RESUMO
Swim training has increased the life span of the transgenic animal model of amyotrophic lateral sclerosis (ALS). Conversely, the progress of the disease is associated with the impairment of iron metabolism and insulin signaling. We used transgenic hmSOD1 G93A (ALS model) and non-transgenic mice in the present study. The study was performed on the muscles taken from trained (ONSET and TERMINAL) and untrained animals at three stages of the disease: BEFORE, ONSET, and TERMINAL. In order to study the molecular mechanism of changes in iron metabolism, we used SH-SY5Y and C2C12 cell lines expression vector pcDNA3.1 and transiently transfected with specific siRNAs. The progress of ALS resulted in decreased P-Akt/Akt ratio, which is associated with increased proteins responsible for iron storage ferritin L, ferritin H, PCBP1, and skeletal muscle iron at ONSET. Conversely, proteins responsible for iron export- TAU significantly decrease. The training partially reverses changes in proteins responsible for iron metabolism. AKT silencing in the SH-SY5Y cell line decreased PCBP2 and ferroportin and increased ferritin L, H, PCBP1, TAU, transferrin receptor 1, and APP. Moreover, silencing APP led to an increase in ferritin L and H. Our data suggest that swim training in the mice ALS model is associated with significant changes in iron metabolism related to AKT activity. Down-regulation of AKT mainly upregulates proteins involved in iron import and storage but decreases proteins involved in iron export.
Assuntos
Esclerose Lateral Amiotrófica , Neuroblastoma , Camundongos , Animais , Humanos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/metabolismo , Superóxido Dismutase-1/metabolismo , Transdução de Sinais , Ferro/metabolismo , Modelos Animais de Doenças , Ferritinas/metabolismo , Proteínas de Ligação a RNA/metabolismoRESUMO
Amyotrophic lateral sclerosis (ALS) may result from the dysfunctions of various mechanisms such as protein accumulation, mitophagy, and biogenesis of mitochondria. The purpose of the study was to evaluate the molecular mechanisms in ALS development and the impact of swim training on these processes. In the present study, an animal model of ALS, SOD1-G93A mice, was used with the wild-type mice as controls. Mice swam five times per week for 30 min. Mice were analyzed before ALS onset (70 days old), at ALS 1 disease onset (116 days old), and at the terminal stage of the disease ALS (130 days old), and compared with the corresponding ALS untrained groups and normalized to the wild-type group. Enzyme activity and protein content were analyzed in the spinal cord homogenates. The results show autophagy disruptions causing accumulation of p62 accompanied by low PGC-1α and IGF-1 content in the spinal cord of SOD1-G93A mice. Swim training triggered a neuroprotective effect, attenuation of NF-l degradation, less accumulated p62, and lower autophagy initiation. The IGF-1 pathway induces pathophysiological adaptation to maintain energy demands through anaerobic metabolism and mitochondrial protection. KEY MESSAGES: The increased protein content of p62 in the spinal cord of SOD1-G93A mice suggests that autophagic clearance and transportation are disrupted. Swim training attenuates neurofilament light destruction in the spinal cord of SOD1-G93A mice. Swim training reducing OGDH provokes suppression of ATP-consuming anabolic pathways. Swim training induces energy metabolic changes and mitochondria protection through the IGF-1 signaling pathway.
Assuntos
Esclerose Lateral Amiotrófica , Animais , Camundongos , Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/metabolismo , Autofagia , Modelos Animais de Doenças , Metabolismo Energético , Fator de Crescimento Insulin-Like I , Camundongos Transgênicos , Mitocôndrias/metabolismo , Neurônios Motores/metabolismo , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Superóxido Dismutase-1/genética , Superóxido Dismutase-1/metabolismoRESUMO
In this study, we aim to verify whether swim training can improve lactate metabolism, NAD+ and NADH levels, as well as modify the activity of glycolytic and NADH shuttle enzymes and monocarboxylate transporters (MCTs) in skeletal muscle of amyotrophic lateral sclerosis (ALS) mice. ALS mice (SOD1G93A) (n = 7 per group) were analyzed before the onset of ALS, at first disease symptoms (trained and untrained), and the last stage of disease (trained and untrained), and then compared with a wild-type (WT) group of mice. The blood lactate and the skeletal muscle concentration of lactate, NAD+ and NADH, MCT1 and MCT4 protein levels, as well as lactate dehydrogenase (LDH) and malate dehydrogenase (MDH) activities in skeletal muscle were determined by fluorometric, Western blotting, liquid chromatography-MS3 spectrometry, and spectrometric methods. In the untrained terminal ALS group, there were decreased blood lactate levels (p < 0.001) and increased skeletal muscle lactate levels (p < 0.05) as compared with a WT group of mice. The amount of nicotinamide adenine dinucleotides in the ALS groups were also significantly reduced as well as LDH activity and the level of MCT1. Swim training increased lactate levels in the blood (p < 0.05 vs. ALS TERMINAL untrained). In addition, cytosolic MDH activity and the cMDH/LDH 2.1 ratio were significantly higher in trained vs. untrained mice (p < 0.05). The data indicate significant dysfunction of lactate metabolism in ALS mice, associated with a reduction in muscle anaerobic metabolism and NADH transporting enzymes, as well as swim-induced compensation of energy demands in the ALS mice.
Assuntos
Esclerose Lateral Amiotrófica , NAD , Adenina/metabolismo , Esclerose Lateral Amiotrófica/metabolismo , Animais , Modelos Animais de Doenças , Ácido Láctico/metabolismo , Malato Desidrogenase/metabolismo , Camundongos , Transportadores de Ácidos Monocarboxílicos/metabolismo , Músculo Esquelético/metabolismo , NAD/metabolismo , Niacinamida/metabolismoRESUMO
This study investigates the effect of Dexamethasone (Dex) treatment on blood and skeletal muscle metabolites level and skeletal muscle activity of enzymes related to energy metabolism after long-duration swimming. To evaluate whether Dex treatment, swimming, and combining these factors act on analyzed data, rats were randomly divided into four groups: saline treatment non-exercise and exercise and Dex treatment non-exercised and exercised. Animals in both exercised groups underwent long-lasting swimming. The concentration of lipids metabolites, glucose, and lactate were measured in skeletal muscles and blood according to standard colorimetric and fluorimetric methods. Also, activities of enzymes related to aerobic and anaerobic metabolism were measured in skeletal muscles. The results indicated that Dex treatment induced body mass loss and increased lipid metabolites in the rats' blood but did not alter these changes in skeletal muscles. Interestingly, prolonged swimming applied after 9 days of Dex treatment significantly intensified changes induced by Dex; however, there was no difference in skeletal muscle enzymatic activities. This study shows for the first time the cumulative effect of exercise and Dex on selected elements of lipid metabolism, which seems to be essential for the patient's health due to the common use of glucocorticoids like Dex.
Assuntos
Dexametasona/farmacologia , Metabolismo Energético/efeitos dos fármacos , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Condicionamento Físico Animal , Natação , Animais , Biomarcadores , Glucose/metabolismo , Ácido Láctico/sangue , Ácido Láctico/metabolismo , Metabolismo dos Lipídeos , Masculino , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Modelos Biológicos , Ratos , Estresse Fisiológico , Fatores de TempoRESUMO
(1) Background: Amyotrophic lateral sclerosis (ALS) is an incurable, neurodegenerative disease. In some cases, ALS causes behavioral disturbances and cognitive dysfunction. Swimming has revealed a neuroprotective influence on the motor neurons in ALS. (2) Methods: In the present study, a SOD1-G93A mice model of ALS were used, with wild-type B6SJL mice as controls. ALS mice were analyzed before ALS onset (10th week of life), at ALS 1 onset (first symptoms of the disease, ALS 1 onset, and ALS 1 onset SWIM), and at terminal ALS (last stage of the disease, ALS TER, and ALS TER SWIM), and compared with wild-type mice. Swim training was applied 5 times per week for 30 min. All mice underwent behavioral tests. The spinal cord was analyzed for the enzyme activities and oxidative stress markers. (3) Results: Pre-symptomatic ALS mice showed increased locomotor activity versus control mice; the swim training reduced these symptoms. The metabolic changes in the spinal cord were present at the pre-symptomatic stage of the disease with a shift towards glycolytic processes at the terminal stage of ALS. Swim training caused an adaptation, resulting in higher glutathione peroxidase (GPx) and protection against oxidative stress. (4) Conclusion: Therapeutic aquatic activity might slow down the progression of ALS.
Assuntos
Esclerose Lateral Amiotrófica/metabolismo , Esclerose Lateral Amiotrófica/fisiopatologia , Glutationa Peroxidase/metabolismo , Locomoção/fisiologia , Neurônios Motores/fisiologia , Medula Espinal/metabolismo , Natação/fisiologia , Animais , Modelos Animais de Doenças , Progressão da Doença , Masculino , Camundongos , Camundongos Transgênicos/metabolismo , Camundongos Transgênicos/fisiologia , Microglia/metabolismo , Microglia/fisiologia , Mitocôndrias/metabolismo , Mitocôndrias/fisiologia , Neurônios Motores/metabolismo , Doenças Neurodegenerativas/metabolismo , Doenças Neurodegenerativas/fisiopatologia , Estresse Oxidativo/fisiologia , Medula Espinal/fisiopatologia , Superóxido Dismutase/metabolismoRESUMO
We tested the hypothesis that swim training reverses the impairment of Akt/FOXO3a signaling, ameliorating muscle atrophy in ALS mice. Transgenic male mice B6SJL-Tg (SOD1G93A) 1Gur/J were used as the ALS model (n = 35), with wild-type B6SJL (WT) mice as controls (n = 7). ALS mice were analyzed before ALS onset, at ALS onset, and at terminal ALS. Levels of insulin/Akt signaling pathway proteins were determined, and the body and tibialis anterior muscle mass and plasma creatine kinase. Significantly increased levels of FOXO3a in ALS groups (from about 13 to 21-fold) compared to WT mice were observed. MuRF1 levels in the ONSET untrained group (12.0 ± 1.7 AU) were significantly higher than in WT mice (1.12 ± 0.2 AU) and in the BEFORE ALS group (3.7 ± 0.9 AU). This was associated with body mass and skeletal muscle mass reduction. Swim training significantly ameliorated the reduction of skeletal muscle mass in both TERMINAL groups (p < 0.001) and partially reversed changes in the levels of Akt signaling pathway proteins. These findings shed light on the swimming-induced attenuation of skeletal muscle atrophy in ALS with possible practical implications for anti-cachexia approaches.
Assuntos
Esclerose Lateral Amiotrófica/metabolismo , Esclerose Lateral Amiotrófica/fisiopatologia , Músculo Esquelético/fisiologia , Atrofia Muscular/fisiopatologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/fisiologia , Natação/fisiologia , Animais , Modelos Animais de Doenças , Proteína Forkhead Box O3/metabolismo , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Músculo Esquelético/metabolismo , Atrofia Muscular/metabolismo , Superóxido Dismutase-1/metabolismo , Proteínas com Motivo Tripartido/metabolismoRESUMO
PURPOSE: Hyperhomocysteinemia is an independent risk factor for cardiovascular diseases and also promotes neuronal death in various neurodegenerative diseases. There is evidence that iron can mediate homocysteine (Hcy) toxicity. Thus, the aim of this study was to investigate the effect of Hcy on iron metabolism in HUVEC and SH-SY5Y cells. METHODS: HUVEC and SH-SY5Y cells were treated with 3 mM Hcy for a defined time. RESULTS: We demonstrate that Hcy induced the upregulation of ferritins type L and H in HUVEC cells in a time-dependent manner and had no effect on the ferritins in SH-SY5Y cells. The change in ferritin expression was preceded by a significant decrease in the cellular level of the active form of Akt kinase in HUVEC but not in SH-SY5Y cells. An increase in ferritin L and H protein levels was observed in the Akt1, Akt2, Akt3 siRNA transfected cells, while in the cells transfected with FOXO3a siRNA, a decrease in both ferritins levels was noticed. Moreover, in the HUVEC cells treated with Hcy for 6 days, the active form of kinase Akt returned to the control level and it was accompanied by a drop in ferritin L and H protein levels. Cytotoxicity of hydrogen peroxide significantly increased in HUVEC cells pre-treated with Hcy for 24 h. CONCLUSIONS: These data indicate that Hcy induces an increase in cellular ferritin level, and the process is mediated by alterations in Akt-FOXO3a signaling pathway.
Assuntos
Homocisteína , Proteínas Proto-Oncogênicas c-akt , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Ferro , Estresse Oxidativo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismoRESUMO
The urinary level of the titin fragment has been considered a non-invasive and sensitive biomarker for muscle damage in clinical cases. However, there is little evidence regarding changes in the urinary titin fragment in response to exercise-induced muscle damage. In this study, we aimed to investigate whether the urinary titin fragment reflects the magnitude of muscle damage induced by two lower-limb eccentric exercises. In this study, healthy young male subjects performed drop jump (n=9) and eccentric ergometer exercise (n=9). Blood and urine samples were collected at various time points before and after the exercises. Although perceived muscle soreness assessed by sit-to-stand tasks was increased at 24 h and 48 h after both drop jump and the eccentric ergometer exercise groups, the pressure pain threshold was not changed. Changes of the urinary titin fragment, plasma myomesin 3 fragments, creatine kinase (CK), and myoglobin (Mb) after the eccentric exercises were increased but not statistically significant. Meanwhile, we found that the changes in the urinary titin fragment levels in response to both drop jump and the eccentric ergometer exercise were correlated with those of plasma CK and Mb levels. These results provide evidence that the urinary titin fragment level is a non-invasive biomarker reflecting the magnitude of eccentric exercise-induced muscle damage.
Assuntos
Conectina/urina , Exercício Físico/fisiologia , Mialgia/urina , Músculo Quadríceps/patologia , Biomarcadores/urina , Conectina/sangue , Creatina Quinase/sangue , Humanos , Masculino , Mialgia/sangue , Mioglobina/sangue , Adulto JovemRESUMO
Targeting neuroendocrine receptors can be considered as another interesting approach to treating fibrotic disorders. Previously, we could demonstrate that tropisetron, a classical serotonin receptor blocker, can modulate collagen synthesis and acts in vitro through the α7 nicotinic acetylcholine receptor (α7nAchR). Here, we used a pharmacologic approach with specific α7nAchR agonists to validate this hypothesis. PHA-543613, an α7nAchR-specific agonist, not only prevented but also reversed established skin fibrosis of mice injected with bleomycin. Interestingly, agonistic stimulation of α7nAchR also attenuated experimental skin fibrosis in the non-inflammation driven adenovirus coding for TGFß receptor Iact mouse model, indicating fibroblast-mediated and not only anti-inflammatory effects of such agents. The fibroblast-mediated effects were confirmed in vitro using human dermal fibroblasts, in which the α7nAchR-specific agonists strongly reduced the impact of TGFß1-mediated expression on collagen and myofibroblast marker expression. These actions were linked to modulation of the redox-sensitive transcription factor JunB and impairment of the mitochondrial respiratory system. Our results indicate that pharmacologic stimulation of the α7nAchR could be a promising target for treatment of patients with skin fibrotic diseases. Moreover, our results suggest a mechanistic axis of collagen synthesis regulation through the mitochondrial respiratory system.
Assuntos
Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Quinuclidinas/farmacologia , Escleroderma Sistêmico/tratamento farmacológico , Pele/patologia , Receptor Nicotínico de Acetilcolina alfa7/agonistas , Adenoviridae/genética , Animais , Bleomicina/toxicidade , Compostos Bicíclicos Heterocíclicos com Pontes/uso terapêutico , Células Cultivadas , Colágeno/metabolismo , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibrose , Vetores Genéticos/administração & dosagem , Vetores Genéticos/genética , Humanos , Masculino , Camundongos , Cultura Primária de Células , Quinuclidinas/uso terapêutico , Receptor do Fator de Crescimento Transformador beta Tipo I/genética , Escleroderma Sistêmico/induzido quimicamente , Escleroderma Sistêmico/genética , Escleroderma Sistêmico/patologia , Pele/citologia , Pele/efeitos dos fármacos , Receptor Nicotínico de Acetilcolina alfa7/metabolismoRESUMO
Metabolic reprogramming in skeletal muscles in the human and animal models of amyotrophic lateral sclerosis (ALS) may be an important factor in the diseases progression. We hypothesized that swim training, a modulator of cellular metabolism via changes in muscle bioenergetics and oxidative stress, ameliorates the reduction in muscle strength in ALS mice. In this study, we used transgenic male mice with the G93A human SOD1 mutation B6SJL-Tg (SOD1G93A) 1Gur/J and wild type B6SJL (WT) mice. Mice were subjected to a grip strength test and isolated skeletal muscle mitochondria were used to perform high-resolution respirometry. Moreover, the activities of enzymes involved in the oxidative energy metabolism and total sulfhydryl groups (as an oxidative stress marker) were evaluated in skeletal muscle. ALS reduces muscle strength (-70% between 11 and 15 weeks, p < 0.05), modulates muscle metabolism through lowering citrate synthase (CS) (-30% vs. WT, p = 0.0007) and increasing cytochrome c oxidase and malate dehydrogenase activities, and elevates oxidative stress markers in skeletal muscle. Swim training slows the reduction in muscle strength (-5% between 11 and 15 weeks) and increases CS activity (+26% vs. ALS I, p = 0.0048). Our findings indicate that swim training is a modulator of skeletal muscle energy metabolism with concomitant improvement of skeletal muscle function in ALS mice.
Assuntos
Esclerose Lateral Amiotrófica/metabolismo , Esclerose Lateral Amiotrófica/fisiopatologia , Metabolismo Energético , Força Muscular , Músculo Esquelético/metabolismo , Natação , Esclerose Lateral Amiotrófica/etiologia , Animais , Biomarcadores , Modelos Animais de Doenças , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Humanos , Camundongos , Camundongos Transgênicos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Músculo Esquelético/fisiopatologia , Estresse Oxidativo , Superóxido Dismutase-1/genética , Superóxido Dismutase-1/metabolismoRESUMO
Melatonin (Mel) is the major biologically active molecule secreted by the pineal gland. Mel and its metabolites, 6-hydroxymelatonin (6(OH)Mel) and 5-methoxytryptamine (5-MT), possess a variety of functions, including the scavenging of free radicals and the induction of protective or reparative mechanisms in the cell. Their amphiphilic character allows them to cross cellular membranes and reach subcellular organelles, including the mitochondria. Herein, the action of Mel, 6(OH)Mel, and 5-MT in human MNT-1 melanoma cells against ultraviolet B (UVB) radiation was investigated. The dose of 50 mJ/cm² caused a significant reduction of cell viability up to 48%, while investigated compounds counteracted this deleterious effect. UVB exposure increased catalase activity and led to a simultaneous Ca++ influx (16%), while tested compounds prevented these disturbances. Additional analysis focused on mitochondrial respiration performed in isolated mitochondria from the liver of BALB/cJ mice where Mel, 6(OH)Mel, and 5-MT significantly enhanced the oxidative phosphorylation at the dose of 10-6 M with lower effects seen at 10-9 or 10-4 M. In conclusion, Mel, 6(OH)Mel and 5-MT protect MNT-1 cells, which express melatonin receptors (MT1 and MT2) against UVB-induced oxidative stress and mitochondrial dysfunction, including the uncoupling of oxidative phosphorylation.
Assuntos
Melanoma/metabolismo , Melatonina/metabolismo , Melatonina/farmacologia , 5-Metoxitriptamina/farmacologia , Animais , Cálcio/metabolismo , Catalase/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Humanos , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/efeitos da radiação , Melatonina/análogos & derivados , Camundongos Endogâmicos BALB C , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias/efeitos da radiação , Fosforilação Oxidativa/efeitos dos fármacos , Fosforilação Oxidativa/efeitos da radiação , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/efeitos da radiação , Raios UltravioletaRESUMO
Recently, in terms of amyotrophic lateral sclerosis (ALS), much attention has been paid to the cell structures formed by the mitochondria and the endoplasmic reticulum membranes (MAMs) that are involved in the regulation of Ca2+ signaling, mitochondrial bioenergetics, apoptosis, and oxidative stress. We assumed that remodeling of these structures via swim training may accompany the prolongation of the ALS lifespan. In the present study, we used transgenic mice with the G93A hmSOD1 gene mutation. We examined muscle energy metabolism, oxidative stress parameters, and markers of MAMs (Caveolin-1 protein level and cholesterol content in crude mitochondrial fraction) in groups of mice divided according to disease progression and training status. The progression of ALS was related to the lowering of Caveolin-1 protein levels and the accumulation of cholesterol in a crude mitochondrial fraction. These changes were associated with aerobic and anaerobic energy metabolism dysfunction and higher oxidative stress. Our data indicated that swim training prolonged the lifespan of ALS mice with accompanying changes in MAM components. Swim training also maintained mitochondrial function and lowered oxidative stress. These data suggest that modification of MAMs might play a crucial role in the exercise-induced deceleration of ALS development.
Assuntos
Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/metabolismo , Colesterol/metabolismo , Metabolismo Energético/fisiologia , Mitocôndrias/metabolismo , Músculo Esquelético/metabolismo , Natação/fisiologia , Esclerose Lateral Amiotrófica/patologia , Animais , Modelos Animais de Doenças , Humanos , Camundongos TransgênicosRESUMO
Purpose: The aim of this study was to compare the effect of applying two different rest recovery times in a 10-s sprint interval training session on aerobic and anaerobic capacities as well as skeletal muscle enzyme activities. Methods: Fourteen physically active but not highly trained male subjects (mean maximal oxygen uptake 50.5 ± 1.0 mlO2·kg-1·min-1) participated in the study. The training protocol involved a series of 10-s sprints separated by either 1-min (SIT10:1) or 4-min (SIT10:4) of recovery. The number of sprints progressed from four to six over six sessions separated by 1-2 days rest. Pre and post intervention anthropometric measurements, assessment of aerobic, anaerobic capacity and muscle biopsy were performed. In the muscle samples maximal activities of citrate synthase (CS), 3-hydroxyacylCoA dehydrogenase (HADH), carnitine palmitoyl-transferase (CPT), malate dehydrogenase (MDH), and its mitochondrial form (mMDH), as well as lactate dehydrogenase (LDH) were determined. Analysis of variance was performed to determine changes between conditions. Results: Maximal oxygen uptake improved significantly in both training groups, by 13.6% in SIT10:1 and 11.9% in SIT10:4, with no difference between groups. Wingate anaerobic test results indicated main effect of time for total work, peak power output and mean power output, which increased significantly and similarly in both groups. Significant differences between training groups were observed for end power output, which increased by 10.8% in SIT10:1, but remained unchanged in SIT10:4. Both training protocols induced similar increase in CS activity (main effect of time p < 0.05), but no other enzymes. Conclusion: Sprint interval training protocols induce metabolic adaptation over a short period of time, and the reduced recovery between bouts may attenuate fatigue during maximal exercise.
RESUMO
BACKGROUND: It has long been suggested that reactive oxygen species (ROS) play a role in oxygen sensing via peripheral chemoreceptors, which would imply their involvement in chemoreflex activation and autonomic regulation of heart rate. We hypothesize that antioxidant affect neurogenic cardiovascular regulation through activation of chemoreflex which results in increased control of sympathetic mechanism regulating heart rhythm. Activity of xanthine oxidase (XO), which is among the major endogenous sources of ROS in the rat has been shown to increase during hypoxia promote oxidative stress. However, the mechanism of how XO inhibition affects neurogenic regulation of heart rhythm is still unclear. AIM: The study aimed to evaluate effects of allopurinol-driven inhibition of XO on autonomic heart regulation in rats exposed to hypoxia followed by hyperoxia, using heart rate variability (HRV) analysis. MATERIAL AND METHODS: 16 conscious male Wistar rats (350 g): control-untreated (N = 8) and pretreated with Allopurinol-XO inhibitor (5 mg/kg, followed by 50 mg/kg), administered intraperitoneally (N = 8), were exposed to controlled hypobaric hypoxia (1h) in order to activate chemoreflex. The treatment was followed by 1h hyperoxia (chemoreflex suppression). Time-series of 1024 RR-intervals were extracted from 4kHz ECG recording for heart rate variability (HRV) analysis in order to calculate the following time-domain parameters: mean RR interval (RRi), SDNN (standard deviation of all normal NN intervals), rMSSD (square root of the mean of the squares of differences between adjacent NN intervals), frequency-domain parameters (FFT method): TSP (total spectral power) as well as low and high frequency band powers (LF and HF). At the end of experiment we used rat plasma to evaluate enzymatic activity of XO and markers of oxidative stress: protein carbonyl group and 8-isoprostane concentrations. Enzymatic activity of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) were measures in erythrocyte lysates. RESULTS: Allopurinol reduced oxidative stress which was the result of hypoxia/hyperoxia, as shown by decreased 8-isoprostane plasma concentration. XO inhibition did not markedly influence HRV parameters in standard normoxia. However, during hypoxia, as well as hyperoxia, allopurinol administration resulted in a significant increase of autonomic control upon the heart as shown by increased SDNN and TSP, with an increased vagal contribution (increased rMSSD and HF), whereas sympathovagal indexes (LF/HF, SDNN/rMSSD) remained unchanged. CONCLUSIONS: Observed regulatory effects of XO inhibition did not confirm preliminary hypothesis which suggested that an antioxidant such as allopurinol might activate chemoreflex resulting in augmented sympathetic discharge to the heart. The HRV regulatory profile of XO inhibition observed during hypoxia as well as post-hypoxic hyperoxia corresponds to reported reduced risk of sudden cardiovascular events. Therefore our data provide a new argument for therapeutical use of allopurinol in hypoxic conditions.
Assuntos
Alopurinol/farmacologia , Frequência Cardíaca/efeitos dos fármacos , Hiperóxia/metabolismo , Hipóxia/metabolismo , Xantina Oxidase/antagonistas & inibidores , Animais , Masculino , Ratos , Ratos WistarRESUMO
BACKGROUND: Recently, skeletal muscle atrophy, impairment of iron metabolism, and insulin signalling have been reported in rats suffering from amyotrophic lateral sclerosis (ALS). However, the interrelationship between these changes has not been studied. We hypothesize that an impaired Akt-FOXO3a signalling pathway triggers changes in the iron metabolism in the muscles of transgenic animals. METHODS: In the present study, we used transgenic rats bearing the G93A hmSOD1 gene and their non-transgenic littermates. The study was performed on the muscles taken from animals at three different stages of the disease: asymptomatic (ALS I), the onset of the disease (ALS II), and the terminal stage of the disease (ALS III). In order to study the molecular mechanism of changes in iron metabolism, we used SH-SY5Y and C2C12 cell lines stably transfected with pcDNA3.1, SOD1 WT and SOD1 G93A, or FOXO3a TM-ER. RESULTS: A significant decrease in P-Akt level and changes in iron metabolism were observed even in the group of ALS I animals. This was accompanied by an increase in the active form of FOXO3a, up-regulation of atrogin-1, and catalase. However, significant muscle atrophy was observed in ALS II animals. An increase in ferritin L and H was accompanied by a rise in PCBP1 and APP protein levels. In SH-SY5Y cells stably expressing SOD1 or SOD1 G93A, we observed elevated levels of ferritin L and H and non-haem iron. Interestingly, insulin treatment significantly down-regulated ferritin L and H proteins in the cell. Conversely, cells transfected with small interfering RNA against Akt 1, 2, 3, respectively, showed a significant increase in the ferritin and FOXO3a levels. In order to assess the role of FOXO3a in the ferritin expression, we constructed a line of SH-SY5Y cells that expressed a fusion protein made of FOXO3a fused at the C-terminus with the ligand-binding domain of the oestrogen receptor (TM-ER) being activated by 4-hydroxytamoxifen. Treatment of the cells with 4-hydroxytamoxifen significantly up-regulated ferritin L and H proteins level. CONCLUSIONS: Our data suggest that impairment of insulin signalling and iron metabolism in the skeletal muscle precedes muscle atrophy and is mediated by changes in Akt/FOXO3a signalling pathways.
Assuntos
Esclerose Lateral Amiotrófica/metabolismo , Proteína Forkhead Box O3/metabolismo , Ferro/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Superóxido Dismutase-1/genética , Esclerose Lateral Amiotrófica/genética , Animais , Linhagem Celular , Modelos Animais de Doenças , Humanos , Insulina/metabolismo , Masculino , Camundongos , Proteínas Musculares/metabolismo , Músculo Esquelético/metabolismo , Mutação , Ratos Sprague-Dawley , Ratos Transgênicos , Proteínas Ligases SKP Culina F-Box/metabolismo , Transdução de SinaisRESUMO
BACKGROUND: Spinning exercise is one of the most popular types of exercise in fitness industry. Its effects on the post exercise metabolism compared to the isocaloric cyclic endurance exercise are not fully understood. The aim of the present study was to compare the effects of isocaloric (299.1±10.8 kcal) spinning vs. endurance exercise on fat and carbohydrate utilization, glucose, lactate, glycerol and NEFA blood concentration during exercise and recovery. METHODS: Six recreationally active males (age: 23.5±0.71) were tested in two conditions: 1) 30-min spinning; 2) isocaloric continuous exercise. Each trial was followed by a 3-h recovery. Rates of carbohydrate and fat oxidation, the blood glucose, lactate, glycerol and NEFA concentration were assessed at rest, during exercise and recovery. RESULTS: Spinning induced significantly higher fat and lower carbohydrate oxidation rate during a recovery period in comparison to isocaloric endurance exercise trial. Spinning induced almost six-fold higher increase in lipid to carbohydrate oxidation rate ratio at the beginning of second hour of postexercise period in comparison to constant intensity trial and reached similar values at 3 hours after exercise. Blood lactate was higher (P<0.01) at the end of exercise in spinning than continuous exercise (8.57±0.9 vs. 0.72±0.1 mmol·L-1), but became similar at the 60 min of recovery. CONCLUSIONS: These data indicate that spinning induces higher metabolic responses during recovery period, and most effectively shifts the pattern of substrate use toward lipids vs. isocaloric endurance exercise.
Assuntos
Metabolismo Energético , Exercício Físico/fisiologia , Metabolismo dos Lipídeos , Consumo de Oxigênio , Glicemia/metabolismo , Metabolismo dos Carboidratos , Terapia por Exercício , Ácidos Graxos não Esterificados/metabolismo , Glicerol/sangue , Humanos , Ácido Láctico/sangue , Masculino , Oxirredução , Descanso , Adulto JovemRESUMO
Studying organelles in isolation has been proven to be indispensable for deciphering the underlying mechanisms of molecular cell biology. However, observing organelles in intact cells with the use of microscopic techniques reveals a new set of different junctions and contact sites between them that contribute to the control and regulation of various cellular processes, such as calcium and lipid exchange or structural reorganization of the mitochondrial network. In recent years, many studies focused their attention on the structure and function of contacts between mitochondria and other organelles. From these studies, findings emerged showing that these contacts are involved in various processes, such as lipid synthesis and trafficking, modulation of mitochondrial morphology, endoplasmic reticulum (ER) stress, apoptosis, autophagy, inflammation and Ca 2 + handling. In this review, we focused on the physical interactions of mitochondria with the endoplasmic reticulum and plasma membrane and summarized present knowledge regarding the role of mitochondria-associated membranes in calcium homeostasis and lipid metabolism.
Assuntos
Cálcio/metabolismo , Membrana Celular/metabolismo , Retículo Endoplasmático/metabolismo , Homeostase , Metabolismo dos Lipídeos , Mitocôndrias/metabolismo , Animais , Apoptose , Transporte Biológico , Membrana Celular/ultraestrutura , Suscetibilidade a Doenças , Retículo Endoplasmático/ultraestrutura , Humanos , Mitocôndrias/ultraestrutura , Dinâmica Mitocondrial , Transporte ProteicoRESUMO
Iron participates in oxygen transport, energetic, metabolic, and immunologic processes. There are 2 main causes of iron overload: hereditary hemochromatosis which is a primary cause, is a metabolic disorder caused by mutations of genes that control iron metabolism and secondary hemochromatosis caused by multitransfusions, chronic hemolysis, and intake of iron rich food. The most common type of hereditary hemochromatosis is caused by HFE gene mutation. In this study, we analyzed iron metabolism in 100 healthy Polish children in relation to their HFE gene status. The wild-type HFE gene was predominant being observed in 60 children (60%). Twenty-five children (25%), presented with heterozygotic H63D mutation, and 15 children (15%), presented with other mutations (heterozygotic C282Y and S65C mutation, compound heterozygotes C282Y/S65C, C282Y/H63D, H63D homozygote). The mean concentration of iron, the level of ferritin, and transferrin saturation were statistically higher in the group of HFE variants compared with the wild-type group. H63D carriers presented with higher mean concentration of iron, ferritin levels, and transferrin saturation compared with the wild-type group. Male HFE carriers presented with higher iron concentration, transferrin saturation, and ferritin levels than females. This preliminary investigation demonstrates allelic impact on potential disease progression from childhood.
Assuntos
Proteína da Hemocromatose/genética , Hemocromatose/epidemiologia , Ferro/metabolismo , Mutação , Adolescente , Criança , Pré-Escolar , Feminino , Ferritinas/sangue , Genótipo , Hemocromatose/sangue , Hemocromatose/genética , Humanos , Ferro/sangue , Masculino , Polônia/epidemiologia , Fatores Sexuais , Transferrina/análiseRESUMO
The reduction in cholesterol in mitochondria, observed after exercise, is related to the inhibition of mitochondrial swelling. Caveolin-1 (Cav-1) plays an essential role in the regulation of cellular cholesterol metabolism and is required by various signalling pathways. Therefore, the aim of this study was to investigate the effect of prolonged swimming on the mitochondrial Cav-1 concentration; additionally, we identified the results of these changes as they relate to the induction of changes in the mitochondrial swelling and cholesterol in rat skeletal muscle and liver. Male Wistar rats were divided into a sedentary control group and an exercise group. The exercised rats swam for 3 hours and were burdened with an additional 3% of their body weight. After the cessation of exercise, their quadriceps femoris muscles and livers were immediately removed for experimentation. The exercise protocol caused an increase in the Cav-1 concentration in crude muscle mitochondria; this was related to a reduction in the cholesterol level and an inhibition of mitochondrial swelling. There were no changes in rat livers, with the exception of increased markers of oxidative stress in mitochondria. These data indicate the possible role of Cav-1 in the adaptive change in the rat muscle mitochondria following exercise.
Assuntos
Caveolina 1/metabolismo , Colesterol/metabolismo , Fígado/metabolismo , Mitocôndrias/metabolismo , Dilatação Mitocondrial , Músculo Esquelético/metabolismo , Condicionamento Físico Animal , Animais , Peso Corporal , Masculino , Estresse Oxidativo , Músculo Quadríceps/metabolismo , Ratos , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismo , Comportamento Sedentário , Transdução de Sinais , NataçãoRESUMO
Impaired mitochondrial capacity may be implicated in the pathology of chronic metabolic diseases. To elucidate the effect of ethyl pyruvate supplementation on skeletal muscles metabolism we examined changes in activities of mitochondrial and antioxidant enzymes, as well as sulfhydryl groups oxidation (an indirect marker of oxidative stress) during the development of obesity. After 6 weeks feeding of control or high fat diet, Wistar rats were divided into four groups: control diet, control diet and ethyl pyruvate, high fat diet, and high fat diet and ethyl pyruvate. Ethyl pyruvate was administered as 0.3% solution in drinking water, for the following 6 weeks. High fat diet feeding induced the increase of activities 3-hydroxyacylCoA dehydrogenase, citrate synthase, and fumarase. Moreover, higher catalase and superoxide dismutase activities, as well as sulfhydryl groups oxidation, were noted. Ethyl pyruvate supplementation did not affect the mitochondrial enzymes' activities, but induced superoxide dismutase activity and sulfhydryl groups oxidation. All of the changes were observed in soleus muscle, but not in extensor digitorum longus muscle. Additionally, positive correlations between fasting blood insulin concentration and activities of catalase (p = 0.04), and superoxide dismutase (p = 0.01) in soleus muscle were noticed. Prolonged ethyl pyruvate consumption elevated insulin concentration, which may cause modifications in oxidative type skeletal muscles.