doses of 5, 50, 300 and 2 000 mg/kg, respectively. After 14 days' observation, the animals were sacrificed for histopathological examination. Irritating action on the oral mucosa was evaluated in 30 Syrian golden hamsters. Seven days later the animals were sacrificed for evaluation. The statistical software GraphPad Prism 5 was used for biological evaluation. Results: Extracts from the different parts of the plant were not found to cause any clinical sign of injury to the lungs, kidneys, heart, spleen or stomach, not even when administered orally at the maximum test dose of 2 000 mg/kg, nor did they have an irritating effect on the mucosa. Conclusions: Changes were not observed in the behavior of albino Wistar rats or Syrian golden hamsters. It is thus concluded that no changes occurred in the clinical signs of the animals studied.
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Animales , Ratas , Croton/toxicidad , Mucosa Bucal , Plantas Medicinales , Cuba , Medicina TradicionalRESUMEN
Ischemic stroke is a major cause of death and disability worldwide. Thrombolysis by tissue plasminogen activator is the only pharmacological treatment approved for clinical practice, but has a narrow therapeutic window and poor efficacy when the cell death cascade is activated. Numerous drugs that are thought to protect neurons against injury have previously failed in human trials despite showing efficacy in experimental models of stroke. Herein, we reviewed the main pre-clinical results of the neuroprotective effects of JM-20, a new hybrid molecule, against brain ischemia. JM-20 appears to protect the brain from ischemic damage by interfering with several elements of the ischemic cascade: antiexcitotoxic, anticalcic, antioxidant, antiapoptotic, and anti-inflammatory. Its ability to protect not only neurons but also glial cells together with its ability to target and preserve mitochondrial function makes JM-20 a promising molecule that may be able to shield the whole neurovascular unit. The multimodal and multi-cell action of JM-20 may explain the high degree of protection observed in a rat model of brain ischemia, as assayed through histological (hematoxylin-eosin, and luxol fast blue staining), neurochemical (glutamate and aspartate levels in cerebrospinal fluid), mitochondrial functionality and behavioural (neurological scale) analysis at doses of 4 and 8mg/kg. Furthermore, the wide therapeutic window of JM-20 of 8h also suggests that this molecule could be of potential interest in situations where brain perfusion is compromised.
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Benzodiazepinas/farmacología , Isquemia Encefálica/prevención & control , Niacina/análogos & derivados , Animales , Evaluación Preclínica de Medicamentos , Fármacos Neuroprotectores/farmacología , Niacina/farmacologíaRESUMEN
Cardiovascular diseases are major causes of death worldwide. Beyond the classical cholesterol risk factor, other conditions such as oxidative stress are well documented to promote atherosclerosis. The Mangifera indica L. extract (Vimang®) was reported to present antioxidant and hypocholesterolemic properties. Thus, here we evaluate the effects of Vimang treatment on risk factors of the atherosclerosis prone model of familial hypercholesterolemia, the LDL receptor knockout mice. Mice were treated with Vimang during 2 weeks and were fed a cholesterol-enriched diet during the second week. The Vimang treated mice presented significantly reduced levels of plasma (15%) and liver (20%) cholesterol, increased plasma total antioxidant capacity (10%) and decreased reactive oxygen species (ROS) production by spleen mononuclear cells (50%), P < 0.05 for all. In spite of these benefits, the average size of aortic atherosclerotic lesions stablished in this short experimental period did not change significantly in Vimang treated mice. Therefore, in this study we demonstrated that Vimang has protective effects on systemic and tissue-specific risk factors, but it is not sufficient to promote a reduction in the initial steps of atherosclerosis development. In addition, we disclosed a new antioxidant target of Vimang, the spleen mononuclear cells that might be relevant for more advanced stages of atherosclerosis.
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Colesterol/sangre , Mangifera/química , Estrés Oxidativo/efectos de los fármacos , Extractos Vegetales/farmacología , Receptores de LDL/genética , Animales , Aorta/patología , Aterosclerosis/metabolismo , Aterosclerosis/patología , Aterosclerosis/veterinaria , Colesterol/análisis , Dieta Alta en Grasa , Leucocitos/citología , Leucocitos/metabolismo , Hígado/efectos de los fármacos , Hígado/metabolismo , Mangifera/metabolismo , Ratones , Ratones Noqueados , Mitocondrias/metabolismo , NADP/química , NADP/metabolismo , Extractos Vegetales/química , Especies Reactivas de Oxígeno/metabolismo , Receptores de LDL/deficiencia , Triglicéridos/análisis , Triglicéridos/sangreRESUMEN
This study aims to examine the effects of a new 1,4-dihydropyridine derivative, VdiE-2N, on cell signaling pathways and mitochondrial events in head and neck squamous cell carcinoma (HNSCC) cells, and on a mice model of xenograft tumor growth/cell proliferation. Four HNSCC cell lines (HN13, HN12, HN6, and CAL27), HEK293 cells (human embryonic kidney 293 cells), and human oral healthy mucosa fibroblasts (OHMF) were used for in vitro assessment of cell viability (resazurin assay) and invasion capacity (modified Boyden chamber assay), and mitochondrial membrane potential (JC-1 fluorescence assay), morphology (transmission electron microscopy), and number of mitochondria (MitoTracker® imaging). SET and pDRP1 proteins were analyzed by immunofluorescence, and proteins involved in cell death/survival pathways were analyzed by Western blotting. HN12 xenograft tumors were established in the flank of Balb/c nude mice, and their characteristics and sensitivity to VdiE-2N were determined by immunohistochemistry and histology. VdiE-2N decreased cell viability in HNSCC cells (IC50 = 9.56 and 22.45µM for HN13 and HN12 cells, respectively) more strongly than it decreased cell viability in OHMF and HEK293 cells (IC50 = 32.90 and > 50µM, respectively). In HN13 cells, VdiE-2N dissipated mitochondrial membrane potential and altered the mitochondria size, shape, and number in a concentration-dependent manner, as well as it induced apoptosis and reduced their invasion capacity. Treatment of mice bearing xenograft tumors with VdiE-2N significantly diminished proliferation of cancer cells. Therefore, VdiE-2N induces HNSCC cell death in vitro through mitochondria-mediated apoptotic pathways and dampens tumor growth in vivo, thus supporting a potential anti-cancer effect.
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Antineoplásicos/química , Antineoplásicos/farmacología , Carcinoma de Células Escamosas/patología , Dihidropiridinas/química , Dihidropiridinas/farmacología , Neoplasias de Cabeza y Cuello/patología , Animales , Apoptosis/efectos de los fármacos , Carcinoma de Células Escamosas/irrigación sanguínea , Carcinoma de Células Escamosas/tratamiento farmacológico , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Genes myc/genética , Neoplasias de Cabeza y Cuello/irrigación sanguínea , Neoplasias de Cabeza y Cuello/tratamiento farmacológico , Humanos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Ratones , Tamaño Mitocondrial/efectos de los fármacos , Invasividad Neoplásica , Neovascularización Patológica/tratamiento farmacológico , Carcinoma de Células Escamosas de Cabeza y Cuello , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Several 1,4-dihydropyridine derivatives overcome the multidrug resistance in tumors, but their intrinsic cytotoxic mechanisms remain unclear. Here we addressed if mitochondria are involved in the cytotoxicity of the novel 1,4-dihydropyridine derivative VE-3N [ethyl 6-chloro-5-formyl-2-methyl-4-(3-nitrophenyl)-1,4-dihydropyridine-3-carboxylate] towards cancer cells by employing hepatic carcinoma (HepG2) cells and isolated rat liver mitochondria. In HepG2 cells, VE-3N induced mitochondrial membrane potential dissipation, ATP depletion, annexin V/propidium iodide double labeling, and Hoechst staining; events indicating apoptosis induction. In isolated rat liver mitochondria, VE-3N promoted mitochondrial uncoupling by exerting protonophoric actions and by increasing membrane fluidity. Mitochondrial uncoupling was evidenced by an increase in resting respiration, dissipation of mitochondrial membrane potential, inhibition of Ca2+ uptake, stimulation of Ca2+ release, decrease in ATP synthesis, and swelling of valinomycin-treated organelles in hyposmotic potassium acetate media. Furthermore, uncoupling concentrations of VE-3N in the presence of Ca2+ plus ruthenium red induced the mitochondrial permeability transition process. These results indicate that mitochondrial uncoupling is potentially involved in the VE-3N cytotoxic actions towards HepG2 cells. Considering that hepatocellular carcinoma is the most common form of liver cancer, our findings may open a new avenue for the development of VE-3N-based cancer therapies, and help to unravel the cytotoxic mechanisms of 1,4-dihydropyridines towards cancer cells.
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Antineoplásicos/farmacología , Dihidropiridinas/farmacología , Mitocondrias Hepáticas/efectos de los fármacos , Desacopladores/farmacología , Adenosina Trifosfato/metabolismo , Animales , Supervivencia Celular/efectos de los fármacos , Metabolismo Energético/efectos de los fármacos , Células Hep G2 , Humanos , Masculino , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Mitocondrias Hepáticas/metabolismo , Ratas , Ratas WistarRESUMEN
OBJECTIVE: Scopolamine (SCO) administration to rats induces molecular features of AD and other dementias, including impaired cognition, increased oxidative stress, and imbalanced cholinergic transmission. Although mitochondrial dysfunction is involved in different types of dementias, its role in cognitive impairment induced by SCO has not been well elucidated. The aim of this work was to evaluate the in vivo effect of SCO on different brain mitochondrial parameters in rats to explore its neurotoxic mechanisms of action. METHODS: Saline (Control) or SCO (1 mg/kg) was administered intraperitoneally 30 min prior to neurobehavioral and biochemical evaluations. Novel object recognition and Y-maze paradigms were used to evaluate the impact on memory, while redox profiles in different brain regions and the acetylcholinesterase (AChE) activity of the whole brain were assessed to elucidate the amnesic mechanism of SCO. Finally, the effects of SCO on brain mitochondria were evaluated both ex vivo and in vitro, the latter to determine whether SCO could directly interfere with mitochondrial function. RESULTS: SCO administration induced memory deficit, increased oxidative stress, and increased AChE activities in the hippocampus and prefrontal cortex. Isolated brain mitochondria from rats administered with SCO were more vulnerable to mitochondrial swelling, membrane potential dissipation, H2O2 generation and calcium efflux, all likely resulting from oxidative damage. The in vitro mitochondrial assays suggest that SCO did not affect the organelle function directly. CONCLUSION: In conclusion, the present results indicate that SCO induced cognitive dysfunction and oxidative stress may involve brain mitochondrial impairment, an important target for new neuroprotective compounds against AD and other dementias.
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Trastornos de la Memoria/metabolismo , Mitocondrias/metabolismo , Acetilcolinesterasa/metabolismo , Animales , Encéfalo/metabolismo , Calcio/metabolismo , Cationes Bivalentes/metabolismo , Modelos Animales de Enfermedad , Peróxido de Hidrógeno/metabolismo , Masculino , Aprendizaje por Laberinto/fisiología , Potencial de la Membrana Mitocondrial/fisiología , Dilatación Mitocondrial/fisiología , Estrés Oxidativo/fisiología , Distribución Aleatoria , Ratas Wistar , Reconocimiento en Psicología/fisiología , EscopolaminaRESUMEN
Oxidative stress resulting from iron and reactive oxygen species (ROS) homeostasis breakdown has been implicated in several diseases. Therefore, molecules capable of binding iron and/or scavenging ROS may be reasonable strategies for protecting cells. Rapanone is a naturally occurring hydroxyl-benzoquinone with a privileged chelating structure. In this work, we addressed the antioxidant properties of rapanone concerning its iron-chelating and scavenging activities, and its protective potential against iron and tert-butyl hydroperoxide-induced damage to mitochondria. Experimental determinations revealed the formation of rapanone-Fe(II)/Fe(III) complexes. Additionally, the electrochemical assays indicated that rapanone oxidized Fe(II) and O2-, thus inhibiting Fenton-Haber-Weiss reactions. Furthermore, rapanone displayed an increased 2,2-diphenyl-1-picrylhydrazyl radical scavenging ability in the presence of Fe(II). The above results explained the capacity of rapanone to provide near-full protection against iron and tert-butyl hydroperoxide induced mitochondrial lipid peroxidation in energized organelles, which fail under non-energized condition. We postulate that rapanone affords protection against iron and reactive oxygen species by means of both iron chelating and iron-stimulated free radical scavenging activity.
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Benzoquinonas/química , Complejos de Coordinación/química , Depuradores de Radicales Libres/química , Hierro/químicaRESUMEN
Cerebral ischemia is the third most common cause of death and a major cause of disability worldwide. Beyond a shortage of essential metabolites, ischemia triggers many interconnected pathophysiological events, including excitotoxicity, oxidative stress, inflammation and apoptosis. Here, we investigated the neuroprotective mechanisms of JM-20, a novel synthetic molecule, focusing on the phosphoinositide-3-kinase (PI3K)/Akt survival pathway and glial cell response as potential targets of JM-20. For this purpose, we used organotypic hippocampal slice cultures exposed to oxygen-glucose deprivation (OGD) to achieve ischemic/reperfusion damage in vitro. Treatment with JM-20 at 0.1 and 10 µM reduced PI incorporation (indicative of cell death) after OGD. OGD decreased the phosphorylation of Akt (pro-survival) and GSK 3ß (pro-apoptotic), resulting in respective inhibition and activation of these proteins. Treatment with JM20 prevented the reduced phosphorylation of these proteins after OGD, representing a shift from pro-apoptotic to pro-survival signaling. The OGD-induced activation of caspase-3 was also attenuated by JM-20 treatment at 10 µM. Moreover, in cultures treated with JM-20 and exposed to OGD conditioning, we observed a decrease in activated microglia, as well as a decrease in interleukin (IL)-1ß, IL-6 and tumor necrosis factor (TNF)-α release into the culture medium, while the level of the anti-inflammatory IL-10 increased. GFAP immunostaining and IB4 labeling showed that JM-20 treatment significantly augmented GFAP immunoreactivity after OGD, when compared with cultures exposed to OGD only, suggesting the activation of astroglial cells. Our results confirm that JM-20 has a strong neuroprotective effect against ischemic injury and suggest that the mechanisms involved in this effect may include the modulation of reactive astrogliosis, as well as neuroinflammation and the anti-apoptotic cell signaling pathway.
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Benzodiazepinas/farmacología , Muerte Celular/efectos de los fármacos , Glucógeno Sintasa Quinasa 3/metabolismo , Hipocampo/efectos de los fármacos , Niacina/análogos & derivados , Oxígeno/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Animales , Animales Recién Nacidos , Glucosa/metabolismo , Glucógeno Sintasa Quinasa 3 beta , Hipocampo/metabolismo , Masculino , Neuronas/efectos de los fármacos , Neuroprotección/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Niacina/farmacología , Fosfatidilinositol 3-Quinasas/metabolismo , Ratas WistarRESUMEN
Three main types of Cuban propolis directly related to their secondary metabolite composition have been identified: brown, red and yellow propolis; the former is majoritarian and is characterized by the presence of nemorosone. In this study, brown Cuban propolis extracts were found cytotoxic against HepG2 cells and primary rat hepatocytes, in close association with the nemorosone contents. In mitochondria isolated from rat liver the extracts displayed uncoupling activity, which was demonstrated by the increase in succinate-supported state 4 respiration rates, dissipation of mitochondrial membrane potential, Ca(2+) release from Ca(2+)-loaded mitochondria, and a marked ATP depletion. As in cells, the degree of such mitotoxic events was closely correlated to the nemorosone content. The propolis extracts that do not contain nemorosone were neither cytotoxic nor mitotoxic, except R-29, whose detrimental effect upon cells and mitochondria could be mediated by its isoflavonoids and chalcones components, well known mitochondrial uncouplers. Our results at least partly unravel the cytotoxic mechanism of Cuban propolis, particularly regarding brown propolis, and raise concerns about the toxicological implication of Cuban propolis consumption.
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Benzofenonas/farmacología , Mitocondrias Hepáticas/efectos de los fármacos , Extractos Vegetales/química , Extractos Vegetales/farmacología , Própolis/química , Própolis/farmacología , Desacopladores/farmacología , Animales , Calcio/metabolismo , Supervivencia Celular/efectos de los fármacos , Cuba , Relación Dosis-Respuesta a Droga , Células Hep G2 , Hepatocitos/efectos de los fármacos , Humanos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Mitocondrias Hepáticas/metabolismo , Ratas , Relación Estructura-Actividad , Desacopladores/químicaRESUMEN
JM-20 (3-ethoxycarbonyl-2-methyl-4-(2-nitrophenyl)-4,11-dihydro-1H-pyrido[2,3-b][1,5]benzodiazepine) is a novel benzodiazepine dihydropyridine hybrid molecule, which has been shown to be a neuroprotective agent in brain disorders involving glutamate receptors. However, the effect of JM-20 on the functionality of the glutamatergic system has not been investigated. In this study, by using different in vitro preparations, we investigated the effects of JM-20 on (i) rat brain synaptic vesicles (L-[(3)H]-glutamate uptake, proton gradient built-up and bafilomycin-sensitive H(+)-ATPase activity), (ii) rat brain synaptosomes (glutamate release) and (iii) primary cultures of rat cortical neurons, astrocytes and astrocyte-neuron co-cultures (L-[(3)H]-glutamate uptake and glutamate release). We observed here that JM-20 impairs H(+)-ATPase activity and consequently reduces vesicular glutamate uptake. This molecule also inhibits glutamate release from brain synaptosomes and markedly increases glutamate uptake in astrocytes alone, and co-cultured neurons and astrocytes. The impairment of vesicular glutamate uptake by inhibition of the H(+)-ATPase caused by JM-20 could decrease the amount of the transmitter stored in synaptic vesicles, increase the cytosolic levels of glutamate, and will thus down-regulate neurotransmitter release. Together, these results contribute to explain the anti-excitotoxic effect of JM-20 and its strong neuroprotective effect observed in different in vitro and in vivo models of brain ischemia.