MiRNA-31-5p inhibits SP-induced autophagy in PAECs. (A) Abundance of LC3B in PAECs treated the combination of SP (10 µM) and miRNA-31-5p mimic, as assessed by Western blotting. (B) Protein expression of LC3B, Beclin-1 and ATG7 in PAECs treated with the combination of SP (10 µM) and miRNA-31-5p mimic, as assessed by immunofluorescence assays (scale bar, 25 µm; magnification, ×20). (C) Autophagic vacuoles in the cellular cytoplasm of PAECs treated the combination of SP (10 µM) and miRNA-31-5p mimic, as evaluated by transmission electron microscopy (scale bar, 2 µm; magnification, ×10,000). Red arrows indicate the autophagic vacuoles. (D) Protein expression of LC3B, Beclin-1 and ATG7 in PAECs treated with the combination of SP (10 µM) and miRNA-31-5p inhibitor, as assessed by Western blotting. (E) The abundance of LC3B in PAECs treated with the combination of SP (10 µM) and miRNA-31-5p inhibitor, as assessed by immunofluorescence assays (scale bar, 25 µm; magnification, ×20). (F) Autophagic vacuoles in the cellular cytoplasm of PAECs treated with the combination of SP (10 µM) and miRNA-31-5p inhibitor, as evaluated by transmission electron microscopy (scale bar, 2 µm; magnification, ×10,000). Red arrows indicate the autophagic vacuoles. *P<0.05. There were at least three replicates in each group available for analysis. SP, spermidine; PAECs, pulmonary artery endothelial cells.