bims-mikwok 2024-08-11 papers

bims-mikwok

Biomed News

on Mitochondrial quality control

Issue of 2024–08–11
thirty-two papers selected by
Gavin McStay, Liverpool John Moores University

Fluorescence Microscopy and Immunoblotting for Mitophagy in Budding Yeast.
Disease-associated mutations in C-terminus of HSP70 interacting protein (CHIP) impair its ability to negatively regulate mitophagy.
Neuro-intestinal acetylcholine signalling regulates the mitochondrial stress response in Caenorhabditis elegans.
Preserving mitochondrial homeostasis protects against drug-induced liver injury via inducing OPTN (optineurin)-dependent Mitophagy.
Monitoring Mitophagy Dynamics in Live Cells.
Dysfunction of Mitochondrial Dynamics Induces Endocytosis Defect and Cell Damage in Drosophila Nephrocytes.
Assessing Neuronal Mitophagy During Development in the Nematode Caenorhabditis elegans.
Real-Time Monitoring of mtDNA Aggregation and Mitophagy Induced by a Fluorescent Platinum Complex in Living Cells.
Methods for Monitoring Mitophagy Using mt-Keima.
Naturally occurring hyperactive variants of human parkin.
Role of Microglial Mitophagy in Alleviating Postoperative Cognitive Dysfunction: a Mechanistic Study.
Assays to Study Mitophagy in Plants.
Analysis of Mitophagy Reporters in Drosophila.
Electroacupuncture improves cognitive impairment after ischemic stroke based on regulation of mitochondrial dynamics through SIRT1/PGC-1α pathway.
Suppression of CDK1/Drp1-mediated mitochondrial fission attenuates dexamethasone-induced extracellular matrix deposition in the trabecular meshwork.
Protocatechuic aldehyde attenuates chondrocyte senescence via the regulation of PTEN-induced kinase 1/Parkin-mediated mitochondrial autophagy.
20S-O-Glc-DM treats left ventricular diastolic dysfunction by modulating cardiomyocyte mitochondrial quality and excess autophagy.
Inhibition of mitophagy via the EIF2S1-ATF4-PRKN pathway contributes to viral encephalitis.
Mitochondrial fragmentation in early differentiation of human MB135 myoblasts: Role of mitochondrial ROS production in the absence of depolarization.
Role of AMBRA1 in mitophagy regulation: emerging evidence in aging-related diseases.
Regulation mechanism of GPS2 on PGC-1α/Drp1-mediated mitochondrial dynamics in inflammation of acute lung injury.
PINK1 insufficiency can be exploited as a specific target for drug combinations inducing mitochondrial pathology-mediated cell death in gastric adenocarcinoma.
Low-dose hexavalent chromium induces mitophagy in rat liver via the AMPK-related PINK1/Parkin signaling pathway.
Mitochondrial dynamics regulate cell morphology in the developing cochlea.
Targeting TGR5 to mitigate liver fibrosis: Inhibition of hepatic stellate cell activation through modulation of mitochondrial fission.
Targeting the PINK1/Parkin Pathway: a new perspective in the prevention and therapy of diabetic retinopathy.
Deciphering the neuroprotective mechanisms of RACK1 in cerebral ischemia-reperfusion injury: Pioneering insights into mitochondrial autophagy and the PINK1/Parkin axis.
Organelle communication maintains mitochondrial and endosomal homeostasis during podocyte lipotoxicity.
Dichloroacetate attenuates brain injury through inhibiting neuroinflammation and mitochondrial fission in a rat model of sepsis-associated encephalopathy.
PFN1 Knockdown Aggravates Mitophagy to Retard Lung Adenocarcinoma Initiation and M2 Macrophage Polarization.
Peroxiredoxin 2 Regulates DAF-16/FOXO Mediated Mitochondrial Remodelling in Response to Exercise that is Disrupted in Ageing.
The role of remifentanil in regulating mitochondrial autophagy in osteoclasts was investigated based on PINK1/Parkin pathway.

Methods Mol Biol. 2024 ;2845 1-14

Fluorescence Microscopy and Immunoblotting for Mitophagy in Budding Yeast.

Yuki Nakayama, Koji Okamoto.

  Selective removal of excess or damaged mitochondria is an evolutionarily conserved process that contributes to mitochondrial quality and quantity control. This catabolic event relies on autophagy, a membrane trafficking system that sequesters cytoplasmic constituents into double membrane-bound autophagosomes and delivers them to lysosomes (vacuoles in yeast) for hydrolytic degradation and is thus termed mitophagy. Dysregulation of mitophagy is associated with various diseases, highlighting its physiological relevance. In budding yeast, the pro-mitophagic single-pass membrane protein Atg32 is upregulated under prolonged respiration or nutrient starvation, anchored on the surface of mitochondria, and activated to recruit the autophagy machinery for the formation of autophagosomes surrounding mitochondria. In this chapter, we provide protocols to assess Atg32-mediated mitophagy using fluorescence microscopy and immunoblotting.

Keywords:  Atg32; Budding yeast; Fluorescence microscopy; Immunoblotting; Mitochondria

DOI:  https://doi.org/10.1007/978-1-0716-4067-8_1
Neurobiol Dis. 2024 Aug 06. pii: S0969-9961(24)00225-0. [Epub ahead of print] 106625

Disease-associated mutations in C-terminus of HSP70 interacting protein (CHIP) impair its ability to negatively regulate mitophagy.

Rebecca Earnshaw, Yu Tong Zhang, Gregory Heymann, Kazuko Fujisawa, Sarah Hui, Minesh Kapadia, Lorraine V Kalia, Suneil K Kalia.

  C-terminus of HSP70 interacting protein (CHIP) is an E3 ubiquitin ligase and HSP70 cochaperone. Mutations in the CHIP encoding gene are the cause of two forms of neurodegenerative conditions: spinocerebellar ataxia autosomal dominant type 48 (SCA48) and autosomal recessive type 16 (SCAR16). The mechanisms underlying CHIP-associated diseases are currently unknown. Mitochondrial dysfunction, specifically dysfunction in mitochondrial autophagy (mitophagy), is increasingly being implicated in neurodegenerative diseases and loss of CHIP has been demonstrated to result in mitochondrial dysfunction in multiple animal models, although how CHIP is involved in mitophagy regulation has been previously unknown. Here, we demonstrate that CHIP acts as a negative regulator of the PTEN-induced kinase 1 (PINK1)/Parkin-mediated mitophagy pathway, promoting the degradation of PINK, impairing Parkin translocation to the mitochondria, and suppressing mitophagy in response to mitochondrial stress. We also show that loss of CHIP enhances neuronal mitophagy in a PINK1 and Parkin dependent manner in Caenorhabditis elegans. Furthermore, we find that multiple disease-associated mutations in CHIP dysregulate mitophagy both in vitro and in vivo in C. elegans neurons, a finding which could implicate mitophagy dysregulation in CHIP-associated diseases.

Keywords:  Ataxia; Mitophagy; Neurodegeneration; SCA48; STUB1

DOI:  https://doi.org/10.1016/j.nbd.2024.106625
Nat Commun. 2024 Aug 03. 15(1): 6594

Neuro-intestinal acetylcholine signalling regulates the mitochondrial stress response in Caenorhabditis elegans.

Rebecca Cornell, Wei Cao, Bernie Harradine, Rasoul Godini, Ava Handley, Roger Pocock.

Neurons coordinate inter-tissue protein homeostasis to systemically manage cytotoxic stress. In response to neuronal mitochondrial stress, specific neuronal signals coordinate the systemic mitochondrial unfolded protein response (UPRmt) to promote organismal survival. Yet, whether chemical neurotransmitters are sufficient to control the UPRmt in physiological conditions is not well understood. Here, we show that gamma-aminobutyric acid (GABA) inhibits, and acetylcholine (ACh) promotes the UPRmt in the Caenorhabditis elegans intestine. GABA controls the UPRmt by regulating extra-synaptic ACh release through metabotropic GABAB receptors GBB-1/2. We find that elevated ACh levels in animals that are GABA-deficient or lack ACh-degradative enzymes induce the UPRmt through ACR-11, an intestinal nicotinic α7 receptor. This neuro-intestinal circuit is critical for non-autonomously regulating organismal survival of oxidative stress. These findings establish chemical neurotransmission as a crucial regulatory layer for nervous system control of systemic protein homeostasis and stress responses.

DOI: https://doi.org/10.1038/s41467-024-50973-y
Autophagy. 2024 Aug 04. 1-20

Preserving mitochondrial homeostasis protects against drug-induced liver injury via inducing OPTN (optineurin)-dependent Mitophagy.

Jiajia Wang, Yueping Qiu, Lijun Yang, Jincheng Wang, Jie He, Chengwu Tang, Zhaoxu Yang, Wenxiang Hong, Bo Yang, Qiaojun He, Qinjie Weng.

  Disruption of mitochondrial function is observed in multiple drug-induced liver injuries (DILIs), a significant global health threat. However, how the mitochondrial dysfunction occurs and whether maintain mitochondrial homeostasis is beneficial for DILIs remains unclear. Here, we show that defective mitophagy by OPTN (optineurin) ablation causes disrupted mitochondrial homeostasis and aggravates hepatocytes necrosis in DILIs, while OPTN overexpression protects against DILI depending on its mitophagic function. Notably, mass spectrometry analysis identifies a new mitochondrial substrate, GCDH (glutaryl-CoA dehydrogenase), which can be selectively recruited by OPTN for mitophagic degradation, and a new cofactor, VCP (valosin containing protein) that interacts with OPTN to stabilize BECN1 during phagophore assembly, thus boosting OPTN-mediated mitophagy initiation to clear damaged mitochondria and preserve mitochondrial homeostasis in DILIs. Then, the accumulation of OPTN in different DILIs is further validated with a protective effect, and pyridoxine is screened and established to alleviate DILIs by inducing OPTN-mediated mitophagy. Collectively, our findings uncover a dual role of OPTN in mitophagy initiation and implicate the preservation of mitochondrial homeostasis via inducing OPTN-mediated mitophagy as a potential therapeutic approach for DILIs.Abbreviation: AILI: acetaminophen-induced liver injury; ALS: amyotrophic lateral sclerosis; APAP: acetaminophen; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CHX: cycloheximide; Co-IP: co-immunoprecipitation; DILI: drug-induced liver injury; FL: full length; GCDH: glutaryl-CoA dehydrogenase; GOT1/AST: glutamic-oxaloacetic transaminase 1; GO: gene ontology; GSEA: gene set enrichment analysis; GPT/ALT: glutamic - pyruvic transaminase; INH: isoniazid; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MMP: mitochondrial membrane potential; MST: microscale thermophoresis; MT-CO2/COX-II: mitochondrially encoded cytochrome c oxidase II; OPTN: optineurin; PINK1: PTEN induced kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; TIMM23: translocase of inner mitochondrial membrane 23; TOMM20: translocase of outer mitochondrial membrane 20; TSN: toosendanin; VCP: valosin containing protein, WIPI2: WD repeat domain, phosphoinositide interacting 2.

Keywords:  Drug-induced liver injury; mitochondrial homeostasis; mitophagy; optineurin; phagophore formation

DOI:  https://doi.org/10.1080/15548627.2024.2384348
Methods Mol Biol. 2024 ;2845 161-175

Monitoring Mitophagy Dynamics in Live Cells.

Christina Karantanou, Sofia-Iris Bibli.

  The purpose of this protocol is to provide a comprehensive, stepwise guide for assessing mitophagy flux utilizing a live-cell mt-KEIMA approach. The proposed protocol is sensitive, reproducible, quantitative, and easy to perform. While mitophagy has been extensively studied, current methodologies primarily focus on terminal measurements, neglecting the dynamic aspect of this process. Hence, the introduction of this straightforward live-cell mitophagy tracing protocol enables real-time monitoring of the dynamics of mitochondrial selective autophagy, thereby enhancing the ability to draw conclusions regarding key regulators and the reversibility of the process. The assay employs a lentiviral approach to induce mt-KEIMA expression in primary or immortalized cell lines. Subsequently, the respective mitophagy reporter cells are observed using a live-cell imaging system at specific time intervals, and further quantification allows the detection of mitophagy flux. This protocol has proven efficacious in investigating mitophagy flux, including responses to chemical inducers or genetically modified cells over time. Notably, this approach is well-suited for large throughput screening of chemicals or appropriate gene-editing libraries that may influence mitophagy responses in cells.

Keywords:  Flux; Live-cell imaging; Mitophagy; Screening; mt-Keima

DOI:  https://doi.org/10.1007/978-1-0716-4067-8_13
Cells. 2024 Jul 25. pii: 1253. [Epub ahead of print]13(15):

Dysfunction of Mitochondrial Dynamics Induces Endocytosis Defect and Cell Damage in Drosophila Nephrocytes.

Jun-Yi Zhu, Jianli Duan, Joyce van de Leemput, Zhe Han.

  Mitochondria are crucial for cellular ATP production. They are highly dynamic organelles, whose morphology and function are controlled through mitochondrial fusion and fission. The specific roles of mitochondria in podocytes, the highly specialized cells of the kidney glomerulus, remain less understood. Given the significant structural, functional, and molecular similarities between mammalian podocytes and Drosophila nephrocytes, we employed fly nephrocytes to explore the roles of mitochondria in cellular function. Our study revealed that alterations in the Pink1-Park (mammalian PINK1-PRKN) pathway can disrupt mitochondrial dynamics in Drosophila nephrocytes. This disruption led to either fragmented or enlarged mitochondria, both of which impaired mitochondrial function. The mitochondrial dysfunction subsequently triggered defective intracellular endocytosis, protein aggregation, and cellular damage. These findings underscore the critical roles of mitochondria in nephrocyte functionality.

Keywords:  Drosophila; Marf (MFN); Parkin; Pink1; endocytosis; mitochondria; nephrocytes; reactive oxygen species (ROS)

DOI:  https://doi.org/10.3390/cells13151253
Methods Mol Biol. 2024 ;2845 55-66

Assessing Neuronal Mitophagy During Development in the Nematode Caenorhabditis elegans.

Christina Ploumi, Antonis Roussos, Konstantinos Palikaras.

  Preserving mitochondrial homeostasis is vital, particularly for the energetically demanding and metabolically active nerve cells. Mitophagy, the selective autophagic removal of mitochondria, stands out as a prominent mechanism for efficient mitochondrial turnover, which is crucial for proper neuronal development and function. Dysfunctional mitochondria and disrupted mitophagy pathways have been linked to a diverse array of neurological disorders. The nematode Caenorhabditis elegans, with its well-defined nervous system, serves as an excellent model to unravel the intricate involvement of mitophagy in developing neurons. This chapter describes the use of Rosella biosensor in C. elegans to monitor neuronal mitophagy, providing a user-friendly platform for screening genes and drugs affecting mitophagic pathways under physiological conditions or in the context of neurodevelopmental pathologies.

Keywords:  Caenorhabditis elegans; Development; Mitochondria; Mitophagy; Neurodevelopmental diseases; Neurons; Rosella biosensor

DOI:  https://doi.org/10.1007/978-1-0716-4067-8_5
Anal Chem. 2024 Aug 07.

Real-Time Monitoring of mtDNA Aggregation and Mitophagy Induced by a Fluorescent Platinum Complex in Living Cells.

Bing Liu, Ting Sun, Yumeng Wang, Xiao-Yu Xia, Shixian Cao, Kang-Nan Wang, Qixin Chen, Zong-Wan Mao.

Mitochondrial DNA (mtDNA) is pivotal for mitochondrial morphology and function. Upon mtDNA damage, mitochondria undergo quality control mechanisms, including fusion, fission, and mitophagy. Real-time monitoring of mtDNA enables a deeper understanding of its effect on mitochondrial function and morphology. Controllable induction and real-time tracking of mtDNA dynamics and behavior are of paramount significance for studying mitochondrial function and morphology, facilitating a deeper understanding of mitochondria-related diseases. In this work, a fluorescent platinum complex was designed and developed that not only induces mitochondrial DNA (mtDNA) aggregation but also triggers mitochondrial autophagy (mitophagy) through the MDV pathway for damaged mtDNA clearance in living cells. Additionally, this complex allows for the real-time monitoring of these processes. This complex may serve as a valuable tool for studying mitochondrial microautophagy and holds promise for broader applications in cellular imaging and disease research.

DOI: https://doi.org/10.1021/acs.analchem.4c01128
Methods Mol Biol. 2024 ;2845 151-160

Methods for Monitoring Mitophagy Using mt-Keima.

Alexandra J Gilsrud, Derek P Narendra.

  Mitochondria-targeted Keima (mt-Keima) is a pH-sensitive, acid-stable fluorescent protein used for the quantification of mitophagy. Mt-Keima contains a mitochondrial matrix targeting sequence and has bimodal excitation with peaks at 440 nM in neutral environments and 586 nM in acidic environments. From this bimodal excitation, a ratiometric signal may be calculated to quantify mitophagy in live cells. This chapter describes procedures for measuring mitophagy by flow cytometry and live cell confocal microscopy with mt-Keima.

Keywords:  Mitochondria; Mitophagy; PINK1; Parkin; Selective autophagy

DOI:  https://doi.org/10.1007/978-1-0716-4067-8_12
Commun Biol. 2024 Aug 08. 7(1): 961

Naturally occurring hyperactive variants of human parkin.

Tahrima Saiha Huq, Jean Luo, Rayan Fakih, Véronique Sauvé, Kalle Gehring.

Parkinson's disease (PD) is the second most common neurodegenerative disease in the world. Although most cases are sporadic and occur later in life, 10-15% of cases are genetic. Loss-of-function mutations in the ring-between-ring E3 ubiquitin ligase parkin, encoded by the PRKN gene, cause autosomal recessive forms of early onset PD. Together with the kinase PINK1, parkin forms a mitochondrial quality control pathway that tags damaged mitochondria for clearance. Under basal conditions, parkin is inhibited and compounds that increase its activity have been proposed as a therapy for PD. Recently, several naturally occurring hyperactive parkin variants were identified, which increased mitophagy in cultured cells. Here, we validate the hyperactivities of these variants in vitro and compare the levels of activity of the variants to those of the wild-type and the well-characterized hyperactive variant, W403A. We also study the effects of mutating the parkin ACT (activating element) on parkin activity in vitro. This work advances our understanding of the pathogenicity of parkin variants and is an important first step in the design of molecules to increase parkin activity.

DOI: https://doi.org/10.1038/s42003-024-06656-x
Mol Neurobiol. 2024 Aug 07.

Role of Microglial Mitophagy in Alleviating Postoperative Cognitive Dysfunction: a Mechanistic Study.

Lina Zhang, Jiaying Li, Chenglong Li, Yujin Wu, Shuai Liu, Qi Li, Sihua Qi.

  Postoperative cognitive dysfunction (POCD), a common complication following anesthesia and surgery, is influenced by hippocampal neuroinflammation and microglial activation. Mitophagy, a process regulating inflammatory responses by limiting the accumulation of damaged mitochondria, plays a significant role. This study aimed to determine whether regulating microglial mitophagy and the cGAS-STING pathway could alleviate cognitive decline after surgery. Exploratory laparotomy was performed to establish a POCD model using mice. Western blotting, immunofluorescence staining, transmission electron microscopy, and mt-Keima assays were used to examine microglial mitophagy and the cGAS-STING pathway. Quantitative polymerase chain reaction (qPCR) was used to detect inflammatory mediators and cytosolic mitochondrial DNA (mtDNA) levels in BV2 cells. Exploratory laparotomy triggered mitophagy and enhanced the cGAS-STING pathway in mice hippocampi. Pharmacological treatment reduced microglial activation, neuroinflammation, and cognitive impairment after surgery. Mitophagy suppressed the cGAS-STING pathway in mice hippocampi. In vitro, microglia-induced inflammation was mediated by mitophagy and the cGAS-STING pathway. Small interfering RNA (siRNA) of PINK1 hindered mitophagy activation and facilitated the cytosolic release of mtDNA, resulting in the initiation of the cGAS-STING pathway and innate immune response. Microglial mitophagy inhibited inflammatory responses via the mtDNA-cGAS-STING pathway inducing microglial mitophagy and inhibiting the mtDNA-cGAS-STING pathway may be an effective therapeutic approach for patients with POCD.

Keywords:  Microglia; Mitophagy; PINK1-Parkin pathway; Postoperative cognitive dysfunction; cGAS-STING pathway

DOI:  https://doi.org/10.1007/s12035-024-04405-z
Methods Mol Biol. 2024 ;2845 39-53

Assays to Study Mitophagy in Plants.

Chengyang Li, Juncai Ma, Tian Zhen, Byung-Ho Kang, Pengwei Wang.

  Like most eukaryotic cells, mitophagy is essential in plant development and stress response. Several recent studies have revealed proteins that regulate this process, such as Friendly (FMT) and TraB family proteins (TRB), which are plant-unique mitophagy regulators so far. Here, we describe methods for studying mitophagy activity in plants through conventional microscopy and the use of loss-of-function mutants, such as using transgenic mitochondrial marker lines followed by image analysis, chemical inhibitor treatment, and plant phenotype studies. These methods can be used in combination to identify the putative mitophagy regulators and understand their functions in mitochondrial-related activities in plants.

Keywords:  Arabidopsis thaliana; De-etiolation; Mitochondria; Mitophagy; Nicotiana benthamiana

DOI:  https://doi.org/10.1007/978-1-0716-4067-8_4
Methods Mol Biol. 2024 ;2845 79-93

Analysis of Mitophagy Reporters in Drosophila.

Alvaro Sanchez-Martinez, Aitor Martinez, Alexander J Whitworth.

  Mitophagy is the degradation of mitochondria via the autophagy-lysosome system, disruption of which has been linked to multiple neurodegenerative diseases. As a flux process involving the identification, tagging, and degradation of subcellular components, the analysis of mitophagy benefits from the microscopy analysis of fluorescent reporters. Studying the pathogenic mechanisms of disease also benefits from analysis in animal models in order to capture the complex interplay of molecular and cell biological phenomena. Here, we describe protocols to analyze mitophagy reporters in Drosophila by light microscopy.

Keywords:  Brain; Drosophila; Light microscopy; Mitochondria; Mitophagy; Muscle; Neurodegeneration; Reporter; mito-QC; mtx-QC

DOI:  https://doi.org/10.1007/978-1-0716-4067-8_7
Brain Res. 2024 Aug 05. pii: S0006-8993(24)00393-7. [Epub ahead of print] 149139

Electroacupuncture improves cognitive impairment after ischemic stroke based on regulation of mitochondrial dynamics through SIRT1/PGC-1α pathway.

Limin Chen, Shuying Chen, Yanjie Bai, Yongchuang Zhang, Xiaoxiao Li, Yan Wang, Yuqian Xiao, Jun Wan, Kexin Sun.

  In recent years, the mechanism of acupuncture in the treatment of post-stroke cognitive impairment (PSCI) has not been fully elucidated. The balance between mitochondrial fission and fusion is important for PSCI. Our previous research demonstrated that electroacupuncture can improve learning and memory in middle cerebral artery ischemia reperfusion (MCAO/R) rats. However, the specific mechanism by which electroacupuncture improves learning and memory in MCAO/R rats by regulating mitochondrial fission and fusion needs to be further investigated. The MCAO/R rats was developed using the line-bolt method. The rats were randomly divided into sham-operated (Sham), model (MCAO/R), electroacupuncture (MCAO/R + EA) and sham-electroacupuncture (MCAO/R + sham EA) groups. Investigating the effects of EA on the expression of Sirtuin1 (SIRT1), peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), Optic atrophy 1R + (OPA1) and Dynamin-related protein 1 (DRP1) in hippocampal neurons and on the morphology and function of hippocampal neurons and mitochondria. EA was able to reduce neurologic deficit scores and cerebral infarct volume and improve new object discrimination in MCAO/R rats, but there were no significant changes in these indices in the sham-electroacupuncture group. Moreover, EA increased the expression of SIRT1, PGC-1α, and OPA1 in hippocampal tissues, inhibited the expression of DRP1, attenuated neuronal and mitochondrial damage, and reduced mitochondrial fragmentation. The mechanism by which EA improves learning memory deficits in MCAO/R rats may be related to the inhibition of SIRT1/PGC-1α expression, the enhancement of mitochondrial fusion and the obstruction of its fission, and the reduction of hippocampal neuronal damage.

Keywords:  Cognitive impairment; Electroacupuncture; Mitochondrial dynamics; SIRT1/PGC-1α; Stroke

DOI:  https://doi.org/10.1016/j.brainres.2024.149139
Antioxid Redox Signal. 2024 Aug 03.

Suppression of CDK1/Drp1-mediated mitochondrial fission attenuates dexamethasone-induced extracellular matrix deposition in the trabecular meshwork.

Xizhi Deng, Min Zhu, Yang Liu, Nan Zhang, Pengyu Zhang, Wen Zeng, Min Ke.

AIMS: Deposition of extracellular matrix (ECM) in the trabecular meshwork (TM), as induced by dexamethasone (DEX), is believed to play an important role in the onset of glucocorticoid-induced glaucoma (GIG). Abnormal ECM deposition is a consequence of mitochondrial dysfunction. We aimed to clarify how mitochondrial dysfunction leads to ECM deposition within the TM and to support the development of novel therapeutic strategies.
RESULTS: In primary human TM cells (pHTMCs) and a dexamethasone acetate-induced murine model of GIG, glucocorticoid administration stimulated both mitochondrial fission and ECM deposition. Excessive mitochondrial fission leads to dysfunction and the overexpression of ECM proteins in pHTMCs. Notably, when pHTMCs were treated with the Drp1 inhibitor Mdivi-1 or with Drp1 siRNA, we observed a marked reduction in DEX-induced mitochondrial damage and ECM proteins in vitro. Furthermore, in C57BL/6J mice, treatment with Mdivi-1 mitigated mitochondrial damage and blocked ECM deposition within the TM. We then employed Ro3306 to inhibit the CDK1-mediated phosphorylation of Drp1 at Ser 616, which restored mitochondrial function and diminished DEX-induced ECM protein expression in pHTMCs.
INNOVATION: This study illuminates the pathogenic mechanism linking mitochondrial dysfunction to ECM deposition in GIG. Our innovative approach revealed that DEX stimulates mitochondrial fission via CDK1-mediated p-Drp1s616 overexpression, which drives ECM accumulation. It offered a novel therapeutic strategy for reducing ECM protein expression by inhibiting excessive mitochondrial fission and restoring mitochondrial function.
CONCLUSION: By targeting the CDK1/Drp1-driven mitochondrial fission process, we can counteract DEX-induced ECM deposition in the TM both in vivo and in vitro.

DOI: https://doi.org/10.1089/ars.2023.0502
Int J Immunopathol Pharmacol. 2024 Jan-Dec;38:38 3946320241271724

Protocatechuic aldehyde attenuates chondrocyte senescence via the regulation of PTEN-induced kinase 1/Parkin-mediated mitochondrial autophagy.

Lishi Jie, Xiaoqing Shi, Junfeng Kang, Houyu Fu, Likai Yu, Di Tian, Wei Mei, Songjiang Yin.

  This study aimed to investigate whether the beneficial effects of PCA on chondrocyte senescence are mediated through the regulation of mitophagy. Chondrocyte senescence plays a significant role in the development and progression of knee osteoarthritis (OA). The compound protocatechuic aldehyde (PCA), which is abundant in the roots of Salvia miltiorrhiza, has been reported to have antioxidant properties and the ability to protect against cellular senescence. To achieve this goal, a destabilization of the medial meniscus (DMM)-induced mouse OA model and a lipopolysaccharide (LPS)-induced chondrocyte senescence model were used, in combination with PINK1 gene knockdown or overexpression. After treatment with PCA, cellular senescence was assessed using Senescence-Associated β-Galactosidase (SA-β-Gal) staining, DNA damage was evaluated using Hosphorylation of the Ser-139 (γH2AX) staining, reactive oxygen species (ROS) levels were measured using Dichlorodihydrofluorescein diacetate (DCFH-DA) staining, mitochondrial membrane potential was determined using a 5,5',6,6'-TETRACHLORO-1,1',3,3'-*. TETRAETHYBENZIMIDA (JC-1) kit, and mitochondrial autophagy was examined using Mitophagy staining. Western blot analysis was also performed to detect changes in senescence-related proteins, PINK1/Parkin pathway proteins, and mitophagy-related proteins. Our results demonstrated that PCA effectively reduced chondrocyte senescence, increased the mitochondrial membrane potential, facilitated mitochondrial autophagy, and upregulated the PINK1/Parkin pathway. Furthermore, silencing PINK1 weakened the protective effects of PCA, whereas PINK1 overexpression enhanced the effects of PCA on LPS-induced chondrocytes. PCA attenuates chondrocyte senescence by regulating PINK1/Parkin-mediated mitochondrial autophagy, ultimately reducing cartilage degeneration.

Keywords:  PTEN-induced kinase 1/parkin pathway; chondrocyte senescence; mitochondrial autophagy; protocatechuic aldehyde

DOI:  https://doi.org/10.1177/03946320241271724
Phytomedicine. 2024 Jul 26. pii: S0944-7113(24)00569-5. [Epub ahead of print]133 155911

20S-O-Glc-DM treats left ventricular diastolic dysfunction by modulating cardiomyocyte mitochondrial quality and excess autophagy.

Xinyi Guo, Zihan Chen, Yanxin Liu, Zhiwei Chen, Modi Lin, Lingzhi Zhang, Ping Zhu, Jinling Yang, Zhe Wang, Jinlan Zhang, Hua Sun.

   BACKGROUND: Left ventricular diastolic dysfunction (LVDD) is a manifestation of heart failure, with both its incidence and prevalence increasing annually. Currently, no pharmacological treatments are available for LVDD, highlighting the urgent need for new therapeutic discoveries. Ginsenosides are commonly used in cardiovascular therapy. Previous research has synthesized the ginsenoside precursor molecule, 20S-O-Glc-DM (C20DM), through biosynthesis. C20DM shows greater bioavailability, eco-friendliness, and cost-effectiveness compared to traditional ginsenosides, positioning it as a promising option for treating LVDD.
PURPOSE: This study firstly documents the therapeutic activity of C20DM against LVDD and unveils its potential mechanisms of action. It provides a pharmacological basis for C20DM as a new cardiovascular therapeutic agent.
METHODS: In this study, models of LVDD in mice and ISO-induced H9C2 cell damage were developed. Cell viability, ROS and Ca2+ levels, mitochondrial membrane potential, and proteins associated with mitochondrial biogenesis and autophagy were evaluated in the in vitro experiments. Animal experiments involved administering medication for 3 weeks to validate the therapeutic effects of C20DM and its impact on mitochondria and autophagy.
RESULTS: Research has shown that C20DM is more effective than Metoprolol in treating LVDD, significantly lowering the E/A ratio, e'/a' ratio, and IVRT, and ameliorating myocardial inflammation and fibrosis. C20DM influences the activity of PGC-1α, downregulates PINK1 and Parkin, thereby enhancing mitochondrial quality control, and restoring mitochondrial oxidative respiration and membrane potential. Furthermore, C20DM reduces excessive autophagy in cardiomyocytes via the AMPK-mTOR-ULK1 pathway, diminishing cardiomyocyte hypertrophy and damage.
CONCLUSIONS: Overall, our research indicates that C20DM has the potential to enhance LVDD through the regulation of mitochondrial quality control and cellular autophagy, making it a promising option for heart failure therapy.

Keywords:  Autophagy; C20DM; Left ventricular diastolic dysfunction; Mitochondrial quality control

DOI:  https://doi.org/10.1016/j.phymed.2024.155911
J Adv Res. 2024 Aug 03. pii: S2090-1232(24)00326-6. [Epub ahead of print]

Inhibition of mitophagy via the EIF2S1-ATF4-PRKN pathway contributes to viral encephalitis.

Xiaowei Song, Yiliang Wang, Weixiangmin Zou, Zexu Wang, Wenyan Cao, Minting Liang, Feng Li, Qiongzhen Zeng, Zhe Ren, Yifei Wang, Kai Zheng.

   INTRODUCTION: Mitophagy, a selective form of autophagy responsible for maintaining mitochondrial homeostasis, regulates the antiviral immune response and acts as viral replication platforms to facilitate infection with various viruses. However, its precise role in herpes simplex virus 1 (HSV-1) infection and herpes simplex encephalitis (HSE) remains largely unknown.
OBJECTIVES: We aimed to investigate the regulation of mitophagy by HSV-1 neurotropic infection and its role in viral encephalitis, and to identify small compounds that regulate mitophagy to affect HSV-1 infection.
METHODS: The antiviral effects of compounds were investigated by Western blot, RT-PCR and plaque assay. The changes of Parkin (PRKN)-mediated mitophagy and Nuclear Factor kappa B (NFKB)-mediated neuroinflammation were examined by TEM, RT-qPCR, Western blot and ELISA. The therapeutic effect of taurine or PRKN-overexpression was confirmed in the HSE mouse model by evaluating survival rate, eye damage, neurodegenerative symptoms, immunohistochemistry analysis and histopathology.
RESULTS: HSV-1 infection caused the accumulation of damaged mitochondria in neuronal cells and in the brain tissue of HSE mice. Early HSV-1 infection led to mitophagy activation, followed by inhibition in the later viral infection. The HSV-1 proteins ICP34.5 or US11 deregulated the EIF2S1-ATF4 axis to suppress PRKN/Parkin mRNA expression, thereby impeding PRKN-dependent mitophagy. Consequently, inhibition of mitophagy by specific inhibitor midiv-1 promoted HSV-1 infection, whereas mitophagy activation by PRKN overexpression or agonists (CCCP and rotenone) attenuated HSV-1 infection and reduced the NF-κB-mediated neuroinflammation. Moreover, PRKN-overexpressing mice showed enhanced resistance to HSV-1 infection and ameliorated HSE pathogenesis. Furthermore, taurine, a differentially regulated gut microbial metabolite upon HSV-1 infection, acted as a mitophagy activator that transcriptionally promotes PRKN expression to stimulate mitophagy and to limit HSV-1 infection both in vitro and in vivo.
CONCLUSION: These results reveal the protective function of mitophagy in HSE pathogenesis and highlight mitophagy activation as a potential antiviral therapeutic strategy for HSV-1-related diseases.

Keywords:  EIF2S1-ATF4; Herpes simplex virus; Neuroinflammation; PRKN-dependent mitophagy; Taurine

DOI:  https://doi.org/10.1016/j.jare.2024.08.003
Life Sci. 2024 Aug 02. pii: S0024-3205(24)00531-9. [Epub ahead of print]354 122941

Mitochondrial fragmentation in early differentiation of human MB135 myoblasts: Role of mitochondrial ROS production in the absence of depolarization.

Daniil A Chernyavskij, Konstantin G Lyamzaev, Olga Yu Pletjushkina, Fei Chen, Anna Karpukhina, Yegor S Vassetzky, Boris V Chernyak, Ekaterina N Popova.

   AIMS: Study of the role of mitochondria-generated reactive oxygen species (mtROS) and mitochondrial polarization in mitochondrial fragmentation at the initial stages of myogenesis.
MAIN METHODS: Mitochondrial morphology, Drp1 protein phosphorylation, mitochondrial electron transport chain components content, mtROS and mitochondrial lipid peroxidation levels, and mitochondrial polarization were evaluated on days 1 and 2 of human MB135 myoblasts differentiation. A mitochondria-targeted antioxidant SkQ1 was used to elucidate the effect of mtROS on mitochondria.
KEY FINDINGS: In immortalized human MB135 myoblasts, mitochondrial fragmentation began on day 1 of differentiation before the myoblast fusion. This fragmentation was preceded by dephosphorylation of p-Drp1 (Ser-637). On day 2, an increase in the content of some mitochondrial proteins was observed, indicating mitochondrial biogenesis stimulation. Furthermore, we found that myogenic differentiation, even on day 1, was accompanied both by an increased production of mtROS, and lipid peroxidation of the inner mitochondrial membrane. SkQ1 blocked these effects and partially reduced the level of mitochondrial fragmentation, but did not affect the dephosphorylation of p-Drp1 (Ser-637). Importantly, mitochondrial fragmentation at early stages of MB135 differentiation was not accompanied by depolarization, as an important stimulus for mitochondrial fragmentation.
SIGNIFICANCE: Mitochondrial fragmentation during early myogenic differentiation depends on mtROS production rather than mitochondrial depolarization. SkQ1 only partially inhibited mitochondrial fragmentation, without significant effects on mitophagy or early myogenic differentiation.

Keywords:  Mitochondrial fragmentation; Mitochondrial membrane potential (ΔΨm); Mitochondrial reactive oxygen species (mtROS); Myogenesis; Myogenic differentiation

DOI:  https://doi.org/10.1016/j.lfs.2024.122941
Autophagy. 2024 Aug 08.

Role of AMBRA1 in mitophagy regulation: emerging evidence in aging-related diseases.

Martina Di Rienzo, Alessandra Romagnoli, Giulia Refolo, Tiziana Vescovo, Fabiola Ciccosanti, Candida Zuchegna, Francesca Lozzi, Luca Occhigrossi, Mauro Piacentini, Gian Maria Fimia.

  Aging is a gradual and irreversible physiological process that significantly increases the risks of developing a variety of pathologies, including neurodegenerative, cardiovascular, metabolic, musculoskeletal, and immune system diseases. Mitochondria are the energy-producing organelles, and their proper functioning is crucial for overall cellular health. Over time, mitochondrial function declines causing an increased release of harmful reactive oxygen species (ROS) and DNA, which leads to oxidative stress, inflammation and cellular damage, common features associated with various age-related pathologies. The impairment of mitophagy, the selective removal of damaged or dysfunctional mitochondria by autophagy, is relevant to the development and progression of age-related diseases. The molecular mechanisms that regulates mitophagy levels in aging remain largely uncharacterized. AMBRA1 is an intrinsically disordered scaffold protein with a unique property of regulating the activity of both proliferation and autophagy core machineries. While the role of AMBRA1 during embryonic development and neoplastic transformation has been extensively investigated, its functions in post-mitotic cells of adult tissues have been limited due to the embryonic lethality caused by AMBRA1 deficiency. Recently, a key role of AMBRA1 in selectively regulating mitophagy in post-mitotic cells has emerged. Here we summarize and discuss these results with the aim of providing a comprehensive view of the mitochondrial roles of AMBRA1, and how defective activity of AMBRA1 has been functionally linked to mitophagy alterations observed in age-related degenerative disorders, including muscular dystrophy/sarcopenia, Parkinson diseases, Alzheimer diseases and age-related macular degeneration.

Keywords:  AMBRA1; Aging; aging-related diseases; autophagy; mitochondria; mitophagy

DOI:  https://doi.org/10.1080/15548627.2024.2389474
Int Immunopharmacol. 2024 Aug 07. pii: S1567-5769(24)01359-6. [Epub ahead of print]140 112838

Regulation mechanism of GPS2 on PGC-1α/Drp1-mediated mitochondrial dynamics in inflammation of acute lung injury.

Liang Zhao, Changping Gu, Yi Zhang, Jie Yan, Lei Qiu, Xiaofeng Qin, Yuelan Wang.

  Acute lung injury (ALI) has been a hot topic in the field of critical care research in recent years. Mitochondrial dynamics consists of mitochondrial fusion and mitochondrial fission. Dynamin-related protein 1 (Drp1), a key molecule that regulates mitochondrial fission, is important in the oxidative stress and inflammatory response to ALI. Peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) is a core protein that mediates mitochondrial biogenesis. G-protein pathway suppressor 2 (GPS2) acts as a transcriptional cofactor with regulatory effects on nuclear-encoded mitochondrial genes. This study aimed to investigate the mechanism of PGC-1α/Drp1-mediated mitochondrial dynamics involved in ALI and to demonstrate the protective mechanism of GPS2 in regulating mitochondrial structure and function and inflammation in ALI. The ALI model was constructed using LPS-induced wild-type mice and human pulmonary microvascular endothelial cells (HPMVECs). It was found that lung injury, oxidative stress and inflammation were exacerbated in the mice ALI model and that mitochondrial structure and function were disrupted in HPMVECs. In vitro studies revealed that LPS led to the upregulated expression of Drp1 and the downregulated expression of PGC-1α and GPS2. Mitochondrial division was reduced and respiratory function was restored in Drp1 knockdown cells, which inhibited oxidative stress and inflammatory response. In addition, the overexpression of PGC-1α and GPS2 significantly inhibited the expression of Drp1, mitochondrial function was restored, and inhibited reactive oxygen species (ROS) production and inflammatory factor release. Moreover, the overexpression of GPS2 promoted the upregulated expression of PGC-1α. This mechanism was also validated in vivo, in which the low expression of GPS2 in mice resulted in the upregulated expression of Drp1 and the downregulated expression of PGC-1α, and further exacerbated LPS-induced ALI. In the present study, we also found that LPS-induced the downregulated expression of GPS2 may be associated with its increased degradation by the proteasome. Therefore, these findings revealed that GPS2 inhibited oxidative stress and inflammation by modulating PGC-1α/Drp1-mediated mitochondrial dynamics to alleviate LPS-induced ALI, which may provide a new approach to the therapeutic orientation for LPS-induced ALI.

Keywords:  Acute lung injury; Endothelium; Inflammation; Lipopolysaccharides; Mitochondrial dynamics; Oxidative stress

DOI:  https://doi.org/10.1016/j.intimp.2024.112838
Arch Biochem Biophys. 2024 Aug 03. pii: S0003-9861(24)00232-7. [Epub ahead of print] 110110

PINK1 insufficiency can be exploited as a specific target for drug combinations inducing mitochondrial pathology-mediated cell death in gastric adenocarcinoma.

Sayak Ghosh, Debapriya Ghatak, Rittick Dutta, Devyani Goswami, Rudranil De.

  There exist very limited non-hazardous therapeutic strategies except for surgical resection and lymphadenectomy against gastric cancer (GC) despite being the third leading cause of cancer deaths worldwide. This study proposes an innovative treatment approach against GC using a drug combination strategy that manipulates mitochondrial dynamics in conjunction with the induction of mitochondrial pathology-mediated cell death. Comparative analysis was done with gastric adenocarcinoma and normal cells by qPCR, western blot, microscopic immunocytochemistry, and live cell imaging. In this study, impairment of dynamin-related protein 1 (Drp1)-mediated mitochondrial fission by Mdivi-1 created an imbalance in mitochondrial structural dynamics in indomethacin-treated AGS cells in which mitophagy-regulator protein PINK1 is downregulated. These drug combinations with the individual sub-lethal doses ultimately led to the activation of cell death machinery upregulating pro-apoptotic proteins like Bax, Puma, and Noxa. Interestingly, this combinatorial therapy did not affect normal gastric epithelial cells significantly and also no significant upregulation of death markers was observed. Moreover, the drug combination strategy also retarded cell migration and reduced stemness in GC cells. In summary, this study offers a pioneering specific therapeutic strategy for GC treatment, sparing normal cells providing opportunities for minimal drug-mediated toxicity utilizing mitochondria as a viable and specific target for anti-cancer therapy in gastric cancer.

Keywords:  Gastric adenocarcinoma; Mitochondria; PINK1; mitochondrial dynamics; mitophagy

DOI:  https://doi.org/10.1016/j.abb.2024.110110
PeerJ. 2024 ;12 e17837

Low-dose hexavalent chromium induces mitophagy in rat liver via the AMPK-related PINK1/Parkin signaling pathway.

Ningning Li, Xiaoying Li, Xiuzhi Zhang, Lixia Zhang, Hui Wu, Yue Yu, Guang Jia, Shanfa Yu.

  Hexavalent chromium (Cr(VI)) is a hazardous metallic compound commonly used in industrial processes. The liver, responsible for metabolism and detoxification, is the main target organ of Cr(VI). Toxicity experiments were performed to investigate the impacts of low-dose exposure to Cr(VI) on rat livers. It was revealed that exposure of 0.05 mg/kg potassium dichromate (K2Cr2O7) and 0.25 mg/kg K2Cr2O7 notably increased malondialdehyde (MDA) levels and the expressions of P-AMPK, P-ULK, PINK1, P-Parkin, and LC3II/LC3I, and significantly reduced SOD activity and P-mTOR and P62 expression levels in liver. Electron microscopy showed that CR(VI) exposure significantly increased mitophagy and the destruction of mitochondrial structure. This study simulates the respiratory exposure mode of CR(VI) workers through intratracheal instillation of CR(VI) in rats. It confirms that autophagy in hepatocytes is induced by low concentrations of CR(VI) and suggest that the liver damage caused by CR(VI) may be associated with the AMPK-related PINK/Parkin signaling pathway.

Keywords:  AMPK; Chromium; Liver injury; Mitophagy; PINK; Parkin

DOI:  https://doi.org/10.7717/peerj.17837
Development. 2024 Aug 01. pii: dev202845. [Epub ahead of print]151(15):

Mitochondrial dynamics regulate cell morphology in the developing cochlea.

James D B O'Sullivan, Stephen Terry, Claire A Scott, Anwen Bullen, Daniel J Jagger, Zoë F Mann.

  In multicellular tissues, the size and shape of cells are intricately linked with their physiological functions. In the vertebrate auditory organ, the neurosensory epithelium develops as a mosaic of sensory hair cells (HCs), and their glial-like supporting cells, which have distinct morphologies and functional properties at different frequency positions along its tonotopic long axis. In the chick cochlea, the basilar papilla (BP), proximal (high-frequency) HCs, are larger than their distal (low-frequency) counterparts, a morphological feature essential for sound perception. Mitochondrial dynamics, which constitute the equilibrium between fusion and fission, regulate differentiation and functional refinement across a variety of cell types. We investigate this as a potential mechanism for regulating the shape of developing HCs. Using live imaging in intact BP explants, we identify distinct remodelling of mitochondrial networks in proximal compared with distal HCs. Manipulating mitochondrial dynamics in developing HCs alters their normal morphology along the proximal-distal (tonotopic) axis. Inhibition of the mitochondrial fusion machinery decreased proximal HC surface area, whereas promotion of fusion increased the distal HC surface area. We identify mitochondrial dynamics as a key regulator of HC morphology in developing inner ear epithelia.

Keywords:  Biogenesis; Chick; Hearing; Metabolism; Mitofusin

DOI:  https://doi.org/10.1242/dev.202845
Int Immunopharmacol. 2024 Aug 06. pii: S1567-5769(24)01352-3. [Epub ahead of print]140 112831

Targeting TGR5 to mitigate liver fibrosis: Inhibition of hepatic stellate cell activation through modulation of mitochondrial fission.

Li Sun, Yuancheng Shao, Zehao Zhuang, Zhixin Liu, Mingjun Liu, Chang Qu, Haojun Yang.

  Chronic hepatitis B virus (HBV) infection continues to be a prominent cause of liver fibrosis and end-stage liver disease in China, necessitating the development of effective therapeutic strategies. This study investigated the potential of targeting TGR5 to alleviate liver fibrosis by impeding the activation of hepatic stellate cells (HSCs), which play a pivotal role in fibrotic progression. Using the human hepatic stellate cell line LX-2 overexpressing hepatitis B virus X protein (HBX), this study revealed that TGR5 activation through INT-777 inhibits HBX-induced LX-2 cell activation, thereby ameliorating liver fibrosis, which is associated with the attenuation of mitochondrial fission and introduces a novel regulatory pathway in liver fibrosis. Additional experiments with mitochondrial fission inducers and inhibitors confirm the crucial role of mitochondrial dynamics in TGR5-mediated effects. In vivo studies using TGR5 knockout mice substantiate these findings, demonstrating exacerbated fibrosis in the absence of TGR5 and its alleviation with the mitochondrial fission inhibitor Mdivi-1. Overall, this study provides insights into TGR5-mediated regulation of liver fibrosis through the modulation of mitochondrial fission in HSCs, suggesting potential therapeutic strategies for liver fibrosis intervention.

Keywords:  Chronic hepatitis B virus; Hepatic stellate cells; Liver fibrosis; Mitochondrial fission; TGR5

DOI:  https://doi.org/10.1016/j.intimp.2024.112831
Exp Eye Res. 2024 Aug 06. pii: S0014-4835(24)00245-8. [Epub ahead of print] 110024

Targeting the PINK1/Parkin Pathway: a new perspective in the prevention and therapy of diabetic retinopathy.

Agata Grazia D'Amico, Grazia Maugeri, Benedetta Magrì, Claudio Bucolo, Velia D'Agata.

  Diabetic retinopathy (DR) is a microvascular complication of diabetes characterized by neurovascular impairment of the retina. The dysregulation of the mitophagy process occurs before apoptotic cell death and the appearance of vascular damage. In particular, mitochondrial alterations happen during DR development, supporting the hypothesis that mitophagy is negatively correlated to disease progression. This process is mainly regulated by the PTEN-induced putative kinase protein 1 (PINK1) /Parkin pathway whose activation promotes mitophagy. In this review, we will summarize the evidence reported in the literature demonstrating the involvement of the PINK1/Parkin pathway in diabetic retinopathy-induced retinal degeneration.

Keywords:  Diabetic Retinopathy; Mitophagy; PINK1; Parkin

DOI:  https://doi.org/10.1016/j.exer.2024.110024
CNS Neurosci Ther. 2024 Aug;30(8): e14836

Deciphering the neuroprotective mechanisms of RACK1 in cerebral ischemia-reperfusion injury: Pioneering insights into mitochondrial autophagy and the PINK1/Parkin axis.

Lanqing Zhao, Yu Chen, Hongxi Li, Xiaoxu Ding, Jinwei Li.

   INTRODUCTION: Cerebral ischemia-reperfusion injury (CIRI) is a common and debilitating complication of cerebrovascular diseases such as stroke, characterized by mitochondrial dysfunction and cell apoptosis. Unraveling the molecular mechanisms behind these processes is essential for developing effective CIRI treatments. This study investigates the role of RACK1 (receptor for activated C kinase 1) in CIRI and its impact on mitochondrial autophagy.
METHODS: We utilized high-throughput transcriptome sequencing and weighted gene co-expression network analysis (WGCNA) to identify core genes associated with CIRI. In vitro experiments used human neuroblastoma SK-N-SH cells subjected to oxygen and glucose deprivation (OGD) to simulate ischemia, followed by reperfusion (OGD/R). RACK1 knockout cells were created using CRISPR/Cas9 technology, and cell viability, apoptosis, and mitochondrial function were assessed. In vivo experiments involved middle cerebral artery occlusion/reperfusion (MCAO/R) surgery in rats, evaluating neurological function and cell apoptosis.
RESULTS: Our findings revealed that RACK1 expression increases during CIRI and is protective by regulating mitochondrial autophagy through the PINK1/Parkin pathway. In vitro, RACK1 knockout exacerbated cell apoptosis, while overexpression of RACK1 reversed this process, enhancing mitochondrial function. In vivo, RACK1 overexpression reduced cerebral infarct volume and improved neurological deficits. The regulatory role of RACK1 depended on the PINK1/Parkin pathway, with RACK1 knockout inhibiting PINK1 and Parkin expression, while RACK1 overexpression restored them.
CONCLUSION: This study demonstrates that RACK1 safeguards against neural damage in CIRI by promoting mitochondrial autophagy through the PINK1/Parkin pathway. These findings offer crucial insights into the regulation of mitochondrial autophagy and cell apoptosis by RACK1, providing a promising foundation for future CIRI treatments.

Keywords:  PINK/Parkin pathway; RACK1; cell apoptosis; cerebral ischemia‐reperfusion injury; mitochondrial autophagy; mitochondrial function; neuroprotection

DOI:  https://doi.org/10.1111/cns.14836
JCI Insight. 2024 Aug 08. pii: e182534. [Epub ahead of print]

Organelle communication maintains mitochondrial and endosomal homeostasis during podocyte lipotoxicity.

Sho Hasegawa, Masaomi Nangaku, Yuto Takenaka, Chigusa Kitayama, Qi Li, Madina Saipidin, Yu Ah Hong, Jin Shang, Yusuke Hirabayashi, Naoto Kubota, Takashi Kadowaki, Reiko Inagi.

  Organelle stress exacerbates podocyte injury, contributing to perturbed lipid metabolism. Simultaneous organelle stresses occur in kidney tissues; therefore, a thorough analysis of organelle communication is crucial for understanding the progression of kidney diseases. Although organelles closely interact with one another at membrane contact sites, limited studies have explored their involvement in kidney homeostasis. The endoplasmic reticulum (ER) protein, PDZ domain-containing 8 (PDZD8), is implicated in multiple organelle tethering processes and cellular lipid homeostasis. In this study, we aimed to elucidate the role of organelle communication in podocyte injury using podocyte-specific Pdzd8-knockout mice. Our findings demonstrated that Pdzd8 deletion exacerbated podocyte injury in an accelerated obesity-related kidney disease model. Proteomic analysis of isolated glomeruli revealed that Pdzd8 deletion exacerbated mitochondrial and endosomal dysfunction during podocyte lipotoxicity. Additionally, electron microscopy revealed the accumulation of "fatty abnormal endosomes" in Pdzd8-deficient podocytes during obesity-related kidney diseases. Lipidomic analysis indicated that glucosylceramide accumulated in Pdzd8-deficient podocytes, owing to accelerated production and decelerated degradation. Thus, the organelle-tethering factor, PDZD8, plays a crucial role in maintaining mitochondrial and endosomal homeostasis during podocyte lipotoxicity. Collectively, our findings highlight the importance of organelle communication at the three-way junction among the ER, mitochondria, and endosomes in preserving podocyte homeostasis.

Keywords:  Chronic kidney disease; Mitochondria; Nephrology; Obesity

DOI:  https://doi.org/10.1172/jci.insight.182534
Int Immunopharmacol. 2024 Aug 05. pii: S1567-5769(24)01361-4. [Epub ahead of print]140 112840

Dichloroacetate attenuates brain injury through inhibiting neuroinflammation and mitochondrial fission in a rat model of sepsis-associated encephalopathy.

Peng Wang, Lian Liang, Qiulin Ge, Siqi Liu, Zhengfei Yang, Longyuan Jiang.

  Sepsis-associated encephalopathy (SAE) is a severe complication of sepsis, characterized by neuroinflammation, mitochondrial dysfunction, and oxidative stress, leading to cognitive decline and high mortality. The effectiveness of dichloroacetate (DCA) in modulating mitochondrial function provides a novel therapeutic strategy for SAE. In this study, we evaluated the neuroprotective effects of DCA in a rat model of SAE induced by cecal ligation and puncture (CLP). Rats treated with DCA exhibited significant improvements in neurological function and survival, as evidenced by less neuron loss from histopathologic analysis, restored neurologic deficit scores, improved Y-maze alternation percentages, and enhanced recognition index performance. Biochemical analyses showed that DCA administration at 25 mg/kg and 100 mg/kg reduced astrocyte and microglial activation, indicating reduced neuroinflammation. Furthermore, DCA simultaneously reduced the production of circulating and cerebral inflammatory cytokines (including TNF-α, IL-1β, and IL-10), concomitant with mitigating oxidative stress through down-regulating expression of 8-Hydroxy-2'-deoxyguanosine (8-OHdG) and reactive oxygen species (ROS) in the brain. Mechanistically, DCA modulated mitochondrial dynamics by suppressing Drp1 and pDrp1 expression, which are indicators of mitochondrial fission. This was corroborated by transmission electron microscopy, quantification of mitochondrial area, and Western blot analyses. Furthermore, DCA treatment improved ATP levels, mitochondrial complex I activity, and NAD+/NADH ratio, indicating a significant attenuation of brain mitochondrial dysfunction. In conclusion, our findings suggest that DCA confers neuroprotection in SAE by curtailing neuroinflammation and mitochondrial fission, outlining a promising therapeutic strategy for treating SAE in critically ill patients.

Keywords:  Cecal ligation and puncture; Dichloroacetate; Mitochondrial fission; Neuroinflammation; Sepsis-associated encephalopathy

DOI:  https://doi.org/10.1016/j.intimp.2024.112840
Mol Biotechnol. 2024 Aug 09.

PFN1 Knockdown Aggravates Mitophagy to Retard Lung Adenocarcinoma Initiation and M2 Macrophage Polarization.

Rongrong Sun, Yang Li, Yu Feng, Xiaoyan Shao, Rantian Li, Hao Li, Sanyuan Sun, Jiangbo Wang.

  Tumor-associated macrophages (TAM) are considered as crucial influencing factors of lung adenocarcinoma (LUAD) carcinogenesis and metastasis. Profilin 1 (PFN1) has been proposed as a potent driver of migration and drug resistance in LUAD. The focus of this work was to figure out the functional mechanism of PFN1 in macrophage polarization in LUAD. PFN1 expression and its significance in patients' survival were detected by ENCORI and Kaplan-Meier Plotter. RT-qPCR and western blotting examined PFN1 expression in LUAD cells. CCK-8 assay and colony formation assay detected cell proliferation. Flow cytometry detected cell apoptosis. Relevant assay kit tested caspase3 concentration. Western blotting analyzed the expression of proliferation- and apoptosis-related proteins. RT-qPCR and immunofluorescence staining measured M1 and M2 macrophages markers. Mitophagy was assessed by MitoTracker Red staining, immunofluorescence staining, and western blotting. PFN1 expression was increased in LUAD tissues and cells and correlated with the poor survival rate of LUAD patients. Deficiency of PFN1 hindered the proliferation, whereas facilitated the apoptosis of LUAD cells. Additionally, PFN1 interference impaired M2 macrophage polarization. Moreover, PFN1 knockdown exacerbated the mitophagy in LUAD cells and mitophagy inhibitor mitochondrial division inhibitor 1 (Mdivi-1) notably reversed the effects of PFN1 down-regulation on the proliferation, apoptosis as well as macrophage polarization in LUAD cells. To sum up, activation of mitophagy initiated by PFN1 depletion might obstruct the occurrence and M2 macrophage polarization in LUAD.

Keywords:  Apoptosis; Lung adenocarcinoma; Macrophage polarization; Mitophagy; PFN1

DOI:  https://doi.org/10.1007/s12033-024-01228-0
Mol Metab. 2024 Aug 06. pii: S2212-8778(24)00134-0. [Epub ahead of print] 102003

Peroxiredoxin 2 Regulates DAF-16/FOXO Mediated Mitochondrial Remodelling in Response to Exercise that is Disrupted in Ageing.

Qin Xia, Penglin Li, José C Casas-Martinez, Antonio Miranda-Vizuete, Emma McDermott, Peter Dockery, Katarzyna Goljanek-Whysall, Brian McDonagh.

  Ageing is associated with mitochondrial dysfunction and increased oxidative stress. Exercise generates endogenous reactive oxygen species (ROS) and promotes rapid mitochondrial remodelling. We investigated the role of Peroxiredoxin 2 (PRDX-2) in mitochondrial adaptations to exercise and ageing using Caenorhabditis elegans as a model system. PRDX-2 was required for the mitochondrial remodelling in response to exercise mediated by DAF-16 transcription factor activation and regulation of mitochondrial fusion gene eat-3. Employing an acute exercise and recovery cycle, we demonstrated exercise-induced mitochondrial ER contact sites (MERCS) assembly and mitochondrial remodelling dependent on PRDX-2 and DAF-16 signalling. There was increased mitochondrial fragmentation, elevated ROS and an altered redox state of PRDX-2 concomitant with impaired DAF-16 nuclear localisation during ageing. Similarly, the prdx-2 mutant strain exhibited increased mitochondrial fragmentation and a failure to activate DAF-16 required for mitochondrial fusion. Collectively, our data highlight the critical role of PRDX-2 in orchestrating mitochondrial remodelling in response to a physiological stress by regulating DAF-16 nuclear localisation.

Keywords:  Ageing; C. elegans; DAF-16; Exercise; Mitochondrial ER Contact Sites; Peroxiredoxin 2

DOI:  https://doi.org/10.1016/j.molmet.2024.102003
Cell Mol Biol (Noisy-le-grand). 2024 Jul 28. 70(7): 186-192

The role of remifentanil in regulating mitochondrial autophagy in osteoclasts was investigated based on PINK1/Parkin pathway.

Pufeng Ye, Guifeng Pan, Yuanfeng Li, Aobo Li, Jianbin Zhang, Mingxiu Xin, Zhenjiang Mai.

This study aimed to explore the regulatory effect of remifentanil-mediated mitochondrial autophagy on osteoclast formation and further investigate its mechanism. Macrophage cell line RAW264.7 was taken and induced to differentiate into mature osteoclasts using nuclear factor kB receptor activating factor ligand (RANKL). The cell model was treated with different concentrations of remifentanil or down-regulated expression of mitochondrial autophagy-related gene PINK1. The survival, death and ROS production of osteoclasts were detected by CCK8 kit and flow cytometry, MMP level was detected by JC-1 method, mitochondrial morphology and autophagy were observed by transmission electron microscopy, and mitochondrial autophagy-related protein expression was detected by Western blot. The number of osteoclasts in the remifentanil-treated group was significantly reduced compared to the control group, accompanied by a reduction in reactive oxygen species (ROS) and mitochondrial membrane potential levels (MMP). Further results showed that remifentanil could significantly up-regulate the activity of PINK1/Parkin pathway, promote the occurrence of mitochondrial autophagy, and damaged mitochondria, and inhibit the formation of osteoclasts. Remifentanil successfully inhibited osteoclast formation by regulating mitochondrial autophagy mediated by PINK1/Parkin pathway. The results of this study revealed that remifentanil plays an important role in the physiology and pathology of osteoclasts, which may provide new ideas and strategies for the clinical treatment of remifentanil in tibial fractures.

DOI: https://doi.org/10.14715/cmb/2024.70.7.27