PM2.5 exposure reprograms cell cycle dynamics in uterine immune cells at single-cell resolution

Abstract

Background: Fine particulate matter (PM2.5) exposure has been associated with adverse effects on reproduction, yet the underlying cellular mechanisms remain poorly understood.

Methods: Using single-cell RNA sequencing, we systematically investigated cell cycle dynamics of immune cell populations in the mouse uterus following PM2.5 exposure. Analysis of 9,000 balanced cells was performed to identify distinct cell populations and characterize changes in cell cycle distribution and gene expression profiles.

Results: PM2.5 exposure induced distinct alterations in immune cell composition and cell cycle distributions. Notably, we observed significant changes in immune cell populations, including reductions in macrophages (510 to 58 cells), NK cells (445 to 91 cells), and granulocytes (1597 to 1 cells). Cell cycle analysis demonstrated cell type-specific responses to PM2.5 exposure: macrophages showed increased G1 phase representation (53.45%, +7.37%) with decreased G2M phase cells (18.97%, -12.79%), while NK cells exhibited relatively modest cell cycle alterations (G1: 28.6%, +2.5%; G2M: 45.1%, +2.6%; S: 26.4%, -5.1%). Differential gene expression analysis further identified crucial regulatory genes involved in cell cycle control, including Cd81 and Nrp1 in macrophages, Vps37b in NK cells. Integration of cell cycle markers with differentially expressed genes revealed distinctive phase-specific perturbations across immune cell types.

Conclusion: PM2.5 exposure induces cell type-specific alterations in cell cycle progression of uterine immune cells, which provides novel mechanistic insights into environmental pollution-induced reproductive dysfunction.

Keywords: cell cycle progression; fine particulate matter; immune cell heterogeneity; reproductive toxicity; single-cell RNA sequencing.

Figure 1

Flow chart of estrual immune cell targeted scRNA sequencing reanalysis.

Figure 2

Uterine cell atlas revealed by single-cell RNA sequencing. (A) tSNE visualization of uterine cell clusters. (B) Bar plot showing the distribution of different cell populations. (C) Heatmap of cell type annotation based on top 3 marker genes for each cluster. (D) Sankey diagram illustrating cell distribution between different groups. Ctrl, control group; PM2.5, PM2.5 exposure group; Fibr, fibroblast; Gran, granulocyte; Epit, epithelial cell; Macr, macrophage; NK, natural killer cell; Meso, mesothelial cell; Endo, endothelial cell; Peri, pericyte; SMC, smooth muscle cell.

Figure 3

PM2.5 exposure disrupts cell cycles of uterine immune cells. (A) Scatter plot showing S phase scores across different uterine cell populations. (B) Scatter plot showing G2M phase scores across different uterine cell populations. (C) Stacked bar chart displaying the percentage distribution of cell cycle phases across different immune cell subpopulations between groups. (D) Stacked bar chart showing the proportion of cell cycle phases within each immune cell type across different groups. Ctrl, control group; PM2.5, PM2.5 exposure group; Macr, macrophage; NK, natural killer cell.

Figure 4

tSNE visualization and volcano plots of differentially expressed genes in macrophages (A) and NK cells (B).

Figure 5

KEGG pathway enrichment analysis based on differentially expressed genes and gene set enrichment analysis (GSEA) based on genome-wide expression for macrophages (A) and NK cells (B).

Figure 6

Venn diagram showing the overlap between differentially expressed genes (DEGs), established marker genes for G1, S, and G2M phases, differentially expressed phase marker (DEPM) genes, and reference genes for S and G2M phases derived from Seurat package for macrophages (A) and NK cells (B). Expression of intersective items are visualized using Ridge plots between groups.