Protocol for generation of a time-resolved cellular interactome during tissue remodeling in adult mice

• Published in: STAR protocols 2023-12, Vol.4 (4), p.102638-102638, Article 102638
• Main Authors: Groppa, Elena, Tung, Lin Wei, Mattevi, Stefania, Ritso, Morten, Rossi, Fabio M.V., Martini, Paolo
• Format: Article
• Language: English
• Subjects: Animals; Bioinformatics; Cell Biology; Cell Communication; Cell isolation; Computer sciences; Flow Cytometry/Mass Cytometry; Mice; Molecular Biology; Muscle, Skeletal; Protocol; RNA; RNAseq; Sequence analysis; Sequence Analysis, RNA; Sequencing; Stem Cells
• ISSN: 2666-1667; 2666-1667
• DOI: 10.1016/j.xpro.2023.102638

Efficient skeletal muscle regeneration necessitates fine-tuned coordination among multiple cell types through an intricate network of intercellular communication. We present a protocol for generation of a time-resolved cellular interactome during tissue remodeling. We describe steps for isolating distinct cell populations from skeletal muscle of adult mice after acute damage and extracting RNA from purified cells prior to the generation of RNA sequencing data. We then detail procedures for generating and deciphering a time- and lineage-resolved model of intercellular crosstalk. For complete details on the use and execution of this protocol, please refer to Groppa et al. (2023).1 [Display omitted] •Multiple staining and gating strategies to purify cell subsets from skeletal muscle•An optimized protocol to extract sufficient and reliable RNA to generate RNAseq data•RNAseq dataset of five cell populations from a time course during muscle regeneration•An informatic analysis based on a high-resolution time course of RNAseq data Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Efficient skeletal muscle regeneration necessitates fine-tuned coordination among multiple cell types through an intricate network of intercellular communication. We present a protocol for generation of a time-resolved cellular interactome during tissue remodeling. We describe steps for isolating distinct cell populations from skeletal muscle of adult mice after acute damage and extracting RNA from purified cells prior to the generation of RNA sequencing data. We then detail procedures for generating and deciphering a time- and lineage-resolved model of intercellular crosstalk.