High-Throughput Image Analysis of Lipid-Droplet-Bound Mitochondria
| Autor: | Nour Alsabeeh, Marc Liesa, Orian S. Shirihai, Kiana Mahdaviani, Nathanael Miller, Mayuko Segawa, Dane M. Wolf |
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| Rok vydání: | 2021 |
| Předmět: |
0301 basic medicine
Chemistry Significant difference Computational biology Mitochondrion Thresholding Mitochondrial fragmentation 03 medical and health sciences Cytosol 030104 developmental biology 0302 clinical medicine Lipid droplet Segmentation Throughput (business) 030217 neurology & neurosurgery |
| Zdroj: | Methods in Molecular Biology ISBN: 9781071612651 |
| DOI: | 10.1007/978-1-0716-1266-8_22 |
| Popis: | Changes to mitochondrial architecture are associated with various adaptive and pathogenic processes. However, quantification of changes to mitochondrial structures is limited by the yet unmet challenge of defining the borders of each individual mitochondrion within an image. Here, we describe a novel method for segmenting primary brown adipocyte (BA) mitochondria images. We describe a granular approach to quantifying subcellular structures, particularly mitochondria in close proximity to lipid droplets: peridroplet mitochondria. In addition, we lay out a novel machine-learning-based mitochondrial segmentation method that eliminates the bias of manual mitochondrial segmentation and improves object recognition compared to conventional thresholding analyses. By applying these methods, we discovered a significant difference between cytosolic and peridroplet BA mitochondrial H2O2 production and validated the machine-learning algorithm in BA via norepinephrine-induced mitochondrial fragmentation and comparing manual analyses to the automated analysis. This approach provides a high-throughput analysis protocol to quantify ratiometric probes in subpopulations of mitochondria in adipocytes. |
| Databáze: | OpenAIRE |
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