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Apoptotic cells were analyzed by TUNEL staining. (a) Apoptotic cells were labeled with fluorescein isothiocyanate (red), and all nuclei were stained with DAPI (blue). (b) Percentage of apoptotic cells relative to total cells. **P < 0.01, ****P < 0.0001. Bar = 50 μm.
Source publication
Ischemic stroke is a common disease with high morbidity and mortality. Remote ischemic preconditioning (RIPC) can stimulate endogenous protection mechanisms by inducing ischemic tolerance to reduce subsequent damage caused by severe or fatal ischemia to non-ischemic organs. This study was designed to assess the therapeutic properties of RIPC in isc...
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Citations
... We then examined whether DCS protected against neuronal death through removal of dysfunctional mitochondria after OGD/R insult (Xie et al., 2024). We found that OGD/R-insulted neurons cocultured with astrocytes that treated by DCS reduced the expression of three mitochondrial cell-survival proteins: the VDAC1 (voltage-dependent anion-selective channel 1) (P = 0.0255, P = 0.1328 and P = 0.0013, respectively) ( Fig. 5C and D) (Stary et al., 2016), the COX4 (P = 0.0106, P = 0.2027 and P = 0.0045, respectively) ( Fig. 5C and D) (Hu, Zhou, et al., 2023) and the HSP60 (P = 0.0267, P = 0.1082 and P = 0.0012, respectively) ( Fig. 5C and D) (Lv et al., 2020). Additionally, double fluorescence staining for VDAC1 and LAMP1 showed that DCS decreased the ratio of VDAC1-positive mitochondria/LAMP1-positive lysosome (P = 0.0063, P = 0.1062 and P = 0.0036, respectively) (Fig. 5E) in cocultured neurons after OGD/R insult. ...
Non‐invasive transcranial direct‐current stimulation (tDCS) is a safe ischaemic stroke therapy. Cathodal bilateral tDCS (BtDCS) is a modified tDCS approach established by us recently. Because selenium (Se) plays a crucial role in cerebral ischaemic injury, we investigated whether cathodal BtDCS conferred neuroprotection via regulating Se‐dependent signalling in rat cerebral ischaemia–reperfusion (I/R) injury. We first showed that the levels of Se and its transport protein selenoprotein P (SEPP1) were reduced in the rat cortical penumbra following I/R, whereas cathodal BtDCS prevented the reduction of Se and SEPP1. Interestingly, direct‐current stimulation (DCS) increased SEPP1 level in cultured astrocytes subjected to oxygen‐glucose deprivation reoxygenation (OGD/R) but had no effect on SEPP1 level in OGD/R‐insulted neurons, indicating that DCS may increase Se in ischaemic neurons by enhancing the synthesis and secretion of SEPP1 in astrocytes. We then revealed that DCS reduced the number of injured mitochondria in OGD/R‐insulted neurons cocultured with astrocytes. DCS and BtDCS prevented the reduction of the mitochondrial quality‐control signalling, vesicle‐associated membrane protein 2 (VAMP2) and syntaxin‐4 (STX4), in OGD/R‐insulted neurons cocultured with astrocytes and the ischaemic brain respectively. Under the same experimental conditions, downregulation of SEPP1 blocked DCS‐ and BtDCS‐induced upregulation of VAMP2 and STX4. Finally, we demonstrated that cathodal BtDCS increased Se to reduce infract volume following I/R. Together, the present study uncovered a molecular mechanism by which cathodal BtDCS confers neuroprotection through increasing SEPP1 in astrocytes and subsequent upregulation of SEPP1/VAMP2/STX4 signalling in ischaemic neurons after rat cerebral I/R injury. image
Key points
Cathodal bilateral transcranial direct‐current stimulation (BtDCS) prevents the reduction of selenium (Se) and selenoprotein P in the ischaemic penumbra.
Se plays a crucial role in cerebral ischaemia injury.
Direct‐current stimulation reduces mitochondria injury and blocks the reduction of vesicle‐associated membrane protein 2 (VAMP2) and syntaxin‐4 (STX4) in oxygen‐glucose deprivation reoxygenation‐insulted neurons following coculturing with astrocytes.
Cathodal BtDCS regulates Se/VAMP2/STX4 signalling to confer neuroprotection after ischaemia.
... This form of degradation is unlike mitophagy, which involves the destruction of the entire organelle. The studies indicate that MDVs act as the first line of defense as they can rapidly remove marked cargo, whereas mitophagy is a last resort [38,46]. Both MDVs and mitophagy are important quality check points for eliminating aged or damaged mitochondrial components [45,47]. ...
Mitochondria are ancient endosymbiotic double membrane organelles that support a wide range of eukaryotic cell functions through energy, metabolism, and cellular control. There are over 1000 known proteins that either reside within the mitochondria or are transiently associated with it. These mitochondrial proteins represent a functional subcellular protein network (mtProteome) that is encoded by mitochondrial and nuclear genomes and significantly varies between cell types and conditions. In neurons, the high metabolic demand and differential energy requirements at the synapses are met by specific modifications to the mtProteome, resulting in alterations in the expression and functional properties of the proteins involved in energy production and quality control, including fission and fusion. The composition of mtProteomes also impacts the localization of mitochondria in axons and dendrites with a growing number of neurodegenerative diseases associated with changes in mitochondrial proteins. This review summarizes the findings on the composition and properties of mtProteomes important for mitochondrial energy production, calcium and lipid signaling, and quality control in neural cells. We highlight strategies in mass spectrometry (MS) proteomic analysis of mtProteomes from cultured cells and tissue. The research into mtProteome composition and function provides opportunities in biomarker discovery and drug development for the treatment of metabolic and neurodegenerative disease.
... For example, the transport of MDVs to lysosomes can be affected by PINK1, Parkin, Tollip or syntaxin-17 (STX17) signalling (McLelland et al., , 2016Peng et al., 2022;Ryan et al., 2020), and transport to peroxisomes can be regulated by Vps35 and mitochondrial-anchored protein ligase (MAPL) (Braschi et al., 2010;Mohanty et al., 2021), while the fusion of MDVs to MVBs and then release into the extracellular space as EVs may be mediated by cluster of differentiation 38 (CD38)/cyclic ADP ribose (cADPR) signalling (Suh et al., 2023), sorting nexin 9 (SNX9) signalling, optic atrophy 1 (OPA1) and inhibition by Parkin (Peng et al., 2022;Todkar et al., 2021), which have been well summarized before (Heyn et al., 2023;Peng et al., 2022;Popov, 2022;Sugiura et al., 2014). Stress conditions are likely to promote the selective incorporation of mitochondrial contents into MDVs, such as oxidative stress Todkar et al., 2021), remote ischemic preconditioning (Lv et al., 2020), hypoxia (Li et al., 2020), cannabidiol treatment (Ramirez et al., 2022), lipopolysaccharide (LPS) (Matheoud et al., 2016) and heat stress (Matheoud et al., 2016). However, whether those factors impact MDVs released into the extracellular space needs further study. ...
Mitochondrial damage plays vital roles in the pathology of many diseases, such as cancers, neurodegenerative diseases, aging, metabolic diseases and many types of organ injury. However, the regulatory mechanism of mitochondrial functions among different cells or organs in vivo is still unclear, and efficient therapies for attenuating mitochondrial damage are urgently needed. Extracellular vesicles (EVs) are cell-derived nanovesicles that can deliver bioactive cargoes among cells or organs. Interestingly, recent evidence shows that diverse mitochondrial contents are enriched in certain EV subpopulations, and such mitoEVs can deliver mitochondrial components to affect the functions of recipient cells under different conditions, which has emerged as a hot topic in this field. However, the overview and many essential questions with respect to this event remain elusive. In this review, we provide a global view of mitoEVs biology and mainly focus on the detailed sorting mechanisms, functional mitochondrial contents, and diverse biological effects of mitoEVs. We also discuss the pathogenic or therapeutic roles of mitoEVs in different diseases and highlight their potential as disease biomarkers or therapies in clinical translation. This review will provide insights into the pathology and drug development for various mitochondrial injury-related diseases.
... Mitochondria, the "energy factory" of cells, play an important role in cellular homeostasis [50]. The brain is an energy dependent organ and performs its functions via aerobic metabolism [51]. In the mitochondrial apoptotic pathway, cerebral ischemia leads to altered mitochondrial membrane potential and increased permeability [52], which then causes the release of cytochrome C (Cyt-C) and apoptosis inducing factor (AIF) and the formation of apoptotic bodies [53], promoting the activation of pro caspase-9. ...
Ischemic stroke and cranial radiotherapy may induce brain inflammatory response, oxidative stress, apoptosis and neuronal loss, and impairment of neurogenesis. Lycium barbarum has anti-oxidation, anti-inflammatory, anti-tumor and anti-aging properties, may produce both neuroprotective and radioprotective effects. In this narrative review paper, we described the neuroprotective effect of Lycium barbarum in different animal models of experimental ischemic stroke and limited studies in irradiated animal models. Relevant molecular mechanisms are also summarized. It has been shown that in experimental ischemic stroke models, Lycium barbarum produces neuroprotective effects by modulating neuroinflammatory factors such as cytokines and chemokines, reactive oxygen species, and neurotransmitter and receptor systems. In irradiation animal models, Lycium barbarum prevents radiation-induced loss of hippocampal interneurons. Given its minimal side-effects, these preclinical studies suggest that Lycium barbarum may be a promising radio-neuro-protective drug that can be used as an adjunct treatment to radiotherapy for brain tumor and in the treatment of ischemic stroke. At molecular levels, Lycium barbarum may regulate PI3K/Akt/GSK-3β, PI3K/Akt/mTOR, PKCε/Nrf2/HO-1, keap1-Nrf2/HO-1, and NR2A and NR2B receptor- related signal transduction pathways to produce neuroprotective effects.
... RIPerC intervention for cerebral infarction has been investigated in human clinical trials as well as animal studies. Although RIPerC was reported to markedly reduce cerebral infarct volume and neurological deficits in animal studies [3][4][5] , human clinical trials in patients with or suspected of having an acute stroke failed to replicate these beneficial effects 6,7 . Factors contributing to a lack of expected clinical outcomes in RIC were discussed at the 11th Hatter Cardiovascular Workshop, and it was concluded that the establishment of an ideal intervention www.nature.com/scientificreports/ ...
... Rats were caged for 1 week after giving birth for pre-experimental acclimation. Fig. 4. The intervention was based on previous studies by Lv et al. 4 Three RIPerC intervention protocols were performed to examine the optimal ischemia and reperfusion times. For RIPerC, elastic pressure cuffs (pediatric sphygmomanometers) were applied to both hindlimb thighs, and one of three patterns of intervention was then performed during cerebral ischemia. ...
... For RIPerC, elastic pressure cuffs (pediatric sphygmomanometers) were applied to both hindlimb thighs, and one of three patterns of intervention was then performed during cerebral ischemia. The applied pressure was 180-200 mmHg and cyanosis appeared in the toes of the rats, confirming ischemia; blocking blood flow in the femoral artery in mice has been reported to cause cyanosis, swelling of the extremities, loss of dorsal pulses in the legs, and a decrease in skin temperature 4 . ...
Remote ischemic perconditioning (RIPerC) is a novel neuroprotective method against cerebral infarction that has shown efficacy in animal studies but has not been consistently neuroprotective in clinical trials. We focused on the temporal regulation of ischemia–reperfusion by RIPerC to establish an optimal method for RIPerC. Rats were assigned to four groups: 10 min ischemia, 5 min reperfusion; 10 min ischemia, 10 min reperfusion; 5 min ischemia, 10 min reperfusion; and no RIPerC. RIPerC interventions were performed during ischemic stroke, which was induced by a 60-min left middle cerebral artery occlusion. Infarct volume, sensorimotor function, neurological deficits, and cellular expressions of brain-derived neurotrophic factor (BDNF), B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax), and caspase 3 were evaluated 48 h after the induction of ischemia. Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling (TUNEL) was also performed. RIPerC of 10 min ischemia/10 min reperfusion, and 5 min ischemia/10 min reperfusion decreased infarct volume, improved sensorimotor function, decreased Bax, caspase 3, and TUNEL-positive cells, and increased BDNF and Bcl-2 expressions. Our findings suggest RIPerC with a reperfusion time of approximately 10 min exerts its neuroprotective effects via an anti-apoptotic mechanism. This study provides important preliminary data to establish more effective RIPerC interventions.
... Radiotracers for mitochondrial complex I such as [ 1a F]-BMS, [ 1a F]-BCPP-EF, [ 1a F]-BCPP-BF, [ 11 C]-BCPP-EM, and ZCM-I-1 have been developed to study the role of mitochondrial death in the ischemic core and caspase activation in the penumbra during reperfusion (Carinci et al., 2021;Xu et al., 2021). Furthermore, an FDG-PET animal study found remote ischemic preconditioning (RIPC) was protective against mitochondrial caspase activation in ischemic stroke (Lv et al., 2020). A PET animal study conducted with the radiotracer [ 11 C]-SAR127303, which is sensitive to the glutamate neuromodulator monoacylglycerol lipase (MAGL), found that administration of minocycline and KML29 could protect the sensorimotor cortex and striatum, respectively (Yamasaki et al., 2021). ...
Ischemic strokes make up 87% of all cerebrovascular events. Intravenous tissue plasminogen activator (tPA), a thrombolytic agent, has been recognized as the only viable option for patients with ischemic stroke if administered within 3.5 h of onset and increases the risk of hemorrhagic transformation if administered beyond the treatment window. Acute treatment strategies are centered around rescuing salvageable penumbra. Molecular imaging using positron emission tomography (PET) has shown higher sensitivity and specificity than CT and MRI in delineating penumbral tissues. In addition, PET imaging has identified the role of key inflammatory mediators in atherosclerosis, cellular damage, and recovery. Recently, a novel PET imaging study has shown the feasibility of investigating synaptic density in subacute stroke. Lastly, novel PET radiotracers have been developed to further explore biochemical mechanisms implicated in stroke pathophysiology. Further investigation with PET is needed to understand stroke mechanisms and advance pharmacologic treatment.
... Some studies showed a strong link between mitochondrial morphology and cancer disease (Maycotte et al., 2017). B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax) play an important role in the regulation of the mitochondrial apoptosis pathway (Lv et al., 2020;Ma et al., 2020). In response to various apoptosis stimulus signals, Bax protein conformation changes and translocates to the outer mitochondrial membrane, forming a homodimer with a microporous structure, stimulating the release of pro-apoptotic factors such as cytochrome C into the cytoplasm, forming a multimeric complex with apoptotic protease-activating factor-1, promoting the self-activation of cysteine aspartate protease-9 (caspase-9) precursor, which activates caspase-3 and ultimately leads to apoptosis (Liu et al., 2020;Montero et al., 2020). ...
Triple-negative breast cancer (TNBC) is a severe threat to women’s health because of its aggressive nature, early age of onset, and high recurrence rate. Therefore, in this study, we aimed to evaluate the anti-tumor effects of Gallic acid (GA) on the TNBC HCC1806 cells in vitro. The cell proliferation was detected by MTT and plate clone formation assays, cell apoptosis, cell cycle, and mitochondrial membrane potential (MMP) were analyzed by flow cytometry and Hoechst 33258 staining assays, and the intracellular reactive oxygen species (ROS) accumulation were also investigated. Real-Time PCR and western blot were examined to explore the mechanism of action. The results indicated that GA suppressed HCC1806 cells proliferation and promoted HCC1806 cells apoptosis. Meanwhile, GA treatment changed the morphology of the HCC1806 cells. In addition, GA blocked the HCC1806 cells cycle in the S phase, and it induced cells apoptosis accompanied by ROS accumulation and MMP depolarization. Real-Time PCR results suggested that GA increased Bax, Caspase-3, Caspase-9, P53, JINK and P38 mRNA expression, and decreased Bcl-2, PI3K, AKT and EGFR mRNA expression. Western blotting results suggested that GA increased Bax, cleaved-Caspase-3, cleaved-Caspase-9, P53, P-ERK1/2, P-JNK, P-P38 proteins expression, and decreased Bcl-2, P-PI3K, P-AKT, P-EGFR proteins expression. Furthermore, molecular docking suggested that GA has the high affinity for PI3K, AKT, EGFR, ERK1/2, JNK, and P38. In conclusion, GA could suppress HCC1806 cells proliferation and promote HCC1806 cells apoptosis through the mitochondrial apoptosis pathway and induces ROS generation which further inhibits PI3K/AKT/EGFR and activates MAPK signaling pathways. Our study will provide some new references for using GA in the treatment of TNBC.
... Oxidative stress is activated after the recovery of cerebral blood ow, resulting in abnormal mitochondrial structure and function, drastic reduction in energy supply, and cell death, an important mechanism for ischemia-reperfusion injury 40 . A study has shown that distal ischemic preconditioning could inhibit apoptosis through mitochondrial pathway 41 , thereby reducing cerebral ischemia-reperfusion injury. In this study, we observed that after ischemia-reperfusion, AIF in rat brain tissue increased compared with sham group. ...
Macrophage migration inhibitory factor (MIF) is an immune mediator associated with inflammation, which is upregulated after ischemia in brain tissue. ISO-1 is a potent inhibitor of MIF tautomerase and can protect against neurons by reducing the permeability of blood brain barrier (BBB). In this study, we investigated the role of ISO-1 in ischemia / reperfusion injury in the brain by establishing a model of middle cerebral artery occlusion / reperfusion in rats. Rats were randomized into four groups: the sham operation group, the ISO-1group, the cerebral I/R group, and the ISO-1 + I/R group. We evaluated the degree of neurological deficit in each group and measured the volume of cerebral infarction. We detected the expression of MIF in the core necrotic area and penumbra. We detected the expression of apoptosis-related proteins, apoptosis-inducing factor (AIF), endonuclease G (EndoG) and cytochrome c oxidase-IV (COX-IV) in the ischemic penumbra region. The results showed that the expression of MIF in the ischemic penumbra area, while ISO-1 injection was able to alleviate nerve function defect and reduce infarction area. In cerebral ischemic penumbra region, ISO-1 could reduc the expression of Bax and Caspase3, and inhibit the displacement of AIF and EndoG to the nucleus simultaneously. Besides, ISO-1 also exhibited the ability to reduce apotosis. In summary, ISO-1 may inhibit neuronal apoptosis through the endogenous mitochondrial pathway and reduce the injury of brain I/R after ischemic stroke.
... Ischemic stroke (IS) refers to brain tissue necrosis resulted from stenosis or occlusion of the blood supply arteries of the brain and insufficient blood supply to the brain (Lv et al. 2020). IS is one of the most important causes of death and disability all round the world (Chen et al. 2017). ...
Background
Bone marrow stromal cells (BMSCs) transplantation is a treatment strategy for ischemic stroke (IS) with great potential. However, the vitality, migration and adhesion of BMSCs are greatly impaired due to the harsh environment of the ischemic area, which affects the therapeutic effects. Herein, we aimed to investigate the roles of nerve growth factor (NGF) in regulating cell behaviors of BMSCs in IS.
Methods
The mRNA and protein expressions were assessed using qRT-PCR and western blot, respectively. To simulate ischemic-like conditions in vitro, Brain microvascular (bEnd.3) cells were exposed to oxygen and glucose deprivation (OGD). Cell viability and cell proliferation were evaluated by MTT assay and BrdU assay, respectively. Transwell migration and cell adhesion assays were carried out to determine cell migration and adhesion of BMSCs, respectively, coupled with flow cytometry to evaluate cell apoptosis of bEnd.3 cells. Finally, angiogenesis assay was performed to assess the angiogenesis ability of bEnd.3 cells.
Results
NGF overexpression resulted in increased cell vitality, adhesion and migration of BMSCs, while NGF knockdown presented the opposite effects. We subsequently discovered that TrkA was a receptor for NGF, and TrkA knockdown significantly inhibited the cell viability, migration and adhesion of BMSCs. Besides, Nrf2 was confirmed as the downstream target of NGF/TrkA to promote the viability, adhesion and migration of BMSC cells. Finally, NGF-silenced BMSCs could not effectively restore the OGD-induced brain microvascular cell damage.
Conclusions
NGF/TrkA promoted the viability, migration and adhesion of BMSCs in IS via activating Nrf2 pathway.
... 10 The role of Vps35 in MDV generation is acknowledged by the fact that its mutation impairs MDVs formation. 28 The restoration of mitochondrial homeostasis by the removal of oxidized proteins via MDVs is upregulated in stress conditions, 8 under remote ischaemic preconditioning, 31 ...
... (i) a player in mitochondrial QC, ensuring the preservation of mitochondrial proteome (containing >1,000 proteins) and the functional integration of mitochondria according to the cellular demands. 8,15,17,26,34,38 Recently, Lv et al. 31 ...
The generation of vesicles is a constitutive attribute of mitochondria inherited from bacterial ancestors. The physiological conditions and mild oxidative stress promote oxidation and dysfunction of certain proteins and lipids within the mitochondrial membranes; these constituents are subsequently packed as small mitochondrial‐derived vesicles (MDVs) (70–150 nm in diameter) and are transported intracellularly to lysosomes and peroxisomes to be degraded. In this way, MDVs remove the damaged mitochondrial components, preserve mitochondrial structural and functional integrity and restore homeostasis. An outline of the current knowledge on MDVs seems to be necessary for understanding the potential impact of this research area in cellular (patho)physiology. The present synopsis is an attempt towards the accomplishment of this demand, highlighting also the still unclear issues related to MDVs. Here, we discuss (i) MDVs budding and generation (molecules and mechanisms), (ii) the distinct cargoes packed and transported by MDVs, (iii) the MDVs trafficking pathways and (iv) the biological role of MDVs, from quality controllers to the involvement in organellar crosstalk, mitochondrial antigen presentation and peroxisome de novo biogenesis. These complex roles uncover also mitochondria integration into the cellular environment. As the therapeutic exploitation of MDVs is currently limited, future insights into MDVs cell biology are expected to direct to novel diagnostic tools and treatments.
