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1.
Cells ; 11(24)2022 12 19.
Artigo em Inglês | MEDLINE | ID: mdl-36552890

RESUMO

PURPOSE: Mitochondrial dysfunction is central to breaking the barrier integrity of retinal endothelial cells (RECs) in various blinding eye diseases such as diabetic retinopathy and retinopathy of prematurity. Therefore, we aimed to investigate the role of different mitochondrial constituents, specifically those of oxidative phosphorylation (OxPhos), in maintaining the barrier function of RECs. METHODS: Electric cell-substrate impedance sensing (ECIS) technology was used to assess in real time the role of different mitochondrial components in the total impedance (Z) of human RECs (HRECs) and its components: capacitance (C) and the total resistance (R). HRECs were treated with specific mitochondrial inhibitors that target different steps in OxPhos: rotenone for complex I, oligomycin for complex V (ATP synthase), and FCCP for uncoupling OxPhos. Furthermore, data were modeled to investigate the effects of these inhibitors on the three parameters that govern the total resistance of cells: Cell-cell interactions (Rb), cell-matrix interactions (α), and cell membrane permeability (Cm). RESULTS: Rotenone (1 µM) produced the greatest reduction in Z, followed by FCCP (1 µM), whereas no reduction in Z was observed after oligomycin (1 µM) treatment. We then further deconvoluted the effects of these inhibitors on the Rb, α, and Cm parameters. Rotenone (1 µM) completely abolished the resistance contribution of Rb, as the Rb became zero immediately after the treatment. Secondly, FCCP (1 µM) eliminated the resistance contribution of Rb only after 2.5 h and increased Cm without a significant effect on α. Lastly, of all the inhibitors used, oligomycin had the lowest impact on Rb, as evidenced by the fact that this value became similar to that of the control group at the end of the experiment without noticeable effects on Cm or α. CONCLUSION: Our study demonstrates the differential roles of complex I, complex V, and OxPhos coupling in maintaining the barrier functionality of HRECs. We specifically showed that complex I is the most important component in regulating HREC barrier integrity. These observed differences are significant since they could serve as the basis for future pharmacological and gene expression studies aiming to improve the activity of complex I and thereby provide avenues for therapeutic modalities in endothelial-associated retinal diseases.


Assuntos
Retinopatia Diabética , Fosforilação Oxidativa , Recém-Nascido , Humanos , Rotenona/farmacologia , Células Endoteliais/metabolismo , Carbonil Cianeto p-Trifluormetoxifenil Hidrazona/metabolismo , Carbonil Cianeto p-Trifluormetoxifenil Hidrazona/farmacologia , Mitocôndrias/metabolismo , Retinopatia Diabética/metabolismo , Oligomicinas/farmacologia
2.
Int J Mol Sci ; 23(22)2022 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-36430329

RESUMO

In the central nervous system (CNS), calcium homeostasis is a critical determinant of neuronal survival. Calpain, a calcium-dependent neutral protease, is widely expressed in the brain, including substantia nigra (SN) dopaminergic (DA) neurons. Though calpain is implicated in human Parkinson's disease (PD) and corresponding animal models, the roles of specific ubiquitous calpain isoforms in PD, calpain-1 and calpain-2, remain poorly understood. In this study, we found that both isoforms are activated in a nigrostriatal pathway with increased phosphorylated synuclein following the administration of rotenone in Lewis rats, but calpain isoforms played different roles in neuronal survival. Although increased expression of calpain-1 and calpain-2 were detected in the SN of rotenone-administered rats, calpain-1 expression was not altered significantly after treatment with calpain inhibitor (calpeptin); this correlated with neuronal survival. By contrast, increased calpain-2 expression in the SN of rotenone rats correlated with neuronal death, and calpeptin treatment significantly attenuated calpain-2 and neuronal death. Calpain inhibition by calpeptin prevented glial (astroglia/microglia) activation in rotenone-treated rats in vivo, promoted M2-type microglia, and protected neurons. These data suggest that enhanced expression of calpain-1 and calpain-2 in PD models differentially affects glial activation and neuronal survival; thus, the attenuation of calpain-2 may be important in reducing SN neuronal loss in PD.


Assuntos
Doença de Parkinson , Rotenona , Ratos , Animais , Humanos , Rotenona/farmacologia , Calpaína/metabolismo , Doença de Parkinson/tratamento farmacológico , Ratos Endogâmicos Lew , Substância Negra/metabolismo , Neurônios Dopaminérgicos/metabolismo
3.
PLoS One ; 17(11): e0277477, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36441709

RESUMO

Rotenone is a commonly used insecticidal chemical in agriculture and it is an inhibitor of mitochondrial complex Ⅰ. Previous studies have found that rotenone induces the production of reactive oxygen species (ROS) by inhibiting electron transport in the mitochondria of somatic and germ cells. However, there is little precise information on the effects of rotenone exposure in porcine oocytes during in vitro maturation, and the mechanisms underlying these effects have not been determined. The Cumulus-oocyte complexes were supplemented with different concentrations of rotenone to elucidate the effects of rotenone exposure on the meiotic maturation of porcine oocytes during in vitro maturation for about 48 hours. First, we found that the maturation rate and expansion of cumulus cells were significantly reduced in the 3 and 5 µM rotenone-treated groups. Subsequently, the concentration of rotenone was determined to be 3 µM. Also, immunofluorescence, western blotting, and image quantification analyses were performed to test the rotenone exposure on the meiotic maturation, total and mitochondrial ROS, mitochondrial function and biogenesis, mitophagy and apoptosis in porcine oocytes. Further experiments showed that rotenone treatment induced mitochondrial dysfunction and failure of mitochondrial biogenesis by repressing the level of SIRT1 during in vitro maturation of porcine oocytes. In addition, rotenone treatment reduced the ratio of active mitochondria to total mitochondria, increased ROS production, and decreased ATP production. The levels of LC3 and active-caspase 3 were significantly increased by rotenone treatment, indicating that mitochondrial dysfunction induced by rotenone increased mitophagy but eventually led to apoptosis. Collectively, these results suggest that rotenone interferes with porcine oocyte maturation by inhibiting mitochondrial function.


Assuntos
Oócitos , Rotenona , Suínos , Animais , Feminino , Rotenona/farmacologia , Espécies Reativas de Oxigênio , Células do Cúmulo , Mitocôndrias
4.
Int J Mol Sci ; 23(22)2022 Nov 17.
Artigo em Inglês | MEDLINE | ID: mdl-36430754

RESUMO

Parkinson's disease (PD), the fastest-growing movement disorder, is still challenged by the unavailability of disease-modifying therapy. Mildly elevated levels of unconjugated bilirubin (UCB, PubChem CID 5280352) have been shown to be protective against several extra-CNS diseases, and the effect is attributed to its well-known anti-oxidant and anti-inflammatory capability. We explored the neuroprotective effect of low concentrations of UCB (from 0.5 to 4 µM) in our PD model based on organotypic brain cultures of substantia nigra (OBCs-SN) challenged with a low dose of rotenone (Rot). UCB at 0.5 and 1 µM fully protects against the loss of TH+ (dopaminergic) neurons (DOPAn). The alteration in oxidative stress is involved in TH+ positive neuron demise induced by Rot, but is not the key player in UCB-conferred protection. On the contrary, inflammation, specifically tumor necrosis factor alpha (TNF-α), was found to be the key to UCB protection against DOPAn sufferance. Further work will be needed to introduce the use of UCB into clinical settings, but determining that TNF-α plays a key role in PD may be crucial in designing therapeutic options.


Assuntos
Neurônios Dopaminérgicos , Doença de Parkinson , Humanos , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/patologia , Fator de Necrose Tumoral alfa/farmacologia , Bilirrubina/farmacologia , Bilirrubina/uso terapêutico , Degeneração Neural/patologia , Dopamina/farmacologia , Rotenona/farmacologia
5.
Int J Mol Sci ; 23(20)2022 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-36293513

RESUMO

Oral rotenone has been proposed as a model for Parkinson's disease (PD) in mice. To establish the model in our lab and study complex behavior we followed a published treatment regimen. C57BL/6 mice received 30 mg/kg body weight of rotenone once daily via oral administration for 4 and 8 weeks. Motor functions were assessed by RotaRod running. Immunofluorescence studies were used to analyze the morphology of dopaminergic neurons, the expression of alpha-Synuclein (α-Syn), and inflammatory gliosis or infiltration in the substantia nigra. Rotenone-treated mice did not gain body weight during treatment compared with about 4 g in vehicle-treated mice, which was however the only robust manifestation of drug treatment and suggested local gut damage. Rotenone-treated mice had no deficits in motor behavior, no loss or sign of degeneration of dopaminergic neurons, no α-Syn accumulation, and only mild microgliosis, the latter likely an indirect remote effect of rotenone-evoked gut dysbiosis. Searching for explanations for the model failure, we analyzed rotenone plasma concentrations via LC-MS/MS 2 h after administration of the last dose to assess bioavailability. Rotenone was not detectable in plasma at a lower limit of quantification of 2 ng/mL (5 nM), showing that oral rotenone had insufficient bioavailability to achieve sustained systemic drug levels in mice. Hence, oral rotenone caused local gastrointestinal toxicity evident as lack of weight gain but failed to evoke behavioral or biological correlates of PD within 8 weeks.


Assuntos
Doença de Parkinson , Transtornos Parkinsonianos , Animais , Camundongos , Rotenona/farmacologia , alfa-Sinucleína/metabolismo , Doença de Parkinson/etiologia , Doença de Parkinson/metabolismo , Transtornos Parkinsonianos/metabolismo , Cromatografia Líquida , Camundongos Endogâmicos C57BL , Espectrometria de Massas em Tandem , Substância Negra/metabolismo , Peso Corporal , Modelos Animais de Doenças
6.
Int J Mol Sci ; 23(19)2022 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-36232895

RESUMO

The mechanisms of mast cell (MC) degranulation and MC-driven skin symptoms are well-described. In contrast, data about the role of mitochondrial respiration for immune functions of human skin MCs are lacking. Oxygen consumption rate (OCR) in primary human skin MCs during IgE-mediated activation in the absence of glucose was examined using a metabolic flux analyzer. Effects of the inhibition of mitochondrial complex I (by rotenone A) and III (by myxothiazol) on degranulation and cytokine secretion (IL-4, IL-5, IL-6, IL-13, TNF-α, and GM-CSF) were explored by the ß-hexosaminidase release assay and multiplex ELISA. IgE-mediated activation rapidly increased the mitochondrial OCR and extracellular acidification; the contribution of non-mitochondrial oxygen consumption remained unchanged at lower levels. Both myxothiazol and rotenone A reduced OCR, the mitochondrial parameters, and extracellular acidification; however, myxothiazol did not affect degranulation and cytokine secretion. In contrast, degranulation and the secretion of IL-6, IL-13, TNF-α, and GM-CSF were reduced by rotenone A, whereas the secretion of IL-4 and IL-5 was not significantly affected. The inhibitors did not affect cell viability. Our results highlight the important role played by mitochondrial respiration in primary human skin MCs and allow for a conclusion on a hierarchy of their effector functions. Drugs targeting specific pathways in mitochondria may provide future options to control MC-driven skin symptoms.


Assuntos
Degranulação Celular , Mastócitos , Transporte de Elétrons , Complexo I de Transporte de Elétrons/metabolismo , Glucose/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Humanos , Imunoglobulina E , Interleucina-13/metabolismo , Interleucina-4/metabolismo , Interleucina-6/metabolismo , Mastócitos/metabolismo , Metacrilatos , Rotenona/metabolismo , Rotenona/farmacologia , Tiazóis , Fator de Necrose Tumoral alfa/metabolismo , beta-N-Acetil-Hexosaminidases/metabolismo
7.
Int J Mol Sci ; 23(20)2022 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-36293562

RESUMO

Mitochondria are an attractive target to fight neurodegenerative diseases due to their important functions for cells and the particularly close relationship between the functional connectivity among brain regions and mitochondrial performance. This work presents a mitochondria-targeted therapy designed to modulate the functionality of the mitochondrial respiratory chain and lipidome, parameters that are affected in neurodegeneration, including in Parkinson's disease (PD). This therapy is supported by SC-Nanophytosomes constructed with membrane polar lipids, from Codium tomentosum, and elderberry anthocyanin-enriched extract, from Sambucus nigra L. SC-Nanophytosomes are nanosized vesicles with a high negative surface charge that preserve their properties, including anthocyanins in the flavylium cation form, under conditions that mimic the gastrointestinal tract pH changes. SC-Nanophytosomes, 3 µM in phospholipid, and 2.5 mg/L of EAE-extract, delivered by drinking water to a rotenone-induced PD rat model, showed significant positive outcomes on disabling motor symptoms associated with the disease. Ex vivo assays were performed with two brain portions, one comprising the basal ganglia and cerebellum (BG-Cereb) and the other with the cerebral cortex (C-Cortex) regions. Results showed that rotenone-induced neurodegeneration increases the α-synuclein levels in the BG-Cereb portion and compromises mitochondrial respiratory chain functionality in both brain portions, well-evidenced by a 50% decrease in the respiratory control rate and up to 40% in complex I activity. Rotenone-induced PD phenotype is also associated with changes in superoxide dismutase and catalase activities that are dependent on the brain portion. Treatment with SC-Nanophytosomes reverted the α-synuclein levels and antioxidant enzymes activity to the values detected in control animals. Moreover, it mitigated mitochondrial dysfunction, with positive outcomes on the respiratory control rate, the activity of individual respiratory complexes, and the fatty acid profile of the membrane phospholipids. Therefore, SC-Nanophytosomes are a promising tool to support mitochondria-targeted therapy for neurodegenerative diseases.


Assuntos
Água Potável , Doença de Parkinson , Animais , Ratos , Rotenona/farmacologia , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/etiologia , Doença de Parkinson/metabolismo , Antocianinas/metabolismo , alfa-Sinucleína/metabolismo , Antioxidantes/farmacologia , Catalase/metabolismo , Mitocôndrias/metabolismo , Encéfalo/metabolismo , Superóxido Dismutase/metabolismo , Fosfolipídeos/metabolismo , Ácidos Graxos/metabolismo , Modelos Animais de Doenças
8.
Behav Pharmacol ; 33(8): 513-526, 2022 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-36094044

RESUMO

Parkinson's disease is a neurodegenerative disease, the etiology of which remains unknown, but some likely causes include oxidative stress, mitochondrial dysfunction and neuroinflammation. Peroxisome-proliferator-activated receptor (PPAR) agonists have been studied in animal models of Parkinson's disease and have shown neuroprotective effects. In this study, we aimed to (1) confirm the neuroprotective effects of PPAR-alpha agonist fenofibrate. To this end, male rats received fenofibrate (100 mg/kg) orally for 15 days, 5 days before the intraperitoneal injections of rotenone (2.5 mg/kg for 10 days). After finishing the treatment with rotenone and fenofibrate, animals were subjected to the open field, the forced swim test and the two-way active avoidance task. Subsequently, rats were euthanized for measurement of dopamine and metabolites levels in the striatum and quantification of tyrosine hydroxylase-immunoreactive neurons in the substantia nigra pars compacta (SNpc). In addition, we aimed to (2) evaluate the neuroprotective effects of fenofibrate on the accumulation of α-synuclein aggregates. Here, rats were treated for 5 days with fenofibrate continuing for over 28 days with rotenone. Then, animals were perfused for immunohistochemistry analysis of α-synuclein. The results showed that fenofibrate reduced depressive-like behavior and memory impairment induced by rotenone. Moreover, fenofibrate diminished the depletion of striatal dopamine and protected against dopaminergic neuronal death in the SNpc. Likewise, the administration of fenofibrate attenuated the aggregation of α-synuclein in the SNpc and striatum in the rotenone-lesioned rats. Our study confirmed that fenofibrate exerted neuroprotective effects because parkinsonian rats exhibited reduced behavioral, neurochemical and immunohistochemical changes, and importantly, a lower number of α-synuclein aggregates.


Assuntos
Fenofibrato , Doenças Neurodegenerativas , Fármacos Neuroprotetores , Doença de Parkinson , Masculino , Ratos , Animais , Rotenona/farmacologia , Doença de Parkinson/metabolismo , Fenofibrato/farmacologia , alfa-Sinucleína , Neuroproteção , Fármacos Neuroprotetores/farmacologia , Dopamina/metabolismo , Receptores Ativados por Proliferador de Peroxissomo/metabolismo , Receptores Ativados por Proliferador de Peroxissomo/farmacologia , Modelos Animais de Doenças , Neurônios Dopaminérgicos , Substância Negra
9.
Biomolecules ; 12(9)2022 08 31.
Artigo em Inglês | MEDLINE | ID: mdl-36139052

RESUMO

Hypoxia plays a crucial role in acute and chronic renal injury, which is attributable to renal tubular and glomerular cell damage. Some studies provide evidence that hypoxia-dependent upregulation of the mitochondrial enzyme arginase type-II (Arg-II) in tubular cells promotes renal tubular injury. It is, however, not known whether Arg-II is also expressed in glomerular cells, particularly podocytes under hypoxic conditions, contributing to hypoxia-induced podocyte injury. The effects of hypoxia on human podocyte cells (AB8/13) in cultures and on isolated kidneys from wild-type (wt) and arg-ii gene-deficient (arg-ii-/-) mice ex vivo, as well as on mice of the two genotypes in vivo, were investigated, respectively. We found that the Arg-II levels were enhanced in cultured podocytes in a time-dependent manner over 48 h, which was dependent on the stabilization of hypoxia-inducible factor 1α (HIF1α). Moreover, a hypoxia-induced derangement of cellular actin cytoskeletal fibers, a decrease in podocin, and an increase in mitochondrial ROS (mtROS) generation-as measured by MitoSOX-were inhibited by adenoviral-mediated arg-ii gene silencing. These effects of hypoxia on podocyte injury were mimicked by the HIFα stabilizing drug DMOG, which inhibits prolyl hydroxylases (PHD), the enzymes involved in HIFα degradation. The silencing of arg-ii prevented the detrimental effects of DMOG on podocytes. Furthermore, the inhibition of mtROS generation by rotenone-the inhibitor of respiration chain complex-I-recapitulated the protective effects of arg-ii silencing on podocytes under hypoxic conditions. Moreover, the ex vivo experiments with isolated kidney tissues and the in vivo experiments with mice exposed to hypoxic conditions showed increased Arg-II levels in podocytes and decreased podocyte markers regarding synaptopodin in wt mice but not in arg-ii-/- mice. While age-associated albuminuria was reduced in the arg-ii-/- mice, the hypoxia-induced increase in albuminuria was, however, not significantly affected in the arg-ii-/-. Our study demonstrates that Arg-II in podocytes promotes cell injury. Arg-ii ablation seems insufficient to protect mice in vivo against a hypoxia-induced increase in albuminuria, but it does reduce albuminuria in aging.


Assuntos
Arginase , Podócitos , Actinas/metabolismo , Albuminúria , Animais , Arginase/genética , Arginase/metabolismo , Humanos , Hipóxia/metabolismo , Camundongos , Podócitos/metabolismo , Prolil Hidroxilases/metabolismo , Prolil Hidroxilases/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Rotenona/farmacologia
10.
Int J Mol Sci ; 23(18)2022 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-36142834

RESUMO

Essential changes in cell metabolism and redox signaling occur during the reprogramming of somatic cells into induced pluripotent stem cells (iPSCs). In this paper, using genetic and pharmacological approaches, we have investigated the role of electron transport chain (ETC) complex-I (CI) of mitochondria in the process of cell reprogramming to pluripotency. Knockdown of NADH-ubiquinone oxidoreductase core subunits S1 (Ndufs1) or subunit B10 (Ndufb10) of the CI or inhibition of this complex with rotenone during mouse embryonic fibroblast (MEF) reprogramming resulted in a significantly decreased number of induced pluripotent stem cells (iPSCs). We have found that mitochondria and ROS levels due course of the reprogramming tightly correlate with each other, both reaching peak by day 3 and significantly declining by day 10 of the process. The transient augmentation of mitochondrial reactive oxygen species (ROS) could be attenuated by antioxidant treatment, which ameliorated overall reprogramming. However, ROS scavenging after day 3 or during the entire course of reprogramming was suppressive for iPSC formation. The ROS scavenging within the CI-deficient iPSC-precursors did not improve, but further suppressed the reprogramming. Our data therefore point to distinct modes of mitochondrial ROS action during the early versus mid and late stages of reprogramming. The data further substantiate the paradigm that balanced levels of oxidative phosphorylation have to be maintained on the route to pluripotency.


Assuntos
Complexo I de Transporte de Elétrons , Células-Tronco Pluripotentes Induzidas , Animais , Antioxidantes/metabolismo , Reprogramação Celular/genética , Complexo I de Transporte de Elétrons/genética , Complexo I de Transporte de Elétrons/metabolismo , Fibroblastos/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Camundongos , Mitocôndrias/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Rotenona/metabolismo , Rotenona/farmacologia
11.
Acta Med Okayama ; 76(4): 373-383, 2022 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-36123151

RESUMO

Parkinson's disease (PD) is a progressive neurodegenerative disease of both the central and peripheral / enteric nervous systems. Oxidative stress and neuroinflammation are associated with the pathogenesis of PD, suggesting that anti-oxidative and anti-inflammatory compounds could be neuroprotective agents for PD. Eucommia ulmoides (EU) is a traditional herbal medicine which exerts neuroprotective effects by anti-inflammatory and anti-oxidative properties. Our previous study showed that treatment with chlorogenic acid, a component of EU, protected against neurodegeneration in the central and enteric nervous systems in a PD model. In this study, we examined the effects of EU extract (EUE) administration on dopaminergic neurodegeneration, glial response and α-synuclein expression in the substantia nigra pars compacta (SNpc), and intestinal enteric neurodegeneration in low-dose rotenone-induced PD model mice. Daily oral administration of EUE ameliorated dopaminergic neurodegeneration and α-synuclein accumulation in the SNpc. EUE treatment inhibited rotenone-induced decreases in the number of total astrocytes and in those expressing the antioxidant molecule metallothionein. EUE also prevented rotenone-induced microglial activation. Furthermore, EUE treatment exerted protective effects against intestinal neuronal loss in the PD model. These results suggest that EU exerts neuroprotective effects in the central and enteric nervous systems of rotenone-induced parkinsonism mice, in part by glial modification.


Assuntos
Eucommiaceae , Doenças Neurodegenerativas , Fármacos Neuroprotetores , Animais , Antioxidantes/metabolismo , Ácido Clorogênico/metabolismo , Ácido Clorogênico/farmacologia , Dopamina/metabolismo , Dopamina/farmacologia , Neurônios Dopaminérgicos/metabolismo , Neurônios Dopaminérgicos/patologia , Eucommiaceae/metabolismo , Metalotioneína/metabolismo , Metalotioneína/farmacologia , Camundongos , Doenças Neurodegenerativas/metabolismo , Doenças Neurodegenerativas/patologia , Fármacos Neuroprotetores/metabolismo , Fármacos Neuroprotetores/farmacologia , Fármacos Neuroprotetores/uso terapêutico , Extratos Vegetais/metabolismo , Extratos Vegetais/farmacologia , Extratos Vegetais/uso terapêutico , Rotenona/metabolismo , Rotenona/farmacologia , alfa-Sinucleína/metabolismo , alfa-Sinucleína/farmacologia
12.
Neuroreport ; 33(15): 635-640, 2022 10 12.
Artigo em Inglês | MEDLINE | ID: mdl-36126260

RESUMO

OBJECTIVE: Strokes represent as one of the leading causes of death and disability in the USA, however, there is no optimal treatment to reduce the occurrence or improve prognosis. Preconditioning of tissues triggers ischemic tolerance, a physiological state that may involve a metabolic switch (i.e. from glycolysis to oxidative phosphorylation or OxPhos) to preserve tissue viability under an ischemic insult. Here, we hypothesized that metabolic switching of energy source from glucose to galactose in cultured mesenchymal stem cells (MSCs) stands as an effective OxPhos-enhancing strategy. METHODS: MSCs were grown under ambient condition (normal MSCs) or metabolic switching paradigm (switched MSCs) and then assayed for oxygen consumption rates (OCR) and extracellular acidification rate (ECAR) using the Seahorse technology to assess mitochondrial respiration. RESULTS: Normal MSCs showed a lower OCR/ECAR ratio than switched MSCs at baseline (P < 0.0001), signifying that there were greater levels of OxPhos compared to glycolysis in switched MSCs. By modulating the mitochondrial metabolism with oligomycin (time points 4-6), carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazone (7-9), and rotenone and antimycin (time points 10-12), switched MSCs greater reliance on OxPhos was further elucidated (time points 5-12; P < 0.0001; time point 4; P < 0.001). CONCLUSION: The metabolic switch from glycolytic to oxidative metabolism amplifies the OxPhos potential of MSCs, which may allow these cells to afford more robust therapeutic effects against neurological disorders that benefit from ischemic tolerance.


Assuntos
Células-Tronco Mesenquimais , Fosforilação Oxidativa , Galactose/metabolismo , Glucose/metabolismo , Glicólise/fisiologia , Células-Tronco Mesenquimais/metabolismo , Oligomicinas/metabolismo , Rotenona/farmacologia
13.
Environ Entomol ; 51(5): 948-957, 2022 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-36130186

RESUMO

Chemicals, including toxic bait and dusts, are the main means of controlling the red imported fire ant Solenopsis invicta (abbreviation 'RIFA'), which is a serious invasive pest. To identify environmentally friendly chemicals for controlling RIFA, we tested the toxicity and horizontal transfer of three botanical insecticides-matrine, rotenone, and pyrethrin-and their impact on aquatic organisms (i.e., three fish and one shrimp). The LD50 value of matrine, rotenone, and pyrethrin was 0.24, 50.929, and 13.590 ng/ant, respectively. Matrine, rotenone, and pyrethrin had effective horizontal transfer and caused significant secondary mortality. After exposure to donor workers, 90.75%, 90.75%, and 100% of recipient workers in the 0.025% matrine, 1.0% rotenone, and 0.3% pyrethrin dust treatments, respectively, died within 48 h. Furthermore, 0.025% matrine dust caused significant tertiary mortality (49.5%). Tertiary mortality caused by 1.0% rotenone and 0.3% pyrethrin dusts was very low, only 7.75% and 18.5%, respectively. In a field trial, the comprehensive mortality effect of 0.05% matrine, 1.50% rotenone, and 0.375% pyrethrin dust was 74.96%, 30.10%, and 29.27%, respectively, after 14 d of treatment. Matrine had low toxicity to the fish Hypophthalmichthys molitrix, and 37.5 mg/L pyrethrin had low toxicity to the fish Cirrhinus molitorella, H. molitrix, and Oreochromis niloticus. However, rotenone was highly toxic to all three fish. The shrimp Neocaridina denticulate was not sensitive to matrine, rotenone, or pyrethrin. Matrine dust has highly effective horizontal transfer, and 0.05% matrine dust has great potential for managing RIFA in organic farms, aquaculture farms of H. molitrix, and water resource regions.


Assuntos
Formigas , Inseticidas , Piretrinas , Animais , Inseticidas/toxicidade , Piretrinas/toxicidade , Rotenona/farmacologia , Organismos Aquáticos , Poeira
14.
Int J Mol Sci ; 23(17)2022 Aug 24.
Artigo em Inglês | MEDLINE | ID: mdl-36076995

RESUMO

Mitochonic Acid 5 (MA-5) enhances mitochondrial ATP production, restores fibroblasts from mitochondrial disease patients and extends the lifespan of the disease model "Mitomouse". Additionally, MA-5 interacts with mitofilin and modulates the mitochondrial inner membrane organizing system (MINOS) in mammalian cultured cells. Here, we used the nematode Caenorhabditis elegans to investigate whether MA-5 improves the Duchenne muscular dystrophy (DMD) model. Firstly, we confirmed the efficient penetration of MA-5 in the mitochondria of C. elegans. MA-5 also alleviated symptoms such as movement decline, muscular tone, mitochondrial fragmentation and Ca2+ accumulation of the DMD model. To assess the effect of MA-5 on mitochondria perturbation, we employed a low concentration of rotenone with or without MA-5. MA-5 significantly suppressed rotenone-induced mitochondria reactive oxygen species (ROS) increase, mitochondrial network fragmentation and nuclear destruction in body wall muscles as well as endogenous ATP levels decline. In addition, MA-5 suppressed rotenone-induced degeneration of dopaminergic cephalic (CEP) neurons seen in the Parkinson's disease (PD) model. Furthermore, the application of MA-5 reduced mitochondrial swelling due to the immt-1 null mutation. These results indicate that MA-5 has broad mitochondrial homing and MINOS stabilizing activity in metazoans and may be a therapeutic agent for these by ameliorating mitochondrial dysfunction in DMD and PD.


Assuntos
Distrofia Muscular de Duchenne , Doença de Parkinson , Trifosfato de Adenosina , Animais , Caenorhabditis elegans/genética , Humanos , Ácidos Indolacéticos , Mamíferos , Distrofia Muscular de Duchenne/tratamento farmacológico , Distrofia Muscular de Duchenne/genética , Doença de Parkinson/tratamento farmacológico , Fenilbutiratos , Rotenona/farmacologia
15.
Chem Res Toxicol ; 35(11): 1974-1982, 2022 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-36178476

RESUMO

In this work, we carried out neurochemical and behavioral analysis of zebrafish (Danio rerio) treated with rotenone, an agent used to chemically induce a syndrome resembling Parkinson's disease (PD). Dopamine release, measured with fast-scan cyclic voltammetry (FSCV) at carbon-fiber electrodes in acutely harvested whole brains, was about 30% of that found in controls. Uptake, represented by the first order rate constant (k) and the half-life (t1/2) determined by nonlinear regression modeling of the stimulated release plots, was also diminished. Behavioral analysis revealed that rotenone treatment increased the time required for zebrafish to reach a reward within a maze by more than 50% and caused fish to select the wrong pathway, suggesting that latent learning was impaired. Additionally, zebrafish treated with rotenone suffered from diminished locomotor activity, swimming shorter distances with lower mean velocity and acceleration. Thus, the neurochemical and behavioral approaches, as applied, were able to resolve rotenone-induced differences in key parameters. This approach may be effective for screening therapies in this and other models of neurodegeneration.


Assuntos
Doença de Parkinson , Rotenona , Animais , Rotenona/farmacologia , Peixe-Zebra/metabolismo , Dopamina/metabolismo , Modelos Animais de Doenças , Cognição
16.
Inflammopharmacology ; 30(5): 1705-1716, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-35931897

RESUMO

Parkinson's disease (PD) remains a disease of little known etiology. In addition to the motor symptoms, depression is present in about 40% of patients, contributing to the loss of quality of life. Recently, the involvement of the autophagy mechanism in the pathogenesis of depression has been studied, in addition to its involvement in PD as well. In this study, we tested the effects of metformin, an antidiabetic drug also with antidepressant effects, on depressive-like behavior in a rotenone-induced PD model and on the autophagy process. Mice 8-week-old male C57BL/6 were induced with rotenone for 20 consecutive days (2.5 mg/kg/day) and treated with metformin (200 mg/kg/day) from the 5th day of induction. All the animals were submitted to rotarod, sucrose preference and tail suspension tests. After euthanasia, the substantia nigra and hippocampus were removed for analysis by western blotting or fixed and analyzed by immunofluorescence. The results show that there was an impairment of autophagy in animals induced by rotenone both in nigral and extranigral regions as well as a depressive-like behavior. Metformin was able to inhibit depressive-like behavior and increase signaling pathway proteins, transcription factors and autophagosome-forming proteins, thus inducing autophagy in both the hippocampus and the substantia nigra. In conclusion, we show that metformin has an antidepressant effect in a rotenone-induced PD model, which may result, at least in part, from the induction of the autophagy process.


Assuntos
Metformina , Doença de Parkinson , Animais , Antidepressivos/farmacologia , Autofagia , Modelos Animais de Doenças , Hipocampo/metabolismo , Hipoglicemiantes/metabolismo , Hipoglicemiantes/farmacologia , Masculino , Metformina/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/metabolismo , Qualidade de Vida , Rotenona/farmacologia , Substância Negra , Sacarose/metabolismo , Sacarose/farmacologia , Fatores de Transcrição/metabolismo , Fatores de Transcrição/farmacologia
17.
Int J Mol Sci ; 23(16)2022 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-36012337

RESUMO

Mitochondrial electron transport chain (ETC) inhibition is a phenomenon interesting in itself and serves as a tool for studying various cellular processes. Despite the fact that searching the term "rotenone" in PubMed returns more than 6900 results, there are many discrepancies regarding the directions of changes reported to be caused by this RTC inhibitor in the delicate redox balance of the cell. Here, we performed a multifaceted study of the popular ETC inhibitors rotenone and antimycin A, involving assessment of mitochondrial membrane potential and the production of hydrogen peroxide and superoxide anions at cellular and mitochondrial levels over a wide range of inhibitor concentrations (1 nmol/dm3-100 µmol/dm3). All measurements were performed with whole cells, with accompanying control of ATP levels. Antimycin A was more potent in hindering HepG2 cells' abilities to produce ATP, decreasing ATP levels even at a 1 nmol/dm3 concentration, while in the case of rotenone, a 10,000-times greater concentration was needed to produce a statistically significant decrease. The amount of hydrogen peroxide produced in the course of antimycin A biological activity increased rapidly at low concentrations and decreased below control level at a high concentration of 100 µmol/dm3. While both inhibitors influenced cellular superoxide anion production in a comparable manner, rotenone caused a greater increase in mitochondrial superoxide anions compared to a modest impact for antimycin A. IC50 values for rotenone and antimycin A with respect to HepG2 cell survival were of the same order of magnitude, but the survival curve of cells treated with rotenone was clearly biphasic, suggesting a concentration-dependent mode of biological action. We propose a clear experimental setup allowing for complete and credible analysis of the redox state of cells under stress conditions which allows for better understanding of the effects of ETC inhibition.


Assuntos
Peróxido de Hidrogênio , Superóxidos , Trifosfato de Adenosina/metabolismo , Antimicina A/farmacologia , Transporte de Elétrons , Peróxido de Hidrogênio/metabolismo , Rotenona/farmacologia , Superóxidos/metabolismo
18.
Int J Mol Sci ; 23(16)2022 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-36012451

RESUMO

INTRODUCTION: Alpha lipoic acid (ALA) is a sulphur-containing organic compound, derived from octanoic acid, and an important cofactor for mitochondrial respiratory enzymes. It has strong antioxidant properties that improve mitochondrial function. We investigated if ALA improves mitochondrial dysfunction in a cellular model of Alzheimer's disease (AD). METHODS: SH-SY5Y-APP695 cells were used as a model for an early stage of AD. Vector-transfected SH-SY5Y-MOCK cells served as controls. Using these cells, we investigated mitochondrial respiration (OXPHOS), mitochondrial membrane potential (MMP), adenosine triphosphate (ATP) production, and citrate synthase activity (CS) in cells treated with ALA. Cells were treated for 24 h with different concentrations of ALA and with or without the complex I inhibitor rotenone. RESULTS: Incubation with ALA showed a significant increase in ATP levels in both SH-SY5Y-APP695 and SH-SY5Y-MOCK cells. MMP levels were elevated in SH-SY5Y-MOCK cells, treatment with rotenone showed a reduction in MMP, which could be partly alleviated after incubation with ALA in SH-SY5Y-MOCK cells. ALA treatment showed significant differences in respiration chain complex activities in SH-SY5Y-MOCK cells. Citrate synthase activity was unaffected. ROS levels were significantly lower in both cell lines treated with ALA. CONCLUSIONS: ALA increased the activity of the different complexes of the respiratory chain, and consequently enhanced the MMP, leading to increased ATP levels indicating improved mitochondrial function. ALA only marginally protects from additional rotenone-induced mitochondrial stress.


Assuntos
Doença de Alzheimer , Neuroblastoma , Ácido Tióctico , Trifosfato de Adenosina/metabolismo , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/metabolismo , Linhagem Celular Tumoral , Citrato (si)-Sintase/metabolismo , Humanos , Potencial da Membrana Mitocondrial , Mitocôndrias/metabolismo , Neuroblastoma/metabolismo , Oxirredução , Rotenona/metabolismo , Rotenona/farmacologia , Ácido Tióctico/metabolismo , Ácido Tióctico/farmacologia
19.
Eur J Med Chem ; 242: 114692, 2022 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-36029560

RESUMO

Parkinson's disease (PD) is a neurodegenerative disorder that causes uncontrollable movements. Although many breakthroughs in PD therapy have been accomplished, there is currently no cure for PD, and only trials to relieve symptoms have been evaluated. Recently, we reported the total synthesis of cudraisoflavone J and its chiral isomers [Lu et al., J. Nat. Prod. 2021, 84, 1359]. In this study, we designed and synthesized a series of novel cudraisoflavone J derivatives and evaluated their neuroprotective activities in neurotoxin-treated PC12 cells. Among these compounds, difluoro-substituted derivative (13m) and prenylated derivative (24) provided significant protection to PC12 cells against toxicity induced by 6-hydroxydopamine (6-OHDA) or rotenone. Both derivatives inhibited 6-OHDA- or rotenone-induced production of reactive oxygen species and partially attenuated lipid peroxidation in rat brain homogenates, indicating their antioxidant properties. They also increased the expression of the antioxidant enzyme, heme oxygenase (HO)-1, and enhanced the nuclear translocation of Nrf2, the transcription factor that regulates the expression of antioxidant proteins. The neuroprotective effects of 13m and 24 were eliminated by Zn(II)-protoporphyrin IX, an HO-1 inhibitor, demonstrating the critical role of HO-1 in their actions. Moreover, upregulation of HO-1 was abolished by nuclear factor erythroid 2-related factor (Nrf2) knockdown, verifying that Nrf2 is an upstream regulator of HO-1. Compounds 13m and 24 triggered phosphorylation of ERK1/2, JNK, and Akt. Most importantly, 13m- and 24-induced enhancement of Nrf2 translocation and HO-1 expression was reversed by U0126 (an ERK inhibitor), SP600125 (a JNK inhibitor), and LY294002 (an Akt inhibitor). Collectively, our results show that compounds 13m and 24 exert neuroprotective and antioxidant effects through the Nrf2/HO-1 pathway mediated by phosphorylation of ERK1/2, JNK, or Akt in PC12 cells. Based on our findings, both derivatives could serve as potential therapeutic candidates for the neuroprotective treatment of PD.


Assuntos
Fármacos Neuroprotetores , Doença de Parkinson , Animais , Antioxidantes/farmacologia , Citocromo P-450 CYP2B1/metabolismo , Citocromo P-450 CYP2B1/farmacologia , Fator de Transcrição de Proteínas de Ligação GA/metabolismo , Heme Oxigenase-1/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Fármacos Neuroprotetores/farmacologia , Neurotoxinas/farmacologia , Estresse Oxidativo , Oxidopamina/farmacologia , Doença de Parkinson/tratamento farmacológico , Inibidores da Agregação Plaquetária/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Espécies Reativas de Oxigênio/metabolismo , Rotenona/farmacologia
20.
Mol Neurobiol ; 59(9): 5874-5890, 2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-35804280

RESUMO

Boswellia serrata gum is a natural product that showed beneficial effects on neurodegenerative diseases in recent studies. In this study, we investigated the effects of Boswellia serrata resin on rotenone-induced dopaminergic neurotoxicity. Firstly, we attempted to see if the resin can induce AMP-activated protein kinase (AMPK) signaling pathway which has been known to have broad neuroprotective effects. Boswellia increased AMPK phosphorylation and reduced phosphorylation of mammalian target of rapamycin (p-mTOR) and α-synuclein (p-α-synuclein) in the striatum while increased the expression level of Beclin1, a marker for autophagy and brain-derived neurotrophic factor. Next, we examined the neuroprotective effects of the Boswellia extract in the rotenone-injected mice. The results showed that Boswellia evidently attenuated the loss of the nigrostriatal dopaminergic neurons and microglial activation caused by rotenone. Moreover, Boswellia ameliorated rotenone-induced decrease in the striatal dopamine and impairment in motor function. Accumulation of α-synuclein meditated by rotenone was significantly ameliorated by Boswellia. Also, we showed that ß-boswellic acid, the active constituents of Boswellia serrata gum, induced AMPK phosphorylation and attenuated α-synuclein phosphorylation in SHSY5 cells. These results suggest that Boswellia protected the dopaminergic neurons from rotenone neurotoxicity via activation of the AMPK pathway which might be associated with attenuation of α-synuclein aggregation and neuroinflammation. Further investigations are warranted to identify specific molecules in Boswellia which are responsible for the neuroprotection.


Assuntos
Boswellia , Fármacos Neuroprotetores , Síndromes Neurotóxicas , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Boswellia/metabolismo , Dopamina/metabolismo , Neurônios Dopaminérgicos/metabolismo , Mamíferos/metabolismo , Metanol/metabolismo , Metanol/farmacologia , Camundongos , Fármacos Neuroprotetores/metabolismo , Fármacos Neuroprotetores/farmacologia , Síndromes Neurotóxicas/metabolismo , Extratos Vegetais/metabolismo , Extratos Vegetais/farmacologia , Rotenona/farmacologia , alfa-Sinucleína/metabolismo
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