| Autor: |
Fabio Bertan, Lena Wischhof, Enzo Scifo, Mihaela Guranda, Joshua Jackson, Anaïs Marsal-Cots, Antonia Piazzesi, Miriam Stork, Michael Peitz, Jochen Herbert Martin Prehn, Dan Ehninger, Pierluigi Nicotera, Daniele Bano |
| Jazyk: |
angličtina |
| Rok vydání: |
2021 |
| Předmět: |
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| Zdroj: |
Cell Reports: Methods, Vol 1, Iss 1, Pp 100002- (2021) |
| Druh dokumentu: |
article |
| ISSN: |
2667-2375 |
| DOI: |
10.1016/j.crmeth.2021.100002 |
| Popis: |
Summary: Mitochondria sustain the energy demand of the cell. The composition and functional state of the mitochondrial oxidative phosphorylation system are informative indicators of organelle bioenergetic capacity. Here, we describe a highly sensitive and reproducible method for a single-cell quantification of mitochondrial CI- and CIV-containing respiratory supercomplexes (CI∗CIV-SCs) as an alternative means of assessing mitochondrial respiratory chain integrity. We apply a proximity ligation assay (PLA) and stain CI∗CIV-SCs in fixed human and mouse brains, tumorigenic cells, induced pluripotent stem cells (iPSCs) and iPSC-derived neural precursor cells (NPCs), and neurons. Spatial visualization of CI∗CIV-SCs enables the detection of mitochondrial lesions in various experimental models, including complex tissues undergoing degenerative processes. We report that comparative assessments of CI∗CIV-SCs facilitate the quantitative profiling of even subtle mitochondrial variations by overcoming the confounding effects that mixed cell populations have on other measurements. Together, our PLA-based analysis of CI∗CIV-SCs is a sensitive and complementary technique for detecting cell-type-specific mitochondrial perturbations in fixed materials. Motivation: The detection of mitochondrial lesions in patient-derived tissues represents a powerful diagnostic tool. However, in complex organs, such as the brain, neurodegenerative processes often alter the cell composition of the tissue, with extensive microgliosis influencing population-based analyses of mitochondrial bioenergetics. To detect cell-type-specific mitochondrial lesions in tissues, we developed a PLA-based method in which complex I, complex IV-containing mitochondrial supercomplexes (CI∗CIV-SCs) can be visualized as dot-like structures. This assay may be an alternative and complementary approach for the detection of mitochondrial perturbation in fixed materials. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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