Characteristics of the Dynamic Evolutionary Pathway of ADSCs Induced Differentiation into Astrocytes Based on scRNA-Seq Analysis.

We employed single-cell transcriptome sequencing to reveal the dynamic gene expression changes during the differentiation of adipose-derived stromal cells (ADSCs) into astrocytes. Single-cell RNA sequencing was conducted on cells from the ADSCs group and the induced groups at 2, 7, 14, and 21 days using the 10 × Chromium platform. Data underwent quality control and dimensionality reduction. Cell differentiation trajectories were constructed using Monocle2, and differentially expressed genes (DEGs) in each cell cluster were identified using differential selection algorithms. DEGs at each time point were annotated using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG), and regulatory intensities of transcription factors were analyzed using SCENIC. Integrating all groups, a total of five samples were divided into 13 cell clusters (0-12 clusters). DEGs between clusters and those compared with ADSCs at various induced time points showed distinct specificities. Monocle2 constructed cell differentiation trajectories; ADSCs can differentiate into mature astrocytes not only through the direct pathway from the 1 branch to the 3 branch but also through an indirect pathway, involving the 1 branch to the 2 branch before progressing to the 3 branch. SCENIC analysis highlighted the critical regulatory roles of STAT1, MYEF2, and SOX6 transcription factors during the differentiation of ADSCs into astrocytes. ADSCs can differentiate into mature astrocytes through two distinct pathways: direct and indirect. By the 14th day of induction, mature astrocytes have formed, characterized by a cell cycle arrest in mitosis. Further induction leads to degenerative senescence changes in differentiated cells.