bims-mireme 2021-08-22 papers

bims-mireme

Biomed News

on Mitochondria in regenerative medicine

Issue of 2021–08–22
seven papers selected by
Brian Spurlock, The University of North Carolina at Chapel Hill

Mitochondrial fatty acid utilization increases chromatin oxidative stress in cardiomyocytes.
Automated Optical Tweezers Manipulation to Transfer Mitochondria from Fetal to Adult MSCs to Improve Antiaging Gene Expressions.
Development-related mitochondrial properties of retinal pigment epithelium cells derived from hEROs.
Dynamic regulation of mitochondrial-endoplasmic reticulum crosstalk during stem cell homeostasis and aging.
Intermittent Starvation Promotes Maturation of Human Embryonic Stem Cell-Derived Cardiomyocytes.
Multiple phosphorylation events of the mitochondrial membrane protein, TTM1, regulate cell death during senescence.
Protocol for rapid manipulation of mitochondrial morphology in living cells using inducible counter mitochondrial morphology (iCMM).

Proc Natl Acad Sci U S A. 2021 Aug 24. pii: e2101674118. [Epub ahead of print]118(34):

Mitochondrial fatty acid utilization increases chromatin oxidative stress in cardiomyocytes.

Ivan Menendez-Montes, Salim Abdisalaam, Feng Xiao, Nicholas T Lam, Shibani Mukherjee, Luke I Szweda, Aroumougame Asaithamby, Hesham A Sadek.

  The inability of adult mammalian cardiomyocytes to proliferate underpins the development of heart failure following myocardial injury. Although the newborn mammalian heart can spontaneously regenerate for a short period of time after birth, this ability is lost within the first week after birth in mice, partly due to increased mitochondrial reactive oxygen species (ROS) production which results in oxidative DNA damage and activation of DNA damage response. This increase in ROS levels coincides with a postnatal switch from anaerobic glycolysis to fatty acid (FA) oxidation by cardiac mitochondria. However, to date, a direct link between mitochondrial substrate utilization and oxidative DNA damage is lacking. Here, we generated ROS-sensitive fluorescent sensors targeted to different subnuclear compartments (chromatin, heterochromatin, telomeres, and nuclear lamin) in neonatal rat ventricular cardiomyocytes, which allowed us to determine the spatial localization of ROS in cardiomyocyte nuclei upon manipulation of mitochondrial respiration. Our results demonstrate that FA utilization by the mitochondria induces a significant increase in ROS detection at the chromatin level compared to other nuclear compartments. These results indicate that mitochondrial metabolic perturbations directly alter the nuclear redox status and that the chromatin appears to be particularly sensitive to the prooxidant effect of FA utilization by the mitochondria.

Keywords:  metabolism; mitochondria; reactive oxygen species

DOI:  https://doi.org/10.1073/pnas.2101674118
Small. 2021 Aug 19. e2103086

Automated Optical Tweezers Manipulation to Transfer Mitochondria from Fetal to Adult MSCs to Improve Antiaging Gene Expressions.

Adnan Shakoor, Bin Wang, Lei Fan, Lingchi Kong, Wendi Gao, Jiayu Sun, Kwan Man, Gang Li, Dong Sun.

  Mitochondrial dysfunction is considered to be an important factor that leads to aging and premature aging diseases. Transferring mitochondria to cells is an emerging and promising technique for the therapy of mitochondrial deoxyribonucleic acid (mtDNA)-related diseases. This paper presents a unique method of controlling the quality and quantity of mitochondria transferred to single cells using an automated optical tweezer-based micromanipulation system. The proposed method can automatically, accurately, and efficiently collect and transport healthy mitochondria to cells, and the recipient cells then take up the mitochondria through endocytosis. The results of the study reveal the possibility of using mitochondria from fetal mesenchymal stem cells (fMSCs) as a potential source to reverse the aging-related phenotype and improve metabolic activities in adult mesenchymal stem cells (aMSCs). The results of the quantitative polymerase chain reaction analysis show that the transfer of isolated mitochondria from fMSCs to a single aMSC can significantly increase the antiaging and metabolic gene expression in the aMSC. The proposed mitochondrial transfer method can greatly promote precision medicine for cell therapy of mtDNA-related diseases.

Keywords:  antiaging; automatic micromanipulation; microfluidics; mitochondrial transfer; optical tweezers

DOI:  https://doi.org/10.1002/smll.202103086
Int J Ophthalmol. 2021 ;14(8): 1138-1150

Development-related mitochondrial properties of retinal pigment epithelium cells derived from hEROs.

Hao-Jue Xu, Qi-You Li, Ting Zou, Zheng-Qin Yin.

   AIM: To explore the temporal mitochondrial characteristics of retinal pigment epithelium (RPE) cells obtained from human embryonic stem cells (hESC)-derived retinal organoids (hEROs-RPE), to verify the optimal period for using hEROs-RPE as donor cells from the aspect of mitochondria and to optimize RPE cell-based therapeutic strategies for age-related macular degeneration (AMD).
METHODS: RPE cells were obtained from hEROs and from spontaneous differentiation (SD-RPE). The mitochondrial characteristics were analyzed every 20d from day 60 to 160. Mitochondrial quantity was measured by MitoTracker Green staining. Transmission electron microscopy (TEM) was adopted to assess the morphological features of the mitochondria, including their distribution, length, and cristae. Mitochondrial membrane potentials (MMPs) were determined by JC-1 staining and evaluated by flow cytometry, reactive oxygen species (ROS) levels were evaluated by flow cytometry, and adenosine triphosphate (ATP) levels were measured by a luminometer. Differences between two groups were analyzed by the independent-samples t-test, and comparisons among multiple groups were made using one-way ANOVA or Kruskal-Wallis H test when equal variance was not assumed.
RESULTS: hEROs-RPE and SD-RPE cells from day 60 to 160 were successfully differentiated from hESCs and expressed RPE markers (Pax6, MITF, Bestrophin-1, RPE65, Cralbp). RPE features, including a cobblestone-like morphology with tight junctions (ZO-1), pigments and microvilli, were also observed in both hEROs-RPE and SD-RPE cells. The mitochondrial quantities of hEROs-RPE and SD-RPE cells both peaked at day 80. However, the cristae of hEROs-RPE mitochondria were less mature and abundant than those of SD-RPE mitochondria at day 80, with hEROs-RPE mitochondria becoming mature at day 100. Both hEROs-RPE and SD-RPE cells showed low ROS levels from day 100 to 140 and maintained a normal MMP during this period. However, hEROs-RPE mitochondria maintained a longer time to produce high levels of ATP (from day 120 to 140) than SD-RPE cells (only day 120).
CONCLUSION: hEROs-RPE mitochondria develop more slowly and maintain a longer time to supply high-level energy than SD-RPE mitochondria. From the mitochondrial perspective, hEROs-RPE cells from day 100 to 140 are an optimal cell source for treating AMD.

Keywords:  human embryonic stem cells, retinal degenerative diseases; mitochondria; retinal organoids; retinal pigment epithelium cells

DOI:  https://doi.org/10.18240/ijo.2021.08.02
Cell Death Dis. 2021 Aug 16. 12(9): 794

Dynamic regulation of mitochondrial-endoplasmic reticulum crosstalk during stem cell homeostasis and aging.

Weiping Lin, Shuxun Chen, Yan Wang, Ming Wang, Wayne Yuk-Wai Lee, Xiaohua Jiang, Gang Li.

Cellular therapy exerts profound therapeutic potential for curing a broad spectrum of diseases. Adult stem cells reside within a specified dynamic niche in vivo, which is essential for continuous tissue homeostatic maintenance through balancing self-renewal with lineage selection. Meanwhile, adult stem cells may be multipotent or unipotent, and are present in both quiescent and actively dividing states in vivo of the mammalians, which may switch to each other state in response to biophysical cues through mitochondria-mediated mechanisms, such as alterations in mitochondrial respiration and metabolism. In general, stem cells facilitate tissue repair after tissue-specific homing through various mechanisms, including immunomodulation of local microenvironment, differentiation into functional cells, cell "empowerment" via paracrine secretion, immunoregulation, and intercellular mitochondrial transfer. Interestingly, cell-source-specific features have been reported between different tissue-derived adult stem cells with distinct functional properties due to the different microenvironments in vivo, as well as differential functional properties in different tissue-derived stem cell-derived extracellular vehicles, mitochondrial metabolism, and mitochondrial transfer capacity. Here, we summarized the current understanding on roles of mitochondrial dynamics during stem cell homeostasis and aging, and lineage-specific differentiation. Also, we proposed potential unique mitochondrial molecular signature features between different source-derived stem cells and potential associations between stem cell aging and mitochondria-endoplasmic reticulum (ER) communication, as well as potential novel strategies for anti-aging intervention and healthy aging.

DOI: https://doi.org/10.1038/s41419-021-03912-4
Front Cell Dev Biol. 2021 ;9 687769

Intermittent Starvation Promotes Maturation of Human Embryonic Stem Cell-Derived Cardiomyocytes.

Jingsi Yang, Nan Ding, Dandan Zhao, Yunsheng Yu, Chunlai Shao, Xuan Ni, Zhen-Ao Zhao, Zhen Li, Jianquan Chen, Zheng Ying, Miao Yu, Wei Lei, Shijun Hu.

  Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) represent an infinite cell source for cardiovascular disease modeling, drug screening and cell therapy. Despite extensive efforts, current approaches have failed to generate hPSC-CMs with fully adult-like phenotypes in vitro, and the immature properties of hPSC-CMs in structure, metabolism and electrophysiology have long been impeding their basic and clinical applications. The prenatal-to-postnatal transition, accompanied by severe nutrient starvation and autophagosome formation in the heart, is believed to be a critical window for cardiomyocyte maturation. In this study, we developed a new strategy, mimicking the in vivo starvation event by Earle's balanced salt solution (EBSS) treatment, to promote hPSC-CM maturation in vitro. We found that EBSS-induced starvation obviously activated autophagy and mitophagy in human embryonic stem cell-derived cardiomyocytes (hESC-CMs). Intermittent starvation, via 2-h EBSS treatment per day for 10 days, significantly promoted the structural, metabolic and electrophysiological maturation of hESC-CMs. Structurally, the EBSS-treated hESC-CMs showed a larger cell size, more organized contractile cytoskeleton, higher ratio of multinucleation, and significantly increased expression of structure makers of cardiomyocytes. Metabolically, EBSS-induced starvation increased the mitochondrial content in hESC-CMs and promoted their capability of oxidative phosphorylation. Functionally, EBSS-induced starvation strengthened electrophysiological maturation, as indicated by the increased action potential duration at 90% and 50% repolarization and the calcium handling capacity. In conclusion, our data indicate that EBSS intermittent starvation is a simple and efficient approach to promote hESC-CM maturation in structure, metabolism and electrophysiology at an affordable time and cost.

Keywords:  autophagy; cardiomyocyte maturation; embryonic stem cells; intermittent starvation; pluripotent stem cells

DOI:  https://doi.org/10.3389/fcell.2021.687769
Plant J. 2021 Aug 19.

Multiple phosphorylation events of the mitochondrial membrane protein, TTM1, regulate cell death during senescence.

Purva Karia, Keiko Yoshioka, Wolfgang Moeder.

  The role of mitochondria in programmed cell death (PCD) during animal growth and development is well documented, but much less is known for plants. We previously showed that the Arabidopsis thaliana triphosphate tunnel metalloenzyme (TTM) proteins TTM1 and TTM2 are tail-anchored proteins that localize in the mitochondrial outer membrane and participate in PCD during senescence and immunity, respectively. Here, we show that TTM1 is specifically involved in senescence induced by abscisic acid (ABA). Moreover, phosphorylation of TTM1 by multiple mitogen-activated protein kinases (MAPKs) regulates its function and turnover. A combination of proteomics and in vitro kinase assays revealed three major phosphorylation sites of TTM1 (Ser10, Ser437, and Ser490). Ser437, which is phosphorylated upon perception of senescence cues such as ABA and prolonged darkness, is phosphorylated by the MAP kinases MPK3 and MPK4, and Ser437 phosphorylation is essential for TTM1 function in senescence. These MPKs, together with three additional MAP kinases (MPK1, MPK7, and MPK6), also phosphorylate Ser10 and Ser490, marking TTM1 for protein turnover, which likely prevents uncontrolled cell death. Taken together, our results show that multiple MPKs regulate the function and turnover of the mitochondrial protein TTM1 during senescence-associated cell death, revealing a novel link between mitochondria and PCD.

Keywords:  Abscisic acid (ABA); Arabidopsis thaliana; MAP kinase; Triphosphate Tunnel Metalloenzyme (TTM); mitochondria; programmed cell death (PCD); protein phosphorylation; senescence

DOI:  https://doi.org/10.1111/tpj.15470
STAR Protoc. 2021 Sep 17. 2(3): 100721

Protocol for rapid manipulation of mitochondrial morphology in living cells using inducible counter mitochondrial morphology (iCMM).

Takafumi Miyamoto, Song-Iee Han, Hitoshi Shimano.

  Disruption of mitochondrial morphology occurs during various diseases, but the biological significance is not entirely clear. Here, we describe a detailed step-by-step protocol for a chemically inducible dimerization system-based synthetic protein device, termed inducible counter mitochondrial morphology. This system allows artificial manipulation of mitochondrial morphology on a timescale of minutes in living mammalian cells. We also describe an AI-assisted imaging processing approach. For complete details on the use and execution of this protocol, please refer to Miyamoto et al., 2021.

Keywords:  Biotechnology and bioengineering; Cell Biology; Molecular/Chemical Probes; Protein Biochemistry

DOI:  https://doi.org/10.1016/j.xpro.2021.100721