Using R, Seurat, and CellChat to Analyze a Single-Cell Transcriptomics Dataset of Mouse Skin Wound Healing.

The process of wound healing is regulated by complex interactions between different cell types across space and time. Through the profiling of individual cells within their complex environment, single-cell transcriptomics methods enable the investigation of cellular heterogeneity, cell communication networks, and cell-cell interactions involved in the wound healing process. However, many single-cell analysis tools are run within a computer coding environment, and their more widespread use by wound healing scientists is thwarted by the apparent lack of bioinformatics expertise. Therefore, a step-by-step workflow is presented showing how to use a graphical coding environment called RStudio to perform a basic single-cell analysis of a temporal mouse excisional skin wound healing dataset. This visual and guided protocol will enable scientists with no bioinformatics background to download a previously published wound healing dataset, perform critical quality control steps, run a standard single-cell analysis workflow including dataset visualizations and cell type annotations using Seurat, run cell subtype analyses, run module scoring analyses, run cell-cell interaction analyses using CellChat, and perform integrative analyses of multiple datasets using Seurat. Narrative explanations are provided for each step in the protocol and graphical results from every line of code are presented to safely guide the user through the workflow. The goal of this visual introduction to a single-cell analysis pipeline is to enable more wound healing scientists to use bioinformatics tools directly in their own laboratories in order to facilitate deeper analyses of their own single-cell datasets as well as more widespread re-analyses of previously published single-cell datasets.