Semi-automated quantitation of mitophagy in cells and tissues.
Mech Ageing Dev
; 185: 111196, 2020 01.
Article
en En
| MEDLINE
| ID: mdl-31843465
ABSTRACT
Mitophagy is a natural phenomenon and entails the lysosomal degradation of mitochondria by the autophagy pathway. In recent years, the development of fluorescent pH-sensitive mitochondrial reporters has greatly facilitated the monitoring of mitophagy by distinguishing between cytosolic mitochondria or those delivered to acidic lysosomes. We recently published the mito-QC reporter, which consists of a mitochondrial outer membrane-localised tandem mCherry-GFP tag. This allows the quantification of mitophagy via the increase in red-only mCherry signal that arises when the GFP signal is quenched upon mitochondrial delivery to lysosomes. Here we develop a macro for FIJI, the mito-QC Counter, and describe its use to allow reliable and consistent semi-automated quantification of mitophagy. In this methods article we describe step-by-step how to detect and quantify mitophagy and show that mitophagy levels can be reliably calculated in different cell lines and under distinct stimuli. Finally, we show that the mito-QC Counter can be used to quantify mitophagy in tissues of mito-QC transgenic mice. We demonstrate that mitophagy levels in skeletal muscle correlates with glycolytic activity. Our present data show that the mito-QC Counter macro for FIJI enables the robust quantification of mitophagy both in vitro and in vivo.
Palabras clave
Texto completo:
1
Banco de datos:
MEDLINE
Asunto principal:
Autofagia
/
Mitofagia
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Lisosomas
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Microscopía Fluorescente
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Mitocondrias
Límite:
Animals
Idioma:
En
Revista:
Mech Ageing Dev
Año:
2020
Tipo del documento:
Article
País de afiliación:
Reino Unido