Integrated spatial multiomics reveals fibroblast fate during tissue repair

Deshka S Foster,Dhananjay Wagh,Clement D Marshall,Michael S Hu,Howard Y Chang,Oscar Da Silva,Shamik Mascharak,Gunsagar S Gulati,Michael Januszyk,Karen Tolentino,Michael T Longaker,Gerlinde Wernig,Ashley L Titan,Jeffrey A Norton,Kathryn E Yost,Derrick C Wan,Geoffrey C Gurtner,Ankit Salhotra,Aaron M Newman,Dominic Henn,Kellen Chen,Heather E Des Jardins-Park,Alan Nguyen ,Ryan C Ransom ,John A Coller ,Austin Burcham ,Malini Chinta ,Ruth Ellen Jones ,Tripp Leavitt

doi:10.1073/pnas.2110025118

Deshka S Foster, Dhananjay Wagh + Show 27 more

Open Access

https://doi.org/10.1073/pnas.2110025118

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Abstract

In the skin, tissue injury results in fibrosis in the form of scars composed of dense extracellular matrix deposited by fibroblasts. The therapeutic goal of regenerative wound healing has remained elusive, in part because principles of fibroblast programming and adaptive response to injury remain incompletely understood. Here, we present a multimodal -omics platform for the comprehensive study of cell populations in complex tissue, which has allowed us to characterize the cells involved in wound healing across both time and space. We employ a stented wound model that recapitulates human tissue repair kinetics and multiple Rainbow transgenic lines to precisely track fibroblast fate during the physiologic response to skin injury. Through integrated analysis of single cell chromatin landscapes and gene expression states, coupled with spatial transcriptomic profiling, we are able to impute fibroblast epigenomes with temporospatial resolution. This has allowed us to reveal potential mechanisms controlling fibroblast fate during migration, proliferation, and differentiation following skin injury, and thereby reexamine the canonical phases of wound healing. These findings have broad implications for the study of tissue repair in complex organ systems.

Highlights

In the skin, tissue injury results in fibrosis in the form of scars composed of dense extracellular matrix deposited by fibroblasts
Among fluorescence activated cell sorting (FACS)-isolated, lineage-negative [29], Rainbow wound fibroblasts (Fig. 2A and SI Appendix, Fig. S3A), we found that most cells fell into the putative category of reticular fibroblasts rather than papillary (CD26+/SCA1−) or hypodermal (DLK1+/−/ SCA1+) (SI Appendix, Fig. S3B)
We found that wound healing fibroblasts showed down-regulation of mechanotransduction- and fibrosis-related pathways with FAK inhibitor (FAKi) treatment (SI Appendix, Fig. S11C)

Summary

Integrated spatial multiomics reveals fibroblast fate during tissue repair

Through integrated analysis of single cell chromatin landscapes and gene expression states, coupled with spatial transcriptomic profiling, we are able to impute fibroblast epigenomes with temporospatial resolution This has allowed us to reveal potential mechanisms controlling fibroblast fate during migration, proliferation, and differentiation following skin injury, and thereby reexamine the canonical phases of wound healing. These findings have broad implications for the study of tissue repair in complex organ systems. The Rainbow mouse model is a four-color reporter system that permits precise clonal analysis and lineage tracing Using this model with phenotypepaired single-cell RNA and ATAC sequencing (scRNA-seq and scATAC-seq), we are able to define the spatial and temporal heterogeneity of wound fibroblasts with unique granularity. This work defines the spatial and temporal dynamics of the fibroblast response to injury and provides a multimodal -omics framework for future studies in tissue repair

Results

Discussion

Materials and Methods