Spatiotemporal transcriptomic mapping of regenerative inflammation in skeletal muscle reveals a dynamic multilayered tissue architecture
Andreas Patsalos, … , H. Lee Sweeney, Laszlo Nagy
Andreas Patsalos, … , H. Lee Sweeney, Laszlo Nagy
Published August 27, 2024
Citation Information: J Clin Invest. 2024;134(20):e173858. https://doi.org/10.1172/JCI173858.
View: Text | PDF
Research Article Inflammation

Spatiotemporal transcriptomic mapping of regenerative inflammation in skeletal muscle reveals a dynamic multilayered tissue architecture

Abstract

Tissue regeneration is orchestrated by macrophages that clear damaged cells and promote regenerative inflammation. How macrophages spatially adapt and diversify their functions to support the architectural requirements of actively regenerating tissue remains unknown. In this study, we reconstructed the dynamic trajectories of myeloid cells isolated from acutely injured and early stage dystrophic muscles. We identified divergent subsets of monocytes/macrophages and DCs and validated markers (e.g., glycoprotein NMB [GPNMB]) and transcriptional regulators associated with defined functional states. In dystrophic muscle, specialized repair-associated subsets exhibited distinct macrophage diversity and reduced DC heterogeneity. Integrating spatial transcriptomics analyses with immunofluorescence uncovered the ordered distribution of subpopulations and multilayered regenerative inflammation zones (RIZs) where distinct macrophage subsets are organized in functional zones around damaged myofibers supporting all phases of regeneration. Importantly, intermittent glucocorticoid treatment disrupted the RIZs. Our findings suggest that macrophage subtypes mediated the development of the highly ordered architecture of regenerative tissues, unveiling the principles of the structured yet dynamic nature of regenerative inflammation supporting effective tissue repair.

Authors

Andreas Patsalos, Laszlo Halasz, Darby Oleksak, Xiaoyan Wei, Gergely Nagy, Petros Tzerpos, Thomas Conrad, David W. Hammers, H. Lee Sweeney, Laszlo Nagy

×

Figure 6

ST organization of dystrophic muscle upon intermittent Pred treatment.

Options: View larger image (or click on image) Download as PowerPoint
ST organization of dystrophic muscle upon intermittent Pred treatment. (...
(A) Left: H&E images of GAST from 2-mo D2.mdx mice treated weekly (Q.W.) for 4 weeks with Pred, used for ST. Histopathological annotation areas: regenerative muscle (yellow), necrotic/inflammatory lesions (green), healthy muscle (blue). Right: Percentage of spots in annotated areas. Each section is from a different biological replicate, and each library was obtained from a separate Visium experiment followed by bioinformatic integration to remove batch effects. (B) Enhanced subspot resolution clustering (BayesSpace) identified 7 meaningful spatial clusters (color coded) unresolved by pathologist annotations. (C) Comparison of the spatial cluster (color coded) subspot composition in untreated D2.mdx and D2.mdx treated Q.W. with Pred. UNT, untreated. (D) Spatial expression of representative glucocorticoid receptor (GR) targets, regenerative muscle, inflammation, and atrophy marker genes is shown. Myh3 indicates newly regenerating fibers, Tsc22d3 GR target engagement, Lyz2 inflammatory myeloid cells, and Trim63 atrophy-inducing pathways. (E) Representative DEGs in untreated D2.mdx versus D2.mdx+Q.W. Pred comparison, grouped in functional categories. Dot size represents the percentage of spots within a treatment group. (F) Unbiased global tissue-wide DGE of all spots in D2.mdx+Q.W. Pred versus D2.mdx-UNT (red dots indicate significant DEGs; P < 0.05, logFC > 1). Top DEG names are indicated. (G) GO pathway enrichment analysis of the DEGs in D2.mdx+Q.W. Pred versus D2.mdx-UNT ST datasets. Top significant up- and downregulated pathways are shown (P < 0.001, fold enrichment > 2). Gray box, enriched GO terms with Pred treatment; red box, downregulated terms with Pred-treatment. (H) Identification of tissue compartments in D2.mdx+Q.W. Pred-treated ST samples using NMF-based decomposition and 2-mo D2.mdx reference immune subtype expression signatures (7). Dot plot of estimated NMF weights of cell subtypes (rows) across 6 predicted NMF components (columns) corresponding to the identified cellular domains. Relative weights normalized across components for every MF subtype are shown. (I) Spatial plots show cell abundance for each immune cell type calculated in H. Scale bars: 1 mm.