Extended Data Fig. 2: LPS-induced change in mitochondrial morphology.
a, BMDMs stimulated with LPS for 24 h or nonstimulated (NS). Representative images left to right: Live cell confocal images showing Mitotracker Deep Red FM, fixed cell super-resolution structured illuminated microscopy images showing TOMM20 signal and live cell super-resolution structured illuminated microscopy images showing Mitotracker green. Scale bars are 20 μm for all confocal images and 5 μm for super-resolution images. Scale bars for inset images are 2 μm. b, BMDMs were stimulated with LPS for 3–24 h LPS or NS and then treated with 5 nM Mitotracker Deep Red FM for 20 min and imaged by confocal live cell microscopy. Mitochondrial object number, length, area and junctions per network were measured in 20 regions of interest (ROIs) (15 μm2) per biological replicates (n = 5 (6 h), n = 6 (3 h, 9 h, 18 h, 24 h), n = 7 (NS)). c, Western blot and quantification of VDAC and TOMM20 normalized to tubulin loading control in BMDMs treated with LPS for 3–24 h or NS (n = 3). Blots show three biological replicates for each condition and data are mean normalized to NS and vinculin loading control. d, Representative images of BMDMs that were NS or stimulated with LPS for 24 h imaged by structured illumination microscopy (Elyra7) showing ATP synthase (magenta), TOMM20 (green) and nucleus (blue). Scale bars are 2 μm for whole cell images and 1 μm for inset image. Quantifications of mitochondrial morphology and ATP synthase density are mean +/- S.E.M. of whole cells from 3 biological replicates (n = 31 (NS), n = 34 (24 h LPS)). All data are mean +/- S.E.M. of biological replicates. P values displayed above graphs calculated using two-tailed Student’s t-test for paired comparisons or one-way ANOVA for multiple comparisons.
