bims-mireme 2021-06-06 papers

bims-mireme

Biomed News

on Mitochondria in regenerative medicine

Issue of 2021–06–06
fourteen papers selected by
Brian Spurlock, University of Alabama at Birmingham

Symmetric and asymmetric activation of hematopoietic stem cells.
Mitochondrial activity of human umbilical cord mesenchymal stem cells.
Mitochondria in neurogenesis: Implications for mitochondrial diseases.
The role of metabolism in directed differentiation versus trans-differentiation of cardiomyocytes.
Studying Mitochondrial Network Formation by In Vivo and In Vitro Reconstitution Assay.
Melatonin reshapes the mitochondrial network and promotes intercellular mitochondrial transfer via tunneling nanotubes (TNTs) after ischemic-like injury in hippocampal HT22 cells.
Assessment of Mitochondrial Reactive Oxygen Species and Redox Regulation in Stem Cells.
MIT-001 Restores Human Placenta-Derived Mesenchymal Stem Cells by Enhancing Mitochondrial Quiescence and Cytoskeletal Organization.
Evaluation of Mitochondria Content and Function in Live Cells by Multicolor Flow Cytometric Analysis.
Mitochondrial Dysfunction in Mitochondrial Medicine: Current Limitations, Pitfalls, and Tomorrow.
Mitochondrial apoptotic priming is a key determinant of cell fate upon p53 restoration.
Mitophagy and Oxidative Stress: The Role of Aging.
Metabolic Analysis of Mouse Hematopoietic Stem and Progenitor Cells.
Generation of three human induced pluripotent stem cell lines, LUMCi024-A, LUMCi025-A, and LUMCi026-A, from two patients with combined oxidative phosphorylation deficiency 8 and a related control.

Curr Opin Hematol. 2021 Jul 01. 28(4): 262-268

Symmetric and asymmetric activation of hematopoietic stem cells.

Dirk Loeffler, Timm Schroeder.

PURPOSE OF REVIEW: Hematopoietic stem cells (HSCs) are in an inactive quiescent state for most of their life. To replenish the blood system in homeostasis and after injury, they activate and divide. HSC daughter cells must then decide whether to return to quiescence and metabolic inactivity or to activate further to proliferate and differentiate and replenish lost blood cells. Although the regulation of HSC activation is not well understood, recent discoveries shed new light on involved mechanisms including asymmetric cell division (ACD).
RECENT FINDINGS: HSC metabolism has emerged as a regulator of cell fates. Recent evidence suggests that cellular organelles mediating anabolic and catabolic processes can be asymmetrically inherited during HSC divisions. These include autophagosomes, mitophagosomes, and lysosomes, which regulate HSC quiescence. Their asymmetric inheritance has been linked to future metabolic and translational activity in HSC daughters, showing that ACD can regulate the balance between HSC (in)activity.
SUMMARY: We discuss recent insights and remaining questions in how HSCs balance activation and quiescence, with a focus on ACD.

DOI: https://doi.org/10.1097/MOH.0000000000000644
Brain Circ. 2021 Jan-Mar;7(1):7(1): 33-36

Mitochondrial activity of human umbilical cord mesenchymal stem cells.

Blaise M Cozene, Eleonora Russo, Rita Anzalone, Giampiero La Rocca, Cesario V Borlongan.

  Human umbilical cord mesenchymal stem cells (hUC-MSCs) serve as a potential cell-based therapy for degenerative disease. They provide immunomodulatory and anti-inflammatory properties, multipotent differentiation potential and are harvested with no ethical concern. It is unknown whether MSCs collected from different areas of the human umbilical cord elicit more favorable effects than others. Three MSC populations were harvested from various regions of the human umbilical cord: cord lining (CL-MSCs), perivascular region (PV-MSCs), and Wharton's jelly (WJ-MSCs). Mesenchymal markers (CD90 and CD73) were expressed by all three cell populations. Stemness marker (OCT4), endothelial cell adhesion molecular marker (CD146), and monocyte-macrophage marker (CD14) were expressed by WJ-MSCs, PV-MSCs, and CL-MSCs, respectively. Stroke presents with oxygen and glucose deprivation and leads to dysfunctional mitochondria and consequently cell death. Targeting the restoration of mitochondrial function in the stroke brain through mitochondrial transfer may be effective in treating stroke. In vitro exposure to ambient and OGD conditions resulted in CL-MSCs number decreasing the least post-OGD/R exposure, and PV-MSCs exhibiting the greatest mitochondrial activity. All three hUC-MSC populations presented similar metabolic activity and survival in normal and pathologic environments. These characteristics indicate hUC-MSCs potential as a potent therapeutic in regenerative medicine.

Keywords:  Bioenergetics; Wharton's Jelly; ischemic diseases; mitochondria; stem cell therapy; stroke; umbilical cord mesenchymal stem cells

DOI:  https://doi.org/10.4103/bc.bc_15_21
Stem Cells. 2021 Jun 05.

Mitochondria in neurogenesis: Implications for mitochondrial diseases.

Dario Brunetti, Werner Dykstra, Stephanie Le, Annika Zink, Alessandro Prigione.

  Mitochondria are organelles with recognized key roles in cellular homeostasis, including bioenergetics, redox, calcium signaling, and cell death. Mitochondria are essential for neuronal function, given the high energy demands of the human brain. Consequently, mitochondrial diseases affecting oxidative phosphorylation (OXPHOS) commonly exhibit neurological impairment. Emerging evidence suggests that mitochondria are important not only for mature postmitotic neurons but also for the regulation of neural progenitor cells (NPCs) during the process of neurogenesis. These recent findings put mitochondria as central regulator of cell fate decisions during brain development. OXPHOS mutations may disrupt the function of NPCs and thereby impair the metabolic programming required for neural fate commitment. Promoting the mitochondrial function of NPCs could therefore represent a novel interventional approach against incurable mitochondrial diseases.

Keywords:  NPCs; iPSCs; mitochondria; mitochondrial diseases; neurogenesis

DOI:  https://doi.org/10.1002/stem.3425
Semin Cell Dev Biol. 2021 May 29. pii: S1084-9521(21)00127-0. [Epub ahead of print]

The role of metabolism in directed differentiation versus trans-differentiation of cardiomyocytes.

James W S Jahng, Mao Zhang, Joseph C Wu.

  The advent of induced pluripotent stem cells (iPSCs) and identification of transcription factors for cardiac reprogramming have raised hope to cure heart disease, the leading cause of death in the world. Our knowledge in heart development and molecular barriers of cardiac reprogramming is advancing, but many hurdles are yet to be overcome for clinical translation. Importantly, we lack a full understanding of molecular mechanisms governing cell fate conversion toward cardiomyocytes. In this review, we will discuss the role of metabolism in directed differentiation versus trans-differentiation of cardiomyocytes. Cardiomyocytes exhibit a unique metabolic feature distinct from PSCs and cardiac fibroblasts, and there are multiple overlapping molecular mechanisms underlying metabolic reprogramming during cardiomyogenesis. We will discuss key metabolic changes occurring during cardiomyocytes differentiation from PSCs and cardiac fibroblasts, and the potential role of metabolic reprogramming in the enhancement strategies for cardiomyogenesis. Only when such details are discovered will more effective strategies to enhance the de novo production of cardiomyocytes be possible.

Keywords:  Cardiomyocytes; Differentiation; Fibroblasts; IPSCs; Metabolism; Reprogramming

DOI:  https://doi.org/10.1016/j.semcdb.2021.05.018
Methods Mol Biol. 2021 ;2276 333-341

Studying Mitochondrial Network Formation by In Vivo and In Vitro Reconstitution Assay.

Wanqing Du, Xiangjun Di, Qian Peter Su.

  Mitochondria change their morphologies from small isolated vesicles to large continuous networks across the cell cycles. The mitochondrial network formation (MNF) plays an important role in maintaining mitochondrial DNA integrity and interchanging mitochondrial materials. The disruption of the mitochondrial network affects mitochondrial functions, such as ATP production, integration of metabolism, calcium homeostasis, and regulation of apoptosis, leading to the abnormal development and several human diseases including neurodegenerative disease. In this unit, we describe the method of studying MNF, which is driven by microtubule-dependent motor protein, by in vivo imaging and single-molecule in vitro reconstitution assays.

Keywords:  In vitro reconstitution system; KIF5B; Mitochondrial network formation (MNF); Single-molecule

DOI:  https://doi.org/10.1007/978-1-0716-1266-8_25
J Pineal Res. 2021 Jun 04. e12747

Melatonin reshapes the mitochondrial network and promotes intercellular mitochondrial transfer via tunneling nanotubes (TNTs) after ischemic-like injury in hippocampal HT22 cells.

Maria Gemma Nasoni, Silvia Carloni, Barbara Canonico, Sabrina Burattini, Erica Cesarini, Stefano Papa, Marica Pagliarini, Patrizia Ambrogini, Walter Balduini, Francesca Luchetti.

  Mitochondrial dysfunction is considered one of the hallmarks of ischemia/reperfusion injury. Mitochondria are plastic organelles that undergo continuous biogenesis, fusion and fission. They can be transferred between cells through tunneling nanotubes (TNTs), dynamic structures that allow the exchange of proteins, soluble molecules and organelles. Maintaining mitochondrial dynamics is crucial to cell function and survival. The present study aimed to assess the effects of melatonin on mitochondrial dynamics, TNT formation and mitochondria transfer in HT22 cells exposed to oxygen/glucose deprivation followed by reoxygenation (OGD/R). The results showed that melatonin treatment during the reoxygenation phase reduced mitochondrial reactive oxygen species (ROS) production, improved cell viability and increased the expression of PGC1α and SIRT3. Melatonin also preserved the expression of the membrane translocase proteins TOM20 and TIM23, and of the matrix protein HSP60, which are involved in mitochondrial biogenesis. Moreover, it promoted mitochondrial fusion and enhanced the expression of MFN2 and OPA1. Remarkably, melatonin also fostered mitochondrial transfer between injured HT22 cells through TNT connections. These results provide new insights into the effect of melatonin on mitochondrial network reshaping and cell survival. Fostering TNTs formation represents a novel mechanism mediating the protective effect of melatonin in ischemia/reperfusion injury.

Keywords:  HT22; Melatonin; mitochondrial network; oxygen-glucose deprivation; tunneling nanotubes

DOI:  https://doi.org/10.1111/jpi.12747
Methods Mol Biol. 2021 ;2277 289-297

Assessment of Mitochondrial Reactive Oxygen Species and Redox Regulation in Stem Cells.

Madhavee Thumiah-Mootoo, Tina Podinic, Mireille Khacho.

  Mitochondrial reactive oxygen species (mtROS) and redox regulation play an important role in stem cell maintenance and cell fate decisions. Although changes in mtROS and redox homeostasis represent a physiological mechanism to drive stem cell commitment and differentiation, dysregulation of this system can lead to defects in stem cell maintenance and regenerative capacity. This chapter explains the methods used to assess mitochondrial superoxide levels and redox regulation in stem cell populations.

Keywords:  Antioxidant; Electron transport chain; Metabolism; Mitochondria; Oxidative stress; Reactive oxygen species (ROS); Redox; Stem cell fate; Stem cells

DOI:  https://doi.org/10.1007/978-1-0716-1270-5_18
Int J Mol Sci. 2021 May 11. pii: 5062. [Epub ahead of print]22(10):

MIT-001 Restores Human Placenta-Derived Mesenchymal Stem Cells by Enhancing Mitochondrial Quiescence and Cytoskeletal Organization.

Won Dong Yu, Yu Jin Kim, Min Jeong Cho, Gi Jin Kim, Soon Ha Kim, Myung Joo Kim, Jung Jae Ko, Jae Ho Lee.

  Inflammation is a major cause of several chronic diseases and is reported to be recovered by the immuno-modulation of mesenchymal stem cells (MSCs). While most studies have focussed on the anti-inflammatory roles of MSCs in stem cell therapy, the impaired features of MSCs, such as the loss of homeostasis by systemic aging or pathologic conditions, remain incompletely understood. In this study, we investigated whether the altered phenotypes of human placenta-derived MSCs (hPD-MSCs) exposed to inflammatory cytokines, including TNF-α and IFN-γ, could be protected by MIT-001, a small anti-inflammatory and anti-necrotic molecule. MIT-001 promoted the spindle-like shape and cytoskeletal organization extending across the long cell axis, whereas hPD-MSCs exposed to TNF-α/IFN-γ exhibited increased morphological heterogeneity with an abnormal cell shape and cytoskeletal disorganization. Importantly, MIT-001 improved mitochondrial distribution across the cytoplasm. MIT-001 significantly reduced basal respiration, ATP production, and cellular ROS levels and augmented the spare respiratory capacity compared to TNF-α/IFN-γ-exposed hPD-MSCs, indicating enhanced mitochondrial quiescence and homeostasis. In conclusion, while TNF-α/IFN-γ-exposed MSCs lost homeostasis and mitochondrial quiescence by becoming over-activated in response to inflammatory cytokines, MIT-001 was able to rescue mitochondrial features and cellular phenotypes. Therefore, MIT-001 has therapeutic potential for clinical applications to treat mitochondrion-related inflammatory diseases.

Keywords:  MIT-001; cytoskeleton; homeostasis; inflammation; mesenchymal stem cells; mitochondria

DOI:  https://doi.org/10.3390/ijms22105062
Methods Mol Biol. 2021 ;2276 203-213

Evaluation of Mitochondria Content and Function in Live Cells by Multicolor Flow Cytometric Analysis.

Hsiu-Han Fan, Tsung-Lin Tsai, Ivan L Dzhagalov, Chia-Lin Hsu.

  To evaluate how a cell responds to the external stimuli, treatment, or alteration of the microenvironment, the quantity and quality of mitochondria are commonly used as readouts. However, it is challenging to apply mitochondrial analysis to the samples that are composed of mixed cell populations originating from tissues or when multiple cell populations are of interest, using methods such as Western blot, electron microscopy, or extracellular flux analysis.Flow cytometry is a technique allowing the detection of individual cell status and its identity simultaneously when used in combination with surface markers. Here we describe how to combine mitochondria-specific dyes or the dyes targeting the superoxide produced by mitochondria with surface marker staining to measure the mitochondrial content and activity in live cells by flow cytometry. This method can be applied to all types of cells in suspension and is particularly useful for analysis of samples composed of heterogeneous cell populations.

Keywords:  FACS; Flow cytometry; Mitochondria; Quantification; Reactive oxygen species

DOI:  https://doi.org/10.1007/978-1-0716-1266-8_15
Methods Mol Biol. 2021 ;2276 1-29

Mitochondrial Dysfunction in Mitochondrial Medicine: Current Limitations, Pitfalls, and Tomorrow.

Naig Gueguen, Guy Lenaers, Pascal Reynier, Volkmar Weissig, Marvin Edeas.

  Until recently restricted to hereditary mitochondrial diseases, mitochondrial dysfunction is now recognized as a key player and strategic factor in the pathophysiological of many human diseases, ranging from the metabolism, vascular, cardiac, and neurodegenerative diseases to cancer. Because of their participation in a myriad of cellular functions and signaling pathways, precisely identifying the cause of mitochondrial "dysfunctions" can be challenging and requires robust and controlled techniques. Initially limited to the analysis of the respiratory chain functioning, these analytical techniques now enlarge to the analyses of mitochondrial and cellular metabolism, based on metabolomic approaches.Here, we address the methods used to assay mitochondrial dysfunction, with a highlight on the techniques used in diagnosis on tissues and cells derived from patients, the information they provide, and their strength and weakness.Targeting mitochondrial dysfunction by various strategies is a huge challenge, requires robust methods of evaluation, and should be able to take into consideration the mitochondria dynamics and localization. The future of mitochondrial medicine is strongly related to a perfect comprehension of its dysfunction.

Keywords:  Bioenergetics; Devices; Metabolomics; Mitochondria evaluation; Mitochondrial dysfunctions

DOI:  https://doi.org/10.1007/978-1-0716-1266-8_1
Proc Natl Acad Sci U S A. 2021 Jun 08. pii: e2019740118. [Epub ahead of print]118(23):

Mitochondrial apoptotic priming is a key determinant of cell fate upon p53 restoration.

Francisco J Sánchez-Rivera, Jeremy Ryan, Yadira M Soto-Feliciano, Mary Clare Beytagh, Lucius Xuan, David M Feldser, Michael T Hemann, Jesse Zamudio, Nadya Dimitrova, Anthony Letai, Tyler Jacks.

  Reactivation of p53 in established tumors typically results in one of two cell fates, cell cycle arrest or apoptosis, but it remains unclear how this cell fate is determined. We hypothesized that high mitochondrial priming prior to p53 reactivation would lead to apoptosis, while low priming would lead to survival and cell cycle arrest. Using a panel of Kras-driven, p53 restorable cell lines derived from genetically engineered mouse models of lung adenocarcinoma and sarcoma (both of which undergo cell cycle arrest upon p53 restoration), as well as lymphoma (which instead undergo apoptosis), we show that the level of mitochondrial apoptotic priming is a critical determinant of p53 reactivation outcome. Cells with high initial priming (e.g., lymphomas) lacked sufficient reserve antiapoptotic capacity and underwent apoptosis after p53 restoration. Forced BCL-2 or BCL-XL expression reduced priming and resulted in survival and cell cycle arrest. Cells with low initial priming (e.g., lung adenocarcinoma and sarcoma) survived and proceeded to arrest in the cell cycle. When primed by inhibition of their antiapoptotic proteins using genetic (BCL-2 or BCL-XL deletion or BAD overexpression) or pharmacologic (navitoclax) means, apoptosis resulted upon p53 restoration in vitro and in vivo. These data demonstrate that mitochondrial apoptotic priming is a key determining factor of cell fate upon p53 activation. Moreover, it is possible to enforce apoptotic cell fate following p53 activation in less primed cells using p53-independent drugs that increase apoptotic priming, including BH3 mimetic drugs.

Keywords:  apoptosis; cell cycle arrest; cell fate; p53

DOI:  https://doi.org/10.1073/pnas.2019740118
Antioxidants (Basel). 2021 May 17. pii: 794. [Epub ahead of print]10(5):

Mitophagy and Oxidative Stress: The Role of Aging.

Anna De Gaetano, Lara Gibellini, Giada Zanini, Milena Nasi, Andrea Cossarizza, Marcello Pinti.

  Mitochondrial dysfunction is a hallmark of aging. Dysfunctional mitochondria are recognized and degraded by a selective type of macroautophagy, named mitophagy. One of the main factors contributing to aging is oxidative stress, and one of the early responses to excessive reactive oxygen species (ROS) production is the induction of mitophagy to remove damaged mitochondria. However, mitochondrial damage caused at least in part by chronic oxidative stress can accumulate, and autophagic and mitophagic pathways can become overwhelmed. The imbalance of the delicate equilibrium among mitophagy, ROS production and mitochondrial damage can start, drive, or accelerate the aging process, either in physiological aging, or in pathological age-related conditions, such as Alzheimer's and Parkinson's diseases. It remains to be determined which is the prime mover of this imbalance, i.e., whether it is the mitochondrial damage caused by ROS that initiates the dysregulation of mitophagy, thus activating a vicious circle that leads to the reduced ability to remove damaged mitochondria, or an alteration in the regulation of mitophagy leading to the excessive production of ROS by damaged mitochondria.

Keywords:  Alzheimer; PINK1; Parkinson; Reactive Oxygen Species; aging; mitochondria; mitophagy

DOI:  https://doi.org/10.3390/antiox10050794
Methods Mol Biol. 2021 ;2308 107-115

Metabolic Analysis of Mouse Hematopoietic Stem and Progenitor Cells.

Liang He, Amina Aouida, Amira Mehtar, Houda Haouas, Fawzia Louache.

  The intrinsic properties of self-renewal and differentiation of hematopoietic stem and progenitor cells (HSPCs) play a critical role in the regeneration of mature hematopoietic cells at steady state and in stress conditions including bleeding, inflammation and aging. Common techniques such as flow cytometry and genetic methods have answered many questions about their intrinsic and extrinsic regulation. Using these approaches, it was demonstrated that HSPCs in the bone marrow demonstrate low rates of proliferation and apoptosis. This dormant phenotype is associated with a low production of reactive oxygen species and low mitochondrial activity.Here, we describe the methodology to characterize the physiologic state of HSPCs isolated from their native hematopoietic organ using flow cytometry-based assays. These protocols allow evaluation of their ROS levels and activated signaling pathways under various conditions.

Keywords:  Bone marrow processing; Cell metabolism; Flow cytometry; HSPCs; ROS levels; ROS signaling

DOI:  https://doi.org/10.1007/978-1-0716-1425-9_9
Stem Cell Res. 2021 May;pii: S1873-5061(21)00220-8. [Epub ahead of print]53 102374

Generation of three human induced pluripotent stem cell lines, LUMCi024-A, LUMCi025-A, and LUMCi026-A, from two patients with combined oxidative phosphorylation deficiency 8 and a related control.

Ruben W J van Helden, Matthew J Birket, Christian Freund, Christaan H Arendzen, Harald M Mikkers, Valeria Orlova, René I de Coo, Christine L Mummery, Milena Bellin.

Combined Oxidative Phosphorylation Deficiency 8 (COXPD8) is an autosomal recessive disorder causing lethal childhood-onset hypertrophic cardiomyopathy. Homozygous or compound heterozygous mutations in the nuclear-encoded mitochondrial alanyl-tRNA synthetase 2 (AARS2) gene underly the pathology. We generated induced pluripotent stem cells (hiPSCs) from two patients carrying the heterozygous compound c.1774 C>T, c.2188 G>A and c.2872 C>T AARS2 mutations, as well as a related healthy control carrying the c.2872 C>T AARS2 mutation. All hiPSC-lines expressed pluripotency markers, maintained a normal karyotype, and differentiated towards the three germ layer derivatives in vitro. These lines can be used to model COXPD8 or mitochondrial dysfunction.

DOI: https://doi.org/10.1016/j.scr.2021.102374