RESUMEN

Mitochondrial impairments in dynamic behavior (fusion/fission balance) associated with mitochondrial dysfunction play a key role in cell lipotoxicity and lipid-induced metabolic diseases. The present work aimed to evaluate dose- and time-dependent effects of the monounsaturated fatty acid oleate on mitochondrial fusion/fission proteins in comparison with the saturated fatty acid palmitate in hepatic cells. To this end, HepG-2 cells were treated with 0, 10 µM, 50 µM, 100 µM, 250 µM or 500 µM of either oleate or palmitate for 8 or 24 h. Cell viability and lipid accumulation were evaluated to assess lipotoxicity. Mitochondrial markers of fusion (mitofusin 2, MFN2) and fission (dynamin-related protein 1, DRP1) processes were evaluated by Western blot analysis. After 8 h, the highest dose of oleate induced a decrease in DRP1 content without changes in MFN2 content in association with cell viability maintenance, whereas palmitate induced a decrease in cell viability associated with a decrease mainly in MFN2 content. After 24 h, oleate induced MFN2 increase, whereas palmitate induced DRP1 increase associated with a higher decrease in cell viability with high doses compared to oleate. This finding could be useful to understand the role of mitochondria in the protective effects of oleate as a bioactive compound.

Asunto(s)

Dinaminas/genética , GTP Fosfohidrolasas/genética , Enfermedades Metabólicas/genética , Dinámicas Mitocondriales/efectos de los fármacos , Proteínas Mitocondriales/genética , Ácido Oléico/metabolismo , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Ácidos Grasos Monoinsaturados/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Células Hep G2 , Humanos , Metabolismo de los Lípidos/efectos de los fármacos , Lípidos/toxicidad , Enfermedades Metabólicas/etiología , Enfermedades Metabólicas/metabolismo , Enfermedades Metabólicas/patología , Mitocondrias/efectos de los fármacos , Mitocondrias/genética , Mitocondrias/patología , Dinámicas Mitocondriales/genética , Ácido Oléico/farmacología , Palmitatos/metabolismo , Palmitatos/farmacología

RESUMEN

Synaptic aging has been associated with neuronal circuit dysfunction and cognitive decline. Reduced mitochondrial function may be an early event that compromises synaptic integrity and neurotransmission in vulnerable brain regions during physiological and pathological aging. Thus, we aimed to measure mitochondrial function in synapses from three brain regions at two different ages in the 3xTg-AD mouse model and in wild mice. We found that aging is the main factor associated with the decline in synaptic mitochondrial function, particularly in synapses isolated from the cerebellum. Accumulation of toxic compounds, such as tau and Aß, that occurred in the 3xTg-AD mouse model seemed to participate in the worsening of this decline in the hippocampus. The changes in synaptic bioenergetics were also associated with increased activation of the mitochondrial fission protein Drp1. These results suggest the presence of altered mechanisms of synaptic mitochondrial dynamics and their quality control during aging and in the 3xTg-AD mouse model; they also point to bioenergetic restoration as a useful therapeutic strategy to preserve synaptic function during aging and at the early stages of Alzheimer's disease (AD).

Asunto(s)

Envejecimiento/genética , Disfunción Cognitiva/genética , Dinaminas/genética , Mitocondrias/metabolismo , Dinámicas Mitocondriales/genética , Envejecimiento/metabolismo , Péptidos beta-Amiloides/genética , Péptidos beta-Amiloides/metabolismo , Animales , Cerebelo/metabolismo , Cerebelo/fisiopatología , Corteza Cerebral/metabolismo , Corteza Cerebral/fisiopatología , Disfunción Cognitiva/metabolismo , Disfunción Cognitiva/fisiopatología , Modelos Animales de Enfermedad , Dinaminas/metabolismo , Femenino , Regulación de la Expresión Génica , Hipocampo/metabolismo , Hipocampo/fisiopatología , Humanos , Potencial de la Membrana Mitocondrial/genética , Ratones , Ratones Transgénicos , Mitocondrias/patología , Neuronas/metabolismo , Neuronas/patología , Especificidad de Órganos , Sinapsis/metabolismo , Sinapsis/patología , Sinaptosomas/metabolismo , Sinaptosomas/patología , Proteínas tau/genética , Proteínas tau/metabolismo

RESUMEN

Amyotrophic lateral sclerosis (ALS) is a multifactorial and progressive neurodegenerative disease of unknown etiology. Due to ALS's unpredictable onset and progression rate, the search for biomarkers that allow the detection and tracking of its development and therapeutic efficacy would be of significant medical value. Considering that alterations of energy supply are one of ALS's main hallmarks and that a correlation has been established between gene expression in human brain tissue and peripheral blood mononuclear cells (PBMCs), the present work investigates whether changes in mitochondrial function could be used to monitor ALS. To achieve this goal, PBMCs from ALS patients and control subjects were used; blood sampling is a quite non-invasive method and is cost-effective. Different parameters were evaluated, namely cytosolic calcium levels, mitochondrial membrane potential, oxidative stress, and metabolic compounds levels, as well as mitochondrial dynamics and degradation. Altogether, we observed lower mitochondrial calcium uptake/retention, mitochondria depolarization, and redox homeostasis deregulation, in addition to a decrease in critical metabolic genes, a diminishment in mitochondrial biogenesis, and an augmentation in mitochondrial fission and autophagy-related gene expression. All of these changes can contribute to the decreased ATP and pyruvate levels observed in ALS PBMCs. Our data indicate that PBMCs from ALS patients show a significant mitochondrial dysfunction, resembling several findings from ALS' neural cells/models, which could be exploited as a powerful tool in ALS research. Our findings can also guide future studies on new pharmacological interventions for ALS since assessments of brain samples are challenging and represent a relevant limited strategy. Graphical abstract.

Asunto(s)

Esclerosis Amiotrófica Lateral/sangre , Esclerosis Amiotrófica Lateral/genética , Biomarcadores/sangre , Leucocitos Mononucleares/metabolismo , Mitocondrias/metabolismo , Dinámicas Mitocondriales , Biogénesis de Organelos , Adulto , Anciano , Antioxidantes/metabolismo , Autofagia/genética , Calcio/metabolismo , Metabolismo Energético , Femenino , Regulación de la Expresión Génica , Humanos , Masculino , Potencial de la Membrana Mitocondrial/genética , Persona de Mediana Edad , Mitocondrias/genética , Dinámicas Mitocondriales/genética , Estrés Oxidativo/genética

RESUMEN

Cellular senescence is an endpoint of chemotherapy, and targeted therapies in melanoma and the senescence-associated secretory phenotype (SASP) can affect tumor growth and microenvironment, influencing treatment outcomes. Metabolic interventions can modulate the SASP, and an enhanced mitochondrial energy metabolism supports resistance to therapy in melanoma cells. Herein, we assessed the mitochondrial function of therapy-induced senescent melanoma cells obtained after exposing the cells to temozolomide (TMZ), a methylating chemotherapeutic agent. Senescence induction in melanoma was accompanied by a substantial increase in mitochondrial basal, ATP-linked, and maximum respiration rates and in coupling efficiency, spare respiratory capacity, and respiratory control ratio. Further examinations revealed an increase in mitochondrial mass and length. Alterations in mitochondrial function and morphology were confirmed in isolated senescent cells, obtained by cell-size sorting. An increase in mitofusin 1 and 2 (MFN1 and 2) expression and levels was observed in senescent cells, pointing to alterations in mitochondrial fusion. Silencing mitofusin expression with short hairpin RNA (shRNA) prevented the increase in mitochondrial length, oxygen consumption rate and secretion of interleukin 6 (IL-6), a component of the SASP, in melanoma senescent cells. Our results represent the first in-depth study of mitochondrial function in therapy-induced senescence in melanoma. They indicate that senescence increases mitochondrial mass, length and energy metabolism; and highlight mitochondria as potential pharmacological targets to modulate senescence and the SASP.

Asunto(s)

Senescencia Celular , Metabolismo Energético , GTP Fosfohidrolasas/metabolismo , Melanoma Experimental/metabolismo , Mitocondrias/metabolismo , Proteínas de Neoplasias/metabolismo , Animales , GTP Fosfohidrolasas/genética , Silenciador del Gen , Interleucina-6/genética , Interleucina-6/metabolismo , Melanoma Experimental/genética , Melanoma Experimental/patología , Ratones , Mitocondrias/genética , Mitocondrias/patología , Dinámicas Mitocondriales/efectos de los fármacos , Dinámicas Mitocondriales/genética , Proteínas de Neoplasias/genética , Temozolomida/farmacología

RESUMEN

DNA barcoding has become a standard method for species identification in taxonomically complex groups. An important step of the barcoding process is the construction of a library of voucher-based material that was properly identified by independent methods, free of inaccurate identification, and paralogs. We provide here a cytochrome oxidase I (mt-Co1) DNA barcode database for species of the genus Oligoryzomys, based on type material and karyotyped specimens, and anchored on the mitochondrial genome of one species of Oligoryzomys, O. stramineus. To evaluate the taxonomic determination of new COI sequences, we assessed species intra/interspecific genetic distances (barcode gap), performed the General Mixed Yule Coalescent method (GMYC) for lineages' delimitation, and identified diagnostic nucleotides for each species of Oligoryzomys. Phylogenetic analyses of Oligoryzomys were performed on 2 datasets including 14 of the 23 recognized species of this genus: a mt-Co1 only matrix, and a concatenated matrix including mt-Co1, cytochrome b (mt-Cytb), and intron 7 of the nuclear fibrinogen beta chain gene (i7Fgb). We recovered nuclear-mitochondrial translocated (Numts) pseudogenes on our samples and identified several published sequences that are cases of Numts. We analyzed the rate of non-synonymous and synonymous substitution, which were higher in Numts in comparison to mtDNA sequences. GMYC delimitations and DNA barcode gap results highlight the need for further work that integrate molecular, karyotypic, and morphological analyses, as well as additional sampling, to tackle persistent problems in the taxonomy of Oligoryzomys.

Asunto(s)

Arvicolinae/genética , Núcleo Celular/genética , Código de Barras del ADN Taxonómico , Bases de Datos Genéticas , Genoma Mitocondrial/genética , Dinámicas Mitocondriales/genética , Animales , Especificidad de la Especie

RESUMEN

Mouse skin mesenchymal stem cells (msMSCs) are dermis CD105(+) CD90(+) CD73(+) CD29(+) CD34(-) mesodermal precursors which, after in vitro induction, undergo chondro, adipo, and osteogenesis. Extensive metabolic reconfiguration has been found to occur during differentiation, and the bioenergetic status of a cell is known to be dependent on the quality and abundance of the mitochondrial population, which may be regulated by fusion and fission. However, little is known regarding the impact of mitochondrial dynamics on the differentiation process. We addressed this knowledge gap by isolating MSCs from Swiss female mice, inducing these cells to differentiate into osteo, chondro, and adipocytes and measuring changes in mass, morphology, dynamics, and bioenergetics. Mitochondrial biogenesis was increased in adipogenesis, as evaluated through confocal microscopy, citrate synthase activity, and mtDNA content. The early steps of adipo and osteogenesis involved mitochondrial elongation, as well as increased expression of mitochondrial fusion proteins Mfn1 and 2. Chondrogenesis involved a fragmented mitochondrial phenotype, increased expression of fission proteins Drp1, Fis1, and 2, and enhanced mitophagy. These events were accompanied by profound bioenergetic alterations during the commitment period. Moreover, knockdown of Mfn2 in adipo and osteogenesis and the overexpression of a dominant negative form of Drp1 during chondrogenesis resulted in a loss of differentiation ability. Overall, we find that mitochondrial morphology and its regulating processes of fission/fusion are modulated early on during commitment, leading to alterations in the bioenergetic profile that are important for differentiation. We thus propose a central role for mitochondrial dynamics in the maintenance/commitment of mesenchymal stem cells.

Asunto(s)

Diferenciación Celular/genética , Dinaminas/biosíntesis , GTP Fosfohidrolasas/biosíntesis , Células Madre Mesenquimatosas , Mitocondrias/metabolismo , Adipogénesis/genética , Animales , Condrogénesis/genética , ADN Mitocondrial/genética , Dinaminas/genética , Femenino , GTP Fosfohidrolasas/genética , Regulación del Desarrollo de la Expresión Génica/genética , Ratones , Mitocondrias/genética , Dinámicas Mitocondriales/genética , Osteogénesis/genética , Piel/citología , Piel/metabolismo

RESUMEN

The rate-limiting step in the biosynthesis of steroid hormones, known as the transfer of cholesterol from the outer to the inner mitochondrial membrane, is facilitated by StAR, the Steroidogenic Acute Regulatory protein. We have described that mitochondrial ERK1/2 phosphorylates StAR and that mitochondrial fusion, through the up-regulation of a fusion protein Mitofusin 2, is essential during steroidogenesis. Here, we demonstrate that mitochondrial StAR together with mitochondrial active ERK and PKA are necessary for maximal steroid production. Phosphorylation of StAR by ERK is required for the maintenance of this protein in mitochondria, observed by means of over-expression of a StAR variant lacking the ERK phosphorylation residue. Mitochondrial fusion regulates StAR levels in mitochondria after hormone stimulation. In this study, Mitofusin 2 knockdown and mitochondrial fusion inhibition in MA-10 Leydig cells diminished StAR mRNA levels and concomitantly mitochondrial StAR protein. Together our results unveil the requirement of mitochondrial fusion in the regulation of the localization and mRNA abundance of StAR. We here establish the relevance of mitochondrial phosphorylation events in the correct localization of this key protein to exert its action in specialized cells. These discoveries highlight the importance of mitochondrial fusion and ERK phosphorylation in cholesterol transport by means of directing StAR to the outer mitochondrial membrane to achieve a large number of steroid molecules per unit of StAR.

Asunto(s)

Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Mitocondrias/metabolismo , Dinámicas Mitocondriales , Fosfoproteínas/metabolismo , Animales , Línea Celular , AMP Cíclico/metabolismo , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , GTP Fosfohidrolasas/metabolismo , Regulación de la Expresión Génica , Masculino , Ratones , Mitocondrias/enzimología , Dinámicas Mitocondriales/genética , Modelos Biológicos , Fosfoproteínas/genética , Fosforilación , Fosfoserina/metabolismo , Transporte de Proteínas , ARN Mensajero/genética , ARN Mensajero/metabolismo , Esteroides/biosíntesis , Transcripción Genética

RESUMEN

Copper is essential in cell physiology, participating in numerous enzyme reactions. In mitochondria, copper is a cofactor for respiratory complex IV, the cytochrome c oxidase. Low copper content is associated with anemia and the appearance of enlarged mitochondria in erythropoietic cells. These findings suggest a connection between copper metabolism and bioenergetics, mitochondrial dynamics and erythropoiesis, which has not been explored so far. Here, we describe that bathocuproine disulfonate-induced copper deficiency does not alter erythropoietic cell proliferation nor induce apoptosis. However it does impair erythroid differentiation, which is associated with a metabolic switch between the two main energy-generating pathways. That is, from mitochondrial function to glycolysis. Switching off mitochondria implies a reduction in oxygen consumption and ROS generation along with an increase in mitochondrial membrane potential. Mitochondrial fusion proteins MFN2 and OPA1 were up-regulated along with the ability of mitochondria to fuse. Morphometric analysis of mitochondria did not show changes in total mitochondrial biomass but rather bigger mitochondria because of increased fusion. Similar results were also obtained with human CD34+, which were induced to differentiate into red blood cells. In all, we have shown that adequate copper levels are important for maintaining proper mitochondrial function and for erythroid differentiation where the energy metabolic switch plus the up-regulation of fusion proteins define an adaptive response to copper deprivation to keep cells alive.

Asunto(s)

Cobre/deficiencia , Metabolismo Energético , Células Eritroides/metabolismo , Eritropoyesis/fisiología , GTP Fosfohidrolasas/biosíntesis , Dinámicas Mitocondriales/genética , Proteínas Mitocondriales/biosíntesis , Regulación hacia Arriba , Apoptosis/genética , Muerte Celular/genética , Diferenciación Celular/genética , Proliferación Celular , Células Cultivadas , Cobre/metabolismo , Metabolismo Energético/genética , Células Eritroides/citología , Células Eritroides/patología , Eritropoyesis/genética , Humanos , Células K562 , Mitocondrias/genética , Mitocondrias/patología , Simulación de Dinámica Molecular , Regulación hacia Arriba/genética

RESUMEN

Recent studies showed that the activation of the retinoid X receptor, which dimerizes with peroxisome proliferator-activated receptors (PPARs), leads to an enhanced clearance of Aß from the brain of transgenic mice model of Alzheimer's disease (AD), because an increased expression of apolipoprotein E and it main transporters. However, the effects observed must involve additional underlying mechanisms that have not been yet explored. Several studies conducted in our laboratory suggest that part of the effects observed for the PPARs agonist might involves mitochondrial function and, particularly, mitochondrial dynamics. In the present study we assessed the effects of oxidative stress challenge on mitochondrial morphology and mitochondrial dynamics-related proteins in hippocampal neurons. Using immunofluorescence, we evaluated the PPARγ co-activator 1α (PGC-1α), dynamin related protein 1 (DRP1), mitochondrial fission protein 1 (FIS1), and mitochondrial length, in order to determine if PPARs agonist pre-treatment is able to protect mitochondrial population from hippocampal neurons through modulation of the mitochondrial fusion-fission events. Our results suggest that both a PPARγ agonist (ciglitazone) and a PPARα agonist (WY 14.643) are able to protect neurons by modulating mitochondrial fusion and fission, leading to a better response of neurons to oxidative stress, suggesting that a PPAR based therapy could acts simultaneously in different cellular components. Additionally, our results suggest that PGC-1α and mitochondrial dynamics should be further studied in future therapy research oriented to ameliorate neurodegenerative disorders, such as AD.

Asunto(s)

Mitocondrias/efectos de los fármacos , Dinámicas Mitocondriales/efectos de los fármacos , PPAR alfa/agonistas , PPAR gamma/agonistas , Pirimidinas/farmacología , Tiazolidinedionas/farmacología , Animales , Dinaminas/genética , Dinaminas/metabolismo , Embrión de Mamíferos , Regulación de la Expresión Génica , Hipocampo/citología , Hipocampo/efectos de los fármacos , Hipocampo/metabolismo , Mitocondrias/genética , Mitocondrias/metabolismo , Mitocondrias/ultraestructura , Dinámicas Mitocondriales/genética , Proteínas Mitocondriales/genética , Proteínas Mitocondriales/metabolismo , Neuronas/citología , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Estrés Oxidativo , PPAR alfa/genética , PPAR alfa/metabolismo , PPAR gamma/genética , PPAR gamma/metabolismo , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma , Cultivo Primario de Células , Ratas , Ratas Sprague-Dawley , Especies Reactivas de Oxígeno/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo