Fig. 3: GelMA crosslinking modulates MÏ• during wound healing. | Nature Communications

Fig. 3: GelMA crosslinking modulates MÏ• during wound healing.

From: Hydrogel crosslinking modulates macrophages, fibroblasts, and their communication, during wound healing

Fig. 3

a Oxidation genes expressed by M4 in wounds with lo- and hi-GelMA. Dot size indicates the proportion of cells expressing each gene; dot color reflects expression levels. b Violin plots showing Mmp12 and Mmp9 expression across all Mϕ (left) and comparing hi- versus lo-GelMA in M4 (right). c Violin plots of Cybb expression in all Mϕ (left) and comparing hi- versus lo-GelMA in M4 (right). d RNAscope images of PWD5 wounds treated with lo versus hi-GelMA, stained for Fcgr1/CD64 (Mϕ marker) and Cybb/Nox2 (oxidation gene). Green box: magnified view and H&E stain; green dashed line: GelMA-dermis border. g: GelMA. Scale bar: 50 µm. Representative images from n = 3 (wounds). e Wounds after a 100 mg/kg luminol injection, with luminescence quantified across n = 5 mice; p = 0.02 (two-sided paired t-test). Scale bar: 2 mm. f Signature scoring for oxidative phosphorylation (GO:0006119) and glycolysis (GO:0045821) in all Mϕ and comparing hi- versus lo-GelMA in M4 Mϕs (right). g BMDM seeded on hi- or lo-GelMA or glass for 24 h, stimulated with 1 µg/ml LPS for 16 h, stained with CellROX-green detecting ROS, with quantification. Scale bar: 25 µm, n = 3 (BMDM harvests from individual mice), p < 0.0001 (one-way ANOVA, Tukey's HSD). h FLIM images of BMDM seeded on hi- versus lo-GelMA or glass for 24 h, stimulated with 1 µg/ml LPS for 16h, with quantification (right). Scale bar: 15 µm. See Supplementary Fig. 7a for FLIM analysis. n = 9, 11, 12, 12, 12, and 11 for lo/V, lo/LPS, hi/V, hi/LPS, G/V, and G/LPS, respectively (cells from 3 mice). lo/hi: p = 0.02, lo/G: p = 0.0009 (one-way ANOVA, Tukey's HSD). i BMDM seeded on hi versus lo-GelMA or glass for 24 h, stimulated with LPS for 16 h, stained with MitoTracker Red CMXRos, with mitochondrial particles and network branches quantified. See Supplementary Fig. 7c for analysis. n = 10 (ROIs from 3 mouse BMDM culture). Scale bar: 25 µm. p < 0.0001 (one-way ANOVA, Tukey's HSD). All data presented as mean ± SD. Source data are provided as a Source Data file.

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