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1.
Int J Mol Sci ; 22(19)2021 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-34638589

RESUMO

Aging is associated with a decline in cognitive function, which can partly be explained by the accumulation of damage to the brain cells over time. Neurons and glia undergo morphological and ultrastructure changes during aging. Over the past several years, it has become evident that at the cellular level, various hallmarks of an aging brain are closely related to mitophagy. The importance of mitochondria quality and quantity control through mitophagy is highlighted by the contribution that defects in mitochondria-autophagy crosstalk make to aging and age-related diseases. In this review, we analyze some of the more recent findings regarding the study of brain aging and neurodegeneration in the context of mitophagy. We discuss the data on the dynamics of selective autophagy in neurons and glial cells during aging and in the course of neurodegeneration, focusing on three mechanisms of mitophagy: non-receptor-mediated mitophagy, receptor-mediated mitophagy, and transcellular mitophagy. We review the role of mitophagy in neuronal/glial homeostasis and in the molecular pathogenesis of neurodegenerative disorders, such as Parkinson's disease, Alzheimer's disease, and other disorders. Common mechanisms of aging and neurodegeneration that are related to different mitophagy pathways provide a number of promising targets for potential therapeutic agents.


Assuntos
Envelhecimento/patologia , Mitofagia/fisiologia , Neuroglia/patologia , Neurônios/patologia , Animais , Autofagia/fisiologia , Homeostase/fisiologia , Humanos , Mitocôndrias/patologia , Doenças Neurodegenerativas/patologia
2.
Cell Mol Life Sci ; 78(21-22): 6851-6867, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34524466

RESUMO

Mitochondria-the intracellular powerhouse in which nutrients are converted into energy in the form of ATP or heat-are highly dynamic, double-membraned organelles that harness a plethora of cellular functions that sustain energy metabolism and homeostasis. Exciting new discoveries now indicate that the maintenance of this ever changing and functionally pleiotropic organelle is particularly relevant in terminally differentiated cells that are highly dependent on aerobic metabolism. Given the central role in maintaining metabolic and physiological homeostasis, dysregulation of the mitochondrial network might therefore confer a potentially devastating vulnerability to high-energy requiring cell types, contributing to a broad variety of hereditary and acquired diseases. In this Review, we highlight the biological functions of mitochondria-localized enzymes from the perspective of understanding-and potentially reversing-the pathophysiology of inherited disorders affecting the homeostasis of the mitochondrial network and cellular metabolism. Using methylmalonic acidemia as a paradigm of complex mitochondrial dysfunction, we discuss how mitochondrial directed-signaling circuitries govern the homeostasis and physiology of specialized cell types and how these may be disturbed in disease. This Review also provides a critical analysis of affected tissues, potential molecular mechanisms, and novel cellular and animal models of methylmalonic acidemia which are being used to develop new therapeutic options for this disease. These insights might ultimately lead to new therapeutics, not only for methylmalonic acidemia, but also for other currently intractable mitochondrial diseases, potentially transforming our ability to regulate homeostasis and health.


Assuntos
Erros Inatos do Metabolismo dos Aminoácidos/metabolismo , Mitocôndrias/metabolismo , Doenças Mitocondriais/metabolismo , Mitofagia/fisiologia , Animais , Metabolismo Energético/fisiologia , Homeostase/fisiologia , Humanos , Organelas/metabolismo , Transdução de Sinais/fisiologia
3.
Sci Rep ; 11(1): 17733, 2021 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-34489512

RESUMO

Macroautophagic recycling of dysfunctional mitochondria, known as mitophagy, is essential for mitochondrial homeostasis and cell viability. Accumulation of defective mitochondria and impaired mitophagy have been widely implicated in many neurodegenerative diseases, and loss-of-function mutations of PINK1 and Parkin, two key regulators of mitophagy, are amongst the most common causes of heritable parkinsonism. This has led to the hypothesis that pharmacological stimulation of mitophagy may be a feasible approach to combat neurodegeneration. Toward this end, we screened ~ 45,000 small molecules using a high-throughput, whole-organism, phenotypic screen that monitored accumulation of PINK-1 protein, a key event in mitophagic activation, in a Caenorhabditis elegans strain carrying a Ppink-1::PINK-1::GFP reporter. We obtained eight hits that increased mitochondrial fragmentation and autophagosome formation. Several of the compounds also reduced ATP production, oxygen consumption, mitochondrial mass, and/or mitochondrial membrane potential. Importantly, we found that treatment with two compounds, which we named PS83 and PS106 (more commonly known as sertraline) reduced neurodegenerative disease phenotypes, including delaying paralysis in a C. elegans ß-amyloid aggregation model in a PINK-1-dependent manner. This report presents a promising step toward the identification of compounds that will stimulate mitochondrial turnover.


Assuntos
Mitocôndrias/metabolismo , Doenças Neurodegenerativas/metabolismo , Proteostase/fisiologia , Animais , Caenorhabditis elegans , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/genética , Mitofagia/efeitos dos fármacos , Mitofagia/fisiologia , Doenças Neurodegenerativas/genética , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteostase/efeitos dos fármacos , Selenito de Sódio/farmacologia , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo
4.
Exp Cell Res ; 407(1): 112758, 2021 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-34437881

RESUMO

Podocytes constitute the outer layer of the renal glomerular filtration barrier. Their energy requirements strongly depend on efficient oxidative respiration, which is tightly connected with mitochondrial dynamics. We hypothesized that hyperglycemia modulates energy metabolism in glomeruli and podocytes and contributes to the development of diabetic kidney disease. We found that oxygen consumption rates were severely reduced in glomeruli from diabetic rats and in human podocytes that were cultured in high glucose concentration (30 mM; HG). In these models, all of the mitochondrial respiratory parameters, including basal and maximal respiration, ATP production, and spare respiratory capacity, were significantly decreased. Podocytes that were treated with HG showed a fragmented mitochondrial network, together with a decrease in expression of the mitochondrial fusion markers MFN1, MFN2, and OPA1, and an increase in the activity of the fission marker DRP1. We showed that markers of mitochondrial biogenesis, such as PGC-1α and TFAM, decreased in HG-treated podocytes. Moreover, PINK1/parkin-dependent mitophagy was inhibited in these cells. These results provide evidence that hyperglycemia impairs mitochondrial dynamics and turnover, which may underlie the remarkable deterioration of mitochondrial respiration parameters in glomeruli and podocytes.


Assuntos
Hiperglicemia/metabolismo , Mitocôndrias/metabolismo , Mitofagia/fisiologia , Podócitos/metabolismo , Animais , Diabetes Mellitus Experimental/metabolismo , Humanos , Rim/metabolismo , Masculino , Proteínas Quinases/metabolismo , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismo
5.
FASEB J ; 35(8): e21796, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34324238

RESUMO

Polycystin-1 (PC1) is a transmembrane protein found in different cell types, including cardiomyocytes. Alterations in PC1 expression have been linked to mitochondrial damage in renal tubule cells and in patients with autosomal dominant polycystic kidney disease. However, to date, the regulatory role of PC1 in cardiomyocyte mitochondria is not well understood. The analysis of mitochondrial morphology from cardiomyocytes of heterozygous PC1 mice (PDK1+/- ) using transmission electron microscopy showed that cardiomyocyte mitochondria were smaller with increased mitochondria density and circularity. These parameters were consistent with mitochondrial fission. We knocked-down PC1 in cultured rat cardiomyocytes and human-induced pluripotent stem cells (iPSC)-derived cardiomyocytes to evaluate mitochondrial function and morphology. The results showed that downregulation of PC1 expression results in reduced protein levels of sub-units of the OXPHOS complexes and less functional mitochondria (reduction of mitochondrial membrane potential, mitochondrial respiration, and ATP production). This mitochondrial dysfunction activates the elimination of defective mitochondria by mitophagy, assessed by an increase of autophagosome adapter protein LC3B and the recruitment of the Parkin protein to the mitochondria. siRNA-mediated PC1 knockdown leads to a loss of the connectivity of the mitochondrial network and a greater number of mitochondria per cell, but of smaller sizes, which characterizes mitochondrial fission. PC1 silencing also deregulates the AKT-FoxO1 signaling pathway, which is involved in the regulation of mitochondrial metabolism, mitochondrial morphology, and processes that are part of cell quality control, such as mitophagy. Together, these data provide new insights about the controls that PC1 exerts on mitochondrial morphology and function in cultured cardiomyocytes dependent on the AKT-FoxO1 signaling pathway.


Assuntos
Proteína Forkhead Box O1/metabolismo , Mitofagia/fisiologia , Miócitos Cardíacos/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Canais de Cátion TRPP/metabolismo , Animais , Animais Recém-Nascidos , Proteína Forkhead Box O1/genética , Regulação da Expressão Gênica/fisiologia , Inativação Gênica , Mitocôndrias/metabolismo , Mitofagia/genética , Proteínas Proto-Oncogênicas c-akt/genética , Ratos , Ratos Sprague-Dawley , Canais de Cátion TRPP/genética
6.
Biomolecules ; 11(7)2021 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-34209255

RESUMO

Various environmental stimuli, including oxidative stress, could lead to granulosa cell (GC) death through mitophagy. Recently, it was reported that melatonin (MEL) has a significant effect on GC survival during oxidative damage. Here, we found that MEL inhibited oxidative stress-induced mitophagy to promote GC survival. The loss of cell viability upon H2O2 exposure was significantly restored after MEL treatment. Concomitantly, MEL inhibited the activation of mitophagy during oxidative stress. Notably, blocking mitophagy repressed GC death caused by oxidative stress. However, MEL cannot further restore viability of cells treated with mitophagy inhibitor. Moreover, PTEN-induced putative kinase 1 (PINK1), a mitochondrial serine/threonine-protein kinase, was inhibited by MEL during oxidative stress. As a result, the E3 ligase Parkin failed to translocate to mitochondria, leading to impaired mitochondria clearance. Using RNAi to knock down PINK1 expression, we further verified the role of the MEL-PINK1-Parkin (MPP) pathway in maintaining GC survival by suppressing mitophagy. Our findings not only clarify the protective mechanisms of MEL against oxidative damage in GCs, but also extend the understanding about how circadian rhythms might influence follicles development in the ovary. These findings reveal a new mechanism of melatonin in defense against oxidative damage to GCs by repressing mitophagy, which may be a potential therapeutic target for anovulatory disorders.


Assuntos
Células da Granulosa/metabolismo , Melatonina/farmacologia , Mitofagia/fisiologia , Animais , Sobrevivência Celular/efeitos dos fármacos , Feminino , Células da Granulosa/fisiologia , Peróxido de Hidrogênio/farmacologia , Masculino , Melatonina/metabolismo , Camundongos , Camundongos Endogâmicos ICR , Mitocôndrias/metabolismo , Mitofagia/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/fisiologia , Substâncias Protetoras/farmacologia , Proteínas Quinases/metabolismo , Ubiquitina-Proteína Ligases/metabolismo
8.
Cell Mol Life Sci ; 78(16): 5925-5951, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34228161

RESUMO

Mitochondrial fidelity is a key determinant of longevity and was found to be perturbed in a multitude of disease contexts ranging from neurodegeneration to heart failure. Tight homeostatic control of the mitochondrial proteome is a crucial aspect of mitochondrial function, which is severely complicated by the evolutionary origin and resulting peculiarities of the organelle. This is, on one hand, reflected by a range of basal quality control factors such as mitochondria-resident chaperones and proteases, that assist in import and folding of precursors as well as removal of aggregated proteins. On the other hand, stress causes the activation of several additional mechanisms that counteract any damage that may threaten mitochondrial function. Countermeasures depend on the location and intensity of the stress and on a range of factors that are equipped to sense and signal the nature of the encountered perturbation. Defective mitochondrial import activates mechanisms that combat the accumulation of precursors in the cytosol and the import pore. To resolve proteotoxic stress in the organelle interior, mitochondria depend on nuclear transcriptional programs, such as the mitochondrial unfolded protein response and the integrated stress response. If organelle damage is too severe, mitochondria signal for their own destruction in a process termed mitophagy, thereby preventing further harm to the mitochondrial network and allowing the cell to salvage their biological building blocks. Here, we provide an overview of how different types and intensities of stress activate distinct pathways aimed at preserving mitochondrial fidelity.


Assuntos
Mitocôndrias/fisiologia , Transdução de Sinais/fisiologia , Animais , Homeostase/fisiologia , Humanos , Mitocôndrias/metabolismo , Mitofagia/fisiologia , Proteoma/metabolismo , Resposta a Proteínas não Dobradas/fisiologia
9.
Metabolism ; 122: 154840, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34331963

RESUMO

OBJECTIVE: Ferroptosis is indicated in cardiovascular diseases. Given the prominent role of mitophagy in the governance of ferroptosis and our recent finding for FUN14 domain containing 1 (FUNDC1) in obesity anomalies, this study evaluated the impact of FUNDC1 deficiency in high fat diet (HFD)-induced cardiac anomalies. METHODS AND MATERIALS: WT and FUNDC1-/- mice were fed HFD (45% calorie from fat) or low fat diet (LFD, 10% calorie from fat) for 10 weeks in the presence of the ferroptosis inhibitor liproxstatin-1 (LIP-1, 10 mg/kg, i.p.). RESULTS: RNAseq analysis for differentially expressed genes (DEGs) reported gene ontology term related to ferroptosis and mitophagy in obese rat hearts, which was validated in obese rodent and human hearts. Although 10-week HFD intake did not alter global metabolism, cardiac geometry and function, ablation of FUNDC1 unmasked metabolic derangement, pronounced cardiac remodeling, contractile, intracellular Ca2+ and mitochondrial anomalies upon HFD challenge, the effects of which with exception of global metabolism were attenuated or mitigated by LIP-1. FUNDC1 ablation unmasked HFD-evoked rises in fatty acid synthase ACSL4, necroptosis, inflammation, ferroptosis, mitochondrial O2- production, and mitochondrial injury as well as dampened autophagy and DNA repair enzyme 8-oxoG DNA glycosylase 1 (OGG1) but not apoptosis, the effect of which except ACSL4 and its regulator SP1 was reversed by LIP-1. In vitro data noted that arachidonic acid, an ACSL4 substrate, provoked cytochrome C release, cardiomyocyte defect, and lipid peroxidation under FUNDC1 deficiency, the effects were interrupted by inhibitors of SP1, ACSL4 and ferroptosis. CONCLUSIONS: These data suggest that FUNDC1 deficiency sensitized cardiac remodeling and dysfunction with short-term HFD exposure, likely through ACSL4-mediated regulation of ferroptosis.


Assuntos
Coenzima A Ligases/metabolismo , Ferroptose/fisiologia , Proteínas de Membrana/metabolismo , Proteínas Mitocondriais/metabolismo , Remodelação Ventricular/fisiologia , Animais , Apoptose/fisiologia , Autofagia/fisiologia , Cálcio/metabolismo , Dieta Hiperlipídica/efeitos adversos , Masculino , Camundongos , Camundongos Knockout , Mitocôndrias/metabolismo , Mitofagia/fisiologia , Miócitos Cardíacos/metabolismo , Obesidade/metabolismo
10.
Sci Rep ; 11(1): 15510, 2021 07 30.
Artigo em Inglês | MEDLINE | ID: mdl-34330933

RESUMO

Ischemia is a major cause of kidney damage. Proximal tubular epithelial cells (PTECs) are highly susceptible to ischemic insults that frequently cause acute kidney injury (AKI), a potentially life-threatening condition with high mortality. Accumulating evidence has identified altered mitochondrial function as a central pathologic feature of AKI. The mitochondrial NAD+-dependent enzyme sirtuin 5 (SIRT5) is a key regulator of mitochondrial form and function, but its role in ischemic renal injury (IRI) is unknown. SIRT5 expression was increased in murine PTECs after IRI in vivo and in human PTECs (hPTECs) exposed to an oxygen/nutrient deprivation (OND) model of IRI in vitro. SIRT5-depletion impaired ATP production, reduced mitochondrial membrane potential, and provoked mitochondrial fragmentation in hPTECs. Moreover, SIRT5 RNAi exacerbated OND-induced mitochondrial bioenergetic dysfunction and swelling, and increased degradation by mitophagy. These findings suggest SIRT5 is required for normal mitochondrial function in hPTECs and indicate a potentially important role for the enzyme in the regulation of mitochondrial biology in ischemia.


Assuntos
Injúria Renal Aguda/metabolismo , Mitocôndrias/metabolismo , Sirtuínas/metabolismo , Injúria Renal Aguda/genética , Animais , Western Blotting , Linhagem Celular , Citrato (si)-Sintase/genética , Citrato (si)-Sintase/metabolismo , Imunofluorescência , Humanos , Imuno-Histoquímica , Masculino , Potencial da Membrana Mitocondrial/genética , Potencial da Membrana Mitocondrial/fisiologia , Camundongos , Mitocôndrias/genética , Mitofagia/genética , Mitofagia/fisiologia , Sirtuínas/genética
11.
FASEB J ; 35(7): e21709, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34143518

RESUMO

Tissues undergo a process of degeneration as the body ages. Mesenchymal stem cells (MSCs) have been found to have major potential in delaying the aging process in tissues and organs. However, the mechanism underlying the anti-aging effects of MSC is not clear which limits clinical applications. In this study, we used adipose-derived mesenchymal stem cells (ADSCs) to perform anti-aging treatments on senescent cells and progeroid animal models. Following intervention with ADSCs, replicative senescence was delayed and metabolic homeostasis was transformed from catabolism to anabolism. Metabolomic tests were used to analyze different metabolites. We found that ADSCs acted to accelerate mitophagy which eliminated intracellular ROS and improved the quality of mitochondria. These processes acted to regulate the cellular metabolic homeostasis and ultimately delayed the process of aging. Allogeneic stem cell therapy in a Progeria animal model (DNA polymerase gamma (POLG) knockin, mitochondrial dysfunction) also showed that ADSC therapy can improve alopecia and kyphosis by promoting mitophagy. Our research confirms for the first time that allogeneic stem cell therapy can improve aging-related symbols and phenotypes through mitochondrial quality control. These results are highly significant for the future applications of stem cells in aging-related diseases.


Assuntos
Adipócitos/metabolismo , Tecido Adiposo/metabolismo , Envelhecimento/metabolismo , Homeostase/fisiologia , Mitofagia/fisiologia , Células-Tronco/metabolismo , Animais , Diferenciação Celular/fisiologia , Proliferação de Células/fisiologia , Células Cultivadas , Senescência Celular/fisiologia , Modelos Animais de Doenças , Feminino , Masculino , Células-Tronco Mesenquimais/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Transplante de Células-Tronco/métodos
12.
Biomed Res Int ; 2021: 5590973, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34095303

RESUMO

Cisplatin is an efficacious anticancer agent, but its use is limited by ototoxicity and resultant irreversible sensorineural hearing loss. Cisplatin ototoxicity is associated with cochlear cell oxidative stress and mitochondrial damage. However, mitophagy is vital for maintaining mitochondrial quality and cellular metabolism. Accordingly, we investigated the role of mitophagy in regulating cisplatin-induced ototoxicity using the auditory cell line HEI-OC1. In this study, HEI-OC1 cells were treated with either cisplatin alone (10 µM, 0, 8, 16, and 24 h); cisplatin (10 µM, 24 h) post transfection with small-interfering (si)RNAs targeting mitophagy-associated mRNAs; cisplatin (10 µM, 24 h) succeeding pretreatment with the mitophagy suppressor, 3-methyladenine (3-MA; 5 or 10 mM, 6 h); or cisplatin (30 µM, 24 h) following pretreatment with the mitophagy promoter, carbonyl cyanide m-chlorophenylhydrazone (CCCP; 1 or 2 µM, 2 h). The viability of cells, expression of mitophagy marker, and mitochondrial functions were then assessed in these cells. Cell viability was determined by a water-soluble tetrazolium assay; expression of mitophagy-associated proteins PINK1, Parkin, BNIP3, FUNDC1, p62, and LC3B was analyzed by Western blotting, mitochondrial membrane potential by flow cytometry, intracellular ATP by spectrophotometry, and mitochondrial degradation by dual staining for mitochondria and autophagosomes or lysosomes. Our results showed that cisplatin gradually reduced the viable cell number over time, induced mitochondrial depolarization, decreased intracellular ATP concentration, and enhanced the expression of PINK1, Parkin, BNIP3, p62, and LC3B. In addition, Parkin and BNIP3 knockdown accelerated cisplatin-induced loss of cell viability, mitochondrial membrane potential, mitophagosome/lysosome formation, and reduction in intracellular ATP production. Pretreatment with 3-MA aggravated the cisplatin-induced cytotoxicity, while that with CCCP reversed this effect. Overall, our findings indicate that mitophagy protects HEI-OC1 cells against cisplatin-induced cell death. Consequently, we strongly believe that targeted promotion of mitophagy may confer protection against cisplatin-induced ototoxicity.


Assuntos
Cisplatino/efeitos adversos , Mitofagia/fisiologia , Ototoxicidade/fisiopatologia , Apoptose/efeitos dos fármacos , Autofagossomos/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cisplatino/farmacologia , Perda Auditiva Neurossensorial/prevenção & controle , Humanos , Potencial da Membrana Mitocondrial/fisiologia , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Mitocôndrias/metabolismo , Ototoxicidade/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo
13.
PLoS Comput Biol ; 17(6): e1009073, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-34106921

RESUMO

Neurons rely on localized mitochondria to fulfill spatially heterogeneous metabolic demands. Mitochondrial aging occurs on timescales shorter than the neuronal lifespan, necessitating transport of fresh material from the soma. Maintaining an optimal distribution of healthy mitochondria requires an interplay between a stationary pool localized to sites of high metabolic demand and a motile pool capable of delivering new material. Interchange between these pools can occur via transient fusion / fission events or by halting and restarting entire mitochondria. Our quantitative model of neuronal mitostasis identifies key parameters that govern steady-state mitochondrial health at discrete locations. Very infrequent exchange between stationary and motile pools optimizes this system. Exchange via transient fusion allows for robust maintenance, which can be further improved by selective recycling through mitophagy. These results provide a framework for quantifying how perturbations in organelle transport and interactions affect mitochondrial homeostasis in neurons, a key aspect underlying many neurodegenerative disorders.


Assuntos
Dinâmica Mitocondrial/fisiologia , Modelos Neurológicos , Neurônios/fisiologia , Animais , Transporte Axonal/fisiologia , Axônios/fisiologia , Biologia Computacional , Dendritos/fisiologia , Homeostase , Humanos , Mitofagia/fisiologia , Proteínas do Tecido Nervoso/biossíntese , Doenças Neurodegenerativas/etiologia , Doenças Neurodegenerativas/fisiopatologia , Processos Estocásticos
14.
EMBO J ; 40(14): e100715, 2021 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-34152608

RESUMO

Clearance of mitochondria following damage is critical for neuronal homeostasis. Here, we investigate the role of Miro proteins in mitochondrial turnover by the PINK1/Parkin mitochondrial quality control system in vitro and in vivo. We find that upon mitochondrial damage, Miro is promiscuously ubiquitinated on multiple lysine residues. Genetic deletion of Miro or block of Miro1 ubiquitination and subsequent degradation lead to delayed translocation of the E3 ubiquitin ligase Parkin onto damaged mitochondria and reduced mitochondrial clearance in both fibroblasts and cultured neurons. Disrupted mitophagy in vivo, upon post-natal knockout of Miro1 in hippocampus and cortex, leads to a dramatic increase in mitofusin levels, the appearance of enlarged and hyperfused mitochondria and hyperactivation of the integrated stress response (ISR). Altogether, our results provide new insights into the central role of Miro1 in the regulation of mitochondrial homeostasis and further implicate Miro1 dysfunction in the pathogenesis of human neurodegenerative disease.


Assuntos
Mitocôndrias/metabolismo , Mitofagia/fisiologia , Neurônios/metabolismo , Proteínas rho de Ligação ao GTP/metabolismo , Animais , Linhagem Celular Tumoral , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Mitocondriais/metabolismo , Doenças Neurodegenerativas/metabolismo , Ubiquitina/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação/fisiologia
15.
Methods Mol Biol ; 2322: 81-92, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34043195

RESUMO

The physiological importance of mitochondrial quality control has been uncovered by the finding that genes for early onset Parkinson's disease (PD), PINK1 and Parkin, regulate mitochondrial autophagy, called mitophagy, and motility. Dopaminergic neurons derived from human-induced pluripotent stem (iPS) cells are a useful tool for analyzing the pathogenesis caused by defects in mitochondrial quality control and for screening candidate drugs for PD. Moreover, dopaminergic neurons could provide new findings not obtained in other cells. In this chapter, we will describe our method for monitoring PINK1-Parkin signaling using iPS cell-derived dopaminergic neurons.


Assuntos
Neurônios Dopaminérgicos/metabolismo , Neurônios Dopaminérgicos/fisiologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Pluripotentes Induzidas/fisiologia , Transdução de Sinais/fisiologia , Ubiquitina-Proteína Ligases/metabolismo , Autofagia/fisiologia , Linhagem Celular , Células HEK293 , Humanos , Mitocôndrias/metabolismo , Mitocôndrias/fisiologia , Mitofagia/fisiologia , Doença de Parkinson/metabolismo , Proteínas Quinases
16.
Biochem Biophys Res Commun ; 561: 143-150, 2021 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-34023779

RESUMO

Retinal pigment epithelium (RPE) cell damage, including mitophagy-associated cell apoptosis, accelerates the pathogenesis of diabetic retinopathy (DR), a common complication of diabetes that causes blindness. Müller cells interact with RPE cells via pro-inflammatory cytokines, such as tumor necrosis factor α (TNF-α). Herein, we investigated the role of the RPE cell epidermal growth factor receptor (EGFR)/p38 mitogen-activated protein kinase (p38)/nuclear factor kappa B (NF-κB) pathway in Müller cell-derived TNF-α-induced mitophagy-associated apoptosis during DR. Our results showed that TNF-α released from Müller cells activated the EGFR/p38/NF-κB/p62 pathway to increase mitophagy and apoptosis in RPE cells under high glucose (HG) conditions. Additionally, blockade of the TNF-α/EGFR axis alleviates blood-retina barrier breakdown in diabetic mice. Our data further illustrate the effects of the Müller cell inflammatory response on RPE cell survival, implying potential molecular targets for DR treatment.


Assuntos
Barreira Hematorretiniana/efeitos dos fármacos , Diabetes Mellitus Experimental/patologia , Retinopatia Diabética/patologia , Células Ependimogliais/patologia , Epitélio Pigmentado da Retina/patologia , Fator de Necrose Tumoral alfa/metabolismo , Animais , Apoptose , Barreira Hematorretiniana/metabolismo , Barreira Hematorretiniana/patologia , Células Cultivadas , Técnicas de Cocultura , Diabetes Mellitus Experimental/metabolismo , Retinopatia Diabética/metabolismo , Dieta Hiperlipídica , Modelos Animais de Doenças , Células Ependimogliais/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mitofagia/fisiologia , Epitélio Pigmentado da Retina/metabolismo
17.
Diabetes ; 70(6): 1229-1241, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-34016598

RESUMO

Insulin-producing pancreatic ß-cells are central to glucose homeostasis, and their failure is a principal driver of diabetes development. To preserve optimal health ß-cells must withstand both intrinsic and extrinsic stressors, ranging from inflammation to increased peripheral insulin demand, in addition to maintaining insulin biosynthesis and secretory machinery. Autophagy is increasingly being appreciated as a critical ß-cell quality control system vital for glycemic control. Here we focus on the underappreciated, yet crucial, roles for selective and organelle-specific forms of autophagy as mediators of ß-cell health. We examine the unique molecular players underlying each distinct form of autophagy in ß-cells, including selective autophagy of mitochondria, insulin granules, lipid, intracellular amyloid aggregates, endoplasmic reticulum, and peroxisomes. We also describe how defects in selective autophagy pathways contribute to the development of diabetes. As all forms of autophagy are not the same, a refined view of ß-cell selective autophagy may inform new approaches to defend against the various insults leading to ß-cell failure in diabetes.


Assuntos
Autofagia/fisiologia , Células Secretoras de Insulina/fisiologia , Animais , Diabetes Mellitus/etiologia , Diabetes Mellitus/patologia , Diabetes Mellitus/fisiopatologia , Humanos , Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/patologia , Ilhotas Pancreáticas/fisiopatologia , Mitofagia/fisiologia , Agregados Proteicos/fisiologia , Fatores de Transcrição/fisiologia , Ubiquitina-Proteína Ligases/fisiologia
18.
Elife ; 102021 05 04.
Artigo em Inglês | MEDLINE | ID: mdl-33942716

RESUMO

Erythropoietin (EPO) drives erythropoiesis and is secreted mainly by the kidney upon hypoxic or anemic stress. The paucity of EPO production in renal EPO-producing cells (REPs) causes renal anemia, one of the most common complications of chronic nephropathies. Although mitochondrial dysfunction is commonly observed in several renal and hematopoietic disorders, the mechanism by which mitochondrial quality control impacts renal anemia remains elusive. In this study, we showed that FUNDC1, a mitophagy receptor, plays a critical role in EPO-driven erythropoiesis induced by stresses. Mechanistically, EPO production is impaired in REPs in Fundc1-/- mice upon stresses, and the impairment is caused by the accumulation of damaged mitochondria, which consequently leads to the elevation of the reactive oxygen species (ROS) level and triggers inflammatory responses by up-regulating proinflammatory cytokines. These inflammatory factors promote the myofibroblastic transformation of REPs, resulting in the reduction of EPO production. We therefore provide a link between aberrant mitophagy and deficient EPO generation in renal anemia. Our results also suggest that the mitochondrial quality control safeguards REPs under stresses, which may serve as a potential therapeutic strategy for the treatment of renal anemia.


Assuntos
Anemia/prevenção & controle , Eritropoetina/metabolismo , Regulação da Expressão Gênica , Nefropatias/prevenção & controle , Proteínas de Membrana/genética , Proteínas Mitocondriais/genética , Mitofagia/genética , Animais , Eritropoese/genética , Eritropoese/fisiologia , Eritropoetina/análise , Eritropoetina/genética , Nefropatias/classificação , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Mitocondriais/metabolismo , Mitofagia/fisiologia , Espécies Reativas de Oxigênio
19.
Mol Med Rep ; 24(1)2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33955500

RESUMO

The acupuncture penetrating line of Baihui (GV20) to Qubin (GB7) spans the parietal, frontal and temporal lobes. The present study aimed to elucidate the mechanism by which electroacupuncture (EA) at GV20­GB7 regulates mitophagy in intracerebral hemorrhage (ICH) and whether it serves a neuroprotective role. A whole blood­induced ICH model was used. Mitophagy­regulating proteins, including BCL/adenovirus E1B 19 kDa­interacting protein 3 (BNIP3), PTEN­induced putative kinase 1 (PINK1), Parkin and apoptosis­associated proteins were detected by western blotting; autophagy following ICH was evaluated by immunofluorescent techniques; morphological characteristics of mitophagy were observed using transmission electron microscopy; and TUNEL assay was performed to determine the number of apoptotic cells. Immunohistochemistry was used to detect p53 expression. The protective role of EA (GV20­GB7) via enhanced mitophagy and suppressed apoptosis in ICH was further confirmed by decreased modified neurological severity score. The results showed that EA (GV20­GB7) treatment upregulated mitochondrial autophagy following ICH and inhibited apoptotic cell death. The mechanism underlying EA (GV20­GB7) treatment may involve inhibition of p53, an overlapping protein of autophagy and apoptosis. EA (GV20­GB7) treatment decreased neurobehavioral deficits following ICH but pretreatment with 3­methyladenine counteracted the beneficial effects of EA (GV20­GB7) treatment. In conclusion, EA (GV20­GB7) improved recovery from ICH by regulating the balance between mitophagy and apoptosis.


Assuntos
Hemorragia Cerebral/terapia , Eletroacupuntura/métodos , Pontos de Acupuntura , Animais , Apoptose/fisiologia , Autofagia/fisiologia , Caspase 3/metabolismo , Hemorragia Cerebral/patologia , Modelos Animais de Doenças , Masculino , Proteínas de Membrana/metabolismo , Mitocôndrias/metabolismo , Proteínas Mitocondriais/metabolismo , Mitofagia/fisiologia , Proteínas Quinases/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Ratos Sprague-Dawley , Índice de Gravidade de Doença , Proteína Supressora de Tumor p53/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Proteína X Associada a bcl-2/metabolismo
20.
Biochem Pharmacol ; 190: 114588, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-33957094

RESUMO

Bladder cancer is one of the most common malignancy in the urinary tract with high recurrence and drug resistance in clinics. Alternative treatments from existing drugs might be a promising strategy. Nitazoxanide (NTZ), an FDA-approved antiprotozoal drug, has got increasingly noticed because of its favorable safety profile and antitumor potential, yet the effects in bladder cancer and underlying mechanisms remain poorly understood. Herein, we find that NTZ induces mitochondrial damage and mitophagy initiation through PINK1-generated phospho-ubiquitin(pS65-Ub) and autophagy receptor-mediated pathway even in the absence of Atg5/Beclin1. Meanwhile, NTZ inhibits lysosomal degradation activity, leading to mitophagy flux impairment at late stage. Mitochondrial reactive oxygen species (ROS) production is critical in this process, as eliminating ROS with N-acetylcysteine (NAC) efficiently inhibits PINK1 signaling-mediated mitophagy initiation and alleviates lysosomal dysfunction. Co-treatment with NTZ and autophagy inhibitor Chloroquine (CQ) to aggravate mitophagy flux impairment promotes NTZ-induced apoptosis, while alleviation of mitophagy flux impairment with ROS scavenger reduces cell death. Moreover, we also discover a similar signaling response in the 3D bladder tumor spheroid after NTZ exposure. In vivo study reveals a significant inhibition of orthotopic bladder tumors with no obvious systemic toxicity. Together, our results uncover the anti-tumor activities of NTZ with the involvement of ROS-mediated mitophagy modulation at different stages and demonstrate it as a potential drug candidate for fighting against bladder tumors.


Assuntos
Antineoplásicos/farmacologia , Lisossomos/metabolismo , Mitofagia/efeitos dos fármacos , Nitrocompostos/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Tiazóis/farmacologia , Neoplasias da Bexiga Urinária/metabolismo , Animais , Antineoplásicos/uso terapêutico , Antiparasitários/farmacologia , Antiparasitários/uso terapêutico , Relação Dose-Resposta a Droga , Feminino , Células HEK293 , Humanos , Lisossomos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Mitofagia/fisiologia , Nitrocompostos/uso terapêutico , Espécies Reativas de Oxigênio/antagonistas & inibidores , Tiazóis/uso terapêutico , Neoplasias da Bexiga Urinária/tratamento farmacológico
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