Mitochondrial transplantation: A promising therapy for mitochondrial disorders.

As a vital energy source for cellular metabolism and tissue survival, the mitochondrion can undergo morphological or positional change and even shuttle between cells in response to various stimuli and energy demands. Multiple human diseases are originated from mitochondrial dysfunction, but the curative succusses by traditional treatments are limited. Mitochondrial transplantation therapy (MTT) is an innovative therapeutic approach that is to deliver the healthy mitochondria either derived from normal cells or reassembled through synthetic biology into the cells and tissues suffering from mitochondrial damages and finally replace their defective mitochondria and restore their function. MTT has already been under investigation in clinical trials for cardiac ischemia-reperfusion injury and given an encouraging performance in animal models of numerous fatal critical diseases including central nervous system disorders, cardiovascular diseases, inflammatory conditions, cancer, renal injury, and pulmonary damage. This review article summarizes the mechanisms and strategies of mitochondrial transfer and the MTT application for types of mitochondrial diseases, and discusses the potential challenge in MTT clinical application, aiming to exhibit the good therapeutic prospects of MTTs in clinics.

Mitochondrial transfer mediates endothelial cell engraftment through mitophagy.

Ischaemic diseases such as critical limb ischaemia and myocardial infarction affect millions of people worldwide1. Transplanting endothelial cells (ECs) is a promising therapy in vascular medicine, but engrafting ECs typically necessitates co-transplanting perivascular supporting cells such as mesenchymal stromal cells (MSCs), which makes clinical implementation complicated2,3. The mechanisms that enable MSCs to facilitate EC engraftment remain elusive. Here we show that, under cellular stress, MSCs transfer mitochondria to ECs through tunnelling nanotubes, and that blocking this transfer impairs EC engraftment. We devised a strategy to artificially transplant mitochondria, transiently enhancing EC bioenergetics and enabling them to form functional vessels in ischaemic tissues without the support of MSCs. Notably, exogenous mitochondria did not integrate into the endogenous EC mitochondrial pool, but triggered mitophagy after internalization. Transplanted mitochondria co-localized with autophagosomes, and ablation of the PINK1-Parkin pathway negated the enhanced engraftment ability of ECs. Our findings reveal a mechanism that underlies the effects of mitochondrial transfer between mesenchymal and endothelial cells, and offer potential for a new approach for vascular cell therapy.

Effects of mitochondrial transplantation on chronic pressure wound healing in a human patient.

BACKGROUND AIMS: Wound healing is a multistage process that requires a concerted effort of various cell types. The intricate processes involved in the healing of wounds result in high energy requirements. Furthermore, mitochondria play a crucial role in the healing process because of their involvement in neo angiogenesis, growth factor synthesis, and cell differentiation. It is unclear how mitochondria transplantation, a promising new approach, influences wound healing. METHODS: In this study, healthy autologous mitochondria obtained from skeletal muscle were injected into chronic pressure wounds as an intervention to promote wound healing. RESULTS: Mitochondrial transplantation accelerated wound healing by reducing wound size, increasing granulation tissue, and hastening epithelialization. CONCLUSIONS: This study is the first to demonstrate the therapeutic efficacy of mitochondrial transplantation in wound healing.

The Therapeutic Potential of Mitochondria Transplantation Therapy in Neurodegenerative and Neurovascular Disorders.

Neurodegenerative and neurovascular disorders affect millions of people worldwide and account for a large and increasing health burden on the general population. Thus, there is a critical need to identify potential disease-modifying treatments that can prevent or slow the disease progression. Mitochondria are highly dynamic organelles and play an important role in energy metabolism and redox homeostasis, and mitochondrial dysfunction threatens cell homeostasis, perturbs energy production, and ultimately leads to cell death and diseases. Impaired mitochondrial function has been linked to the pathogenesis of several human neurological disorders. Given the significant contribution of mitochondrial dysfunction in neurological disorders, there has been considerable interest in developing therapies that can attenuate mitochondrial abnormalities and proffer neuroprotective effects. Unfortunately, therapies that target specific components of mitochondria or oxidative stress pathways have exhibited limited translatability. To this end, mitochondrial transplantation therapy (MTT) presents a new paradigm of therapeutic intervention, which involves the supplementation of healthy mitochondria to replace the damaged mitochondria for the treatment of neurological disorders. Prior studies demonstrated that the supplementation of healthy donor mitochondria to damaged neurons promotes neuronal viability, activity, and neurite growth and has been shown to provide benefits for neural and extra-neural diseases. In this review, we discuss the significance of mitochondria and summarize an overview of the recent advances and development of MTT in neurodegenerative and neurovascular disorders, particularly Parkinson's disease, Alzheimer's disease, and stroke. The significance of MTT is emerging as they meet a critical need to develop a diseasemodifying intervention for neurodegenerative and neurovascular disorders.

Mesenchymal stem cell-mediated transfer of mitochondria: mechanisms and functional impact.

There is a steadily growing interest in the use of mitochondria as therapeutic agents. The use of mitochondria derived from mesenchymal stem/stromal cells (MSCs) for therapeutic purposes represents an innovative approach to treat many diseases (immune deregulation, inflammation-related disorders, wound healing, ischemic events, and aging) with an increasing amount of promising evidence, ranging from preclinical to clinical research. Furthermore, the eventual reversal, induced by the intercellular mitochondrial transfer, of the metabolic and pro-inflammatory profile, opens new avenues to the understanding of diseases' etiology, their relation to both systemic and local risk factors, and also leads to new therapeutic tools for the control of inflammatory and degenerative diseases. To this end, we illustrate in this review, the triggers and mechanisms behind the transfer of mitochondria employed by MSCs and the underlying benefits as well as the possible adverse effects of MSCs mitochondrial exchange. We relay the rationale and opportunities for the use of these organelles in the clinic as cell-based product.

Mitochondrial Transplantation Attenuates Cerebral Ischemia-Reperfusion Injury: Possible Involvement of Mitochondrial Component Separation.

BACKGROUND: Mitochondrial dysfunctions play a pivotal role in cerebral ischemia-reperfusion (I/R) injury. Although mitochondrial transplantation has been recently explored for the treatment of cerebral I/R injury, the underlying mechanisms and fate of transplanted mitochondria are still poorly understood. METHODS: Mitochondrial morphology and function were assessed by fluorescent staining, electron microscopy, JC-1, PCR, mitochondrial stress testing, and metabolomics. Therapeutic effects of mitochondria were evaluated by cell viability, reactive oxygen species (ROS), and apoptosis levels in a cellular hypoxia-reoxygenation model. Rat middle cerebral artery occlusion model was applied to assess the mitochondrial therapy in vivo. Transcriptomics was performed to explore the underlying mechanisms. Mitochondrial fate tracking was implemented by a variety of fluorescent labeling methods. RESULTS: Neuro-2a (N2a) cell-derived mitochondria had higher mitochondrial membrane potential, more active oxidative respiration capacity, and less mitochondrial DNA copy number. Exogenous mitochondrial transplantation increased cellular viability in an oxygen-dependent manner, decreased ROS and apoptosis levels, improved neurobehavioral deficits, and reduced infarct size. Transcriptomic data showed that the differential gene enrichment pathways are associated with metabolism, especially lipid metabolism. Mitochondrial tracking indicated specific parts of the exogenous mitochondria fused with the mitochondria of the host cell, and others were incorporated into lysosomes. This process occurred at the beginning of internalization and its efficiency is related to intercellular connection. CONCLUSIONS: Mitochondrial transplantation may attenuate cerebral I/R injury. The mechanism may be related to mitochondrial component separation, altering cellular metabolism, reducing ROS, and apoptosis in an oxygen-dependent manner. The way of isolated mitochondrial transfer into the cell may be related to intercellular connection.

Investigation of the effect of isolated mitochondria transplantation on renal ischemia-reperfusion injury in rats.

Ischemia/Reperfusion (I/R) injury is clinically important in many surgical practice including kidney transplantation. It is known that mitochondria have a key role in the intracellular and extracellular signaling pathways of ischemia and reperfusion injury. In this respect, we pointed to explore the probable effects of isolated mitochondria transplantation from MSCs (mesenchymal stem cells), to alleviate ischemia/reperfusion-induced renal injury. Experiments were held on the 48 male Sprague Dawley rats. Groups were divided as Control (C1), I/R-Control (C2), Vehicle-1 (V1), Vehicle-2 (V2), Transplantation-1 (T1) and Transplantation-2 (T2) group. Unilaterally nephrectomy was performed in all groups. In the groups except the control, the left kidneys ischemized for 45 min and then reperfusion was carried out. According to the study groups, isolated mitochondria or vehicle infused into the renal cortex and rats were monitored for 48 h. Following that mentioned procedure, animals were sacrificed and biological samples were taken for physiological, histological and biochemical examinations. The results of present study show that mitochondrial transplantation promoted proliferation and regeneration of tubular cells after renal injury. Moreover, mitochondrial transplantation reduced mitochondrial dynamics-DRP-1 fission protein of tubular cells and reversed renal deficits. Mitochondrial transplantation diminished apoptotic markers including TUNEL and Caspase-3 levels in injured renal cells. Our results provide a direct link between mitochondria dysfunction and ischemia/reperfusion-induced renal injury and suggest a therapeutic effect of transplanting isolated mitochondria obtained from MSCs against renal injury.

Mitochondrial aspartate regulates TNF biogenesis and autoimmune tissue inflammation.

Misdirected immunity gives rise to the autoimmune tissue inflammation of rheumatoid arthritis, in which excess production of the cytokine tumor necrosis factor (TNF) is a central pathogenic event. Mechanisms underlying the breakdown of self-tolerance are unclear, but T cells in the arthritic joint have a distinctive metabolic signature of ATPlo acetyl-CoAhi proinflammatory effector cells. Here we show that a deficiency in the production of mitochondrial aspartate is an important abnormality in these autoimmune T cells. Shortage of mitochondrial aspartate disrupted the regeneration of the metabolic cofactor nicotinamide adenine dinucleotide, causing ADP deribosylation of the endoplasmic reticulum (ER) sensor GRP78/BiP. As a result, ribosome-rich ER membranes expanded, promoting co-translational translocation and enhanced biogenesis of transmembrane TNF. ERrich T cells were the predominant TNF producers in the arthritic joint. Transfer of intact mitochondria into T cells, as well as supplementation of exogenous aspartate, rescued the mitochondria-instructed expansion of ER membranes and suppressed TNF release and rheumatoid tissue inflammation.

Mitochondrial Dysfunction in Diseases, Longevity, and Treatment Resistance: Tuning Mitochondria Function as a Therapeutic Strategy.

Mitochondria are very important intracellular organelles because they have various functions. They produce ATP, are involved in cell signaling and cell death, and are a major source of reactive oxygen species (ROS). Mitochondria have their own DNA (mtDNA) and mutation of mtDNA or change the mtDNA copy numbers leads to disease, cancer chemo/radioresistance and aging including longevity. In this review, we discuss the mtDNA mutation, mitochondrial disease, longevity, and importance of mitochondrial dysfunction in cancer first. In the later part, we particularly focus on the role in cancer resistance and the mitochondrial condition such as mtDNA copy number, mitochondrial membrane potential, ROS levels, and ATP production. We suggest a therapeutic strategy employing mitochondrial transplantation (mtTP) for treatment-resistant cancer.

Mitotherapy: Unraveling a Promising Treatment for Disorders of the Central Nervous System and Other Systemic Conditions.

Mitochondria are key players of aerobic respiration and the production of adenosine triphosphate and constitute the energetic core of eukaryotic cells. Furthermore, cells rely upon mitochondria homeostasis, the disruption of which is reported in pathological processes such as liver hepatotoxicity, cancer, muscular dystrophy, chronic inflammation, as well as in neurological conditions including Alzheimer's disease, schizophrenia, depression, ischemia and glaucoma. In addition to the well-known spontaneous cell-to-cell transfer of mitochondria, a therapeutic potential of the transplant of isolated, metabolically active mitochondria has been demonstrated in several in vitro and in vivo experimental models of disease. This review explores the striking outcomes achieved by mitotherapy thus far, and the most relevant underlying data regarding isolated mitochondria transplantation, including mechanisms of mitochondria intake, the balance between administration and therapy effectiveness, the relevance of mitochondrial source and purity and the mechanisms by which mitotherapy is gaining ground as a promising therapeutic approach.

Requirements for successful mitochondrial transplantation.

Maintenance of mitochondrial oxidative phosphorylation capacity and other mitochondrial functions are essential for the prevention of mitochondrial dysfunction-related diseases such as neurodegenerative, cardiovascular, and liver diseases. To date, no well-known treatment modality has been developed to prevent or reduce mitochondrial dysfunction. However, a novel approach that transplants fully functional mitochondria directly into defective cells has recently caught the attention of scientists. In this review, we provide an overview of the cell/tissue source of the mitochondria to prompt cell regeneration or tissue repair in vitro and in vivo applications. The animal and human models entail that effective procedures should be used in the isolation and confirmation of mitochondrial membrane potential and function. We believe that these procedures for mitochondrial transplantation for tissue or cell culture will confirm intact, viable, and free from contamination isolated mitochondria from the appropriate sources.

The effects of mitochondrial transplantation in acetaminophen-induced liver toxicity in rats.

AIMS: Acetaminophen (APAP) toxicity is one of the leading causes of acute liver injury-related death and liver failure worldwide. In many studies, mitochondrial dysfunction has been identified as an important cause of damage in APAP toxicity. Therefore, our study aimed to investigate the possible effects of mitochondrial transplantation on liver damage due to APAP toxicity. MAIN METHODS: APAP toxicity model was implemented by administering a toxic dose of APAP. To demonstrate the efficiency of mitochondria transplantation, it was compared with N-acetylcysteine (NAC) application, which is now clinically accepted. Mitochondrial transplantation was carried out by delivering mitochondria to the liver via the portal circulation, which was injected into the spleen. In our study, the rats were randomly divided into 6 groups as Sham, APAP, Control 1, APAP+mito, Control 2, and APAP+NAC. In the end of the experiment, histological and biochemical analysis were performed and the biodistribution of the transplanted mitochondria to target cells were also shown. KEY FINDINGS: Successful mitochondrial transplantation was confirmed and mitochondrial transplantation improved the liver histological structure to a similar level with healthy rats. Moreover, plasma ALT levels, apoptotic cells, and total oxidant levels were decreased. It was also observed that NAC treatment increased GSH levels to the highest level among the groups. However, mitochondrial transplantation was more effective than NAC application in terms of histological and functional improvement. SIGNIFICANCE: It has been evaluated that mitochondrial transplantation can be used as an important alternative or adjunctive treatment method in liver damage caused by toxic dose APAP intake.

Mitochondria transfer from early stages of erythroblasts to their macrophage niche via tunnelling nanotubes.

Adult erythropoiesis entails a series of well-coordinated events that produce mature red blood cells. One of such events is the mitochondria clearance that occurs cell-autonomously via autophagy-dependent mechanisms. Interestingly, recent studies have shown mitochondria transfer activities between various cell types. In the context of erythropoiesis, macrophages are known to interact closely with the early stages of erythroblasts to provide a specialized niche, termed erythroblastic islands (EBI). However, whether mitochondria transfer can occur in the EBI niche has not been explored. Here, we report that mitochondria transfer in the EBI niche occurs in vivo. We observed mitochondria transfer activities from the early stages of erythroblasts to macrophages in the reconstituted in vitro murine EBI via different modes, including tunnelling nanotubes (TNT). Moreover, we demonstrated that Wiskott-Aldrich syndrome protein (WASp) in macrophages mediates TNT formation and mitochondria transfer via the modulation of F-actin filamentation, thus promoting mitochondria clearance from erythroid cells, to potentially enhance their differentiation. Taken together, our findings provide novel insight into the mitochondria clearance machineries that mediate erythroid maturation.

Mitochondrial Transplantation as a Novel Therapeutic Strategy for Mitochondrial Diseases.

Mitochondria are the major source of intercellular bioenergy in the form of ATP. They are necessary for cell survival and play many essential roles such as maintaining calcium homeostasis, body temperature, regulation of metabolism and apoptosis. Mitochondrial dysfunction has been observed in variety of diseases such as cardiovascular disease, aging, type 2 diabetes, cancer and degenerative brain disease. In other words, the interpretation and regulation of mitochondrial signals has the potential to be applied as a treatment for various diseases caused by mitochondrial disorders. In recent years, mitochondrial transplantation has increasingly been a topic of interest as an innovative strategy for the treatment of mitochondrial diseases by augmentation and replacement of mitochondria. In this review, we focus on diseases that are associated with mitochondrial dysfunction and highlight studies related to the rescue of tissue-specific mitochondrial disorders. We firmly believe that mitochondrial transplantation is an optimistic therapeutic approach in finding a potentially valuable treatment for a variety of mitochondrial diseases.

Mitochondria transplantation protects traumatic brain injury via promoting neuronal survival and astrocytic BDNF.

Traumatic brain injury (TBI) is one of the leading causes of disability and paralysis around the world. Secondary injury, characterized by progressive neuronal loss and astrogliosis, plays important roles in the post-TBI cognitive impairment and mood disorder. Unfortunately, there still lacks effective treatments, particularly surgery interferences for it. Recent findings of intercellular mitochondria transfer implies a potential therapeutic value of mitochondria transplantation for TBI, which has not been tested yet. In the present study, we demonstrated a quick dysfunction of mitochondria, up-regulation of Tom20 in the injured cortex and subsequent cognitive and mood impairment. Our data demonstrated that mitochondria derived from allogeneic liver or autogeneic muscle stimulated similar microglial activation in brain parenchyma. In vitro experiments showed that exogenous mitochondria could be easily internalized by neurons, astrocytes, and microglia, except for oligodendrocytes. Mitochondria transplantation effectively rescued neuronal apoptosis, restored the expression of Tom20 and the phosphorylation of JNK. Further analysis revealed that mitochondria transplantation in injured cortex induced a significant up-regulation of BDNF in reactive astrocytes, improved animals' spatial memory and alleviated anxiety. In together, our data indicate that mitochondria transplantation may has the potential of clinical translation for TBI treatment, in combination with surgery.

Is It Possible to Treat Infertility with Stem Cells?

Infertility is a major health problem, and despite improved treatments over the years, there are still some conditions that cannot be treated successfully using a conventional approach. Therefore, new options are being considered and one of them is cell therapy using stem cells. Stem cell treatments for infertility can be divided into two major groups, the first one being direct transplantation of stem cells or their paracrine factors into reproductive organs and the second one being in vitro differentiation into germ cells or gametes. In animal models, all of these approaches were able to improve the reproductive potential of tested animals, although in humans there is still too little evidence to suggest successful use. The reasons for lack of evidence are unavailability of proper material, the complexity of explored biological processes, and ethical considerations. Despite all of the above-mentioned hurdles, researchers were able to show that in women, it seems to be possible to improve some conditions, but in men, no similar clinically important improvement was achieved. To conclude, the data presented in this review suggest that the treatment of infertility with stem cells seems plausible, because some types of treatments have already been tested in humans, achieving live births, while others show great potential only in animal studies, for now.

Mitochondrial Transfer Improves Cardiomyocyte Bioenergetics and Viability in Male Rats Exposed to Pregestational Diabetes.

Offspring born to diabetic or obese mothers have a higher lifetime risk of heart disease. Previously, we found that rat offspring exposed to late-gestational diabetes mellitus (LGDM) and maternal high-fat (HF) diet develop mitochondrial dysfunction, impaired cardiomyocyte bioenergetics, and cardiac dysfunction at birth and again during aging. Here, we compared echocardiography, cardiomyocyte bioenergetics, oxidative damage, and mitochondria-mediated cell death among control, pregestational diabetes mellitus (PGDM)-exposed, HF-diet-exposed, and combination-exposed newborn offspring. We hypothesized that PGDM exposure, similar to LGDM, causes mitochondrial dysfunction to play a central, pathogenic role in neonatal cardiomyopathy. We found that PGDM-exposed offspring, similar to LGDM-exposed offspring, have cardiac dysfunction at birth, but their isolated cardiomyocytes have seemingly less bioenergetics impairment. This finding was due to confounding by impaired viability related to poorer ATP generation, more lipid peroxidation, and faster apoptosis under metabolic stress. To mechanistically isolate and test the role of mitochondria, we transferred mitochondria from normal rat myocardium to control and exposed neonatal rat cardiomyocytes. As expected, transfer provides a respiratory boost to cardiomyocytes from all groups. They also reduce apoptosis in PGDM-exposed males, but not in females. Findings highlight sex-specific differences in mitochondria-mediated mechanisms of developmentally programmed heart disease and underscore potential caveats of therapeutic mitochondrial transfer.

Study of Mesenchymal Stem Cell-Mediated Mitochondrial Transfer in In Vitro Models of Oxidant-Mediated Airway Epithelial and Smooth Muscle Cell Injury.

Mesenchymal stem cells (MSCs) have emerged as an attractive candidate for cell-based therapy. In the past decade, many animal and pilot clinical studies have demonstrated that MSCs are therapeutically beneficial for the treatment of obstructive lung diseases such as asthma and chronic obstructive pulmonary disease (COPD). However, due to the scarcity of adult human MSCs, human-induced pluripotent stem cells mesenchymal stem cells (iPSCs) are now increasingly used as a source of MSCs. iPSCs are derived by reprogramming somatic cells from a wide variety of tissues such as skin biopsies and then differentiating them into iPSC-MSCs. One of the mechanisms through which MSCs exert their protective effects is mitochondrial transfer. Specifically, transfer of mitochondria from iPSC-MSCs to lung cells was shown to protect lung cells against oxidative stress-induced mitochondrial dysfunction and apoptosis and to reduce lung injury and inflammation in in vivo models of lung disease. In this chapter, we detail our methods to visualize and quantify iPSC-MSC-mediated mitochondrial transfer and to study its effects on oxidant-induced airway epithelial and smooth muscle cell models of acute airway cell injury.

One step forward: extracellular mitochondria transplantation.

Mitochondria play a key role in cellular energy production and contribute to cell metabolism, homeostasis, intracellular signalling and organelle's quality control, among other roles. Viable, respiratory-competent mitochondria exist also outside the cells. Such extracellular/exogenous mitochondria occur in the bloodstream, being released by platelets, activated monocytes and endothelial progenitor cells. In the nervous system, the cerebrospinal fluid contains mitochondria discharged by astrocytes. Various pathologies, including the cardiovascular and neurodegenerative diseases, are associated with mitochondrial dysfunction. A strategy to reverse dysfunction and restore cell normality is the transplantation of mitochondria (freshly isolated from a healthy tissue) into the zone at risk, such as the ischemic heart and/or damaged nervous tissue. The functional exogenous mitochondria will replace the harmed ones, ensuing cardioprotective and neuroprotective effects. The diversity of transplantation settings (in vitro, in animal models and patients) offered variable answers (including lack of consensus) on efficacy of this strategy. Therefore, a critical overview of the current and future trends in mitochondrial transplantation seems to be required. Here, we outline the recent developments on (i) extracellular mitochondria types and roles, (ii) transplantation protocols, (iii) mechanisms of mitochondrial incorporation, (iv) the benefit of extracellular mitochondria transplantation in human health and diseases and (v) open questions that deserve urgent answers.