Mitochondria as a therapeutic target for ischemic stroke

Journal Pre-proof Mitochondria as a therapeutic target for ischemic stroke Zhi He, Niya Ning, Qiongxiu Zhou, Seyed Esmaeil Khoshnam, Maryam Farzaneh PII:

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https://doi.org/10.1016/j.freeradbiomed.2019.11.005

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Free Radical Biology and Medicine

Received Date: 19 August 2019 Revised Date:

7 October 2019

Accepted Date: 3 November 2019

Please cite this article as: Z. He, N. Ning, Q. Zhou, S.E. Khoshnam, M. Farzaneh, Mitochondria as a therapeutic target for ischemic stroke, Free Radical Biology and Medicine (2019), doi: https:// doi.org/10.1016/j.freeradbiomed.2019.11.005. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. © 2019 Published by Elsevier Inc.

Graphical abstract

Ischemic stroke

Mitochondrial transfer Stem cell therapy

Mitochondrial dysfunction MPTP induction NLRP3 Inflammasome

ROS formation

Mitophagy Cytochrome C

Caspase-1 activation

Mitochondrial DNA damage Apoptosis

Pro-IL-1β Pro-IL-18

IL-1β IL-18

Cell death

Inflammatory response

Title: Mitochondria as a therapeutic target for ischemic stroke Running title: Mitochondria and ischemic stroke Zhi He1, Niya Ning2, Qiongxiu Zhou3*, Seyed Esmaeil Khoshnam4*, Maryam Farzaneh5 1

Department of pharmacy, Luohe Medical College, Luohe, 462000, China

2

Department of Obstetrics and Gynecology, Shaoling District People's Hospital of Luohe City,

Luohe, 462300, China 3

Institute of Blood Transfusion, Chinese Academy of Medical Sciences, Chengdu, 610052,

China 4

Physiology Research Center, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran

5

Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan

Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran

Corresponding author: Qiongxiu Zhou Institute of Blood Transfusion, Chinese Academy of Medical Sciences, Chengdu, 610052, China E-mail: [email protected] Seyed Esmaeil Khoshnam Physiology Research center, Ahvaz Jundishapur University of Medical Science, Ahvaz, Iran Zip code: 6135715794 Tel: +989171491729 Email: [email protected]

Mitochondria as a therapeutic target for ischemic stroke

Abstract Stroke is the leading cause of death and physical disability worldwide. Mitochondrial dysfunction has been considered as one of the hallmarks of ischemic stroke and contributes to the pathology of ischemia and reperfusion. Mitochondria is essential in promoting neural survival and neurological improvement following ischemic stroke. Therefore, mitochondria represent an important drug target for stroke treatment. This review discusses the mitochondrial molecular mechanisms underlying cerebral ischemia and involved in reactive oxygen species generation, mitochondrial electron transport dysfunction, mitochondria-mediated regulation of inflammasome activation, mitochondrial dynamics and biogenesis, and apoptotic cell death. We highlight the potential of mitochondrial transfer by stem cells as a therapeutic target for stroke treatment and provide valuable insights for clinical strategies. A better understanding of the roles of mitochondria in ischemia-induced cell death and protection may provide a rationale design of novel therapeutic interventions in the ischemic stroke.

Key words: Ischemic stroke, Mitochondria, Apoptosis, Mitochondrial biogenesis, Mitochondrial dynamics, Inflammasome, Stem cell.

1. Introduction Stroke is one of the leading causes of death and long-term disability worldwide [1] and accounts for 5.5 million death with 44 million physical disabilities annually [2]. Stroke especially in lowand middle-income countries causes a high burden to both the patient and society [3]. More than 80% of strokes are ischemic and are caused by an occlusion of cerebral arteries [4]. Lack of blood supply deprives the brain cells of necessary glucose and oxygen and disturbs cellular homeostasis, which triggers pathophysiological processes including excitotoxicity, oxidative stress, inflammation, apoptosis and cell death [5]. Currently, tissue plasminogen activator (tPA) and endovascular therapy are the only approved therapies for acute ischemic stroke [6-8]. However, tPA therapy remains limited due to the narrow therapeutic window [9]. Hence, in patients with acute cerebral large‐vessel occlusion endovascular therapy could be useful, but real‐world efficacies are unknown [8]. Neuroprotective agents as an alternative approach for treating ischemic models were failed due to their low efficacy [10-13]. Therefore, it is essential to find a novel therapeutic strategy for ischemic stroke [14, 15]. Mitochondria, as the powerhouse of the cell, perform a key role in the cell energy homeostasis and are thus certainly involved in the neuronal death following ischemic stroke. During ischemia, energy balance is disrupted due to the reduction of blood supply, and adenosine triphosphate (ATP) synthesis is disturbed. In addition to their essential role in energy generation, mitochondria are involved in the regulation of different mechanisms of cell death including apoptosis and autophagy. One of the hallmarks of ischemia/reperfusion (I/R) injury is mitochondrial dysfunction which induces neuronal death [16] and preserving of the mitochondrial function is essential for cell survival and neurological improvement after ischemia. Therefore, one of the promising therapeutic strategies for ischemic stroke is targeting of the mitochondria [17]. In this review, we will focus on the role of mitochondria in cell survival and cell death after ischemia, and highlight the improvement of mitochondria-based therapy in ischemic stroke. 2. Pathophysiology of ischemic stroke Cerebral artery occlusion caused severe oxygen and glucose deprivation (OGD) and triggered a cascade of cellular and molecular events, which result in the irretrievable cerebral injury [18]. Neurons are more vulnerable to ischemic conditions and become quickly dysfunctional or die after ischemic stroke [19-21]. Following ischemia, OGD- induced mitochondrial dysfunction,

leading to the ATP depletion and overproduction of reactive oxidative species (ROS). Neurons have higher energy demand compared to the other brain cells, but their energy reserves are inadequate. ATP depletion triggers ischemic cascades including membrane ion pump failure, an influx of sodium, chloride, and water, efflux of cellular potassium, and membrane depolarization [22-24]. Neurons are unable to maintain their normal transmembrane ionic gradient and homeostasis after ischemic stroke. This elicits several pathophysiological processes including excitotoxicity, mitochondrial response, oxidative and nitrative stress, acidotoxity, protein misfolding, inflammation, and apoptosis. These pathophysiological processes are deleterious to the neural, glial and endothelial cells, and are intertwisted, activating each other in a positive feedback loop that terminates in cell death [5, 18, 25-27]. Following ischemic stroke, a spectrum of severity is observed in the ischemic brain, which includes infarct core and the ischemic penumbra. The blood flow to the “infarct core” is declined below the threshold of energy failure (15%-20%), and undergoes irreversible neuronal damage due to necrotic cell death [28]. While, the ischemic penumbra is the surrounding tissues with impaired functions, which contains salvageable and metabolically active cells and cell death occurs less rapidly in this area [29]. During the early hours of ischemia, cells are salvageable in the penumbra area if reperfusion was established or neuronal demand for oxygen and glucose was prepared by collateral circulation [25]. Therefore, most therapeutic methods focused on safeguarding neurons from the pathological mechanisms occur in the penumbra zone, and this area is the pharmacological target for the treatment of acute ischemic stroke [30]. Neuronal cells in the infarct core undergo necrotic cell death, which accompanied by excitotoxic cell damage to the neighboring areas [31]. Several types of cell death are can be initiated by ischemia, including autophagy, apoptosis, necroptosis, and pyroptosis which can cross-regulate each other [32, 33]. Following ischemia, mitochondrial impairments trigger a cascade of events that leads to neuronal death. Ischemia caused mitochondrial depolarization that initiates excessive ROS production, reduced ATP generation, and increased PINK1 (PTEN-induced putative kinase 1) accumulation [34]. It was demonstrated that a great part of cerebral ATP is consumed for the electrogenic activity of the neurons [35]. Mitochondria is thus essential for the excitability and survival of the neuronal cells. Additionally, the major sources of ROS and serve as apoptotic regulators [36, 37]. Both these roles of the mitochondria have been associated with the pathogenesis of neurodegenerative diseases and ischemic stroke [37, 38].

Mitochondrial dysfunction leads to apoptotic cell death through increasing of the ROS generation, calcium accumulation, the opening of mitochondrial permeability transition pore (MPTP), and releasing of cytochrome c (cytc) [39]. Furthermore, mitochondrial impairment trigger mitophagy following increasing of PINK1 [40]. 3. Ischemia and reperfusion injury Disruption of blood flow to an organ is ischemia, which depriving cells of oxygen and metabolites and triggering a build-up of lactate and succinate as the metabolic products [41-43]. The ratio ATP: ADP is reduced following OGD, which results in adenine nucleotide breakdown [44]. The apparent therapy for ischemia is to return blood flow as soon as possible to the ischemic tissue. For example, the standard of care for ischemic stroke is to remove the blockage from the cerebral artery through thrombolysis by tPA [45] or by angiographic revascularization [46]. Despite the promotion of the reperfusion by these interventions, a major cause of ischemia injuries caused by the restoration of blood flow to the ischemic tissue which known as ischemia and reperfusion (IR) injury [41-43, 47], which is a key driver of pathology in stroke [41, 42]. Therefore, there is now considerable concern in developing therapies that minimizing the inevitable IR injury following ischemia [43]. 4. Mitochondria roles in IR injury It was demonstrated that mitochondria contribute to the pathologies of ischemic stroke, which cause by its aberrant production of ROS, calcium accumulation, defective mitochondrial biogenesis, triggering cell death by induction of the MPTP and activation of apoptosis, disruption in the ATP: ADP ratio, and reducing NAD+ levels [34, 48]. ROS production in the mitochondrial respiratory chain is the initiating factor of IR injury, which triggers a cascade of tissue damage during the reperfusion phase [42, 43]. It was evidenced that succinate, a citric acid cycle (CAC) metabolite, dramatically builds up during ischemia and rapidly oxidized upon reperfusion. Consequently, oxidized succinate drives ROS production at complex I by reverse electron transport (RET), which leads to oxidative damage and disruption of the mitochondrial functions [42]. Therefore, during reperfusion, oxygen is restored, which can employ as a substrate to generate oxygen free radicals through the pro-oxidant enzymic system and mitochondria [49]. Oxidative damage and calcium accumulation lead to the MPTP induction during ischemia, which caused organ dysfunction and cell death [50-52]. MPTP induction leads to the release of succinate and mitochondrial damage-associated molecular patterns (DAMPs),

resulting in the activation of an inflammatory response that will contribute to tissue damage following IR injury [53] (Figure 1). Therefore, inhibiting RET at complex I could suppress the ROS burst during reperfusion [54-56]. Protecting mitochondria from oxidative damage using exogenous antioxidants appears to be an appropriate intervention during IR injury [57]. During IR injury, the next point to protect mitochondria is inhibiting of the MPTP induction [50-52]. MPTP has been demonstrated that involved in both apoptosis and necrosis cell death, and its opening causes a decrease in mitochondrial membrane potential (∆ψm), depolarization of mitochondria, swelling and the translocation of the mitochondrial substances to the cytoplasm [58]. The opening of the MPTP is a response to mitochondrial calcium concentrations [59]. Hence, several other elements can trigger MPTP opening and mitochondrial injury, including Ca2+ accumulation, production of ROS and reactive nitrogen species (RNS) and loss of ∆ψm [60, 61]. Inhibiting the MPTP can protect the integrity and function of mitochondria after stroke insult [62]. The endogenous antioxidant system can neutralize a small change in the reactive intermediate products. However, severe oxidative stress can result in apoptotic or necrotic cell death [63]. The major source of intracellular ROS is mitochondrial organelle [64, 65]. Following IR, superoxide anion (O2-) generated in the mitochondria by the reaction of free electrons with molecular oxygen. Subsequently, the active O2- reacts with the nitrogen oxide (NO-) to form peroxynitrite anion (NO3-), which consecutively causes the formation of the cytotoxic radicals and modifications in the cellular main structures such as DNA, proteins, and lipids [66]. Therefore, blocking ROS formation is essential for the normal function of the cells, while the prolonged activity of the mitochondria has an intrinsic risk of ROS production. The balance between ROS formation and its clearance is compromised during cerebral ischemia, leading to ROS-induced cell injury. Other studies also reviewed the pathogenic role of free radicals following ischemic stroke [67, 68]. 5. Proteins involved in the mitochondrion-dependent apoptosis ROS formation increased after cerebral ischemia, which triggers a variety of molecular signaling pathways that increases injuring to the lipids, proteins, and nucleic acids. Therefore, disruption of the mitochondria inducing cell death by releasing cytc or apoptosis-inducing factor as the proapoptotic proteins [37]. It has been shown that apoptosis has a key role in ischemia-induced cerebral injuries in both human and animal models [69-72].

Cell death induced by several conditions and factors, which include ischemia severity, metabolic deregulation, bioenergetics failure, aging, and genetic factors [73, 74]. It was proved that intracellular ATP concentration is an important determinant of cell death. Since mitochondrial structure and function determined ATP production. Necrosis is associated with ATP deficiency, while apoptosis required ATP [75]. Mitochondria have associations with several apoptosis-related proteins, which play an important role in cell death [76, 77]. Several studies have shown that neuronal death in cerebral ischemic stroke profoundly regulated by a group of proteins of the B-cell lymphoma (BCL-2) family [7882], which have an essential role in the apoptotic cascade [83]. Two main classes of the BCL-2 family are anti-apoptotic proteins such as Bcl-2, Bcl-xL, and Bcl-w, and apoptotic proteins, which include Bax, Bak, Bim, Bid, Bad, and Noxa [37, 83]. Accumulating evidence suggests that after cerebral ischemia the pro-apoptotic BH3-only BCL-2 subfamily is upregulated, showing that ischemic stroke triggers several apoptotic pathways comprising mitochondria [8486]. The function of the mitochondria and the ROS formation are associated with the activities of anti- and pro-apoptotic proteins. Beside BCL-2 pathway, some other apoptotic pathways initiated from mitochondria, which involved releasing of the pro-apoptotic factors such as AIF (apoptosis-inducing factor), cytc, HtrA2/ OMI (high-temperature requirement protein A), endonuclease-G, SMAC/DIABLO (second mitochondrion-derived activator of caspase/direct inhibitor of apoptosis-binding protein with low pI), mitochondrial electron transport chain (ETC) changing, homeostasis cellular redox alteration, and losing the transmembrane potential of the mitochondria [76, 87]. Moreover, MPTPs is another important apoptotic cascade in the mitochondrial inner membrane [88], which its opening is triggered by cellular stress, and leads to the apoptotic cascade by a collapse of the mitochondrial transmembrane potential. The latter event stimulates releasing of the cytc that complemented by other proapoptotic molecules. At the cytosol, cytc interacts with the Apaf-1 (apoptotic-protease-activating factor-1), which initiates the apoptosome formation and triggers the apoptotic cascade. Apaf-1 in a complex with cytc and deoxy ATP stimulates the inactive procaspase-9, which subsequently activates caspase-3 [89-91]. It was demonstrated that the anti-apoptotic activity of the XIAP (X chromosome-linked inhibitor-of-apoptosis protein) suppressed by SMAC protein, which leaked from mitochondria. Therefore, SMAC protein activates apoptosis and inhibits serial procaspase activation following

cerebral ischemia [92, 93]. The AIF is a mitochondrial effector of apoptosis after translocation to the nucleus and suggested to function as a caspase-independent mediator of the degradation phase of apoptotic cell death [94]. AIF has been shown to accelerating the apoptotic process by inhibition of the poly(ADP-ribose) polymerase [95]. Translocation of the AIF to the nucleus in an animal model of ischemic stroke occurs before or at the time of cytc release from mitochondria [96], and also is responsible for neuronal death in vitro model of ischemic stroke [95]. 6. Mitochondrial dysfunction and regulation of the NLRP3 inflammasome Inflammation is a devastating pathophysiological process following ischemic stroke. It has been suggested that activation of the NOD-like receptor protein (NLRP3)-inflammasome mediate inflammatory responses during ischemic stroke [97], and DAMPs are the critical initial stimulus to activate NLRP3 [98]. Briefly, activation of the NLRP3 inflammasome converts pro-caspase-1 into caspase-1[99, 100], which subsequently cleaves both pro-IL-1 and pro-IL-18 into their active pro-inflammatory cytokines that are eventually released into the extracellular environment [101], leading to autophagy and pyroptosis [102], which could release DAMPs to induce more inflammation [103]. Findings provide evidence that the NLRP3 inflammasome is very important in mediating neuronal cell death and behavioral deficits after stroke [104]. The involvement of mitochondria in the NLRP3 inflammasome activation and pyroptotic cell death has evolved in the last decade. It has been proposed that mitochondrial dysfunction regulates NLRP3 inflammation activation by several distinct pathways (Figure 2) [105, 106]. Recently, it was demonstrated that mitochondrial dysfunction is pivotal in the NLRP3 inflammasome activation in the microglia after OGD and cerebral IR injury [107]. Studies have demonstrated that NLRP3 inflammasome is activated by releasing of mitochondrial ROS, mitochondrial DNA (mtDNA) damage and releasing of phospholipid cardiolipin [108-110], which discussed below. ROS and mtDNA may participate in a feed-forward signaling circuitry whereby mitochondrial danger signals are first translated into adaptive responses and then if homeostasis cannot be re-established, execute cell death [34]. 6.1. Mitochondrial ROS Mitochondria are the major source of cellular ROS, and induction of ROS production was shown to activates inflammasomes [111, 112]. Mitochondrial ROS is needed or the activation of the pro-survival NF-κB pathway and the NLRP3 inflammasome [34]. It was reported that ROS

generation was increased following mitochondrial dysfunction, which manifested as a decrease in ∆ψm, induced by the opening of the MPTP, and could promote NLRP3 inflammasome activation [113] (Figure

1). NLRP3 inflammasome activated in macrophages during

phagocytosis of silica and asbestos particles and ROS generation [114]. Conversely, treatment of macrophages with the ROS inhibitors can inhibit silica and asbestos-induced NLRP3 inflammasome activation [114, 115]. Furthermore, ROS production increased through inhibition of mitochondrial complex-I or complex-III [116, 117], which is sufficient to activate NLRP3 inflammasome, suggesting mitochondrial ROS as a direct activator of the NLRP3 inflammasome [109]. It was reported that NLRP3 activators induce the generation of mitochondrial ROS, which in turn activates the NLRP3 inflammasome [109]. This conclusion is inconsistent with several studies that based on the treatments of ROS scavengers or inhibitors of ROS generation [118121]. 6.2. MtDNA Following intrinsic cell death signaling, mtDNA is released into the cytoplasm, which acts as a mitochondrial danger signal and promotes the activation of the NLRP3 inflammasomes [122, 123]. It has been suggested that mitochondrial ROS has a key role in the releasing of mtDNA and triggering of the NLRP3 inflammasome [124]. It has been established that mtDNA is released by mitochondria in response to ROS-mediated inflammasome activation [124]. Furthermore, mtDNA act as a bona fide inflammasome stimulator (in its oxidized form) following mitochondrial outer membrane permeabilization [125]. Activation of the NLRP3 inflammasome by mtDNA is established [126-128]. However, NLRP3 is required for releasing of mtDNA into the cytoplasm [124], which suggests a positive feedback loop to potentiate activation of the NLRP3 inflammasome. In contrast to NLRP3 inflammasome activation by apoptosis-induced mtDNA, a report showed that mitochondrial apoptosis is dispensable for NLRP3-dependent apoptosis [129, 130]. More studies are required to clarify the involvement of mitochondrial apoptosis in the triggering of the NLRP3 inflammasome. 6.3. Mitochondrial Ca2+ overload Calcium (Ca2+) is essential in the regulation of several signaling pathways within cells, and its intracellular levels precisely controlled using multiple strategies. In particular, mitochondrion is a critical player in regulating Ca2+ levels [131]. However, Ca2+ overload can cause mitochondrial dysfunction. Inhibition of extracellular Ca2+ entry or depletion of Ca2+ stores in the endoplasmic

reticulum (ER) attenuates NLRP3 inflammasome activation [132]. Incubation of macrophages with Ca2+-chelating agent inhibited NLRP3 inflammasome activation in LPS-primed macrophages [133, 134]. Mounting evidence now demonstrates that an influx of Ca2+ into the cytoplasm detected by the Ca2+-sensing receptor (CaSR) [135, 136], and promotes the activation of the NLRP3 inflammasome [132, 136]. It was proposed that K+ efflux promotes Ca2+ influx, and could be upstream of Ca2+- induced NLRP3 inflammasome activation [137]. Furthermore, it was shown that K+ efflux is the common trigger of NLRP3 inflammasome activation, and high extracellular K+ abolished NLRP3 activation by Ca2+ [137]. 6.4. Cardiolipin Cardiolipins are lipids that localized in the inner mitochondrial membrane and facilitate optimal oxidative phosphorylation in mitochondria [138]. After mitochondrial destabilization, cardiolipin externalized to the outer membrane of the mitochondria and induced cell death [139]. Interestingly, cardiolipin deficiency in the cells leading to cell death resistance [140]. It was showed that cardiolipin is a critical modulator of the NLRP3 inflammasome activation, which directly binds to the leucine-rich repeats (LRRs) of NLRP3 [141]. 6.5. Mitofusins Mitofusins 2 (Mfn2) is a mitochondrial outer membrane guanosine triphosphatase (GTPase) that is essential for mitochondrial fusion and facilitates the maintenance of cell homeostasis [142, 143]. It has been reported that mitofusins are implicated in the activation of the NLRP3 inflammasome, and Mfn2 interacts with NLRP3 by their hydrophobic heptad repeat region, which promotes NLRP3 recruitment IL-1β secretion. The knockdown of Mfn2 expression reduced NLRP3-dependent caspase-1 activation and IL-1b production. Even though Mitofusins 1 (Mfn1) was also observed to associate with NLRP3, its role in the activation of the NLRP3 inflammasome remains obscure [144]. Mfn1/2 was reported to provide a location for NLRP3 inflammasome assembly [109, 132]. 7. Mitochondrial dynamics following ischemic stroke It has been documented that mitochondrial dynamics are involved in apoptosis following stress signals [145, 146]. Two opposing processes that contribute to the radical morphological transformation of the mitochondria are fission and fusion. The fusion process caused tethering and joining two mitochondrial organelles, whereas fission includes cleavage and constrictions [147-149] (Figure 3). Accumulating precise evidence shown the mitochondrial dynamic state is

changing constantly, which involves dividing and elongating (i.e., mitochondrial fission and fusion, respectively). The balance of these two events has a key role in mitochondrial homeostasis, cell stability, and cell survival [150, 151]. Dysfunctional mitochondria contain impaired proteins, damaged membranes, and mutated or damaged mtDNA, leading to the segregation of mitochondria through the fission process [152-154]. In contrast, fusion contributes to the balance of matrix metabolites, and aid in the equilibration of mitochondrial membrane contents including complex I of the ETC [155-157]. A family of dynamin-related proteins (DRPs), especially Drp1, are reported to regulate fission and fusion [158, 159]. Drp1, a key regulator of fission, is recruited from the cytosol to the mitochondrial membrane, and interact with Drp1 receptors including mitochondrial dynamics proteins of 49 and 51 kDa (Mid 49 and Mid51), mitochondrial fission factor (Mff), as well as interact with mitochondrial fission protein (Fis1) to create the fission complex. Subsequently, dynamin 2 contributes with Drp1 to form a ring-like structure through oligomerization and split the mitochondrial membrane by GTP hydrolysis and self-assembly in the final step of fission [160-164]. Fission caused cleavage of mitochondria into several small parts just before apoptosis, and inhibiting mitochondrial fission can prevent releasing of the cytc and delay the apoptotic process [146]. It has been shown that Drp1 expression upregulated during mitochondrial oxidative stress, resulting in an imbalance of mitochondrial fission and fusion, which leads to mitochondria dysfunction and disintegrate, and cell death [165]. Drp1 expression and mitochondrial fragmentation can be reduced by antioxidants such as vitamin E or MitoQ [166, 167]. While oxidative stress and mitochondrial ROS production reduced after knockdown of Drp1 [167-169]. Drp1 has been reported to have an essential role in ischemic stroke, and infarct volume was reduced following Drp1 downregulation [170-172]. Three different GTPases mediate fusion, including optic atrophy 1 (Opa1) and Mitofusins 1 and 2 (Mfn1/2) [157]. Mfn1/2 and Opa1 mediates the fusion of the outer and inner mitochondrial membrane, respectively [173-175] (Figure 3). Mfn1/2 is anchored in the outer mitochondrial membrane, initiate the joining of two mitochondrial membranes by formation, and grows a hemifusion stalk [152, 176], which creates a lipidic hole as well as a hemifusion diaphragm to reestablish membrane continuity. Finally, a pore is made through the lipid-binding domain in Opa1 that is specific for cardiolipin [176, 177]. Fusion has been documented to augments

mitochondrial integrity in the normal condition [178], but deficits in the fusion may result in neurodegenerative disorders [179, 180]. Mitochondrial fusion proteins are less studied in ischemic stroke [181-184]. Mfn2 has been demonstrated to exert an anti-apoptotic effect, and its expression decreased in the hypoxic conditions [181, 182]. The Opa1 can attenuate brain edema in ischemic stroke, and its expression increased after exercise [183]. Cell survival and the pathobiology of stroke have essential connections with mitochondria dynamic. Fission and fusion, as mitochondrial dynamics processes, are crucial to the mitochondrial function. Fusion presuming that is appropriate to cell survival, but studies show that fission facilitates cell death [185]. Fission and fusion are also involved in inhibiting or progressing programmed cell death [186]. Fission can lead to degradation of mitochondria, and involved in the mitochondrial quality control signaling [187]. Furthermore, fission contributes to the renewal and redistribution of mitochondria [188]. Mitochondrial dynamics have a key role in cell fate; especially, fission of the mitochondria is an initial incident in apoptotic cell death following ischemic stroke [170-172]. During cerebral ischemia, mitochondrial function play a crucial role in cell destiny via production of ATP, which can trigger apoptotic signaling by releasing

of pro-apoptotic factors including AIF, cytc,

endonuclease G, mitochondrial serine protease HtrA2/Omi, and SMAC/ DIABLO [189, 190]. Therefore, precise quality control processes are essential to preserving a healthy mitochondrial network. Such processes contain mitochondrial dynamic and mitophagy [171]. 8. Mitophagy and ischemic stroke Mitochondria autophagy, termed as mitophagy, is a process that selected mitochondrial cargos are engulfed by mitochondrial-derived vesicles and delivered to the lysosome or peroxisome for degradation process, which is essential to the mitochondrial content and metabolism homeostasis. Mitophagy plays an important role in the cellular and organismal physiology and fine-tunes the mitochondrial homeostasis and biogenesis, as well as controls the number and quality of the mitochondrial organelle. Mitophagy reducing excessive or dysfunctional mitochondria that produced ROS and caused cell death [191]. Impairments in the mitophagy may be causing mitochondrial accumulation, increasing oxygen consumption, and excessive ROS formation, which finally resulting in the cellular degeneration and triggering of cell death cascades [192]. Several cascades may lead to mitophagy, including excessive ROS formation, MPTP opening, loss of ∆ψm, mitochondrial fission and fusion [193] (Figure 1). Studies showed

that MPTP is essential to the initiation of mitophagy [194], and involved in the selective removal of damaged mitochondria by autophagy [195]. Excessive ROS formation may cause depolarization of the mitochondrial, resulting in mitophagy, a process that has an important role in the ROS clearance by attenuating oxidative stress [196]. Mitophagy is a neuroprotective mechanism by removing ROS generation, and insufficient mitophagy (autophagy) leads to cell death. Mitophagy can regulate ROS generation by a negative feedback mechanism [197]. Following the stroke, ROS generation contributes to brain injury by inducing apoptosis or injury to the blood-brain barrier (BBB) [198]. It was reported that melatonin administration reduced ROS generation by increasing mitophagy, and inhibiting the NLRP3 inflammasome activation in a stroke rat model [199]. Besides, mitophagy was shown to reduced ROS accumulation in an OGD condition [200]. 9. Intercellular mitochondrial transfer and its application in ischemic stroke Mitochondrial organelle was considered to remain in the intracellular space for their lifetime. Recently, it has been evidenced that vesicles and organelles can be exchanged between cells through structures including tunneling nanotubes (TNT) or extracellular vesicles (EVs) [201, 202], and mitochondria are transported between cells via these intercellular quality control structures [203] (Figure 4). It has been demonstrated that healthy mitochondria transferred from adjacent cells using TNTs to rescues cells containing damaged mitochondria [204-209], and it can be bidirectional [210]. Mitochondrial rho GTPase 1 (MIRO1) has been reported to plays a pivotal role in the TNTs- dependent intercellular transport of mitochondria [211], and its knockdown prevents mitochondrial transport from MSCs (mesenchymal stem cells) to epithelial cells. While its overexpression leads to an increase in the transport of mitochondria [211]. It has been demonstrated that EVs can transport mitochondria between cells [212, 213]. EVs containing mitochondria released from the astrocytes, which rescue neurons from OGD or ischemic stroke conditions, and mediated by CD38- cADPR (cyclic ADP ribose)-Ca2+ signaling [214]. Triggering of CD38 and addition of cADPR was reported to increase the releasing of mitochondria from astrocytes. While mitochondria release from astrocyte reduced following knockdown of CD38 or BAPTA-AM (an intracellular Ca2+chelator) [214]. Intercellular transfer of mitochondria was reported in cardiovascular injury model, and also observed in stroke models [214-216]. The mitochondrial transfer was shown in the MSCs, which are the most popular for the transfer of mitochondria [212]. Additionally, the transfer of

mitochondria was reported in different cell types [206, 209, 214-219]. Mitochondrial transfer is a protective mechanism for salvage of damaged cells from mitochondrial dysfunction in response to the stress cascades [214, 220, 221]. It has been documented that MSCs repressed apoptosis in the endothelial cells following IR injury, which caused by the rescue of aerobic respiration by the transfer of mitochondria to damaged endothelial cells using TNT-like structures [216]. These studies confirm that the transfer of mitochondria triggers metabolic cross-talk among healthy and damaged cells, and is a promising and novel therapeutic approach for ischemic stroke [222]. Injection of astrocyte-derived mitochondria in the middle carotid artery occlusion (MCAO) animal model increased the level of phosphorylated AKT and BCL-XL, which promote cell survival [214]. In the MCAO rats, injected exogenous mitochondria were taken by neurons, astrocytes, and microglia, which improved mitochondrial function and increased motor performance [223]. It was demonstrated that functional extracellular mitochondria produce in the cultured astrocytes after OGD condition, which increased the ATP level and neural viabilities. Hence, mitochondrial transfer in the MCAO mice led to the triggering of cell-survival-related signals [214]. Following ischemia, neural cells were assumed to transfer mitochondria not only for disposal but also for recovering ATP level and quickly decreasing energy deficits caused by ischemic stroke [222]. Recently it was suggested that restoration of the imbalanced mitochondrial dynamics via mitochondrial transplantation might potentially be a way to attenuate stroke-induced neuronal death. Application of the mitochondrial transplantation depends on the source and quality of isolated mitochondria, a delivery protocol, and cellular uptake of supplemented mitochondria. [224]. 9.1. Mitochondrial transfer and stem cells Recently, apart from antioxidant and pharmacological strategies, stem cell therapy has also come up as an alternative for targeting mitochondria following ischemic stroke [225]. Several reports have shown utilizing stem cells as therapeutic agents in diseases where mitochondrial dysfunction plays an essential role [226-231]. It has been approved that mitochondrial transfer is possible from astrocytes into the adjacent neurons which increased cell survival following transient focal ischemia [214]. Therefore, stem cells can also be beneficial in a similar manner following transplantation into the injury site [232]. An intercellular mitochondrial transfer can restore mitochondrial function, amplified cellular survival signals and reprogram differentiated cells [209, 211, 212, 217, 233-235]. The first evidence for mitochondrial transfer was

demonstrated between human stem cells and mitochondria-depleted cells [212], and most of the reports have utilized stem cells as mitochondria donors [236]. Liu et al. provide evidence of mitochondrial transfer from MSCs to HUVE (human umbilical vein endothelial) cells subjected to IR injury in vitro, which restore the aerobic respiration in the cells. They also indicate that the injured cells expressed phosphatidylserine, activating MSCs towards the formation of TNTs [216]. Besides, MSCs repaired cellular damage and improved cell survival in cardiomyocytes by the formation of TNTs following OGD and reperfusion [237]. It was showed that MSCs engulfed and degraded damaged mitochondria, which trigger mitochondrial biogenesis and the stimulation of haeme-oxygenase-1 (HO-1) as a cytoprotective enzyme, thereby improving the ability of MSCs to donate mitochondria to the damaged cells for overcoming oxidative stress [238]. Mitochondrial transfer contributed to tissue repair, mainly by enhancing function and improving cellular bioenergetics [209]. It was demonstrated that connexins regulated mitochondrial transfer between the stem cells and alveolar cells by the formation of nanotubes and vesicles [209]. MSCs were reported to increase mitochondrial transfer and improved expression of Miro1, which attenuates neurological deficit following stroke. Moreover, mitochondria transfer through TNTs observed in MSCs in the OGD model of ischemia [239]. Transfer of mitochondria confronts important questions, which regarding the degree of cellular injuries that are required for initiating mitochondria transfer, and the mechanism by which neighbor cells sense the stress of the damaged cells to trigger mitochondrial transfer [240]. Additional studies and experiments are essential to finding these challenges. 10. Conclusions and perspectives Mitochondrial dysfunction associated with the pathology of ischemic stroke and targeting mitochondria by new therapeutic strategies may be used to attenuate the devastating consequences of ischemic stroke. This raises the opportunity of treating cerebral ischemia with novel mitochondria-targeted therapies. An interesting prospect is raised by the current knowledge that mitochondria contribute to the pathology of ischemic stroke through elevated ROS formation, oxidative stress, calcium accumulation, MPTP induction, controlling cell death and inflammation, and regulating NLRP3 inflammasome activation [105, 241]. Mitochondrial dysfunction is an initial event following ischemic stroke [242]. Defected bioenergetics, aberrant mitochondrial morphology and structure, and abnormal mitochondrial dynamics play a crucial role in triggering cell death [243].

Preclinical studies showed that targeting mitochondrial quality control and mitochondrial dynamics via pharmacological factors or genetic interventions have been neuroprotective [244, 245]. Mitochondrial targeting by pharmacological agents is still challenging in the clinic. Currently, the reports of mitochondrial transfer exposed a completely new perspective in intercellular communication. Emerging findings show that different extracellular stimuli can initiate “help-me” signaling in the injured mitochondria, which cause the recruitment of the adjacent cells to rescue the impaired cells. Recent therapeutic approaches to treat hypoxia/ischemia related diseases are replenishing healthy mitochondria and eliminating the injured mitochondria, which are especially important in the CNS that mitochondrial organelles are plenty in the neuronal synapses and dendrites [246, 247]. Currently, stem cells have shown the ability to protect mitochondria in numerous pre-clinical settings. Stem cells are thought to transfer mitochondria to the injured cells via

TNTs,

extracellular vesicles or simply by cellular fusion, which helps to revive cell energetics in the recipient cell [225]. Recently, in the patients who suffered from myocardial IR injury, mitochondrial transplantation has been performed successfully, and researchers should focus on utilizing stem cells for the therapy of ischemic stroke. However, challenges remain to implement mitochondrial transfer in clinics [248]. Understanding the main mechanisms of the mitochondrial release from donor cells and their recognition by recipient cells needs to be investigated in future studies.

Conflict of interest The authors declare that the research was conducted in the absence of any commercial/ noncommercial

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construed

as

Table. Drugs and Chemical agents targeting mitochondrial function. Agents

Mechanisms

CsA

Binding to the matrix protein CYD and prevents cell death [249] caused by formation of the MPTP

S1QELs and S3QELs

Directly binds to respiratory chain complexes I and III, [250, 251] respectively, and inhibits ROS production

DNP

Decreasing mitochondrial ROS production

[252]

NAM, NR and NMN

Attenuates bioenergetic defects by replenishing NAD+ levels

[253-260]

Drp1 siRNA

Inhibition of fission via DRP1 down-regulation

[171, 261263]

Ginkgolide K

Inhibition of fission by increasing Drp1 phosphorylation and [264] inhibiting Drp1 recruitment

mdivi-1

Inhibition of fission by inhibiting the GTPase activity of Drp1

P110

References

[261, 263]

Inhibition of fission by inhibiting Drp1 enzyme activity and [265, 266] blocking Drp1/ Fis1 interaction CsA; cyclosporin A , CYD; cyclophilin D, MPTP; mitochondrial permeability transition pore, ROS ; reactive oxygen species, S1QELs; suppressors of site IQ electron leak, S3QELs; suppressors of site IIIQo electron leak, DNP; dinitrophenol, NAM; nicotinamide, NR; NAM riboside, NMN; nicotinamide mononucleotide, DRP1; dynamin-related protein 1

Figure 1. Mitochondria play a central role in pathological conditions following ischemic stroke. During ischemia, OGD leads to the ATP depletion and failure of Na+/K+ ATPase pump, which results in the neuronal membrane depolarization and excessive releases of glutamate [5]. Glutamate receptors including NMDA and AMPA are over-activated, which leads to the influx and accumulation of calcium ions into the cells. Excessive Ca2+ influx leads ROS production and mitochondrial

dysfunction,

which

trigger

several

pathological

processes

including

mitochondrial-induced apoptosis, mitochondrion-dependent fission and fusion, and mitophagy. Furthermore, ischemia triggers the depolarization of mitochondrial membrane potential (∆Ψm), reduction of ATP production, and induction of the MPTP, which these cellular processes eventually leading to neuronal death [222, 267]. Several elements can trigger MPTP opening and mitochondrial injury, including Ca2+ accumulation, production of ROS and RNS and loss of mitochondrial membrane potential [60, 61]. MPTP induction leads to the release of succinate and mitochondrial DAMPs, resulting in the activation of an inflammatory response that will contribute to tissue damage following IR injury [53]. Hence, MPTP is essential to the initiation of mitophagy [194]. Mitophagy can regulate ROS generation by a negative feedback mechanism [197]. Following ischemic insults, decreasing of the oxygen availability induced mitochondrial anaerobic glycolysis and excessive production of lactic acid and acidosis, which subsequently activates NLRP3 inflammasome. Furthermore, failing of the mitochondrial ETC increased ROS formation in the cytoplasm and activation of PKR and TXNIP, which triggers NLRP3 inflammasome.

NMDA;

N-methyl-D-aspartate,

AMPA;

α-amino-3-hydroxy-5-methyl-4-

isoxazole-propionic acid, DAMPs; mitochondrial damage-associated molecular patterns, MAPK; mitogen- activated protein kinase, AP-1; activator protein-1, NLRP; NOD (nucleotide-binding oligomerization domain)-like receptor (NLR) Pyrin domain containing 3), ASC; apoptosisassociated speck-like protein containing a caspase recruitment domain, ASIC; acid-sensing ion channel, ATP; adenosine triphosphate, ROS; reactive oxygen species, RNS; reactive nitrogen species, TXNIP; thioredoxin-interacting protein, PKR; protein kinase R, ∆ψ; membrane potential, MPTP; mitochondrial permeability transition pore. Figure 2. Mechanism of mitochondria-mediated signaling in NLRP3 inflammasome activity. Mitochondria are central regulators of NLRP3 inflammasome activation. Mitochondrial ROS, Ca2+ overload, reduced NAD+, cardiolipin, mitofusin, and mtDNA have all been presented to trigger NLRP3 inflammasome activation [105]. These stimuli trigger the NLRP3

inflammasome complex by facilitating the oligomerization of NLRP3, ASC, and procaspase-1. This complex is responsible for activating procaspase-1 into cleaved caspase-1, which cleaves pro-IL-1β and pro-IL-18 into their active forms and also provokes an inflammatory form of cell death that is referred to as pyroptosis [97]. Parkin/PINK1 dependent mitophagy is involved in the clearance of dysfunctional mitochondria, and thus negatively regulates NLRP3 inflammasome activation [268]. ROS; reactive oxygen species, NAD; nicotinamide adenine dinucleotide, mtDNA; mitochondrial DNA, PINK1; PTEN induced putative kinase 1, LC3; microtubuleassociated protein 1A/1B light chain 3. IL; interleukin, Pre; Precursor. Figure 3. Schematic representation of mitochondrial dynamics. A) Fusion is triggered through the Mfn1/2 and Opa1, which mediates fusion of the outer and inner mitochondrial membrane, respectively. Mfn1/2 are anchored to the outer membrane of the mitochondria and interact with each other to form a hemifusion stalk, which grows into a lipidic hole and finally reestablishes membrane continuity. Inner membrane fusion is mediated by heterotypic interaction of Opa1 and cardiolipin [269, 270]. B) During fission, Drp1 is recruited from the cytosol to the outer membrane of the mitochondria and interacts with its receptor proteins and Fis1 to create the fission complex in the outer membrane. Drp1 is then oligomerized into filaments that wrap around the mitochondria, leading to constriction of mitochondria and sequential separation of the inner and outer membrane [269, 270].

Drp; dynamin-related

proteins, Mfn1/2; mitofusins, Opa1; optic atrophy 1 Figure 4. Schematic of intercellular mitochondrial transfer and quality control of mitochondria. Mitochondria are transported between cells by the actin-based tunneling nanotubes between donor cells and injured neurons, and extracellular vesicles that released from healthy neurons (or donor cells) and then internalized into injured neurons. Mitochondrial transfer mechanism help to rescue cells containing damaged mitochondria by transporting healthy mitochondria from the adjacent cells [203, 224]. Furthermore, this schematic represents intracellular quality control of mitochondria to maintain functions of mitochondria and consists of fusion, fission, and degradation (mitophagy). Dysfunctional and injured mitochondria are dissociated by fission. Drp1 translocated from the cytosol to the outer mitochondrial membrane and drives dynamin-like scission of mitochondria. Subsequently, two unequal mitochondria are born, which dysfunctional mitochondria are degraded (mitophagy) by autophagosome, and

healthy mitochondria continue to fuse to preserve the network which is organized by Mfn1/2 and Opa1, separately participating in outer and inner mitochondrial membrane fusion. Mitochondrial fusion prevents the accumulation of damaged contents in a single mitochondrion through the exchange of mtDNA, proteins, and metabolites between healthy and damaged mitochondria [203, 271]. mtDNA; mitochondrial DNA, Drp1; dynamin-related protein-1, Mfn1/2; Mitofusin1/2, Opa1; optic atrophy 1.

References [1] V.L. Feigin, B. Norrving, G.A. Mensah, Global burden of stroke, Circulation research 120(3) (2017) 439-448. [2] H.-L. Huang, C.-C. Lin, K.-C.G. Jeng, P.-W. Yao, L.-T. Chuang, S.-L. Kuo, C.-W. Hou, Fresh green tea and gallic acid ameliorate oxidative stress in kainic acid-induced status epilepticus, Journal of agricultural and food chemistry 60(9) (2012) 2328-2336. [3] O. Karimi-Khouzani, E. Heidarian, S.A. Amini, Anti-inflammatory and ameliorative effects of gallic acid on fluoxetine-induced oxidative stress and liver damage in rats, Pharmacological Reports 69(4) (2017) 830-835. [4] P.A. Lapchak, J.H. Zhang, The high cost of stroke and stroke cytoprotection research, Translational stroke research 8(4) (2017) 307-317. [5] S.E. Khoshnam, W. Winlow, M. Farzaneh, Y. Farbood, H.F. Moghaddam, Pathogenic mechanisms following ischemic stroke, Neurological Sciences 38(7) (2017) 1167-1186. [6] D. Keizman, P. Huang, M.A. Eisenberger, R. Pili, J.J. Kim, E.S. Antonarakis, H. Hammers, M.A. Carducci, Angiotensin system inhibitors and outcome of sunitinib treatment in patients with metastatic renal cell carcinoma: a retrospective examination, European journal of cancer 47(13) (2011) 1955-1961. [7] A.P. Jadhav, T.G. Jovin, Endovascular therapy for acute ischemic stroke: The standard of care, Brain Circulation 2(4) (2016) 178. [8] S. Yoshimura, N. Sakai, K. Uchida, H. Yamagami, M. Ezura, Y. Okada, K. Kitagawa, K. Kimura, M. Sasaki, N. Tanahashi, Endovascular Therapy in Ischemic Stroke With Acute Large-Vessel Occlusion: Recovery by Endovascular Salvage for Cerebral Ultra-Acute Embolism Japan Registry 2, Journal of the American Heart Association 7(9) (2018) e008796. [9] B. Diop-Frimpong, V.P. Chauhan, S. Krane, Y. Boucher, R.K. Jain, Losartan inhibits collagen I synthesis and improves the distribution and efficacy of nanotherapeutics in tumors, Proceedings of the National Academy of Sciences 108(7) (2011) 2909-2914. [10] S.E. Khoshnam, Y. Farbood, H.F. Moghaddam, A. Sarkaki, M. Badavi, L. Khorsandi, Vanillic acid attenuates cerebral hyperemia, blood-brain barrier disruption and anxiety-like behaviors in rats following transient bilateral common carotid occlusion and reperfusion, Metabolic Brain Disease (2018) 1-9. [11] S.E. Khoshnam, A. Sarkaki, L. Khorsandi, W. Winlow, M. Badavi, H.F. Moghaddam, Y. Farbooda, Vanillic acid attenuates effects of transient bilateral common carotid occlusion and reperfusion in rats, Biomedicine & Pharmacotherapy 96 (2017) 667-674. [12] H. Van der Worp, L. Kappelle, A. Algra, P. Bär, J. Orgogozo, E. Ringelstein, P. Bath, J. Van Gijn, TESS, T.I. Investigators, The effect of tirilazad mesylate on infarct volume of patients with acute ischemic stroke, Neurology 58(1) (2002) 133-135. [13] S.E. Khoshnam, A. Sarkaki, M. Rashno, Y. Farbood, Memory deficits and hippocampal inflammation in cerebral hypoperfusion and reperfusion in male rats: neuroprotective role of vanillic acid, Life sciences 211 (2018) 126-132. [14] C. Xing, K. Hayakawa, E.H. Lo, Mechanisms, imaging, and therapy in stroke recovery, Springer, 2017. [15] S.-H. Yang, M. Lou, B. Luo, W.-J. Jiang, R. Liu, Precision medicine for ischemic stroke, let us move beyond time is brain, Translational stroke research 9(2) (2018) 93-95. [16] P.S. Vosler, S.H. Graham, L.R. Wechsler, J. Chen, Mitochondrial targets for stroke: focusing basic science research toward development of clinically translatable therapeutics, Stroke 40(9) (2009) 31493155. [17] J.-l. Huang, A. Manaenko, Z.-h. Ye, X.-j. Sun, Q. Hu, Hypoxia therapy--a new hope for the treatment of mitochondrial dysfunctions, Medical gas research 6(3) (2016) 174.

[18] S.E. Khoshnam, W. Winlow, M. Farzaneh, The interplay of MicroRNAs in the inflammatory mechanisms following ischemic stroke, Journal of Neuropatholgy & Experimental Neurology 76(7) (2017) 548-561. [19] S.E. Khoshnam, W. Winlow, Y. Farbood, H.F. Moghaddam, M. Farzaneh, Emerging roles of microRNAs in ischemic stroke: as possible therapeutic agents, Journal of stroke 19(2) (2017) 166. [20] E. Heydari, M. Alishahi, F. Ghaedrahmati, W. Winlow, S.E. Khoshnam, A. Anbiyaiee, The role of noncoding RNAs in neuroprotection and angiogenesis following ischemic stroke, Metabolic brain disease (2019) 1-13. [21] M. Alishahi, F. Ghaedrahmati, T.A. Kolagar, W. Winlow, N. Nikkar, M. Farzaneh, S.E. Khoshnam, Long non-coding RNAs and cell death following ischemic stroke, Metabolic brain disease (2019) 1-9. [22] J. Hofmeijer, M.J. van Putten, Ischemic cerebral damage: an appraisal of synaptic failure, Stroke 43(2) (2012) 607-615. [23] J.-M. Lee, M.C. Grabb, G.J. Zipfel, D.W. Choi, Brain tissue responses to ischemia, The Journal of clinical investigation 106(6) (2000) 723-731. [24] P.A. Dharmasaroja, Fluid intake related to brain edema in acute middle cerebral artery infarction, Translational stroke research 7(1) (2016) 49-53. [25] H.B. Van Der Worp, M.R. Macleod, R. Kollmar, E.S.R.N.f. Hypothermia, Therapeutic hypothermia for acute ischemic stroke: ready to start large randomized trials?, Journal of Cerebral Blood Flow & Metabolism 30(6) (2010) 1079-1093. [26] P.-y. Li, X. Wang, R.A. Stetler, J. Chen, W.-f. Yu, Anti-inflammatory signaling: the point of convergence for medical gases in neuroprotection against ischemic stroke, Medical gas research 6(4) (2016) 227. [27] T. Luo, Y. Park, X. Sun, C. Liu, B. Hu, Protein misfolding, aggregation, and autophagy after brain ischemia, Translational stroke research 4(6) (2013) 581-588. [28] H. Ahnstedt, J. Sweet, P. Cruden, N. Bishop, M.J. Cipolla, Effects of early post-ischemic reperfusion and tPA on cerebrovascular function and nitrosative stress in female rats, Translational stroke research 7(3) (2016) 228-238. [29] J. Astrup, B.K. Siesjö, L. Symon, Thresholds in cerebral ischemia-the ischemic penumbra, Stroke 12(6) (1981) 723-725. [30] S. Jung, M. Gilgen, J. Slotboom, M. El-Koussy, C. Zubler, C. Kiefer, R. Luedi, M.-L. Mono, M.R. Heldner, A. Weck, Factors that determine penumbral tissue loss in acute ischaemic stroke, Brain 136(12) (2013) 3554-3560. [31] A. Pisani, P. Bonsi, P. Calabresi, Calcium signaling and neuronal vulnerability to ischemia in the striatum, Cell Calcium 36(3-4) (2004) 277-284. [32] V. Nikoletopoulou, M. Markaki, K. Palikaras, N. Tavernarakis, Crosstalk between apoptosis, necrosis and autophagy, Biochimica et Biophysica Acta (BBA)-Molecular Cell Research 1833(12) (2013) 34483459. [33] H. Wei, Y. Li, S. Han, S. Liu, N. Zhang, L. Zhao, S. Li, J. Li, cPKCγ-modulated autophagy in neurons alleviates ischemic injury in brain of mice with ischemic stroke through Akt-mTOR pathway, Translational stroke research 7(6) (2016) 497-511. [34] L. Galluzzi, O. Kepp, G. Kroemer, Mitochondria: master regulators of danger signalling, Nature reviews Molecular cell biology 13(12) (2012) 780. [35] A. Ames III, CNS energy metabolism as related to function, Brain research reviews 34(1-2) (2000) 4268. [36] J.L. Franklin, Redox regulation of the intrinsic pathway in neuronal apoptosis, Antioxidants & redox signaling 14(8) (2011) 1437-1448.

[37] K. Niizuma, H. Yoshioka, H. Chen, G.S. Kim, J.E. Jung, M. Katsu, N. Okami, P.H. Chan, Mitochondrial and apoptotic neuronal death signaling pathways in cerebral ischemia, Biochimica et Biophysica Acta (BBA)-Molecular Basis of Disease 1802(1) (2010) 92-99. [38] M.T. Lin, M.F. Beal, Mitochondrial dysfunction and oxidative stress in neurodegenerative diseases, Nature 443(7113) (2006) 787. [39] N.R. Sims, H. Muyderman, Mitochondria, oxidative metabolism and cell death in stroke, Biochimica et Biophysica Acta (BBA)-Molecular Basis of Disease 1802(1) (2010) 80-91. [40] D.P. Narendra, S.M. Jin, A. Tanaka, D.-F. Suen, C.A. Gautier, J. Shen, M.R. Cookson, R.J. Youle, PINK1 is selectively stabilized on impaired mitochondria to activate Parkin, PLoS biology 8(1) (2010) e1000298. [41] T.H. Sanderson, C.A. Reynolds, R. Kumar, K. Przyklenk, M. Hüttemann, Molecular mechanisms of ischemia–reperfusion injury in brain: pivotal role of the mitochondrial membrane potential in reactive oxygen species generation, Molecular neurobiology 47(1) (2013) 9-23. [42] E.T. Chouchani, V.R. Pell, A.M. James, L.M. Work, K. Saeb-Parsy, C. Frezza, T. Krieg, M.P. Murphy, A unifying mechanism for mitochondrial superoxide production during ischemia-reperfusion injury, Cell metabolism 23(2) (2016) 254-263. [43] E.J. Lesnefsky, Q. Chen, B. Tandler, C.L. Hoppel, Mitochondrial dysfunction and myocardial ischemiareperfusion: implications for novel therapies, Annual review of pharmacology and toxicology 57 (2017) 535-565. [44] R. Jennings, H.K. Hawkins, J.E. Lowe, M.L. Hill, S. Klotman, K.A. Reimer, Relation between high energy phosphate and lethal injury in myocardial ischemia in the dog, The American journal of pathology 92(1) (1978) 187. [45] O. Adeoye, R. Hornung, P. Khatri, D. Kleindorfer, Recombinant tissue-type plasminogen activator use for ischemic stroke in the United States: a doubling of treatment rates over the course of 5 years, Stroke 42(7) (2011) 1952-1955. [46] O.O. Zaidat, A.J. Yoo, P. Khatri, T.A. Tomsick, R. Von Kummer, J.L. Saver, M.P. Marks, S. Prabhakaran, D.F. Kallmes, B.-F.M. Fitzsimmons, Recommendations on angiographic revascularization grading standards for acute ischemic stroke: a consensus statement, Stroke 44(9) (2013) 2650-2663. [47] T.M. Dawson, V.L. Dawson, Mitochondrial mechanisms of neuronal cell death: potential therapeutics, Annual review of pharmacology and toxicology 57 (2017) 437-454. [48] N. Sun, R.J. Youle, T. Finkel, The mitochondrial basis of aging, Molecular cell 61(5) (2016) 654-666. [49] H. Chen, H. Yoshioka, G.S. Kim, J.E. Jung, N. Okami, H. Sakata, C.M. Maier, P. Narasimhan, C.E. Goeders, P.H. Chan, Oxidative stress in ischemic brain damage: mechanisms of cell death and potential molecular targets for neuroprotection, Antioxidants & redox signaling 14(8) (2011) 1505-1517. [50] A.C. Schinzel, O. Takeuchi, Z. Huang, J.K. Fisher, Z. Zhou, J. Rubens, C. Hetz, N.N. Danial, M.A. Moskowitz, S.J. Korsmeyer, Cyclophilin D is a component of mitochondrial permeability transition and mediates neuronal cell death after focal cerebral ischemia, Proceedings of the National Academy of Sciences 102(34) (2005) 12005-12010. [51] T. Nakagawa, S. Shimizu, T. Watanabe, O. Yamaguchi, K. Otsu, H. Yamagata, H. Inohara, T. Kubo, Y. Tsujimoto, Cyclophilin D-dependent mitochondrial permeability transition regulates some necrotic but not apoptotic cell death, Nature 434(7033) (2005) 652. [52] C.P. Baines, R.A. Kaiser, N.H. Purcell, N.S. Blair, H. Osinska, M.A. Hambleton, E.W. Brunskill, M.R. Sayen, R.A. Gottlieb, G.W. Dorn, Loss of cyclophilin D reveals a critical role for mitochondrial permeability transition in cell death, Nature 434(7033) (2005) 658. [53] J. Lutz, K. Thürmel, U. Heemann, Anti-inflammatory treatment strategies for ischemia/reperfusion injury in transplantation, Journal of Inflammation 7(1) (2010) 27. [54] E.T. Chouchani, T.R. Hurd, S.M. Nadtochiy, P.S. Brookes, I.M. Fearnley, K.S. Lilley, R.A. Smith, M.P. Murphy, Identification of S-nitrosated mitochondrial proteins by S-nitrosothiol difference in gel

electrophoresis (SNO-DIGE): implications for the regulation of mitochondrial function by reversible Snitrosation, Biochemical Journal 430(1) (2010) 49-59. [55] T.A. Prime, F.H. Blaikie, C. Evans, S.M. Nadtochiy, A.M. James, C.C. Dahm, D.A. Vitturi, R.P. Patel, C.R. Hiley, I. Abakumova, A mitochondria-targeted S-nitrosothiol modulates respiration, nitrosates thiols, and protects against ischemia-reperfusion injury, Proceedings of the National Academy of Sciences 106(26) (2009) 10764-10769. [56] E.T. Chouchani, C. Methner, S.M. Nadtochiy, A. Logan, V.R. Pell, S. Ding, A.M. James, H.M. Cochemé, J. Reinhold, K.S. Lilley, Cardioprotection by S-nitrosation of a cysteine switch on mitochondrial complex I, Nature medicine 19(6) (2013) 753. [57] N.S. Dhalla, A.B. Elmoselhi, T. Hata, N. Makino, Status of myocardial antioxidants in ischemia– reperfusion injury, Cardiovascular research 47(3) (2000) 446-456. [58] A. Rasola, P. Bernardi, The mitochondrial permeability transition pore and its involvement in cell death and in disease pathogenesis, Apoptosis 12(5) (2007) 815-833. [59] P. Varanyuwatana, A.P. Halestrap, The roles of phosphate and the phosphate carrier in the mitochondrial permeability transition pore, Mitochondrion 12(1) (2012) 120-125. [60] M. Picard, K. Csukly, M.-E. Robillard, R. Godin, A. Ascah, C. Bourcier-Lucas, Y. Burelle, Resistance to Ca2+-induced opening of the permeability transition pore differs in mitochondria from glycolytic and oxidative muscles, American Journal of Physiology-Regulatory, Integrative and Comparative Physiology 295(2) (2008) R659-R668. [61] A.P. Halestrap, G.P. McStay, S.J. Clarke, The permeability transition pore complex: another view, Biochimie 84(2-3) (2002) 153-166. [62] C.P. Baines, The mitochondrial permeability transition pore and ischemia-reperfusion injury, Basic research in cardiology 104(2) (2009) 181-188. [63] S. Lennon, S. Martin, T. Cotter, Dose-dependent induction of apoptosis in human tumour cell lines by widely diverging stimuli, Cell proliferation 24(2) (1991) 203-214. [64] S.-D. Chen, H.-Y. Wu, D.-I. Yang, S.-Y. Lee, F.-Z. Shaw, T.-K. Lin, C.-W. Liou, Y.-C. Chuang, Effects of rosiglitazone on global ischemia-induced hippocampal injury and expression of mitochondrial uncoupling protein 2, Biochemical and biophysical research communications 351(1) (2006) 198-203. [65] E. Novo, M. Parola, Redox mechanisms in hepatic chronic wound healing and fibrogenesis, Fibrogenesis & tissue repair 1(1) (2008) 5. [66] A. Kunz, L. Park, T. Abe, E.F. Gallo, J. Anrather, P. Zhou, C. Iadecola, Neurovascular protection by ischemic tolerance: role of nitric oxide and reactive oxygen species, Journal of Neuroscience 27(27) (2007) 7083-7093. [67] S.-D. Chen, D.-I. Yang, T.-K. Lin, F.-Z. Shaw, C.-W. Liou, Y.-C. Chuang, Roles of oxidative stress, apoptosis, PGC-1α and mitochondrial biogenesis in cerebral ischemia, International journal of molecular sciences 12(10) (2011) 7199-7215. [68] T. Kalogeris, Y. Bao, R.J. Korthuis, Mitochondrial reactive oxygen species: a double edged sword in ischemia/reperfusion vs preconditioning, Redox biology 2 (2014) 702-714. [69] H. Endo, H. Kamada, C. Nito, T. Nishi, P.H. Chan, Mitochondrial translocation of p53 mediates release of cytochrome c and hippocampal CA1 neuronal death after transient global cerebral ischemia in rats, Journal of Neuroscience 26(30) (2006) 7974-7983. [70] M. Fang, J. Li, S. Tiu, L. Zhang, M. Wang, D.T. Yew, N-methyl-D-aspartate receptor and apoptosis in Alzheimer's disease and multiinfarct dementia, Journal of neuroscience research 81(2) (2005) 269-274. [71] B.I. Lee, P.H. Chan, G.W. Kim, Metalloporphyrin-based superoxide dismutase mimic attenuates the nuclear translocation of apoptosis-inducing factor and the subsequent DNA fragmentation after permanent focal cerebral ischemia in mice, Stroke 36(12) (2005) 2712-2717.

[72] T. Sairanen, R. Szepesi, M.-L. Karjalainen-Lindsberg, J. Saksi, A. Paetau, P.J. Lindsberg, Neuronal caspase-3 and PARP-1 correlate differentially with apoptosis and necrosis in ischemic human stroke, Acta neuropathologica 118(4) (2009) 541-552. [73] J.P. MacMANUS, A.M. Buchan, Apoptosis after experimental stroke: fact or fashion?, Journal of neurotrauma 17(10) (2000) 899-914. [74] H. Zhao, G.K. Steinberg, R.M. Sapolsky, General versus specific actions of mild-moderate hypothermia in attenuating cerebral ischemic damage, Journal of Cerebral Blood Flow & Metabolism 27(12) (2007) 1879-1894. [75] M. Leist, B. Single, A.F. Castoldi, S. Kühnle, P. Nicotera, Intracellular adenosine triphosphate (ATP) concentration: a switch in the decision between apoptosis and necrosis, Journal of Experimental Medicine 185(8) (1997) 1481-1486. [76] D.R. Green, G. Kroemer, The pathophysiology of mitochondrial cell death, Science 305(5684) (2004) 626-629. [77] G. Kroemer, Mitochondrial control of apoptosis: an overview, Biochemical Society Symposia, Portland Press Limited, 1999, pp. 1-15. [78] T. Kitayama, K. Ogita, Y. Yoneda, Sustained potentiation of AP1 DNA binding is not always associated with neuronal death following systemic administration of kainic acid in murine hippocampus, Neurochemistry international 35(6) (1999) 453-462. [79] D. Liu, C. Lu, R. Wan, W.W. Auyeung, M.P. Mattson, Activation of mitochondrial ATP-dependent potassium channels protects neurons against ischemia-induced death by a mechanism involving suppression of Bax translocation and cytochrome c release, Journal of Cerebral Blood Flow & Metabolism 22(4) (2002) 431-443. [80] A. Rami, I. Bechmann, J.H. Stehle, Exploiting endogenous anti-apoptotic proteins for novel therapeutic strategies in cerebral ischemia, Progress in neurobiology 85(3) (2008) 273-296. [81] Y.S. Song, Y.-S. Lee, P. Narasimhan, P.H. Chan, Reduced oxidative stress promotes NF-κB-mediated neuroprotective gene expression after transient focal cerebral ischemia: lymphocytotrophic cytokines and antiapoptotic factors, Journal of Cerebral Blood Flow & Metabolism 27(4) (2007) 764-775. [82] H. Zhao, M.A. Yenari, D. Cheng, R.M. Sapolsky, G.K. Steinberg, Bcl-2 overexpression protects against neuron loss within the ischemic margin following experimental stroke and inhibits cytochrome c translocation and caspase-3 activity, Journal of neurochemistry 85(4) (2003) 1026-1036. [83] D.T. Chao, S.J. Korsmeyer, BCL-2 family: regulators of cell death, Annual review of immunology 16(1) (1998) 395-419. [84] T. Engel, N. Plesnila, J.H. Prehn, D.C. Henshall, In vivo contributions of BH3-only proteins to neuronal death following seizures, ischemia, and traumatic brain injury, Journal of Cerebral Blood Flow & Metabolism 31(5) (2011) 1196-1210. [85] I. Inta, S. Paxian, I. Maegele, W. Zhang, M. Pizzi, P. Spano, I. Sarnico, S. Muhammad, O. Herrmann, D. Inta, Bim and Noxa are candidates to mediate the deleterious effect of the NF-κB subunit RelA in cerebral ischemia, Journal of Neuroscience 26(50) (2006) 12896-12903. [86] K.A. Webster, R.M. Graham, J.W. Thompson, M.-G. Spiga, D.P. Frazier, A. Wilson, N.H. Bishopric, Redox stress and the contributions of BH3-only proteins to infarction, Antioxidants & redox signaling 8(9-10) (2006) 1667-1676. [87] X. Saelens, N. Festjens, L.V. Walle, M. Van Gurp, G. Van Loo, P. Vandenabeele, Toxic proteins released from mitochondria in cell death, Oncogene 23(16) (2004) 2861. [88] M. Crompton, Mitochondrial intermembrane junctional complexes and their role in cell death, The Journal of physiology 529(1) (2000) 11-21. [89] P. Li, D. Nijhawan, I. Budihardjo, S.M. Srinivasula, M. Ahmad, E.S. Alnemri, X. Wang, Cytochrome c and dATP-dependent formation of Apaf-1/caspase-9 complex initiates an apoptotic protease cascade, Cell 91(4) (1997) 479-489.

[90] H. Yoshida, Y.-Y. Kong, R. Yoshida, A.J. Elia, A. Hakem, R. Hakem, J.M. Penninger, T.W. Mak, Apaf1 is required for mitochondrial pathways of apoptosis and brain development, Cell 94(6) (1998) 739-750. [91] H. Zou, W.J. Henzel, X. Liu, A. Lutschg, X. Wang, Apaf-1, a human protein homologous to C. elegans CED-4, participates in cytochrome c–dependent activation of caspase-3, Cell 90(3) (1997) 405-413. [92] I. Ferrer, B. Friguls, E. Dalfo, C. Justicia, A. Planas, Caspase-dependent and caspase-independent signalling of apoptosis in the penumbra following middle cerebral artery occlusion in the adult rat, Neuropathology and applied neurobiology 29(5) (2003) 472-481. [93] A. Saito, T. Hayashi, S. Okuno, M. Ferrand-Drake, P.H. Chan, Interaction between XIAP and Smac/DIABLO in the mouse brain after transient focal cerebral ischemia, Journal of Cerebral Blood Flow & Metabolism 23(9) (2003) 1010-1019. [94] S.A. Susin, H.K. Lorenzo, N. Zamzami, I. Marzo, B.E. Snow, G.M. Brothers, J. Mangion, E. Jacotot, P. Costantini, M. Loeffler, Molecular characterization of mitochondrial apoptosis-inducing factor, Nature 397(6718) (1999) 441. [95] C. Culmsee, C. Zhu, S. Landshamer, B. Becattini, E. Wagner, M. Pellecchia, K. Blomgren, N. Plesnila, Apoptosis-inducing factor triggered by poly (ADP-ribose) polymerase and Bid mediates neuronal cell death after oxygen-glucose deprivation and focal cerebral ischemia, Journal of Neuroscience 25(44) (2005) 10262-10272. [96] N. Plesnila, C. Zhu, C. Culmsee, M. Gröger, M.A. Moskowitz, K. Blomgren, Nuclear translocation of apoptosis-inducing factor after focal cerebral ischemia, Journal of Cerebral Blood Flow & Metabolism 24(4) (2004) 458-466. [97] M. Alishahi, M. Farzaneh, F. Ghaedrahmati, A. Nejabatdoust, A. Sarkaki, S.E. Khoshnam, NLRP3 inflammasome in ischemic stroke: as possible therapeutic target, International Journal of Stroke (2019) 1747493019841242. [98] V. Hornung, E. Latz, Critical functions of priming and lysosomal damage for NLRP3 activation, European journal of immunology 40(3) (2010) 620-3. [99] L. Agostini, F. Martinon, K. Burns, M.F. McDermott, P.N. Hawkins, J. Tschopp, NALP3 forms an IL-1βprocessing inflammasome with increased activity in Muckle-Wells autoinflammatory disorder, Immunity 20(3) (2004) 319-325. [100] F. Martinon, K. Burns, J. Tschopp, The inflammasome: a molecular platform triggering activation of inflammatory caspases and processing of proIL-β, Molecular cell 10(2) (2002) 417-426. [101] F. Bauernfeind, A. Ablasser, E. Bartok, S. Kim, J. Schmid-Burgk, T. Cavlar, V. Hornung, Inflammasomes: current understanding and open questions, Cellular and Molecular Life Sciences 68(5) (2011) 765-783. [102] K. Schroder, J. Tschopp, The inflammasomes, Cell 140(6) (2010) 821-32. [103] M. Lamkanfi, V.M. Dixit, Manipulation of host cell death pathways during microbial infections, Cell host & microbe 8(1) (2010) 44-54. [104] D.Y.-W. Fann, S. Lee, S. Manzanero, S. Tang, M. Gelderblom, P. Chunduri, C. Bernreuther, M. Glatzel, Y. Cheng, J. Thundyil, Intravenous immunoglobulin suppresses NLRP1 and NLRP3 inflammasome-mediated neuronal death in ischemic stroke, Cell death & disease 4(9) (2013) e790. [105] P. Gurung, J.R. Lukens, T.-D. Kanneganti, Mitochondria: diversity in the regulation of the NLRP3 inflammasome, Trends in molecular medicine 21(3) (2015) 193-201. [106] Q. Liu, D. Zhang, D. Hu, X. Zhou, Y. Zhou, The role of mitochondria in NLRP3 inflammasome activation, Molecular immunology 103 (2018) 115-124. [107] Z. Gong, J. Pan, Q. Shen, M. Li, Y. Peng, Mitochondrial dysfunction induces NLRP3 inflammasome activation during cerebral ischemia/reperfusion injury, Journal of neuroinflammation 15(1) (2018) 242. [108] R. Zhou, A. Tardivel, B. Thorens, I. Choi, J. Tschopp, Thioredoxin-interacting protein links oxidative stress to inflammasome activation, Nature immunology 11(2) (2010) 136.

[109] R. Zhou, A.S. Yazdi, P. Menu, J. Tschopp, A role for mitochondria in NLRP3 inflammasome activation, Nature 469(7329) (2011) 221. [110] N. Subramanian, K. Natarajan, M.R. Clatworthy, Z. Wang, R.N. Germain, The adaptor MAVS promotes NLRP3 mitochondrial localization and inflammasome activation, Cell 153(2) (2013) 348-361. [111] C.M. Cruz, A. Rinna, H.J. Forman, A.L. Ventura, P.M. Persechini, D.M. Ojcius, ATP activates a reactive oxygen species-dependent oxidative stress response and secretion of proinflammatory cytokines in macrophages, Journal of Biological Chemistry 282(5) (2007) 2871-2879. [112] V. Petrilli, S. Papin, C. Dostert, A. Mayor, F. Martinon, J. Tschopp, Activation of the NALP3 inflammasome is triggered by low intracellular potassium concentration, Cell death and differentiation 14(9) (2007) 1583. [113] R. Zhou, A.S. Yazdi, P. Menu, J. Tschopp, A role for mitochondria in NLRP3 inflammasome activation, Nature 469(7329) (2011) 221-5. [114] C. Dostert, V. Pétrilli, R. Van Bruggen, C. Steele, B.T. Mossman, J. Tschopp, Innate immune activation through Nalp3 inflammasome sensing of asbestos and silica, Science 320(5876) (2008) 674677. [115] D.J. Klionsky, F.C. Abdalla, H. Abeliovich, R.T. Abraham, A. Acevedo-Arozena, K. Adeli, L. Agholme, M. Agnello, P. Agostinis, J.A. Aguirre-Ghiso, Guidelines for the use and interpretation of assays for monitoring autophagy, Autophagy 8(4) (2012) 445-544. [116] L.-s. Huang, D. Cobessi, E.Y. Tung, E.A. Berry, Binding of the respiratory chain inhibitor antimycin to the mitochondrial bc1 complex: a new crystal structure reveals an altered intramolecular hydrogenbonding pattern, Journal of molecular biology 351(3) (2005) 573-597. [117] N. Li, K. Ragheb, G. Lawler, J. Sturgis, B. Rajwa, J.A. Melendez, J.P. Robinson, Mitochondrial complex I inhibitor rotenone induces apoptosis through enhancing mitochondrial reactive oxygen species production, Journal of Biological Chemistry 278(10) (2003) 8516-8525. [118] S. Alfonso-Loeches, J.R. Ureña-Peralta, M.J. Morillo-Bargues, C. Guerri, Role of mitochondria ROS generation in ethanol-induced NLRP3 inflammasome activation and cell death in astroglial cells, Frontiers in cellular neuroscience 8 (2014) 216. [119] S.-Y. Chuang, C.-H. Yang, C.-C. Chou, Y.-P. Chiang, T.-H. Chuang, L.-C. Hsu, TLR-induced PAI-2 expression suppresses IL-1β processing via increasing autophagy and NLRP3 degradation, Proceedings of the National Academy of Sciences 110(40) (2013) 16079-16084. [120] J. Dai, X. Zhang, Y. Wang, H. Chen, Y. Chai, ROS-activated NLRP3 inflammasome initiates inflammation in delayed wound healing in diabetic rats, Int J Clin Exp Pathol 10(9) (2017) 9902-9909. [121] D. Liu, M. Xu, L.-H. Ding, L.-L. Lv, H. Liu, K.-L. Ma, A.-H. Zhang, S.D. Crowley, B.-C. Liu, Activation of the Nlrp3 inflammasome by mitochondrial reactive oxygen species: a novel mechanism of albumininduced tubulointerstitial inflammation, The international journal of biochemistry & cell biology 57 (2014) 7-19. [122] K. Shimada, T.R. Crother, J. Karlin, J. Dagvadorj, N. Chiba, S. Chen, V.K. Ramanujan, A.J. Wolf, L. Vergnes, D.M. Ojcius, A. Rentsendorj, M. Vargas, C. Guerrero, Y. Wang, K.A. Fitzgerald, D.M. Underhill, T. Town, M. Arditi, Oxidized mitochondrial DNA activates the NLRP3 inflammasome during apoptosis, Immunity 36(3) (2012) 401-14. [123] K. Nakahira, J.A. Haspel, V.A. Rathinam, S.J. Lee, T. Dolinay, H.C. Lam, J.A. Englert, M. Rabinovitch, M. Cernadas, H.P. Kim, K.A. Fitzgerald, S.W. Ryter, A.M. Choi, Autophagy proteins regulate innate immune responses by inhibiting the release of mitochondrial DNA mediated by the NALP3 inflammasome, Nature immunology 12(3) (2011) 222-30. [124] K. Nakahira, J.A. Haspel, V.A. Rathinam, S.-J. Lee, T. Dolinay, H.C. Lam, J.A. Englert, M. Rabinovitch, M. Cernadas, H.P. Kim, Autophagy proteins regulate innate immune responses by inhibiting the release of mitochondrial DNA mediated by the NALP3 inflammasome, Nature immunology 12(3) (2011) 222.

[125] K. Shimada, T.R. Crother, J. Karlin, J. Dagvadorj, N. Chiba, S. Chen, V.K. Ramanujan, A.J. Wolf, L. Vergnes, D.M. Ojcius, Oxidized mitochondrial DNA activates the NLRP3 inflammasome during apoptosis, Immunity 36(3) (2012) 401-414. [126] D.N. Bronner, B.H. Abuaita, X. Chen, K.A. Fitzgerald, G. Nuñez, Y. He, X.-M. Yin, M.X. O’Riordan, Endoplasmic reticulum stress activates the inflammasome via NLRP3-and caspase-2-driven mitochondrial damage, Immunity 43(3) (2015) 451-462. [127] M.E. Heid, P.A. Keyel, C. Kamga, S. Shiva, S.C. Watkins, R.D. Salter, Mitochondrial reactive oxygen species induces NLRP3-dependent lysosomal damage and inflammasome activation, The Journal of Immunology 191(10) (2013) 5230-5238. [128] Y. Zhuang, G. Ding, M. Zhao, M. Bai, L. Yang, J. Ni, R. Wang, Z. Jia, S. Huang, A. Zhang, NLRP3 inflammasome mediates albumin-induced renal tubular injury through impaired mitochondrial function, Journal of Biological Chemistry 289(36) (2014) 25101-25111. [129] R. Allam, K.E. Lawlor, E.C.W. Yu, A.L. Mildenhall, D.M. Moujalled, R.S. Lewis, F. Ke, K.D. Mason, M.J. White, K.J. Stacey, Mitochondrial apoptosis is dispensable for NLRP3 inflammasome activation but nonapoptotic caspase-8 is required for inflammasome priming, EMBO reports 15(9) (2014) 982-990. [130] V. Sagulenko, S.J. Thygesen, D.P. Sester, A. Idris, J.A. Cridland, P.R. Vajjhala, T.L. Roberts, K. Schroder, J.E. Vince, J.M. Hill, AIM2 and NLRP3 inflammasomes activate both apoptotic and pyroptotic death pathways via ASC, Cell death and differentiation 20(9) (2013) 1149. [131] C. Chinopoulos, V. Adam-Vizi, Mitochondrial Ca2+ sequestration and precipitation revisited, The FEBS journal 277(18) (2010) 3637-3651. [132] T. Murakami, J. Ockinger, J. Yu, V. Byles, A. McColl, A.M. Hofer, T. Horng, Critical role for calcium mobilization in activation of the NLRP3 inflammasome, Proceedings of the National Academy of Sciences 109(28) (2012) 11282-11287. [133] D. Brough, R.A. Le Feuvre, R.D. Wheeler, N. Solovyova, S. Hilfiker, N.J. Rothwell, A. Verkhratsky, Ca2+ stores and Ca2+ entry differentially contribute to the release of IL-1β and IL-1α from murine macrophages, The Journal of Immunology 170(6) (2003) 3029-3036. [134] H.M. Lee, J.M. Yuk, K.H. Kim, J. Jang, G. Kang, J.B. Park, J.W. Son, E.K. Jo, Mycobacterium abscessus activates the NLRP3 inflammasome via Dectin-1–Syk and p62/SQSTM1, Immunology and cell biology 90(6) (2012) 601-610. [135] M. Rossol, M. Pierer, N. Raulien, D. Quandt, U. Meusch, K. Rothe, K. Schubert, T. Schöneberg, M. Schaefer, U. Krügel, Extracellular Ca 2+ is a danger signal activating the NLRP3 inflammasome through G protein-coupled calcium sensing receptors, Nature communications 3 (2012) 1329. [136] G.-S. Lee, N. Subramanian, A.I. Kim, I. Aksentijevich, R. Goldbach-Mansky, D.B. Sacks, R.N. Germain, D.L. Kastner, J.J. Chae, The calcium-sensing receptor regulates the NLRP3 inflammasome through Ca 2+ and cAMP, Nature 492(7427) (2012) 123. [137] R. Muñoz-Planillo, P. Kuffa, G. Martínez-Colón, B.L. Smith, T.M. Rajendiran, G. Núñez, K+ efflux is the common trigger of NLRP3 inflammasome activation by bacterial toxins and particulate matter, Immunity 38(6) (2013) 1142-1153. [138] R. Arias-Cartin, S. Grimaldi, P. Arnoux, B. Guigliarelli, A. Magalon, Cardiolipin binding in bacterial respiratory complexes: structural and functional implications, Biochimica et Biophysica Acta (BBA)Bioenergetics 1817(10) (2012) 1937-1949. [139] C.T. Chu, J. Ji, R.K. Dagda, J.F. Jiang, Y.Y. Tyurina, A.A. Kapralov, V.A. Tyurin, N. Yanamala, I.H. Shrivastava, D. Mohammadyani, Cardiolipin externalization to the outer mitochondrial membrane acts as an elimination signal for mitophagy in neuronal cells, Nature cell biology 15(10) (2013) 1197. [140] Z. Huang, J. Jiang, V.A. Tyurin, Q. Zhao, A. Mnuskin, J. Ren, N.A. Belikova, W. Feng, I.V. Kurnikov, V.E. Kagan, Cardiolipin deficiency leads to decreased cardiolipin peroxidation and increased resistance of cells to apoptosis, Free Radical Biology and Medicine 44(11) (2008) 1935-1944.

[141] S.S. Iyer, Q. He, J.R. Janczy, E.I. Elliott, Z. Zhong, A.K. Olivier, J.J. Sadler, V. Knepper-Adrian, R. Han, L. Qiao, Mitochondrial cardiolipin is required for Nlrp3 inflammasome activation, Immunity 39(2) (2013) 311-323. [142] H. Chen, A. Chomyn, D.C. Chan, Disruption of fusion results in mitochondrial heterogeneity and dysfunction, Journal of Biological Chemistry 280(28) (2005) 26185-26192. [143] H. Chen, S.A. Detmer, A.J. Ewald, E.E. Griffin, S.E. Fraser, D.C. Chan, Mitofusins Mfn1 and Mfn2 coordinately regulate mitochondrial fusion and are essential for embryonic development, The Journal of cell biology 160(2) (2003) 189-200. [144] T. Ichinohe, T. Yamazaki, T. Koshiba, Y. Yanagi, Mitochondrial protein mitofusin 2 is required for NLRP3 inflammasome activation after RNA virus infection, Proceedings of the National Academy of Sciences 110(44) (2013) 17963-17968. [145] J.-C. Martinou, S. Herzig, Mitochondrial dynamics: to be in good shape to survive, Current molecular medicine 8(2) (2008) 131-137. [146] D.-F. Suen, K.L. Norris, R.J. Youle, Mitochondrial dynamics and apoptosis, Genes & development 22(12) (2008) 1577-1590. [147] D.C. Chan, Mitochondrial fusion and fission in mammals, Annu. Rev. Cell Dev. Biol. 22 (2006) 7999. [148] B. Westermann, Molecular machinery of mitochondrial fusion and fission, Journal of Biological Chemistry 283(20) (2008) 13501-13505. [149] K. Okamoto, J.M. Shaw, Mitochondrial morphology and dynamics in yeast and multicellular eukaryotes, Annu. Rev. Genet. 39 (2005) 503-536. [150] G. Twig, O.S. Shirihai, The interplay between mitochondrial dynamics and mitophagy, Antioxidants & redox signaling 14(10) (2011) 1939-1951. [151] L. Calo, Y. Dong, R. Kumar, K. Przyklenk, T. H Sanderson, Mitochondrial dynamics: an emerging paradigm in ischemia-reperfusion injury, Current pharmaceutical design 19(39) (2013) 6848-6857. [152] S. Hoppins, L. Lackner, J. Nunnari, The machines that divide and fuse mitochondria, Annu. Rev. Biochem. 76 (2007) 751-780. [153] G. Twig, B. Hyde, O.S. Shirihai, Mitochondrial fusion, fission and autophagy as a quality control axis: the bioenergetic view, Biochimica et Biophysica Acta (BBA)-Bioenergetics 1777(9) (2008) 10921097. [154] L.C. Gomes, L. Scorrano, High levels of Fis1, a pro-fission mitochondrial protein, trigger autophagy, Biochimica et Biophysica Acta (BBA)-Bioenergetics 1777(7-8) (2008) 860-866. [155] T. Ono, K. Isobe, K. Nakada, J.-I. Hayashi, Human cells are protected from mitochondrial dysfunction by complementation of DNA products in fused mitochondria, Nature genetics 28(3) (2001) 272. [156] K.B. Busch, J. Bereiter-Hahn, I. Wittig, H. Schagger, M. Jendrach, Mitochondrial dynamics generate equal distribution but patchwork localization of respiratory Complex I, Molecular membrane biology 23(6) (2006) 509-520. [157] K.S. Dimmer, L. Scorrano, (De) constructing mitochondria: what for?, Physiology 21(4) (2006) 233241. [158] I.R. Boldogh, L.A. Pon, Interactions of mitochondria with the actin cytoskeleton, Biochimica et Biophysica Acta (BBA)-Molecular Cell Research 1763(5-6) (2006) 450-462. [159] P.J. Hollenbeck, W.M. Saxton, The axonal transport of mitochondria, Journal of cell science 118(23) (2005) 5411-5419. [160] J.E. Lee, L.M. Westrate, H. Wu, C. Page, G.K. Voeltz, Multiple dynamin family members collaborate to drive mitochondrial division, Nature 540(7631) (2016) 139. [161] O.C. Losón, Z. Song, H. Chen, D.C. Chan, Fis1, Mff, MiD49, and MiD51 mediate Drp1 recruitment in mitochondrial fission, Molecular biology of the cell 24(5) (2013) 659-667.

[162] H. Otera, C. Wang, M.M. Cleland, K. Setoguchi, S. Yokota, R.J. Youle, K. Mihara, Mff is an essential factor for mitochondrial recruitment of Drp1 during mitochondrial fission in mammalian cells, The Journal of cell biology 191(6) (2010) 1141-1158. [163] C.S. Palmer, L.D. Osellame, D. Laine, O.S. Koutsopoulos, A.E. Frazier, M.T. Ryan, MiD49 and MiD51, new components of the mitochondrial fission machinery, EMBO reports 12(6) (2011) 565-573. [164] E. Smirnova, L. Griparic, D.-L. Shurland, A.M. Van Der Bliek, Dynamin-related protein Drp1 is required for mitochondrial division in mammalian cells, Molecular biology of the cell 12(8) (2001) 22452256. [165] S. Wu, F. Zhou, Z. Zhang, D. Xing, Mitochondrial oxidative stress causes mitochondrial fragmentation via differential modulation of mitochondrial fission–fusion proteins, The FEBS journal 278(6) (2011) 941-954. [166] G. De Arriba, M. Calvino, S. Benito, T. Parra, Cyclosporine A-induced apoptosis in renal tubular cells is related to oxidative damage and mitochondrial fission, Toxicology letters 218(1) (2013) 30-38. [167] L.F. Ferrari, A. Chum, O. Bogen, D.B. Reichling, J.D. Levine, Role of Drp1, a key mitochondrial fission protein, in neuropathic pain, Journal of Neuroscience 31(31) (2011) 11404-11410. [168] S. Kobashigawa, K. Suzuki, S. Yamashita, Ionizing radiation accelerates Drp1-dependent mitochondrial fission, which involves delayed mitochondrial reactive oxygen species production in normal human fibroblast-like cells, Biochemical and biophysical research communications 414(4) (2011) 795-800. [169] L. Peng, X. Men, W. Zhang, H. Wang, S. Xu, M. Xu, Y. Xu, W. Yang, J. Lou, Dynamin-related protein 1 is implicated in endoplasmic reticulum stress-induced pancreatic β-cell apoptosis, International journal of molecular medicine 28(2) (2011) 161-169. [170] M.J. Barsoum, H. Yuan, A.A. Gerencser, G. Liot, Y. Kushnareva, S. Gräber, I. Kovacs, W.D. Lee, J. Waggoner, J. Cui, Nitric oxide-induced mitochondrial fission is regulated by dynamin-related GTPases in neurons, The EMBO journal 25(16) (2006) 3900-3911. [171] J. Grohm, S. Kim, U. Mamrak, S. Tobaben, A. Cassidy-Stone, J. Nunnari, N. Plesnila, C. Culmsee, Inhibition of Drp1 provides neuroprotection in vitro and in vivo, Cell death and differentiation 19(9) (2012) 1446. [172] Y.X. Zhao, M. Cui, S.F. Chen, Q. Dong, X.Y. Liu, Amelioration of ischemic mitochondrial injury and Bax-dependent outer membrane permeabilization by Mdivi-1, CNS neuroscience & therapeutics 20(6) (2014) 528-538. [173] A. Santel, M.T. Fuller, Control of mitochondrial morphology by a human mitofusin, Journal of cell science 114(5) (2001) 867-874. [174] M. Escobar-Henriques, F. Anton, Mechanistic perspective of mitochondrial fusion: tubulation vs. fragmentation, Biochimica et Biophysica Acta (BBA)-Molecular Cell Research 1833(1) (2013) 162-175. [175] N. Ishihara, Y. Fujita, T. Oka, K. Mihara, Regulation of mitochondrial morphology through proteolytic cleavage of OPA1, The EMBO journal 25(13) (2006) 2966-2977. [176] L.V. Chernomordik, M.M. Kozlov, Membrane hemifusion: crossing a chasm in two leaps, Cell 123(3) (2005) 375-382. [177] G. Meglei, G.A. McQuibban, The dynamin-related protein Mgm1p assembles into oligomers and hydrolyzes GTP to function in mitochondrial membrane fusion, Biochemistry 48(8) (2009) 1774-1784. [178] G. Benard, M. Karbowski, Mitochondrial fusion and division: Regulation and role in cell viability, Seminars in cell & developmental biology, Elsevier, 2009, pp. 365-374. [179] S. Züchner, P. De Jonghe, A. Jordanova, K.G. Claeys, V. Guergueltcheva, S. Cherninkova, S.R. Hamilton, G. Van Stavern, K.M. Krajewski, J. Stajich, Axonal neuropathy with optic atrophy is caused by mutations in mitofusin 2, Annals of Neurology: Official Journal of the American Neurological Association and the Child Neurology Society 59(2) (2006) 276-281.

[180] S. Züchner, I.V. Mersiyanova, M. Muglia, N. Bissar-Tadmouri, J. Rochelle, E.L. Dadali, M. Zappia, E. Nelis, A. Patitucci, J. Senderek, Mutations in the mitochondrial GTPase mitofusin 2 cause Charcot-MarieTooth neuropathy type 2A, Nature genetics 36(5) (2004) 449. [181] S. Li, X. Sun, L. Xu, R. Sun, Z. Ma, X. Deng, B. Liu, Q. Fu, R. Qu, S. Ma, Baicalin attenuates in vivo and in vitro hyperglycemia-exacerbated ischemia/reperfusion injury by regulating mitochondrial function in a manner dependent on AMPK, European journal of pharmacology 815 (2017) 118-126. [182] C. Peng, W. Rao, L. Zhang, K. Wang, H. Hui, L. Wang, N. Su, P. Luo, Y.-l. Hao, Y. Tu, Mitofusin 2 ameliorates hypoxia-induced apoptosis via mitochondrial function and signaling pathways, The international journal of biochemistry & cell biology 69 (2015) 29-40. [183] L. Zhang, Z. He, Q. Zhang, Y. Wu, X. Yang, W. Niu, Y. Hu, J. Jia, Exercise pretreatment promotes mitochondrial dynamic protein OPA1 expression after cerebral ischemia in rats, International journal of molecular sciences 15(3) (2014) 4453-4463. [184] S. Kumari, L. Anderson, S. Farmer, S.L. Mehta, P.A. Li, Hyperglycemia alters mitochondrial fission and fusion proteins in mice subjected to cerebral ischemia and reperfusion, Translational stroke research 3(2) (2012) 296-304. [185] S.L. Archer, Mitochondrial dynamics—mitochondrial fission and fusion in human diseases, New England Journal of Medicine 369(23) (2013) 2236-2251. [186] P. Mishra, D.C. Chan, Mitochondrial dynamics and inheritance during cell division, development and disease, Nature reviews Molecular cell biology 15(10) (2014) 634. [187] G. Twig, A. Elorza, A.J. Molina, H. Mohamed, J.D. Wikstrom, G. Walzer, L. Stiles, S.E. Haigh, S. Katz, G. Las, Fission and selective fusion govern mitochondrial segregation and elimination by autophagy, The EMBO journal 27(2) (2008) 433-446. [188] R.X. Santos, S.C. Correia, X. Wang, G. Perry, M.A. Smith, P.I. Moreira, X. Zhu, A synergistic dysfunction of mitochondrial fission/fusion dynamics and mitophagy in Alzheimer's disease, Journal of Alzheimer's Disease 20(s2) (2010) S401-S412. [189] M. Van Gurp, N. Festjens, G. Van Loo, X. Saelens, P. Vandenabeele, Mitochondrial intermembrane proteins in cell death, Biochemical and biophysical research communications 304(3) (2003) 487-497. [190] X. Wang, The expanding role of mitochondria in apoptosis, Genes & development 15(22) (2001) 2922-2933. [191] R. Guan, W. Zou, X. Dai, X. Yu, H. Liu, Q. Chen, W. Teng, Mitophagy, a potential therapeutic target for stroke, Journal of biomedical science 25(1) (2018) 87. [192] K. Palikaras, N. Tavernarakis, Mitochondrial homeostasis: the interplay between mitophagy and mitochondrial biogenesis, Experimental gerontology 56 (2014) 182-8. [193] R. Guan, W. Zou, X. Dai, X. Yu, H. Liu, Q. Chen, W. Teng, Mitophagy, a potential therapeutic target for stroke, Journal of biomedical science 25(1) (2018) 87. [194] S. Rodriguez-Enriquez, Y. Kai, E. Maldonado, R.T. Currin, J.J. Lemasters, Roles of mitophagy and the mitochondrial permeability transition in remodeling of cultured rat hepatocytes, Autophagy 5(8) (2009) 1099-1106. [195] S.P. Elmore, T. Qian, S.F. Grissom, J.J. Lemasters, The mitochondrial permeability transition initiates autophagy in rat hepatocytes, The FASEB Journal 15(12) (2001) 2286-2287. [196] M.P. Murphy, How mitochondria produce reactive oxygen species, Biochemical journal 417(1) (2009) 1-13. [197] D.J. Wible, S.B. Bratton, Reciprocity in ROS and autophagic signaling, Current opinion in toxicology 7 (2018) 28-36. [198] J. Qu, W. Chen, R. Hu, H. Feng, The injury and therapy of reactive oxygen species in intracerebral hemorrhage looking at mitochondria, Oxidative medicine and cellular longevity 2016 (2016).

[199] S. Cao, S. Shrestha, J. Li, X. Yu, J. Chen, F. Yan, G. Ying, C. Gu, L. Wang, G. Chen, Melatoninmediated mitophagy protects against early brain injury after subarachnoid hemorrhage through inhibition of NLRP3 inflammasome activation, Scientific reports 7(1) (2017) 2417. [200] Y. Di, Y.-L. He, T. Zhao, X. Huang, K.-W. Wu, S.-H. Liu, Y.-Q. Zhao, M. Fan, L.-Y. Wu, L.-L. Zhu, Methylene blue reduces acute cerebral ischemic injury via the induction of mitophagy, Molecular medicine 21(1) (2015) 420-429. [201] A. Rustom, R. Saffrich, I. Markovic, P. Walther, H.-H. Gerdes, Nanotubular highways for intercellular organelle transport, Science 303(5660) (2004) 1007-1010. [202] S. Mathivanan, H. Ji, R.J. Simpson, Exosomes: extracellular organelles important in intercellular communication, Journal of proteomics 73(10) (2010) 1907-1920. [203] Y. Kiriyama, H. Nochi, Intra-and intercellular quality control mechanisms of mitochondria, Cells 7(1) (2018) 1. [204] M. Koyanagi, R.P. Brandes, J. Haendeler, A.M. Zeiher, S. Dimmeler, Cell-to-cell connection of endothelial progenitor cells with cardiac myocytes by nanotubes: a novel mechanism for cell fate changes?, Circulation research 96(10) (2005) 1039-1041. [205] B. Önfelt, S. Nedvetzki, R.K. Benninger, M.A. Purbhoo, S. Sowinski, A.N. Hume, M.C. Seabra, M.A. Neil, P.M. French, D.M. Davis, Structurally distinct membrane nanotubes between human macrophages support long-distance vesicular traffic or surfing of bacteria, The Journal of Immunology 177(12) (2006) 8476-8483. [206] X. Wang, H.-H. Gerdes, Transfer of mitochondria via tunneling nanotubes rescues apoptotic PC12 cells, Cell death and differentiation 22(7) (2015) 1181. [207] I. Sáenz-de-Santa-María, C. Bernardo-Castiñeira, E. Enciso, I. García-Moreno, J.L. Chiara, C. Suarez, M.-D. Chiara, Control of long-distance cell-to-cell communication and autophagosome transfer in squamous cell carcinoma via tunneling nanotubes, Oncotarget 8(13) (2017) 20939. [208] K. Yasuda, H.-C. Park, B. Ratliff, F. Addabbo, A.K. Hatzopoulos, P. Chander, M.S. Goligorsky, Adriamycin nephropathy: a failure of endothelial progenitor cell-induced repair, The American journal of pathology 176(4) (2010) 1685-1695. [209] M.N. Islam, S.R. Das, M.T. Emin, M. Wei, L. Sun, K. Westphalen, D.J. Rowlands, S.K. Quadri, S. Bhattacharya, J. Bhattacharya, Mitochondrial transfer from bone-marrow–derived stromal cells to pulmonary alveoli protects against acute lung injury, Nature medicine 18(5) (2012) 759. [210] K. He, X. Shi, X. Zhang, S. Dang, X. Ma, F. Liu, M. Xu, Z. Lv, D. Han, X. Fang, Long-distance intercellular connectivity between cardiomyocytes and cardiofibroblasts mediated by membrane nanotubes, Cardiovascular research 92(1) (2011) 39-47. [211] T. Ahmad, S. Mukherjee, B. Pattnaik, M. Kumar, S. Singh, R. Rehman, B.K. Tiwari, K.A. Jha, A.P. Barhanpurkar, M.R. Wani, Miro1 regulates intercellular mitochondrial transport & enhances mesenchymal stem cell rescue efficacy, The EMBO journal 33(9) (2014) 994-1010. [212] J.L. Spees, S.D. Olson, M.J. Whitney, D.J. Prockop, Mitochondrial transfer between cells can rescue aerobic respiration, Proceedings of the National Academy of Sciences 103(5) (2006) 1283-1288. [213] D.G. Phinney, M. Di Giuseppe, J. Njah, E. Sala, S. Shiva, C.M. St Croix, D.B. Stolz, S.C. Watkins, Y.P. Di, G.D. Leikauf, Mesenchymal stem cells use extracellular vesicles to outsource mitophagy and shuttle microRNAs, Nature communications 6 (2015) 8472. [214] K. Hayakawa, E. Esposito, X. Wang, Y. Terasaki, Y. Liu, C. Xing, X. Ji, E.H. Lo, Transfer of mitochondria from astrocytes to neurons after stroke, Nature 535(7613) (2016) 551. [215] E. Plotnikov, T. Khryapenkova, A. Vasileva, M. Marey, S. Galkina, N. Isaev, E. Sheval, V. Polyakov, G. Sukhikh, D. Zorov, Cell-to-cell cross-talk between mesenchymal stem cells and cardiomyocytes in coculture, Journal of cellular and molecular medicine 12(5a) (2008) 1622-1631.

[216] K. Liu, K. Ji, L. Guo, W. Wu, H. Lu, P. Shan, C. Yan, Mesenchymal stem cells rescue injured endothelial cells in an in vitro ischemia–reperfusion model via tunneling nanotube like structuremediated mitochondrial transfer, Microvascular research 92 (2014) 10-18. [217] A. Acquistapace, T. Bru, P.F. Lesault, F. Figeac, A.E. Coudert, O. Le Coz, C. Christov, X. Baudin, F. Auber, R. Yiou, Human mesenchymal stem cells reprogram adult cardiomyocytes toward a progenitorlike state through partial cell fusion and mitochondria transfer, Stem cells 29(5) (2011) 812-824. [218] K.C. Vallabhaneni, H. Haller, I. Dumler, Vascular smooth muscle cells initiate proliferation of mesenchymal stem cells by mitochondrial transfer via tunneling nanotubes, Stem cells and development 21(17) (2012) 3104-3113. [219] E. Lou, S. Fujisawa, A. Morozov, A. Barlas, Y. Romin, Y. Dogan, S. Gholami, A.L. Moreira, K. ManovaTodorova, M.A. Moore, Tunneling nanotubes provide a unique conduit for intercellular transfer of cellular contents in human malignant pleural mesothelioma, PloS one 7(3) (2012) e33093. [220] I. Melentijevic, M.L. Toth, M.L. Arnold, R.J. Guasp, G. Harinath, K.C. Nguyen, D. Taub, J.A. Parker, C. Neri, C.V. Gabel, C. elegans neurons jettison protein aggregates and mitochondria under neurotoxic stress, Nature 542(7641) (2017) 367. [221] D. Jiang, F. Gao, Y. Zhang, D.S.H. Wong, Q. Li, H.-f. Tse, G. Xu, Z. Yu, Q. Lian, Mitochondrial transfer of mesenchymal stem cells effectively protects corneal epithelial cells from mitochondrial damage, Cell death & disease 7(11) (2016) e2467. [222] F. Liu, J. Lu, A. Manaenko, J. Tang, Q. Hu, Mitochondria in Ischemic stroke: New insight and implications, Aging and disease 9(5) (2018) 924. [223] P.-J. Huang, C.-C. Kuo, H.-C. Lee, C.-I. Shen, F.-C. Cheng, S.-F. Wu, J.-C. Chang, H.-C. Pan, S.-Z. Lin, C.-S. Liu, Transferring xenogenic mitochondria provides neural protection against ischemic stress in ischemic rat brains, Cell transplantation 25(5) (2016) 913-927. [224] C.-Y. Chang, M.-Z. Liang, L. Chen, Current progress of mitochondrial transplantation that promotes neuronal regeneration, Translational neurodegeneration 8(1) (2019) 17. [225] D. Sarmah, H. Kaur, J. Saraf, K. Vats, K. Pravalika, M. Wanve, K. Kalia, A. Borah, A. Kumar, X. Wang, Mitochondrial dysfunction in stroke: implications of stem cell therapy, Translational stroke research 10(2) (2019) 121-136. [226] A. Trounson, C. McDonald, Stem cell therapies in clinical trials: progress and challenges, Cell stem cell 17(1) (2015) 11-22. [227] C.V. Borlongan, Age of PISCES: stem-cell clinical trials in stroke, The Lancet 388(10046) (2016) 736738. [228] K. Prasad, A. Sharma, A. Garg, S. Mohanty, S. Bhatnagar, S. Johri, K.K. Singh, V. Nair, R.S. Sarkar, S.P. Gorthi, Intravenous autologous bone marrow mononuclear stem cell therapy for ischemic stroke: a multicentric, randomized trial, Stroke 45(12) (2014) 3618-3624. [229] L. Hao, Z. Zou, H. Tian, Y. Zhang, H. Zhou, L. Liu, Stem cell-based therapies for ischemic stroke, BioMed research international 2014 (2014). [230] M. Farzaneh, F. Rahimi, M. Alishahi, S.E. Khoshnam, Paracrine mechanisms involved in mesenchymal stem cell differentiation into cardiomyocytes, Current stem cell research & therapy 14(1) (2019) 9-13. [231] T. Kolagar, M. Farzaneh, N. Nikkar, A. Anbiyaiee, E. Heydari, S. Khoshnam, Human Pluripotent Stem Cells in Neurodegenerative Diseases: Potentials, Advances, and Limitations, Current stem cell research & therapy (2019). [232] D. Sarmah, H. Kaur, J. Saraf, K. Pravalika, A. Goswami, K. Kalia, A. Borah, X. Wang, K.R. Dave, D.R. Yavagal, Getting closer to an effective intervention of ischemic stroke: the big promise of stem cell, Translational stroke research 9(4) (2018) 356-374.

[233] Y.M. Cho, J.H. Kim, M. Kim, S.J. Park, S.H. Koh, H.S. Ahn, G.H. Kang, J.-B. Lee, K.S. Park, H.K. Lee, Mesenchymal stem cells transfer mitochondria to the cells with virtually no mitochondrial function but not with pathogenic mtDNA mutations, PloS one 7(3) (2012) e32778. [234] H.-Y. Lin, C.-W. Liou, S.-D. Chen, T.-Y. Hsu, J.-H. Chuang, P.-W. Wang, S.-T. Huang, M.-M. Tiao, J.-B. Chen, T.-K. Lin, Mitochondrial transfer from Wharton's jelly-derived mesenchymal stem cells to mitochondria-defective cells recaptures impaired mitochondrial function, Mitochondrion 22 (2015) 3144. [235] X. Li, Y. Zhang, S.C. Yeung, Y. Liang, X. Liang, Y. Ding, M.S. Ip, H.-F. Tse, J.C. Mak, Q. Lian, Mitochondrial transfer of induced pluripotent stem cell–derived mesenchymal stem cells to airway epithelial cells attenuates cigarette smoke–induced damage, American journal of respiratory cell and molecular biology 51(3) (2014) 455-465. [236] M.V. Berridge, M.J. McConnell, C. Grasso, M. Bajzikova, J. Kovarova, J. Neuzil, Horizontal transfer of mitochondria between mammalian cells: beyond co-culture approaches, Current opinion in genetics & development 38 (2016) 75-82. [237] H. Han, J. Hu, Q. Yan, J. Zhu, Z. Zhu, Y. Chen, J. Sun, R. Zhang, Bone marrow‑derived mesenchymal stem cells rescue injured H9c2 cells via transferring intact mitochondria through tunneling nanotubes in an in vitro simulated ischemia/reperfusion model, Molecular medicine reports 13(2) (2016) 1517-1524. [238] M. Mahrouf-Yorgov, L. Augeul, C.C. Da Silva, M. Jourdan, M. Rigolet, S. Manin, R. Ferrera, M. Ovize, A. Henry, A. Guguin, Mesenchymal stem cells sense mitochondria released from damaged cells as danger signals to activate their rescue properties, Cell death and differentiation 24(7) (2017) 1224. [239] V. Babenko, D. Silachev, V. Popkov, L. Zorova, I. Pevzner, E. Plotnikov, G. Sukhikh, D. Zorov, Miro1 enhances mitochondria transfer from multipotent mesenchymal stem cells (MMSC) to neural cells and improves the efficacy of cell recovery, Molecules 23(3) (2018) 687. [240] D. Torralba, F. Baixauli, F. Sánchez-Madrid, Mitochondria know no boundaries: mechanisms and functions of intercellular mitochondrial transfer, Frontiers in cell and developmental biology 4 (2016) 107. [241] M.P. Murphy, R.C. Hartley, Mitochondria as a therapeutic target for common pathologies, Nature reviews. Drug discovery 17(12) (2018) 865-886. [242] L. Catanese, J. Tarsia, M. Fisher, Acute ischemic stroke therapy overview, Circulation research 120(3) (2017) 541-558. [243] A. Suomalainen, B.J. Battersby, Mitochondrial diseases: the contribution of organelle stress responses to pathology, Nature reviews Molecular cell biology 19(2) (2018) 77. [244] D. Tucker, Y. Lu, Q. Zhang, From mitochondrial function to neuroprotection—an emerging role for methylene blue, Molecular neurobiology 55(6) (2018) 5137-5153. [245] S.V. Narayanan, K.R. Dave, I. Saul, M.A. Perez-Pinzon, Resveratrol preconditioning protects against cerebral ischemic injury via nuclear erythroid 2–related factor 2, Stroke 46(6) (2015) 1626-1632. [246] Z. Li, K.-I. Okamoto, Y. Hayashi, M. Sheng, The importance of dendritic mitochondria in the morphogenesis and plasticity of spines and synapses, Cell 119(6) (2004) 873-887. [247] N. Egawa, J. Lok, K. Washida, K. Arai, Mechanisms of axonal damage and repair after central nervous system injury, Translational stroke research 8(1) (2017) 14-21. [248] S.M. Emani, B.L. Piekarski, D. Harrild, J. Pedro, J.D. McCully, Autologous mitochondrial transplantation for dysfunction after ischemia-reperfusion injury, The Journal of thoracic and cardiovascular surgery 154(1) (2017) 286-289. [249] A.P. Halestrap, A.M. Davidson, Inhibition of Ca2+-induced large-amplitude swelling of liver and heart mitochondria by cyclosporin is probably caused by the inhibitor binding to mitochondrial-matrix peptidyl-prolyl cis-trans isomerase and preventing it interacting with the adenine nucleotide translocase, Biochemical Journal 268(1) (1990) 153-160.

[250] A.L. Orr, L. Vargas, C.N. Turk, J.E. Baaten, J.T. Matzen, V.J. Dardov, S.J. Attle, J. Li, D.C. Quackenbush, R.L. Goncalves, Suppressors of superoxide production from mitochondrial complex III, Nature chemical biology 11(11) (2015) 834. [251] M.D. Brand, R.L. Goncalves, A.L. Orr, L. Vargas, A.A. Gerencser, M.B. Jensen, Y.T. Wang, S. Melov, C.N. Turk, J.T. Matzen, Suppressors of superoxide-H2O2 production at site IQ of mitochondrial complex I protect against stem cell hyperplasia and ischemia-reperfusion injury, Cell metabolism 24(4) (2016) 582592. [252] J. Harper, K. Dickinson, M. Brand, Mitochondrial uncoupling as a target for drug development for the treatment of obesity, Obesity Reviews 2(4) (2001) 255-265. [253] S.J. Yang, J.M. Choi, L. Kim, S.E. Park, E.J. Rhee, W.Y. Lee, K.W. Oh, S.W. Park, C.-Y. Park, Nicotinamide improves glucose metabolism and affects the hepatic NAD-sirtuin pathway in a rodent model of obesity and type 2 diabetes, The Journal of nutritional biochemistry 25(1) (2014) 66-72. [254] A.N. Long, K. Owens, A.E. Schlappal, T. Kristian, P.S. Fishman, R.A. Schuh, Effect of nicotinamide mononucleotide on brain mitochondrial respiratory deficits in an Alzheimer’s disease-relevant murine model, BMC neurology 15(1) (2015) 19. [255] V. Sorrentino, M. Romani, L. Mouchiroud, J.S. Beck, H. Zhang, D. D’amico, N. Moullan, F. Potenza, A.W. Schmid, S. Rietsch, Enhancing mitochondrial proteostasis reduces amyloid-β proteotoxicity, Nature 552(7684) (2017) 187. [256] D. Ryu, H. Zhang, E.R. Ropelle, V. Sorrentino, D.A. Mázala, L. Mouchiroud, P.L. Marshall, M.D. Campbell, A.S. Ali, G.M. Knowels, NAD+ repletion improves muscle function in muscular dystrophy and counters global PARylation, Science translational medicine 8(361) (2016) 361ra139-361ra139. [257] J. Yoshino, K.F. Mills, M.J. Yoon, S.-i. Imai, Nicotinamide mononucleotide, a key NAD+ intermediate, treats the pathophysiology of diet-and age-induced diabetes in mice, Cell metabolism 14(4) (2011) 528-536. [258] J. Camacho-Pereira, M.G. Tarragó, C.C. Chini, V. Nin, C. Escande, G.M. Warner, A.S. Puranik, R.A. Schoon, J.M. Reid, A. Galina, CD38 dictates age-related NAD decline and mitochondrial dysfunction through an SIRT3-dependent mechanism, Cell metabolism 23(6) (2016) 1127-1139. [259] K. Gariani, K.J. Menzies, D. Ryu, C.J. Wegner, X. Wang, E.R. Ropelle, N. Moullan, H. Zhang, A. Perino, V. Lemos, Eliciting the mitochondrial unfolded protein response by nicotinamide adenine dinucleotide repletion reverses fatty liver disease in mice, Hepatology 63(4) (2016) 1190-1204. [260] J.B. Lin, S. Kubota, N. Ban, M. Yoshida, A. Santeford, A. Sene, R. Nakamura, N. Zapata, M. Kubota, K. Tsubota, NAMPT-mediated NAD+ biosynthesis is essential for vision in mice, Cell reports 17(1) (2016) 69-85. [261] N. Zhang, S. Wang, Y. Li, L. Che, Q. Zhao, A selective inhibitor of Drp1, mdivi-1, acts against cerebral ischemia/reperfusion injury via an anti-apoptotic pathway in rats, Neuroscience letters 535 (2013) 104109. [262] Y. Tang, X. Liu, J. Zhao, X. Tan, B. Liu, G. Zhang, L. Sun, D. Han, H. Chen, M. Wang, Hypothermiainduced ischemic tolerance is associated with Drp1 inhibition in cerebral ischemia-reperfusion injury of mice, Brain research 1646 (2016) 73-83. [263] J. Wang, P. Wang, S. Li, S. Wang, Y. Li, N. Liang, M. Wang, Mdivi-1 prevents apoptosis induced by ischemia–reperfusion injury in primary hippocampal cells via inhibition of reactive oxygen species– activated mitochondrial pathway, Journal of Stroke and Cerebrovascular Diseases 23(6) (2014) 14911499. [264] X. Zhou, H.-Y. Wang, B. Wu, C.-Y. Cheng, W. Xiao, Z.-Z. Wang, Y.-Y. Yang, P. Li, H. Yang, Ginkgolide K attenuates neuronal injury after ischemic stroke by inhibiting mitochondrial fission and GSK-3βdependent increases in mitochondrial membrane permeability, Oncotarget 8(27) (2017) 44682. [265] X. Qi, N. Qvit, Y.-C. Su, D. Mochly-Rosen, A novel Drp1 inhibitor diminishes aberrant mitochondrial fission and neurotoxicity, J Cell Sci 126(3) (2013) 789-802.

[266] X. Guo, H. Sesaki, X. Qi, Drp1 stabilizes p53 on the mitochondria to trigger necrosis under oxidative stress conditions in vitro and in vivo, Biochemical Journal 461(1) (2014) 137-146. [267] J.-L. Yang, S. Mukda, S.-D. Chen, Diverse roles of mitochondria in ischemic stroke, Redox biology 16 (2018) 263-275. [268] E. Villa, S. Marchetti, J.-E. Ricci, No Parkin zone: mitophagy without Parkin, Trends in cell biology 28(11) (2018) 882-895. [269] A.R. Anzell, R. Maizy, K. Przyklenk, T.H. Sanderson, Mitochondrial quality control and disease: insights into ischemia-reperfusion injury, Molecular neurobiology 55(3) (2018) 2547-2564. [270] S. Kameoka, Y. Adachi, K. Okamoto, M. Iijima, H. Sesaki, Phosphatidic acid and cardiolipin coordinate mitochondrial dynamics, Trends in cell biology 28(1) (2018) 67-76. [271] Q. Wu, C.L. Luo, L.Y. Tao, Dynamin-related protein 1 (Drp1) mediating mitophagy contributes to the pathophysiology of nervous system diseases and brain injury, Histology and histopathology 32(6) (2017) 551-559.

Figure 1

Membrane depolarization

Ischemic stroke

Failure of Na+/ K+ ATPase

Glucose/ O2 deprivation

Ischemic neuron

Ion imbalance and calcium influx Glutamate

NMDA/ AMPA receptor

Ca2+

Decreased ∆Ψm Ca2+

Inflammatory response

ROS

Succinate MPTP

DAMPs

Necrosis

Mitochondria

Mitophagy

Apoptosis

Cytochrome C ↑Lactic acid

Oxidative DNA damage

Survival Cell death

Acidosis Fission Mitochondrial DNA damage

Mitophagy Fusion

PKR ↑ROS TXNIP

NLRP3 Inflammasome

Figure 2

Ischemic stroke

Parkin

Ubiquitin

PINK1

Mitochondrial dysfunction

P P

P

P

P

Mitofusin 2

Cardiolipin

ROS

Ubiquitin

Mt DNA

Parkin

Mitochondria

↓NAD+

Ca2+

Mitophagy

P

NLRP3 Mitophagy Adaptor ASC LC3

NLRP3 Inflammasome

Pro-caspase-1 Autophagy machinery Caspase-1 activation LRR NACHT PYD

NLRP3

PYD

CARD

CARD ASC

P20/10

Procaspase-1

Pro-IL-1β Pro-IL-18

IL-1β IL-18

Pyroptosis

Figure 3 A. Fusion

Drp1 Drp1 receptor Mitofusin 1/2 Fis 1 Opa 1 Cardiolipin

B. Fission

Figure 4

Cardiolipin

Healthy mitochondria

Mfn1/2

Opa1

Healthy mitochondria Autophagosome

Injured mitochondria Degradation (Mitophagy)

Drp1

Microvesicles

Healthy neuron

Tunneling nanotubes Healthy mitochondria Injured neuron Dysfunctional mitochondria

Microvesicles

Highlights • •

Mitochondria contribute to the pathology of ischemic stroke by aberrant production of ROS. Mitochondrial dysfunction regulate NLRP3 inflammation activation by several distinct pathways

•

Mitochondrial dynamics have a key role in cell fate following ischemic stroke

•

The mitochondrial transfer might potentially be a way to attenuate stroke-induced neuronal death Stem cells help to revive cell energetics by the transfer of mitochondria to the recipient cells

•