Highly-purified rapidly expanding clones, RECs, are superior for functional-mitochondrial transfer
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Published:2023-03-16
Issue:1
Volume:14
Page:
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ISSN:1757-6512
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Container-title:Stem Cell Research & Therapy
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language:en
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Short-container-title:Stem Cell Res Ther
Author:
Yang JiahaoORCID, Liu Lu, Oda Yasuaki, Wada Keisuke, Ago Mako, Matsuda Shinichiro, Hattori Miho, Goto Tsukimi, Kawashima Yuki, Matsuzaki Yumi, Taketani Takeshi
Abstract
Abstract
Background
Mitochondrial dysfunction caused by mutations in mitochondrial DNA (mtDNA) or nuclear DNA, which codes for mitochondrial components, are known to be associated with various genetic and congenital disorders. These mitochondrial disorders not only impair energy production but also affect mitochondrial functions and have no effective treatment. Mesenchymal stem cells (MSCs) are known to migrate to damaged sites and carry out mitochondrial transfer. MSCs grown using conventional culture methods exhibit heterogeneous cellular characteristics. In contrast, highly purified MSCs, namely the rapidly expanding clones (RECs) isolated by single-cell sorting, display uniform MSCs functionality. Therefore, we examined the differences between RECs and MSCs to assess the efficacy of mitochondrial transfer.
Methods
We established mitochondria-deficient cell lines (ρ0 A549 and ρ0 HeLa cell lines) using ethidium bromide. Mitochondrial transfer from RECs/MSCs to ρ0 cells was confirmed by PCR and flow cytometry analysis. We examined several mitochondrial functions including ATP, reactive oxygen species, mitochondrial membrane potential, and oxygen consumption rate (OCR). The route of mitochondrial transfer was identified using inhibition assays for microtubules/tunneling nanotubes, gap junctions, or microvesicles using transwell assay and molecular inhibitors.
Results
Co-culture of ρ0 cells with MSCs or RECs led to restoration of the mtDNA content. RECs transferred more mitochondria to ρ0 cells compared to that by MSCs. The recovery of mitochondrial function, including ATP, OCR, mitochondrial membrane potential, and mitochondrial swelling in ρ0 cells co-cultured with RECs was superior than that in cells co-cultured with MSCs. Inhibition assays for each pathway revealed that RECs were sensitive to endocytosis inhibitor, dynasore.
Conclusions
RECs might serve as a potential therapeutic strategy for diseases linked to mitochondrial dysfunction by donating healthy mitochondria.
Funder
JST Support for Pioneering Research Initiated by the Next Generation: SPRING SU Capacity Building Project Towards a Sustainable Society JSPS KAKENHI
Publisher
Springer Science and Business Media LLC
Subject
Cell Biology,Biochemistry, Genetics and Molecular Biology (miscellaneous),Molecular Medicine,Medicine (miscellaneous)
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