fig4

Figure 4. EV-NID1-treated DCs promote the proliferation and Treg differentiation of CD4⁺ T cells. (A) Schematic diagram of the co-culture of CD4⁺ T cells and BMDCs. CD4⁺ T cells isolated by magnetic bead sorting were co-cultured with BMDCs subjected to different treatments (ratio, 1:1), and cells were harvested after 72 h for flow cytometry analysis; (B) Analysis of CD4⁺ T cell proliferation by flow cytometry after co-culture with NID1 NC EV (10 μg/mL), NID1 KD EV (10 μg/mL), or LPS (50 ng/mL); (C) Analysis of Treg proportion in CD4⁺ T cells by flow cytometry after co-culture with NID1 NC EV (10 μg/mL), NID1 KD EV (10 μg/mL), or LPS (50 ng/mL). Data are presented as the mean ± SD, with statistical significance determined by one-way ANOVA (Tukey’s test). Biological replicates are represented as individual data points. ^*P < 0.05; ^**P < 0.01; ^***P < 0.001; ^****P < 0.0001. NID1: Nidogen 1; NC: negative control; KD: knockdown; EV: extracellular vesicle; DC: dendritic cell; BMDC: bone marrow-derived dendritic cell; CD4: cluster of differentiation 4; Treg: regulatory T cell; FOXP3: forkhead box protein P3; IL-2: interleukin-2; CFSE: carboxyfluorescein succinimidyl ester; CTV: CellTrace Violet; LPS: lipopolysaccharide; SD: standard deviation; ANOVA: analysis of variance; ns: not significant.