RESUMO
Photobiomodulation (PBM), an emerging and non-invasive intervention, has been shown to benefit the nervous system by modifying the mitochondrial cytochrome c-oxidase (CCO) enzyme, which has red (620-680 nm) or infrared (760-825 nm) spectral absorption peaks. The effect of a single 810-nm wavelength with a combination of 810 nm and 660 nm lights in the brain metabolic activity of male and female rats was compared. PBM, with a wavelength of 810 nm and a combination of 810 nm and 660 nm, was applied for 5 days on the prefrontal cortex. Then, brain metabolic activity in the prefrontal area, hippocampus, retrosplenial, and parietal cortex was explored. Sex differences were found in cortical and subcortical regions, indicating higher male brain oxidative metabolism, regardless of treatment. CCO activity in the cingulate and prelimbic area, dentate gyrus, retrosplenial and parietal cortex was enhanced in both treatments (810 + 660 nm and 810 nm). Moreover, using the combination of waves, CCO increased in the infralimbic area, and in CA1 and CA3 of the hippocampus. Thus, employment of a single NIR treatment or a combination of red to NIR treatment led to slight differences in CCO activity across the limbic system, suggesting that a combination of lights of the spectrum may be relevant.
Assuntos
Terapia com Luz de Baixa Intensidade , Ratos , Masculino , Feminino , Animais , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Oxirredução , Encéfalo/metabolismo , Hipocampo/metabolismoRESUMO
BACKGROUND: Recent studies have demonstrated that copper (Cu) plays an important role in the progression of tumor diseases. Metastasis associated with colon cancer protein 1 (MACC1) promotes the transcription and expression of various tumor-related genes. Cytochrome c oxidase (COX) 19, present in the cytoplasm and intermembrane space of mitochondria, may transport Cu within the mitochondria. However, the mechanism through which MACC1 regulates the Cu homeostasis mediated by COX19 remains unclear. OBJECTIVES: The aim of this study was to elucidate the mechanism through which MACC1 initiates the transcription and expression of COX19, and promotes malignant behavior in tumor cells. METHODS: Immunohistochemistry, western blotting, and real-time polymerase chain reaction (PCR) analyses were conducted to analyze the expression of MACC1 and COX19 proteins and genes in tumor and normal tissues. RNA-chromatin immunoprecipitation was used to detect the transcriptional initiation of COX19 by MACC1. The effects of MACC1 and COX19 on mitochondrial activity were determined using an ATP assay kit and Cytochrome c Oxidase Assay Kit. A Cell Counting Kit-8 kit was used to detect the effect of high-dose Cu or overexpression of MACC1 and COX19 on tumor cell proliferation. A xenograft mouse model was used to analyze the effect of the COX19 overexpression on the malignant behavior of the tumors. RESULTS: Cu enhanced the proliferation, invasion, and migration and inhibited apoptosis of SW480 cells. MACC1 was highly expressed in colorectal cancer tissues and activated the expression of COX19 by binding to its promoter region of COX19. The overexpression of COX19 increased mitochondrial Cu content and enhanced the activity of mitochondrial COX and ATP content, and inhibited apoptosis, promoted tumor growth of mice. CONCLUSIONS: Our results indicate that COX19 functions as a target gene of MACC1 and regulates mitochondrial activity and promotes the progression of colorectal cancer. MACC1/COX19 may provide a novel therapeutic target for colorectal cancer.
Assuntos
Neoplasias do Colo , Neoplasias Colorretais , Animais , Humanos , Camundongos , Trifosfato de Adenosina/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Neoplasias do Colo/metabolismo , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Cobre/metabolismo , Complexo IV da Cadeia de Transporte de Elétrons/genética , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Regulação Neoplásica da Expressão Gênica , Mitocôndrias/metabolismo , Transativadores/genética , Transativadores/metabolismoRESUMO
The aim of this study was to investigate the effect of dietary tryptophan (Trp) supplementation on serum biochemical indices, antioxidant indices, cytokine levels, mitochondrial biosynthesis, and mitochondrial morphology of heat-stressed broilers. A total of 180 female Arbor Acres broilers (18-day-old) were randomly allocated into three groups with six replicates of 10 broilers each. Broilers in thermoneutral (TN) (23 ± 1 °C) group were fed a basal diet; the other two groups were fed the basal diet supplemented with 0 or 0.18% Trp under heat stress (HS) (34 ± 1 °C for 8 h/day (h/day) and 23 ± 1°C for the remaining time) condition. The heat stress lasted for 21 days (days 21 to 42). The results indicated that heat stress reduced serum total protein content (TP) and decreased the activities of serum superoxide dismutase (SOD) and total antioxidant capacity (T-AOC), but increased the levels of serum uric acid (UA), interleukin (IL)-1ß, IL-6, and IL-18 (P < 0.05) compared to the TN group. However, dietary supplementation with 0.18% Trp enhanced serum TP content, glutathione peroxidase (GSH-Px), SOD, catalase (CAT) activities, and T-AOC; decreased aspartate aminotransferase (AST) activities (P < 0.05); and lowered serum IL-1ß, IL-6, IL-18 contents (P < 0.05). Meanwhile, heat stress exposure downregulated the mRNA expression of mitochondrial transcription factor A (TFAM), cytochrome c oxidase subunit 1 (COX1), and cytochrome c oxidase subunit 5A (COX5A) in ileum (P < 0.05) as compared to the TN group. Dietary Trp supplementation enhanced the mitochondrial membrane potential (MMP) and the mRNA expression of TFAM, COX1 in ileum mucosa (P < 0.05) and ameliorated the damage of mitochondrial structure. Collectively, dietary supplementation with Trp could improve antioxidant capacity and mitochondrial structure and regulate mitochondrial function-related genes and decrease inflammatory response in heat-stressed broilers. Dietary Trp supplementation might be an effective nutritional strategy to protect against heat stress impairment.
Assuntos
Antioxidantes , Galinhas , Feminino , Animais , Antioxidantes/metabolismo , Galinhas/fisiologia , Interleucina-18/metabolismo , Triptofano/metabolismo , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Interleucina-6/metabolismo , Ácido Úrico , Suplementos Nutricionais , Dieta/veterinária , Resposta ao Choque Térmico/fisiologia , Mitocôndrias , Superóxido Dismutase/metabolismo , RNA Mensageiro/metabolismo , Ração Animal/análiseRESUMO
In this paper, we present a multi-layer, activity-dependent model for the joint development of ocular dominance (OD) columns and cytochrome oxidase (CO) blobs in primate V1. For simplicity, we focus on layers 4C and 2/3 with both layers receiving direct thalamic inputs and layer 4C sending vertical projections to layer 2/3. Both the thalamic and the vertical connections are taken to be modifiable by activity. Using a correlation-based Hebbian learning rule with subtractive normalization, we show how the formation of an OD map in layer 4C is inherited by layer 2/3 via the vertical projections. Competition between these feedforward projections and the direct thalamic input to layer 2/3 then results in the formation of CO blobs superimposed upon the ocular dominance map. The spacing of the OD columns is determined by the spatial profile of the intralaminar connections within layer 4, while the spacing of CO blobs depends both on the width of the OD columns inherited from layer 4 and the spatial distribution of intralaminar connections within the superficial layer. The resulting CO blob distribution is shown to be consistent with experimental data. In addition, we numerically simulate monocular deprivation and find that while the CO blob distribution is unaltered, the OD pattern undergoes modification. The OD stripes of the deprived eye narrow, whereas the OD stripes for the remaining open eye widen.
Assuntos
Dominância Ocular , Córtex Visual , Animais , Córtex Visual/metabolismo , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Córtex Visual Primário , TálamoRESUMO
Cannabis has been used for decades as a palliative therapy in the treatment of cancer. This is because of its beneficial effects on the pain and nausea that patients can experience as a result of chemo/radiotherapy. Tetrahydrocannabinol and cannabidiol are the main compounds present in Cannabis sativa, and both exert their actions through a receptor-mediated mechanism and through a non-receptor-mediated mechanism, which modulates the formation of reactive oxygen species. These oxidative stress conditions might trigger lipidic changes, which would compromise cell membrane stability and viability. In this sense, numerous pieces of evidence describe a potential antitumor effect of cannabinoid compounds in different types of cancer, although controversial results limit their implementation. In order to further investigate the possible mechanism involved in the antitumoral effects of cannabinoids, three extracts isolated from Cannabis sativa strains with high cannabidiol content were analyzed. Cell mortality, cytochrome c oxidase activity and the lipid composition of SH-SY5Y cells were determined in the absence and presence of specific cannabinoid ligands, with and without antioxidant pre-treatment. The cell mortality induced by the extracts in this study appeared to be related to the inhibition of the cytochrome c oxidase activity and to the THC concentration. This effect on cell viability was similar to that observed with the cannabinoid agonist WIN55,212-2. The effect was partially blocked by the selective CB1 antagonist AM281, and the antioxidant α-tocopherol. Moreover, certain membrane lipids were affected by the extracts, which demonstrated the importance of oxidative stress in the potential antitumoral effects of cannabinoids.
Assuntos
Cannabis , Neuroblastoma , Extratos Vegetais , Humanos , Canabidiol/análise , Canabinoides/análise , Cannabis/química , Dronabinol/farmacologia , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Neuroblastoma/tratamento farmacológico , Extratos Vegetais/química , Extratos Vegetais/uso terapêuticoRESUMO
Low-level laser therapy, or photobiomodulation, utilizes red or near-infrared light for the treatment of pathological conditions due to the presence of intracellular photoacceptors, such as mitochondrial cytochrome c oxidase, that serve as intermediates for the therapeutic effects. We present an in-detail analysis of the effect of low-intensity LED red light irradiation on the respiratory chain of brain mitochondria. We tested whether low-level laser therapy at 650 nm could alleviate the brain mitochondrial dysfunction in the model of acute hypobaric hypoxia in mice. The irradiation of the mitochondrial fraction of the left cerebral cortex with low-intensity LED red light rescued Complex I-supported respiration during oxidative phosphorylation, normalized the initial polarization of the inner mitochondrial membrane, but has not shown any significant effect on the activity of Complex IV. In comparison, the postponed effect (in 24 h) of the similar transcranial irradiation following hypoxic exposure led to a less pronounced improvement of the mitochondrial functional state, but normalized respiration related to ATP production and membrane polarization. In contrast, the similar irradiation of the mitochondria isolated from control healthy animals exerted an inhibitory effect on CI-supported respiration. The obtained results provide significant insight that can be beneficial for the development of non-invasive phototherapy.
Assuntos
Encéfalo , Hipóxia , Terapia com Luz de Baixa Intensidade , Mitocôndrias , Animais , Camundongos , Encéfalo/metabolismo , Encéfalo/efeitos da radiação , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Hipóxia/complicações , Hipóxia/metabolismo , Hipóxia/radioterapia , Raios Infravermelhos/uso terapêutico , Mitocôndrias/metabolismo , Mitocôndrias/efeitos da radiação , Pressão/efeitos adversos , Respiração Celular/efeitos da radiaçãoRESUMO
Although hydrogen sulfide (H2S) is an endogenous signaling molecule with antioxidant properties, it is also cytotoxic by potently inhibiting cytochrome c oxidase and mitochondrial respiration. Paradoxically, the primary route of H2S detoxification is thought to occur inside the mitochondrial matrix via a series of relatively slow enzymatic reactions that are unlikely to compete with its rapid inhibition of cytochrome c oxidase. Therefore, alternative or complementary cellular mechanisms of H2S detoxification are predicted to exist. Here, superoxide dismutase [Cu-Zn] (SOD1) is shown to be an efficient H2S oxidase that has an essential role in limiting cytotoxicity from endogenous and exogenous sulfide. Decreased SOD1 expression resulted in increased sensitivity to H2S toxicity in yeast and human cells, while increased SOD1 expression enhanced tolerance to H2S. SOD1 rapidly converted H2S to sulfate under conditions of limiting sulfide; however, when sulfide was in molar excess, SOD1 catalyzed the formation of per- and polysulfides, which induce cellular thiol oxidation. Furthermore, in SOD1-deficient cells, elevated levels of reactive oxygen species catalyzed sulfide oxidation to per- and polysulfides. These data reveal that a fundamental function of SOD1 is to regulate H2S and related reactive sulfur species.
Assuntos
Complexo IV da Cadeia de Transporte de Elétrons , Sulfeto de Hidrogênio , Superóxido Dismutase-1 , Humanos , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Sulfeto de Hidrogênio/metabolismo , Sulfeto de Hidrogênio/toxicidade , Sulfetos/metabolismo , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Superóxido Dismutase-1/genética , Superóxido Dismutase-1/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismoRESUMO
Photobiomodulation is a term for using low-power red to near-infrared light to stimulate a variety of positive biological effects. Though the scientific and clinical acceptance of PBM as a therapeutic intervention has increased dramatically in recent years, the molecular underpinnings of the effect remain poorly understood. The putative chromophore for PBM effects is cytochrome c oxidase. It is postulated that light absorption at cytochrome c oxidase initiates a signaling cascade involving ATP and generation of reactive oxygen species (ROS), which subsequently results in improved cellular robustness. However, this hypothesis is largely based on inference and indirect evidence, and the precise molecular mechanisms that govern how photon absorption leads to these downstream effects remain poorly understood. We conducted low-power PBM-type light exposures of isolated mitochondria to 808 nm NIR light, at a number of irradiances. NIR exposure was found to enhance the activity of complex IV, depress the activity of complex III, and had no effect on the activity of complex II. Further, examining the dose-response of complex IV we found NIR enhancement did not exhibit irradiance reciprocity, indicating the effect on complex IV may not have direct photochemical basis. In summary, this research presents a novel method to interrogate the earliest stages of PBM in the mitochondria, and a unique window into the corresponding molecular mechanism(s) of induction.
Assuntos
Complexo IV da Cadeia de Transporte de Elétrons , Terapia com Luz de Baixa Intensidade , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Transporte de Elétrons , Terapia com Luz de Baixa Intensidade/métodos , Mitocôndrias/metabolismo , Membranas Mitocondriais/metabolismoRESUMO
Acute myeloid leukemia (AML)-the most frequent form of adult blood cancer-is characterized by heterogeneous mechanisms and disease progression. Developing an effective therapeutic strategy that targets metabolic homeostasis and energy production in immature leukemic cells (blasts) is essential for overcoming relapse and improving the prognosis of AML patients with different subtypes. With respect to metabolic regulation, fructose-1,6-bisphosphatase 1 (FBP1) is a gluconeogenic enzyme that is vital to carbohydrate metabolism, since gluconeogenesis is the central pathway for the production of important metabolites and energy necessary to maintain normal cellular activities. Beyond its catalytic activity, FBP1 inhibits aerobic glycolysis-known as the "Warburg effect"-in cancer cells. Importantly, while downregulation of FBP1 is associated with carcinogenesis in major human organs, restoration of FBP1 in cancer cells promotes apoptosis and prevents disease progression in solid tumors. Recently, our large-scale sequencing analyses revealed FBP1 as a novel inducible therapeutic target among 17,757 vitamin-D-responsive genes in MV4-11 or MOLM-14 blasts in vitro, both of which were derived from AML patients with FLT3 mutations. To investigate FBP1's anti-leukemic function in this study, we generated a new AML cell line through lentiviral overexpression of an FBP1 transgene in vitro (named FBP1-MV4-11). Results showed that FBP1-MV4-11 blasts are more prone to apoptosis than MV4-11 blasts. Mechanistically, FBP1-MV4-11 blasts have significantly increased gene and protein expression of P53, as confirmed by the P53 promoter assay in vitro. However, enhanced cell death and reduced proliferation of FBP1-MV4-11 blasts could be reversed by supplementation with post-glycolytic metabolites in vitro. Additionally, FBP1-MV4-11 blasts were found to have impaired mitochondrial homeostasis through reduced cytochrome c oxidase subunit 2 (COX2 or MT-CO2) and upregulated PTEN-induced kinase (PINK1) expressions. In summary, this is the first in vitro evidence that FBP1-altered carbohydrate metabolism and FBP1-activated P53 can initiate leukemic death by activating mitochondrial reprogramming in AML blasts, supporting the clinical potential of FBP1-based therapies for AML-like cancers.
Assuntos
Metabolismo dos Carboidratos , Células Precursoras de Granulócitos , Leucemia Mieloide Aguda , Mitocôndrias , Proteína Supressora de Tumor p53 , Apoptose , Metabolismo dos Carboidratos/efeitos dos fármacos , Metabolismo dos Carboidratos/genética , Dióxido de Carbono/metabolismo , Linhagem Celular Tumoral , Ciclo-Oxigenase 2/metabolismo , Progressão da Doença , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Frutose/farmacologia , Frutose-Bifosfatase/genética , Frutose-Bifosfatase/metabolismo , Glicólise , Células Precursoras de Granulócitos/metabolismo , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Proteínas Quinases/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Vitamina D/farmacologia , Vitaminas/farmacologia , Tirosina Quinase 3 Semelhante a fms/genética , Tirosina Quinase 3 Semelhante a fms/metabolismoRESUMO
Lyophyllum decastes is a common mushroom that is prone to browning during prolonged storage. In this study, the effects of ultrasonic treatment on metabolic gene expression, enzyme activity, and metabolic compounds related to L. decastes browning were investigated. Treatment of the fruiting body at 35 kHz and 300 W for 10 min reduced the browning index of L. decastes by 21.0 % and increased the L* value by 11.1 %. Ultrasonic treatment of the fruiting body resulted in higher levels of total phenols, flavonoids, and 9 kinds of amino acid with catalase (CAT) and peroxidase (POD) activities maintained at high levels. Higher cytochrome c oxidase (CCO), succinate dehydrogenase (SDH), phosphofructokinase (PFK), and pyruvate kinase (PK) activities may be ascribed to increased antioxidant capacity. Moreover, ultrasonication retained higher adenosine triphosphate (ATP) concentrations with an increased energy charge, while there were lower levels of adenosine diphosphate (ADP) and reduced and oxidized nicotinamide adenine dinucleotide (NADH and NAD+), respectively. Meanwhile, lower lignin contents were observed, along with retarded polyphenol oxidase (PPO) and lipoxygenase (LOX) activities. Lower PPO activity reduced the fruiting body enzymatic browning rate through decreased expression of LdPpo1, LdPpo2, and LdPpo3 during storage at 4 °C for 16 days. This activity may be used to determine the effectiveness of ultrasonication.
Assuntos
NAD , Succinato Desidrogenase , Difosfato de Adenosina/metabolismo , Trifosfato de Adenosina/metabolismo , Agaricales , Aminoácidos/metabolismo , Antioxidantes/metabolismo , Catalase/metabolismo , Catecol Oxidase/metabolismo , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Metabolismo Energético , Flavonoides , Lignina/metabolismo , Lipoxigenases/metabolismo , NAD/metabolismo , Fenóis/química , Fosfofrutoquinases/metabolismo , Piruvato Quinase/metabolismo , Succinato Desidrogenase/metabolismo , UltrassomRESUMO
Copper (Cu) is a cofactor of around 300 Arabidopsis proteins, including photosynthetic and mitochondrial electron transfer chain enzymes critical for adenosine triphosphate (ATP) production and carbon fixation. Plant acclimation to Cu deficiency requires the transcription factor SQUAMOSA PROMOTER-BINDING PROTEIN-LIKE7 (SPL7). We report that in the wild type (WT) and in the spl7-1 mutant, respiratory electron flux via Cu-dependent cytochrome c oxidase is unaffected under both normal and low-Cu cultivation conditions. Supplementing Cu-deficient medium with exogenous sugar stimulated growth of the WT, but not of spl7 mutants. Instead, these mutants accumulated carbohydrates, including the signaling sugar trehalose 6-phosphate, as well as ATP and NADH, even under normal Cu supply and without sugar supplementation. Delayed spl7-1 development was in agreement with its attenuated sugar responsiveness. Functional TARGET OF RAPAMYCIN and SNF1-RELATED KINASE1 signaling in spl7-1 argued against fundamental defects in these energy-signaling hubs. Sequencing of chromatin immunoprecipitates combined with transcriptome profiling identified direct targets of SPL7-mediated positive regulation, including Fe SUPEROXIDE DISMUTASE1 (FSD1), COPPER-DEFICIENCY-INDUCED TRANSCRIPTION FACTOR1 (CITF1), and the uncharacterized bHLH23 (CITF2), as well as an enriched upstream GTACTRC motif. In summary, transducing energy availability into growth and reproductive development requires the function of SPL7. Our results could help increase crop yields, especially on Cu-deficient soils.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Cobre/química , Trifosfato de Adenosina/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Cromatina/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Regulação da Expressão Gênica de Plantas , Crescimento e Desenvolvimento , NAD/metabolismo , Fosfatos/metabolismo , Sirolimo , Solo , Superóxidos/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Trealose/metabolismoRESUMO
Cytochrome c oxidase (CcO) is a multimeric copper-containing enzyme of the mitochondrial respiratory chain that powers cellular energy production. The two core subunits of cytochrome c oxidase, Cox1 and Cox2, harbor the catalytic CuB and CuA sites, respectively. Biogenesis of each copper site occurs separately and requires multiple proteins that constitute the mitochondrial copper delivery pathway. Currently, the identity of all the members of the pathway is not known, though several evolutionarily conserved twin CX9C motif-containing proteins have been implicated in this process. Here, we performed a targeted yeast suppressor screen that placed Coa4, a twin CX9C motif-containing protein, in the copper delivery pathway to the Cox1 subunit. Specifically, we show that overexpression of Cox11, a copper metallochaperone required for the formation of CuB site, can restore Cox1 abundance, cytochrome c oxidase assembly, and mitochondrial respiration in coa4Δ cells. This rescue is dependent on the copper-coordinating cysteines of Cox11. The abundance of Coa4 and Cox11 in mitochondria is reciprocally regulated, further linking Coa4 to the CuB site biogenesis. Additionally, we find that coa4Δ cells have reduced levels of copper and exogenous copper supplementation can partially ameliorate its respiratory-deficient phenotype, a finding that connects Coa4 to cellular copper homeostasis. Finally, we demonstrate that human COA4 can replace the function of yeast Coa4 indicating its evolutionarily conserved role. Our work provides genetic evidences for the role of Coa4 in the copper delivery pathway to the CuB site of cytochrome c oxidase.
Assuntos
Complexo IV da Cadeia de Transporte de Elétrons , Saccharomyces cerevisiae , Cobre , Complexo IV da Cadeia de Transporte de Elétrons/genética , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Humanos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Saccharomyces cerevisiae/metabolismoRESUMO
Advanced age is the main risk factor for the manifestation of late onset neurodegenerative diseases. Metformin, an anti-diabetic drug, was shown to extend longevity, and to ameliorate the activity of recognized aging hallmarks. Here, we compared the clinical, pathologic and biochemical effects of Metformin to those of Nano-PSO (Granagard), a brain targeted anti-oxidant shown by us to delay disease advance in transgenic mice mimicking for genetic Creutzfeldt Jacob disease (CJD) linked to the E200KPrP mutation. We demonstrate that both Metformin and Nano-PSO reduced aging hallmarks activities such as activated AMPK, the main energy sensor of cells as well as Nrf2 and COX IV1, regulators of oxidation, and mitochondrial activity. Both compounds reduced inflammation and increased stem cells production, however did not decrease PrP accumulation. As opposed to Nano-PSO, Metformin neither delayed clinical disease advance in these mice nor reduced the accumulation of sulfated glycosaminoglycans, a pathologic feature of prion disease. We conclude that elevation of anti-aging markers may not be sufficient to delay the fatal advance of genetic CJD.
Assuntos
Síndrome de Creutzfeldt-Jakob/genética , Síndrome de Creutzfeldt-Jakob/prevenção & controle , Metformina/farmacologia , Metformina/uso terapêutico , Óleos de Plantas/farmacologia , Óleos de Plantas/uso terapêutico , Adenilato Quinase/metabolismo , Animais , Antioxidantes , Modelos Animais de Doenças , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Camundongos , Camundongos Transgênicos , Mutação , Fator 2 Relacionado a NF-E2/metabolismo , Proteínas PrPSc/genética , Proteínas PrPSc/metabolismoRESUMO
Cryopreservation, the most common method of preserving stem cells, requires post-processing because it produces trauma to the cells. Post-thawing trauma typically induces cell death, elevates reactive oxygen species (ROS) concentration, and lowers mitochondrial membrane potential (MMP). Although this trauma has been solved using antioxidants, we attempted to use photobiomodulation (PBM) instead of chemical treatment. We used a 950-nm near-infrared LED to create a PBM device and chose a pulsed-wave mode of 30 Hz and a 30% duty cycle. Near-infrared radiation (NIR) at 950 nm was effective in reducing cell death caused by hydrogen peroxide induced-oxidative stress. Cryodamage also leads to apoptosis of cells, which can be avoided by irradiation at 950 nm NIR. Irradiation as post-processing for cryopreservation had an antioxidant effect that reduced both cellular and mitochondrial ROS. It also increased mitochondrial mass and activated mitochondrial activity, resulting in increased MMP, ATP generation, and increased cytochrome c oxidase activity. In addition, NIR increased alkaline phosphatase (ALP) activity, a biomarker of differentiation. As a result, we identified that 950 nm NIR PBM solves cryodamage in human stem cells from the apical papilla, indicating its potential as an alternative to antioxidants for treatment of post-thawing trauma, and further estimated its mechanism.
Assuntos
Antioxidantes/metabolismo , Potencial da Membrana Mitocondrial , Fototerapia/métodos , Células-Tronco/citologia , Fosfatase Alcalina/metabolismo , Apoptose , Biomarcadores/metabolismo , Biofísica , Biologia Celular , Morte Celular , Proliferação de Células/efeitos da radiação , Sobrevivência Celular/efeitos dos fármacos , Criopreservação , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Desenho de Equipamento , Humanos , Peróxido de Hidrogênio/metabolismo , Peróxido de Hidrogênio/farmacologia , Luminescência , Mitocôndrias/metabolismo , Estresse Oxidativo , Espécies Reativas de Oxigênio , Espectroscopia de Luz Próxima ao InfravermelhoRESUMO
The tremendous therapeutic potential of photobiomodulation therapy in different branches of medicine has been described in the literature. One of the molecular mechanisms for this treatment implicates the mitochondrial enzyme, cytochrome C oxidase. However, the efficacy and consistency of clinical outcomes with photobiomodulation treatments has been fiercely debated. This work was motivated by this need to improve photobiomodulation devices and delivery approaches. We designed a novel hand-piece with a flat-top beam profile of irradiation. We compared the beam profile versus a standard hand-piece and a fibre probe. We utilized isolated mitochondria and performed treatments at various spots within the beam, namely, the centre, left and right edge. We examined mitochondrial activity by assessing ATP synthesis with the luciferin/luciferase chemiluminescent method as a primary endpoint, while mitochondrial damage was assessed as the secondary endpoint. We observed a uniform distribution of the power density with the flat-top prototype compared to a wide Gaussian beam profile with the standard fibre and standard hand-piece. We noted increased production of ATP in the centre of all three beams with respect to the non-treated controls (p < 0.05). Both the fibre and standard hand-piece demonstrated less increase in ATP synthesis at the edges than the centre (p < 0.05). In contrast, ATP synthesis was increased homogenously in the flat-top handpiece, both in the centre and the edges of the beam. Fibre, standard hand-piece and the flat-top hand-piece prototype have discrete beam distribution characteristics. This significantly affected the mitochondrial activity with respect to their position within the treated areas. Flat-top hand-piece enhances the uniformity of photobiomodulation treatments and can improve the rigour and reproducibility of PBM clinical outcomes.
Assuntos
Trifosfato de Adenosina/metabolismo , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Lasers Semicondutores/estatística & dados numéricos , Mitocôndrias/enzimologia , Consumo de Oxigênio , Humanos , Mitocôndrias/efeitos da radiaçãoRESUMO
White adipose tissue (WAT) is a dynamic endocrine organ that can play a significant role in thermoregulation. WAT has the capacity to adopt structural and functional characteristics of the more metabolically active brown adipose tissue (BAT) and contribute to non-shivering thermogenesis under specific stimuli. Non-shivering thermogenesis was previously thought to be uncoupling protein 1 (UCP1)-dependent however, recent evidence suggests that UCP1-independent mechanisms of thermogenesis exist. Namely, futile creatine cycling has been identified as a contributor to WAT thermogenesis. The purpose of this study was to examine the efficacy of creatine supplementation to alter mitochondrial markers as well as adipocyte size and multilocularity in inguinal (iWAT), gonadal (gWAT), and BAT. Thirty-two male and female Sprague-Dawley rats were treated with varying doses (0 g/L, 2.5 g/L, 5 g/L, and 10 g/L) of creatine monohydrate for 8 weeks. We demonstrate that mitochondrial markers respond in a sex and depot specific manner. In iWAT, female rats displayed significant increases in COXIV, PDH-E1alpha, and cytochrome C protein content. Male rats exhibited gWAT specific increases in COXIV and PDH-E1alpha protein content. This study supports creatine supplementation as a potential method of UCP1-independant thermogenesis and highlights the importance of taking a sex-specific approach when examining the efficacy of browning therapeutics in future research.
Assuntos
Tecido Adiposo Branco/efeitos dos fármacos , Regulação da Temperatura Corporal/efeitos dos fármacos , Creatina/farmacologia , Suplementos Nutricionais , Mitocôndrias/efeitos dos fármacos , Tecido Adiposo Marrom/efeitos dos fármacos , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Branco/metabolismo , Animais , Citocromos c/metabolismo , Relação Dose-Resposta a Droga , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Feminino , Masculino , Mitocôndrias/metabolismo , Piruvato Desidrogenase (Lipoamida) , Ratos Sprague-Dawley , Fatores Sexuais , Proteína Desacopladora 1/metabolismoRESUMO
Objective: To study the effects of 670 and 830 nm irradiation on oxygen consumption by cytochrome c oxidase (CCO) in a Clark electrode type reaction chamber. To explore the effect of irradiation on the nitric oxide (NO) donor-induced inhibition of oxygen consumption. Background: Most theories of photobiomodulation (PBM) involve the enzyme CCO as a cellular target for red-to-near infrared light (R-NIR) irradiation. Attempts to measure the effect of irradiation on the kinetics of CCO have failed to demonstrate a significant effect. It remains to explore the effects of irradiation on the consumption of oxygen. NO has been proposed as a possible mediator for PBM due to its inhibitory effects on CCO. Studying the effect of R-NIR on NO-induced inhibition of oxygen consumption is needed to explore this thesis. Methods: Oxygen consumption assays at 22°C were performed in a Mitocell MT200A system equipped with a 1302 oxygen electrode. R-NIR irradiation at 670 nm (41 mW/cm2) or 830 nm (31 mW/cm2) was provided to the reaction mixture. Calculated second-order rate constants were compared with control runs at four cytochrome c concentrations. Assays were also performed with or without NO donor and/or light for two substrate concentrations. Results: Kinetics constants for oxygen consumption with or without R-NIR showed no significant differences with either wavelength at any substrate concentration. The NO donor showed significant inhibition that was not relieved by irradiation. Conclusions: This lack of effect by R-NIR calls into question both the CCO activity model and the NO inhibition relief model of PBM.
Assuntos
Complexo IV da Cadeia de Transporte de Elétrons , Terapia com Luz de Baixa Intensidade , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Raios Infravermelhos , Doadores de Óxido Nítrico/farmacologia , Consumo de OxigênioRESUMO
Food and feed are daily exposed to mycotoxin contamination which effects may be counteracted by antioxidants like carotenoids. Some mycotoxins as well as carotenoids penetrate the blood brain barrier (BBB) inducing alterations related to redox balance in the mitochondria. Therefore, the in vitro BBB model ECV304 was subcultured for 7 days and exposed to beauvericine, enniatins, ochratoxin A, zearalenone (100 nM each), individually and combined, and pumpkin extract (500 nM). Reactive oxygen species were measured by fluorescence using the dichlorofluorescein diacetate probe at 0 h, 2 h and 4 h. Intracellular ROS generation reported was condition dependent. RNA extraction was performed and gene expression was analyzed by qPCR after 2 h exposure. The selected genes were related to the Electron Transport Chain (ETC) and mitochondrial activity. Gene expression reported upregulation for exposures including mycotoxins plus pumpkin extract versus individual mycotoxins. Beauvericin and Beauvericin-Enniatins exposure significantly downregulated Complex I and pumpkin addition reverted the effect upregulating Complex I. Complex IV was the most downregulated structure of the ETC. Thioredoxin Interacting Protein was the most upregulated gene. These data confirm that mitochondrial processes in the BBB could be compromised by mycotoxin exposure and damage could be modulated by dietary antioxidants like carotenoids.
Assuntos
Carotenoides/farmacologia , Expressão Gênica/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Micotoxinas/toxicidade , Estresse Oxidativo/efeitos dos fármacos , Extratos Vegetais/farmacologia , Barreira Hematoencefálica/efeitos dos fármacos , Proteínas de Transporte/metabolismo , Linhagem Celular , Cucurbita/química , Depsipeptídeos/toxicidade , Regulação para Baixo/efeitos dos fármacos , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Genes Mitocondriais/efeitos dos fármacos , Humanos , Oxirredutases atuantes sobre Doadores de Grupo Enxofre/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteína Desacopladora 2/metabolismo , Regulação para Cima/efeitos dos fármacosRESUMO
BACKGROUND AND OBJECTIVES: Photobiomodulation (PBM) therapy uses light at various wavelengths to stimulate wound healing, grow hair, relieve pain, and more-but there is no consensus about optimal wavelengths or dosimetry. PBM therapy works through putative, wavelength-dependent mechanisms including direct stimulation of mitochondrial respiration, and/or activation of transmembrane signaling channels by changes in water activity. A common wavelength used in the visible red spectrum is ~660 nm, whereas recently ~980 nm is being explored and both have been proposed to work via different mechanisms. We aimed to gain more insight into identifying treatment parameters and the putative mechanisms involved. STUDY DESIGN/MATERIALS AND METHODS: Fluence-response curves were measured in cultured keratinocytes and fibroblasts exposed to 660 or 980 nm from LED sources. Metabolic activity was assessed using the MTT assay for reductases. ATP production, a major event triggered by PBM therapy, was assessed using a luminescence assay. To measure the role of mitochondria, we used an ELISA to measure COX-1 and SDH-A protein levels. The respective contributions of cytochrome c oxidase and ATP synthase to the PBM effects were gauged using specific inhibitors. RESULTS: Keratinocytes and fibroblasts responded differently to exposures at 660 nm (red) and 980 nm (NIR). Although 980 nm required much lower fluence for cell stimulation, the resulting increase in ATP levels was short-term, whereas 660 nm stimulation elevated ATP levels for at least 24 hours. COX-1 protein levels were increased following 660 nm treatment but were unaffected by 980 nm. In fibroblasts, SDH-A levels were affected by both wavelengths, whereas in keratinocytes only 660 nm light impacted SDH-A levels. Inhibition of ATP synthase nearly completely abolished the effects of both wavelengths on ATP synthesis. Interestingly, inhibiting cytochrome c oxidase did not prevent the rise in ATP levels in response to PBM treatment. CONCLUSION: To the best of our knowledge, this is the first demonstration of differing kinetics in response to PBM therapy at red versus NIR wavelength. We also found cell-type-specific differences in PBM therapy response to the two wavelengths studied. These findings confirm that different response pathways are involved after 660 and 980 nm exposures and suggest that 660 nm causes a more durable response. © 2021 Wiley Periodicals LLC.