Fig. 6

S1PR1 was a functional target of miR-301a-3p from TiO₂-induced EVs in HUVECs. a Volcano plot of the DE-miRNAs. The black dots represent miRNAs that are not differentially expressed between PBS-EVs and TiO₂-EVs, and the red dots and green dots represent the upregulated and downregulated miRNAs, respectively. b Heat map of the eleven upregulated miRNAs compared to PBS-EVs. c RT-PCR analysis of differentially expressed miRNAs in PBS-EVs and TiO₂-EVs. Data are shown as mean ± SD (n = 3). d The relative luciferase activity of wild-type (wt) and mutant-type (mt) 3′-UTR of S1PR1 was determined. Data are shown as mean ± SD (n = 3). e Western blotting analysis of VE-cadherin, S1PR1, and VEGFR2. f RT-PCR analysis of S1PR1 expression in HUVECs which were treated with a miR-301a-3p mimic. Data are shown as mean ± SD (n = 3). g Immunofluorescence analysis of VE-cadherin, S1PR1, and VEGFR2. miR-301a-3p was overexpressed via a miR-301a-3p mimic or downregulated using a miR-301a-3p inhibitor in HUVECs, respectively. Scale bars = 10 μm. Nuclei, Blue (DAPI); S1PR1, VE-cadherin or VEGFR2, Red. Statistical analysis in (c, d and f) was measured by one-way ANOVA