Figure 5.

Single-cell transcriptomics implicates senescence pathways in PTEN-regulated mechanical ventilation-associated pulmonary fibrosis. (A) Schematic representation of the single-cell RNA sequencing (scRNA-seq) workflow. Following HCl and mechanical ventilation (MV), lung tissues were extracted, single cells were isolated, and next-generation sequencing was performed using the 10x Genomics platform. (B) Proportion of different cell types in lung tissue from PTEN^F/F and PTEN^CKO (knockout) mice under HCl+MV condition. (C) Uniform Manifold Approximation and Projection (UMAP) plot showing distinct cell populations in lung tissue from mice under HCl+MV condition. (D) UMAP plot comparing PTEN^F/F and PTEN^CKO mice under HCl+MV condition, highlighting changes in cellular distribution patterns. (E) Heatmap of the top 15 differentially expressed genes (DEGs) in lung epithelial cells (ECs) following PTEN knockout. (F) Pathway analysis in lung ECs showing significant enrichment in pathways related to cellular senescence. (G) Heatmap of the top 15 DEGs in immune cells (macrophages, monocytes, dendritic cells, mast cells, and neutrophils) following PTEN knockout. (H) Pathway analysis in immune cells showing significant enrichment in pathways related to interleukin-1 and interleukin-1 β signaling (cellular senescence-related pathways). (I) Senescence-associated secretory phenotype (SASP) scores in 11 major lung cell types (AddModuleScore), with the highest scores observed in mast cells and fibroblasts. Differences in SASP scores were observed only in lung ECs, indicating a protective effect of PTEN knockout on MV-PF through downregulation of cellular senescence in lung ECs. Comparisons between two groups were performed using independent sample t tests.