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1.
Proc Natl Acad Sci U S A ; 120(5): e2215575120, 2023 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-36696445

RESUMO

Chloroplast division involves the coordination of protein complexes from the stroma to the cytosol. The Min system of chloroplasts includes multiple stromal proteins that regulate the positioning of the division site. The outer envelope protein PLASTID DIVISION1 (PDV1) was previously reported to recruit the cytosolic chloroplast division protein ACCUMULATION AND REPLICATION OF CHLOROPLAST5 (ARC5). However, we show here that PDV1 is also important for the stability of the inner envelope chloroplast division protein PARALOG OF ARC6 (PARC6), a component of the Min system. We solved the structure of both the C-terminal domain of PARC6 and its complex with the C terminus of PDV1. The formation of an intramolecular disulfide bond within PARC6 under oxidized conditions prevents its interaction with PDV1. Interestingly, this disulfide bond can be reduced by light in planta, thus promoting PDV1-PARC6 interaction and chloroplast division. Interaction with PDV1 can induce the dimerization of PARC6, which is important for chloroplast division. Magnesium ions, whose concentration in chloroplasts increases upon light exposure, also promote the PARC6 dimerization. This study highlights the multilayer regulation of the PDV1-PARC6 interaction as well as chloroplast division.


Assuntos
Proteínas de Arabidopsis , Arabidopsis , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Plastídeos/metabolismo , Cloroplastos/metabolismo , Dissulfetos/metabolismo , Dinaminas/metabolismo
2.
Int J Mol Sci ; 24(2)2023 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-36674740

RESUMO

Ovarian cancer (OC) represents the main cause of death from gynecological malignancies in western countries. Altered cellular and mitochondrial metabolism are considered hallmarks in cancer disease. Several mitochondrial aspects have been found altered in OC, such as the oxidative phosphorylation system, oxidative stress and mitochondrial dynamics. Mitochondrial dynamics includes cristae remodeling, fusion, and fission processes forming a dynamic mitochondrial network. Alteration of mitochondrial dynamics is associated with metabolic change in tumour development and, in particular, the mitochondrial shaping proteins appear also to be responsible for the chemosensitivity and/or chemoresistance in OC. In this review a focus on the mitochondrial dynamics in OC cells is presented.


Assuntos
Dinâmica Mitocondrial , Neoplasias Ovarianas , Humanos , Feminino , Mitocôndrias/metabolismo , Membranas Mitocondriais/metabolismo , Neoplasias Ovarianas/metabolismo , Proteínas Mitocondriais/metabolismo , Dinaminas/metabolismo
3.
J Mol Cell Cardiol ; 174: 115-132, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36509022

RESUMO

RATIONAL: Excessive mitochondrial fission is considered key process involved in myocardial ischemia/reperfusion (I/R) injury. However, the upstream mechanism remains largely unclear. Decreased level of Kruppel Like Factor 4 (KLF4) has been implicated in the pathogenesis of mitochondrial dysfunction and heart's adaption to stress. However, the role of Klf4 in I/R process is not fully elucidated. This study aims to investigate how Klf4 regulates mitochondrial dynamics and further clarify its underlying mechanism during cardiac I/R injury. METHODS: Loss-of-function and gain-of-function strategies were applied to investigate the role of Klf4 in cardiac I/R injury via genetic ablation or intra-myocardial adenovirus injection. Mitochondrial dynamics was analyzed by confocal microscopy in vitro and transmission electron microscopy in vivo. Chromatin immunoprecipitation and luciferase reporter assay were performed to explore the underlying mechanisms. RESULTS: KLF4 was downregulated in I/R heart. Cardiac-specific Klf4 knockout significantly exacerbated cardiac dysfunction in I/R mice. Mechanistically, Klf4 deficiency aggravated mitochondrial apoptosis, reduced ATP generation and boosted ROS overproduction via enhancing DRP1-dependent mitochondrial fission. ROCK1 was identified as a kinase regulating DRP1 activity at Ser616. Klf4 deficiency upregulated the expression of ROCK1 at transcriptional level, thus increasing S616-DRP1-mediated mitochondrial fission during I/R. Finally, reconstitution of Klf4 inhibited mitochondrial fission, restored mitochondrial function and alleviated I/R injury. CONCLUSION: Our study provides the first evidence that Klf4 deficiency exacerbates myocardial I/R injury through regulating ROCK1 expression at transcriptional level to induce DRP1-mediated mitochondrial fission. Targeting mitochondrial dynamics by restoring Klf4 might be potentially cardio-protective strategies attenuating I/R injury.


Assuntos
Traumatismo por Reperfusão Miocárdica , Animais , Camundongos , Apoptose/genética , Dinaminas/metabolismo , Coração , Mitocôndrias/metabolismo , Dinâmica Mitocondrial , Traumatismo por Reperfusão Miocárdica/metabolismo , Miocárdio/metabolismo
4.
Biochem Biophys Res Commun ; 643: 16-23, 2023 Feb 05.
Artigo em Inglês | MEDLINE | ID: mdl-36584588

RESUMO

Understanding GPER biology in breast cancer is rather limited in compassion to the classic estrogen receptors. Mitochondrial dynamics play a critical role in determining cell survival and death under various microenvironmental conditions. We present evidence that GPER-induce mitochondrial fission in breast cancer cells. GPER mediated mitochondrial fission through activating Drp1 by phosphorylating S616 residue and down-regulates fusion proteins Mfn1 and Mfn2 levels. GPER-induced Drp1 activation mediated by p44/42 MAPK and inhibition of this signalling axis completely reverse the mitochondrial fission induced by GPER. Further, mitochondrial fission is required for GPER-induced cell death in breast cancer cells. To conclude, GPER induces mitochondrial fission through p44/42 MAPK - Drp1 signalling, and mitochondrial fission is critical for GPER-induced cell death in breast cancer cells. GENERAL SIGNIFICANCE: First time we report GPER's role in mitochondrial dynamics in cancer cells. Mitochondrial dynamics play a critical role in cancer progression including tamoxifen resistance. Exploring a detailed mechanistic understanding of GPER signalling may help to design new therapy for advanced cancers.


Assuntos
Neoplasias da Mama , Dinaminas , Humanos , Feminino , Dinaminas/metabolismo , GTP Fosfo-Hidrolases/metabolismo , Neoplasias da Mama/metabolismo , Dinâmica Mitocondrial/fisiologia , Receptores de Estrogênio , Proteínas Mitocondriais/metabolismo
5.
Gene ; 855: 147130, 2023 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-36543307

RESUMO

Stroke had emerged as one of the leading causes of death and long-term disability across the globe. Emerging evidence suggests a significant increase in the incidence of stroke with age, which is further expected to increase dramatically owing to an ever-expanding elderly population. The current situation imposes a significant burden on the healthcare system and requires a deeper understanding of the underlying mechanisms and development of novel interventions. It is well established that mitochondrial dysfunction plays a pivotal role in the onset of stroke. Dynamin-related protein 1 (Drp1), is a key regulator of mitochondria fission, and plays a crucial role during the pathogenesis of stroke. Drp1 protein levels significantly increase after stroke potentially in a p38 mitogen-activated protein kinases (MAPK) dependent manner. Protein phosphatase 2A (PP2A) facilitate mitochondrial fission and cell death by dephosphorylating the mitochondrial fission enzyme Drp1 at the inhibitory phosphorylation site serine 637. Outer mitochondrial membrane A-Kinase Anchoring Proteins 1 (AKAP 1) and protein kinase A complex (PKA) complex inhibits Drp1-dependent mitochondrial fission by phosphorylating serine 637. Drp1 activation promotes the release of cytochrome C from mitochondria and therefore leads to apoptosis. In addition, Drp1 activation inhibits mitochondrial glutathione dependent free radical scavenging, which further enhances the ROS level and exacerbate mitochondrial dysfunction. Drp1 translocate p53 to mitochondrial membrane and leads to mitochondria-related necrosis. The current review article discusses the possible mechanistic pathways by which Drp1 can influence the pathogenesis of stroke. Besides, it will describe various inhibitors for Drp1 and their potential role as therapeutics for stroke in the future.


Assuntos
Dinaminas , Acidente Vascular Cerebral , Idoso , Humanos , Fosforilação , Dinaminas/metabolismo , Mitocôndrias/metabolismo , Acidente Vascular Cerebral/metabolismo , Apoptose , Serina/metabolismo , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo
6.
Pharmacol Res ; 187: 106608, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36566000

RESUMO

Mitochondrial metabolism plays a pivotal role in various cellular processes and fibrosis. However, the mechanism underlying mitochondrial metabolic function and liver fibrosis remains poorly understood. In this study, we determined whether mitochondrial metabolism mediates liver fibrosis using cells, animal models, and clinical samples to elucidate the potential effects and underlying mechanism of mitochondrial metabolism in liver fibrosis. We report that AlkB Homolog 5 (ALKBH5) decreases mitochondrial membrane potential (MMP) and oxygen consumption rate (OCR), suppresses mitochondrial fission and hepatic stellate cell (HSC) proliferation and migration and ameliorates liver fibrosis. Enhancement of mitochondrial fission, an essential event during HSC proliferation and migration, is dependent on decreased ALKBH5 expression. Furthermore, we reveal that low ALKBH5 expression is associated with elevated N6-methyladenosine (m6A) mRNA levels. Mechanistically, ALKBH5 mediates m6A demethylation in the 3'UTR of Drp1 mRNA and induces its translation in a YTH domain family proteins 1 (YTHDF1)-independent manner. Subsequently, in transforming growth factor-ß1 (TGF-ß1) induced HSC, Dynamin-related protein 1 (Drp1) mediates mitochondrial fission and increases cell proliferation and migration. Decreased Drp1 expression inhibits mitochondrial fission and suppresses HSC proliferation and migration. Notably, human fibrotic liver and heart tissue exhibited enhanced mitochondrial fission; increased YTHDF1, Drp1, alpha-smooth muscle actin (α-SMA) and collagen I expression; decreased ALKBH5 expression and increased liver fibrosis. Our results highlight a novel mechanism by which ALKBH5 suppresses mitochondrial fission and HSC proliferation and migration by reducing Drp1 methylation in an m6A-YTHDF1-dependent manner, which may indicate a demethylation-based approach for liver fibrosis diagnosis and therapy.


Assuntos
Cirrose Hepática , Dinâmica Mitocondrial , Animais , Humanos , Metilação , Cirrose Hepática/metabolismo , Dinaminas/metabolismo , RNA Mensageiro/metabolismo , Células Estreladas do Fígado , Homólogo AlkB 5 da RNA Desmetilase/genética , Homólogo AlkB 5 da RNA Desmetilase/metabolismo
7.
Nat Commun ; 13(1): 7732, 2022 12 13.
Artigo em Inglês | MEDLINE | ID: mdl-36513638

RESUMO

Chromosome segregation is initiated by cohesin degradation, which is driven by anaphase-promoting complex/cyclosome (APC/C). Chromosome cohesin is removed by activated separase, with the degradation of securin and cyclinB1. Dynamin-related protein 1 (DRP1), a component of the mitochondrial fission machinery, is related to cyclin dynamics in mitosis progression. Here, we show that DRP1 is recruited to the kinetochore by centromeric Centromere protein F (CENP-F) after nuclear envelope breakdown in mouse oocytes. Loss of DRP1 during prometaphase leads to premature cohesin degradation and chromosome segregation. Importantly, acute DRP1 depletion activates separase by initiating cyclinB1 and securin degradation during the metaphase-to-anaphase transition. Finally, we demonstrate that DRP1 is bound to APC2 to restrain the E3 ligase activity of APC/C. In conclusion, DRP1 is a CENP-F-dependent atypical spindle assembly checkpoint (SAC) protein that modulates metaphase-to-anaphase transition by controlling APC/C activity during meiosis I in oocytes.


Assuntos
Segregação de Cromossomos , Meiose , Animais , Camundongos , Ciclossomo-Complexo Promotor de Anáfase/metabolismo , Proteínas de Ciclo Celular/metabolismo , Dinaminas/metabolismo , Cinetocoros/metabolismo , Oócitos/metabolismo , Securina/genética , Securina/metabolismo , Separase/metabolismo
8.
Cell Rep ; 41(11): 111818, 2022 12 13.
Artigo em Inglês | MEDLINE | ID: mdl-36516772

RESUMO

Oncogenic KRas activates mitochondrial fission through Erk-mediated phosphorylation of the mitochondrial fission GTPase Drp1. Drp1 deletion inhibits tumorigenesis of KRas-driven pancreatic cancer, but the role of mitochondrial dynamics in other Ras-driven malignancies is poorly defined. Here we show that in vitro and in vivo growth of KRas-driven lung adenocarcinoma is unaffected by deletion of Drp1 but is inhibited by deletion of Opa1, the GTPase that regulates inner membrane fusion and proper cristae morphology. Mechanistically, Opa1 knockout disrupts cristae morphology and inhibits electron transport chain (ETC) assembly and activity, which inhibits tumor cell proliferation through loss of NAD+ regeneration. Simultaneous inactivation of Drp1 and Opa1 restores cristae morphology, ETC activity, and cell proliferation indicating that mitochondrial fission activity drives ETC dysfunction induced by Opa1 knockout. Our results support a model in which mitochondrial fission events disrupt cristae structure, and tumor cells with hyperactive fission activity require Opa1 activity to maintain ETC function.


Assuntos
Adenocarcinoma de Pulmão , NAD , Humanos , NAD/metabolismo , Mitocôndrias/metabolismo , GTP Fosfo-Hidrolases/genética , GTP Fosfo-Hidrolases/metabolismo , Membranas Mitocondriais/metabolismo , Dinâmica Mitocondrial , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/metabolismo , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Dinaminas/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/metabolismo
9.
Int J Mol Sci ; 23(24)2022 Dec 08.
Artigo em Inglês | MEDLINE | ID: mdl-36555170

RESUMO

Mitochondria can fuse or divide, a phenomenon known as mitochondrial dynamics, and their distribution within a cell changes according to the physiological status of the cell. However, the functions of mitochondrial dynamics during spermatogenesis in animals other than mammals and fruit flies are poorly understood. In this study, we analyzed mitochondrial distribution and morphology during spermiogenesis in Sipuncula (Phascolosoma esculenta) and investigated the expression dynamics of mitochondrial fusion-related protein MFN2 and fission-related protein DRP1 during spermiogenesis. The mitochondria, which were elliptic with abundant lamellar cristae, were mainly localized near the nucleus and distributed unilaterally in cells during most stages of spermiogenesis. Their major axis length, average diameter, cross-sectional area, and volume are significantly changed during spermiogenesis. mfn2 and drp1 mRNA and proteins were most highly expressed in coelomic fluid, a spermatid development site for male P. esculenta, and highly expressed in the breeding stage compared to in the non-breeding stage. MFN2 and DRP1 expression levels were higher in components with many spermatids than in spermatid-free components. Immunofluorescence revealed that MFN2 and DRP1 were consistently expressed and that MFN2 co-localizes with mitochondria during spermiogenesis. The results provide evidence for an important role of mitochondrial dynamics during spermiogenesis from morphology and molecular biology in P. esculenta, broadening insights into the role of mitochondrial dynamics in animal spermiogenesis.


Assuntos
GTP Fosfo-Hidrolases , Mitocôndrias , Animais , Masculino , GTP Fosfo-Hidrolases/genética , GTP Fosfo-Hidrolases/metabolismo , Mitocôndrias/genética , Mitocôndrias/metabolismo , Membranas Mitocondriais/metabolismo , Espermatogênese/genética , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Hidrolases/metabolismo , Dinâmica Mitocondrial , Dinaminas/genética , Dinaminas/metabolismo , Mamíferos/metabolismo
10.
Int J Mol Sci ; 23(23)2022 Nov 25.
Artigo em Inglês | MEDLINE | ID: mdl-36499050

RESUMO

Mitochondria have many forms and can change their shape through fusion and fission of the outer and inner membranes, called "mitochondrial dynamics". Mitochondrial outer membrane proteins, such as mitochondrial fission protein 1 (FIS1), mitochondrial fission factor (MFF), mitochondrial 98 dynamics proteins of 49 kDa (MiD49), and mitochondrial dynamics proteins of 51 kDa (MiD51), can aggregate at the outer mitochondrial membrane and thus attract Dynamin-related protein 1 (DRP1) from the cytoplasm to the outer mitochondrial membrane, where DRP1 can perform a scissor-like function to cut a complete mitochondrion into two separate mitochondria. Other organelles can promote mitochondrial fission alongside mitochondria. FIS1 plays an important role in mitochondrial-lysosomal contacts, differentiating itself from other mitochondrial-fission-associated proteins. The contact between the two can also induce asymmetric mitochondrial fission. The kidney is a mitochondria-rich organ, requiring large amounts of mitochondria to produce energy for blood circulation and waste elimination. Pathological increases in mitochondrial fission can lead to kidney damage that can be ameliorated by suppressing their excessive fission. This article reviews the current knowledge on the key role of mitochondrial-fission-associated proteins in the pathogenesis of kidney injury and the role of their various post-translational modifications in activation or degradation of fission-associated proteins and targeted drug therapy.


Assuntos
Nefropatias , Dinâmica Mitocondrial , Humanos , Proteínas Mitocondriais/metabolismo , Fatores de Alongamento de Peptídeos/metabolismo , Dinaminas/metabolismo , Lisossomos/metabolismo
11.
Nat Commun ; 13(1): 7641, 2022 12 10.
Artigo em Inglês | MEDLINE | ID: mdl-36496453

RESUMO

Eps15-homology domain containing proteins (EHDs) are eukaryotic, dynamin-related ATPases involved in cellular membrane trafficking. They oligomerize on membranes into filaments that induce membrane tubulation. While EHD crystal structures in open and closed conformations were previously reported, little structural information is available for the membrane-bound oligomeric form. Consequently, mechanistic insights into the membrane remodeling mechanism have remained sparse. Here, by using cryo-electron tomography and subtomogram averaging, we determined structures of nucleotide-bound EHD4 filaments on membrane tubes of various diameters at an average resolution of 7.6 Å. Assembly of EHD4 is mediated via interfaces in the G-domain and the helical domain. The oligomerized EHD4 structure resembles the closed conformation, where the tips of the helical domains protrude into the membrane. The variation in filament geometry and tube radius suggests a spontaneous filament curvature of approximately 1/70 nm-1. Combining the available structural and functional data, we suggest a model for EHD-mediated membrane remodeling.


Assuntos
Dinaminas , Tomografia com Microscopia Eletrônica , Dinaminas/metabolismo , Adenosina Trifosfatases/metabolismo , Membrana Celular/metabolismo , Membranas/metabolismo , Microscopia Crioeletrônica
12.
Nat Commun ; 13(1): 7234, 2022 Nov 24.
Artigo em Inglês | MEDLINE | ID: mdl-36433988

RESUMO

Caveolae are small coated plasma membrane invaginations with diverse functions. Caveolae undergo curvature changes. Yet, it is unclear which proteins regulate this process. To address this gap, we develop a correlative stimulated emission depletion (STED) fluorescence and platinum replica electron microscopy imaging (CLEM) method to image proteins at single caveolae. Caveolins and cavins are found at all caveolae, independent of curvature. EHD2 is detected at both low and highly curved caveolae. Pacsin2 associates with low curved caveolae and EHBP1 with mostly highly curved caveolae. Dynamin is absent from caveolae. Cells lacking dynamin show no substantial changes to caveolae, suggesting that dynamin is not directly involved in caveolae curvature. We propose a model where caveolins, cavins, and EHD2 assemble as a cohesive structural unit regulated by intermittent associations with pacsin2 and EHBP1. These coats can flatten and curve to enable lipid traffic, signaling, and changes to the surface area of the cell.


Assuntos
Cavéolas , Caveolinas , Cavéolas/metabolismo , Membrana Celular/metabolismo , Caveolinas/metabolismo , Endocitose , Dinaminas/metabolismo , Proteínas/metabolismo
13.
Zhong Nan Da Xue Xue Bao Yi Xue Ban ; 47(9): 1191-1199, 2022.
Artigo em Inglês, Chinês | MEDLINE | ID: mdl-36411702

RESUMO

OBJECTIVES: Methotrexate (MTX) is the most common therapeutic agent that may have the risk of drug-induced liver injury. Its pathogenic mechanism is related to oxidative stress caused by mitochondrial dysfunction. Superoxide dismutase (SOD), including manganese-containing SOD (Mn-SOD), can exert its effect of anti-oxidative stress by scavenging superoxide free radicals. Accordingly, this study is performed to explore the underlying molecular mechanism via observing whether Mn-SOD could affect the damage of MTX to hepatocytes. METHODS: Human hepatocyte cell line L-02 was cultured in vitro and divided into 4 groups, including a blank group with the addition of the same volume of serum-free medium, a MTX group (40 µg/well MTX drug-treatment), a MTX+NC group (40 µg/well MTX drug-treatment+blank plasmid), and a MTX+SOD group (40 µg/well MTX drug-treatment+Mn-SOD plasmid). The levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), and microRNA-122 (miR-122) in the supernatant of cell culture were respectively detected by automatic biochemical analytical instrument and real-time RT-PCR to evaluate the degree of hepatocyte damage in each group. MitoSOX fluorescent probe was used to label intracellular superoxide in each group, and cell apoptosis was detected by flow cytometry. Meanwhile, the contents of glycogen synthase kinase-3 beta (GSK-3ß), hemeoxygenase-1 (HO-1), mitochondrial fission-mediated protein of dynamin-related protein 1 (Drp1), and Mn-SOD were detected by Western blotting. RESULTS: Compared with the blank group, the levels of ALT, AST, and miR-122 in the supernatant of hepatocyte culture of the MTX group and MTX+NC group were significantly elevated (all P<0.05), and that in the MTX+SOD group were significantly decreased (P<0.05) and equivalent to that in the blank group. MitoSOX staining revealed that the MTX group and MTX+NC had the most abundant superoxide; and the amount was significantly reduced in the MTX+SOD group, without a significant difference when compared with the blank group. Furthermore, the results of flow cytometry indicated that compared with the blank group, the MTX group and MTX+NC group showed significantly increased cell apoptosis (P<0.05); while there was obviously reduced cell apoptosis in the MTX+SOD group than that in the MTX group and MTX+NC group (P<0.05). According to the results of Western blotting, the blank group and MTX+SOD group had higher expressions of Mn-SOD, p-GSK-3ß, and HO-1; while the MTX group and MTX+NC group exhibited remarkably lower levels of Mn-SOD, p-GSK-3ß, and HO-1 than those in the blank group (P<0.05). Besides, a completely opposite trend was found in the expression of Drp1, which was highly expressed in the MTX group and MTX+NC group, but lowly expressed in the blank group and the MTX+SOD group. CONCLUSIONS: MTX may induce hepatocyte damage, and one of the mechanisms may be due to the decrease of intracellular Mn-SOD level, which can cause the accumulation of superoxide, affect the levels of HO-1 and Drp1 through GSK-3ß leading to mitochondrial damage and cell apoptosis. High expression of Mn-SOD intracellularly through exogenous introduction can scavenge drug-produced superoxide, affect HO-1 and Drp1 levels through GSK-3ß, activate mitochondria, protect cells against damage from oxidative stress, and inhibit hepatocyte apoptosis eventually. So exogenous introduction of SOD may be a potential therapeutic approach to block or reverse MTX-related hepatocyte injury.


Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , MicroRNAs , Humanos , Dinaminas/metabolismo , Dinaminas/farmacologia , Glicogênio Sintase Quinase 3 beta/genética , Glicogênio Sintase Quinase 3 beta/metabolismo , Glicogênio Sintase Quinase 3 beta/farmacologia , Metotrexato/efeitos adversos , MicroRNAs/metabolismo , Estresse Oxidativo , Superóxido Dismutase/metabolismo , Superóxidos/farmacologia
14.
Oxid Med Cell Longev ; 2022: 5064494, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36338340

RESUMO

Excessive generation of reactive oxygen species (ROS) has great impacts on MSU crystal-induced inflammation. Drp1-dependent mitochondrial fission is closely associated with mitochondrial ROS levels. However, whether Drp1 signaling contributes to MSU crystal-induced inflammation remains unclear. Mice bone marrow-derived macrophages (BMDMs) were primed with LPS and then stimulated with MSU suspensions for 12 h. The protein levels associated with mitochondrial dynamics, oxidative stress, and mitophagy were detected by Western blot. BMDMs were loaded with MitoTracker Green probe to detect mitochondrial morphology. To measure mitochondrial reactive oxygen species (ROS) and total ROS levels, cells were loaded, respectively, with MitoSOX and DHE probes. The effects of Mito-TEMPO, an antioxidant that targets the mitochondria or DRP1 inhibitor (Mdivi-1) on MSU crystal-induced peritonitis and arthritis mouse models, were evaluated. Our study revealed that MSU crystal stimulation resulted in elevation of mitochondrial fragmentation of BMDMs. Treatment with Mito-TEMPO or Drp1 knockdown significantly ameliorated the mitochondrial damage induced by MSU crystals. BMDMs exposure to MSU crystals increased the expression of auto/mitophagy marker proteins and promoted the fusion of mitophagosomes with lysosomes, leading to accumulation of mitolysosomes. Drp1 knockdown alleviated defective mitophagy and activation of the NLRP3 inflammasome in MSU crystal-treated BMDMs. This study indicates that there is crosstalk between mitochondrial ROS and Drp1 signaling in MSU crystal-induced inflammation. Drp1 signaling is involved in MSU crystal-induced mitochondrial damage, impaired mitophagy and NLRP3 inflammasome activation.


Assuntos
Inflamassomos , Mitofagia , Camundongos , Animais , Inflamassomos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Dinaminas/metabolismo , Dinâmica Mitocondrial , Inflamação
15.
Int J Mol Sci ; 23(21)2022 Nov 07.
Artigo em Inglês | MEDLINE | ID: mdl-36362420

RESUMO

Slow and progressive loss of retinal ganglion cells (RGCs) is the main characteristic of glaucoma, the second leading cause of blindness worldwide. Previous studies have shown that impaired mitochondrial dynamics could facilitate retinal neurodegeneration. Mitochondrial dynamics are regulated directly (fission) or more indirectly (fusion) by dynamin-like protein 1 (DNML1). Therefore, DNM1L might be a promising target for an antibody-based approach to treat glaucoma. The consequences of targeting endogenous DNM1L by antibodies in a glaucoma animal model have not been investigated yet. Here, we show that the intravitreal application of an anti-DNM1L antibody showed protective effects regarding the survival of RGCs and their axons in the retinal nerve fiber layer (RNFL). Antibody treatment also improved retinal functionality, as observed by electroretinography (Ganzfeld ERG). Western blot analysis revealed altered DNM1L phosphorylation and altered expression of proteins related to apoptosis suggesting a decreased apoptosis rate. Mass spectrometry analysis revealed 28 up-regulated and 21 down-regulated proteins (p < 0.05) in both experimental groups. Protein pathway analysis showed that many proteins interacted directly with the target protein DNM1L and could be classified into three main protein clusters: Vesicle traffic-associated (NSF, SNCA, ARF1), mitochondrion-associated (HSP9A, SLC25A5/ANT2, GLUD1) and cytoskeleton-associated (MAP1A) signaling pathway. Our results demonstrate that DNM1L is a promising target for an antibody-based approach to glaucoma therapy.


Assuntos
Glaucoma , Animais , Glaucoma/tratamento farmacológico , Glaucoma/metabolismo , Dinaminas/metabolismo , Células Ganglionares da Retina/metabolismo , Dinâmica Mitocondrial , Modelos Animais de Doenças , Imunoterapia
16.
PLoS One ; 17(10): e0275428, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36315519

RESUMO

Oxaliplatin (OXA) is a third-generation platinum compound with clinical activity in multiple solid tumors. Due to the repetition of chemotherapy cycle, OXA-induced chronic neuropathy presenting as paresthesia and pain. This study explored the neuropathy of chemotherapy pain and investigated the analgesic effect of 2-bromopalmitate (2-BP) on the pain behavior of OXA-induced rats. The chemotherapy pain rat model was established by the five consecutive administration of OXA (intraperitoneal, 4 mg/kg). After the establishment of OXA-induced rats, the pain behavior test, inflammatory signal analysis and mitochondrial function measurement were conducted. OXA-induced rats exhibited mechanical allodynia and spinal inflammatory infiltration. Our fluorescence and western blot analysis revealed spinal astrocytes were activated in OXA rats with up-regulation of astrocytic markers. In addition, NOD-, LRR- and pyrin domain-containing 3 (NLRP3) inflammasome mediated inflammatory signal cascade was also activated. Inflammation was triggered by dysfunctional mitochondria which represented by increase in cyclooxygenase-2 (COX-2) level and manganese superoxide dismutase (Mn-SOD) activity. Intrathecally injection of 2-BP significantly attenuated dynamin-related protein 1 (Drp1) mediated mitochondrial fission, recovered mitochondrial function, suppressed NLRP3 inflammasome cascade, and consequently decreased mechanical pain sensitivity. For cell research, 2-BP treatment significantly reversed tumor necrosis factor-α (TNF-α) induced mitochondria membrane potential deficiency and high reactive oxygen species (ROS) level. These findings indicate 2-BP decreases spinal inflammation and relieves OXA-induced neuropathic pain via reducing Drp1-mediated mitochondrial dysfunction.


Assuntos
Inflamassomos , Neuralgia , Ratos , Animais , Oxaliplatina/efeitos adversos , Inflamassomos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Ratos Sprague-Dawley , Neuralgia/induzido quimicamente , Neuralgia/tratamento farmacológico , Neuralgia/metabolismo , Dinaminas/metabolismo , Inflamação/patologia , Mitocôndrias/metabolismo
17.
Curr Oncol Rep ; 24(12): 1751-1763, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36181612

RESUMO

PURPOSE OF REVIEW: This study is aimed at reviewing the recent progress in Drp1 inhibition as a novel approach for reducing doxorubicin-induced cardiotoxicity and for improving cancer treatment. RECENT FINDINGS: Anthracyclines (e.g. doxorubicin) are one of the most common and effective chemotherapeutic agents to treat a variety of cancers. However, the clinical usage of doxorubicin has been hampered by its severe cardiotoxic side effects leading to heart failure. Mitochondrial dysfunction is one of the major aetiologies of doxorubicin-induced cardiotoxicity. The morphology of mitochondria is highly dynamic, governed by two opposing processes known as fusion and fission, collectively known as mitochondrial dynamics. An imbalance in mitochondrial dynamics is often reported in tumourigenesis which can lead to adaptive and acquired resistance to chemotherapy. Drp1 is a key mitochondrial fission regulator, and emerging evidence has demonstrated that Drp1-mediated mitochondrial fission is upregulated in both cancer cells to their survival advantage and injured heart tissue in the setting of doxorubicin-induced cardiotoxicity. Effective treatment to prevent and mitigate doxorubicin-induced cardiotoxicity is currently not available. Recent advances in cardio-oncology have highlighted that Drp1 inhibition holds great potential as a targeted mitochondrial therapy for doxorubicin-induced cardiotoxicity.


Assuntos
Proteínas Mitocondriais , Neoplasias , Humanos , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Proteínas Mitocondriais/farmacologia , Cardiotoxicidade/prevenção & controle , Dinaminas/metabolismo , Dinaminas/farmacologia , Mitocôndrias/metabolismo , Doxorrubicina/efeitos adversos , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo
18.
Oxid Med Cell Longev ; 2022: 3119649, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36254232

RESUMO

Mitophagy, the selective removal of damaged mitochondria through autophagy, is crucial for mitochondrial turnover and quality control. Docosahexaenoic acid (DHA), an essential omega-3 fatty acid, protects mitochondria in various diseases. This study aimed to investigate the neuroprotective role of DHA in ischaemic stroke models in vitro and in vivo and its involvement in mitophagy and mitochondrial dysfunction. A mouse model of ischaemic stroke was established through middle cerebral artery occlusion (MCAO). To simulate ischaemic stroke in vitro, PC12 cells were subjected to oxygen-glucose deprivation (OGD). Immunofluorescence analysis, western blotting (WB), electron microscopy (EM), functional behavioural tests, and Seahorse assay were used for analysis. DHA treatment significantly alleviated the brain infarction volume, neuronal apoptosis, and behavioural dysfunction in mice with ischaemic stroke. In addition, DHA enhanced mitophagy by significantly increasing the number of autophagosomes and LC3-positive mitochondria in neurons. The Seahorse assay revealed that DHA increased glutamate and succinate metabolism in neurons after ischaemic stroke. JC-1 and MitoSox staining, and evaluation of ATP levels indicated that DHA-induced mitophagy alleviated reactive oxygen species (ROS) accumulation and mitochondrial injury. Mechanistically, DHA improved mitochondrial dynamics by increasing the expression of dynamin-related protein 1 (Drp1), LC3, and the mitophagy clearance protein Pink1/Parkin. Mdivi-1, a specific mitophagy inhibitor, abrogated the neuroprotective effects of DHA, indicating that DHA protected neurons by enhancing mitophagy. Therefore, DHA can protect against neuronal apoptosis after stroke by clearing the damaged mitochondria through Pink1/Parkin-mediated mitophagy and by alleviating mitochondrial dysfunction.


Assuntos
Lesões Encefálicas , Isquemia Encefálica , AVC Isquêmico , Fármacos Neuroprotetores , Acidente Vascular Cerebral , Trifosfato de Adenosina/farmacologia , Animais , Encéfalo/metabolismo , Isquemia Encefálica/tratamento farmacológico , Ácidos Docosa-Hexaenoicos/farmacologia , Ácidos Docosa-Hexaenoicos/uso terapêutico , Dinaminas/metabolismo , Glucose/farmacologia , Glutamatos/farmacologia , Camundongos , Mitofagia , Fármacos Neuroprotetores/farmacologia , Fármacos Neuroprotetores/uso terapêutico , Oxigênio/farmacologia , Proteínas Quinases/metabolismo , Ratos , Espécies Reativas de Oxigênio/metabolismo , Acidente Vascular Cerebral/tratamento farmacológico , Succinatos/farmacologia , Ubiquitina-Proteína Ligases/metabolismo
19.
Mol Med ; 28(1): 126, 2022 10 25.
Artigo em Inglês | MEDLINE | ID: mdl-36284300

RESUMO

BACKGROUND: Pyroptosis is a form of programmed cell death involved in the pathophysiological progression of hypoxic pulmonary hypertension (HPH). Emerging evidence suggests that N6-methyladenosine (m6A)-modified transcripts of long noncoding RNAs (lncRNAs) are important regulators that participate in many diseases. However, whether m6A modified transcripts of lncRNAs can regulate pyroptosis in HPH progression remains unexplored. METHODS: The expression levels of FENDRR in hypoxic pulmonary artery endothelial cells (HPAECs) were detected by using quantitative real-time polymerase chain reaction (qRT-PCR) and fluorescence in situ hybridization (FISH). Western blot, Lactate dehydrogenase (LDH) release assay, Annexin V-FITC/PI double staining, Hoechst 33342/PI fluorescence staining and Caspase-1 activity assay were used to detect the role of FENDRR in HPAEC pyroptosis. The relationship between FENDRR and dynamin-related protein 1 (DRP1) was explored using bioinformatics analysis, Chromatin Isolation by RNA Purification (CHIRP), Electrophoretic mobility shift assay (EMSA) and Methylation-Specific PCR (MSP) assays. RNA immunoprecipitation (RIP) and m6A dot blot were used to detect the m6A modification levels of FENDRR. A hypoxia-induced mouse model of pulmonary hypertension (PH) was used to test preventive effect of conserved fragment TFO2 of FENDRR. RESULTS: We found that FENDRR was significantly downregulated in the nucleus of hypoxic HPAECs. FENDRR overexpression inhibited hypoxia-induced HPAEC pyroptosis. Additionally, DRP1 is a downstream target gene of FENDRR, and FENDRR formed an RNA-DNA triplex with the promoter of DRP1, which led to an increase in DRP1 promoter methylation that decreased the transcriptional level of DRP1. Notably, we illustrated that the m6A reader YTHDC1 plays an important role in m6A-modified FENDRR degradation. Additionally, conserved fragment TFO2 of FENDEE overexpression prevented HPH in vivo. CONCLUSION: In summary, our results demonstrated that m6A-induced decay of FENDRR promotes HPAEC pyroptosis by regulating DRP1 promoter methylation and thereby provides a novel potential target for HPH therapy.


Assuntos
Hipertensão Pulmonar , RNA Longo não Codificante , Camundongos , Animais , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Metilação de DNA , Células Endoteliais/metabolismo , Piroptose , Artéria Pulmonar , Hipertensão Pulmonar/genética , Hibridização in Situ Fluorescente , Hipóxia/genética , Dinaminas/genética , Dinaminas/metabolismo , Cromatina , Lactato Desidrogenases/genética , Lactato Desidrogenases/metabolismo , Caspases
20.
Int J Mol Sci ; 23(19)2022 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-36233236

RESUMO

(1) Background: Heart failure with preserved ejection fraction (HFpEF) is a major subtype of HF with no effective treatments. Mitochondrial dysfunctions relevant to the imbalance of fusion and fission occur in HFpEF. Drp1 is a key protein regulating mitochondrial fission, and PINK1 is the upstream activator of Drp1, but their relationship with HF has not been clarified. The aim of the study is to investigate molecular mechanisms of mitochondrial dysfunctions in animals with hypertension-induced HFpEF. (2) Methods and Results: The hypertension-induced HFpEF model was established by feeding Dahl/SS rats with high salt, showing risk factors such as hypertension, mitochondrial dysfunctions, and so on. Physiological and biological measurements showed a decrease in the expression of mitochondrial function-related genes, ATP production, and mitochondrial fission index. PINK1 knockout in H9C2 cardiomyocytes showed similar effects. Moreover, PINK1 myocardium-specific overexpression activated Drp1S616 phosphorylation and enhanced mitochondrial fission to slow the progression of hypertension-induced HFpEF. (3) Conclusions: PINK1 could phosphorylate Drp1S616 to improve mitochondrial fission and relieve mitochondrial dysfunctions, which highlights potential treatments of HFpEF.


Assuntos
Dinaminas , Insuficiência Cardíaca , Hipertensão , Proteínas Quinases , Trifosfato de Adenosina/metabolismo , Animais , Dinaminas/genética , Dinaminas/metabolismo , Insuficiência Cardíaca/genética , Hipertensão/genética , Dinâmica Mitocondrial , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Ratos , Ratos Endogâmicos Dahl , Volume Sistólico
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