Figure 8. Syrosingopine or MCT4 knockdown suppresses the expression of EC adhesion molecules in HPAECs activated by Hypoxia/Reoxygenation and IL-1β.

(A) Experimental scheme for HPAECs subjected to 0.5% O₂ for 18 hours in the presence of syrosingopine (5 µM) or MCT4 siRNA followed by reoxygenation for 8 hours in the presence of IL-1β (1 ng/ml). (B) mRNA levels of VCAM1 and ICAM1 in syrosingopine-vs vehicle-treated HPAECs, that were activated by Hypoxia/Reoxygenation and IL-1β. (C) THP1 monocyte adhesion to inflamed ECs. THP1 monocyte cells, labeled with Green CMFDA Dye, were introduced on a monolayer of HPAECs that had been subjected to the indicated experimental conditions. Following a 90-minute incubation period, floating cells were washed away and adhered THP1 cells were visualized using a fluorescent microscope and subsequently quantified. Representative images of fluorescent THP1 cells attached to ECs in different experimental groups are presented. Scale bar, 200 μm. (D) mRNA expression of VCAM1 and ICAM1 in HPAECs transfected with control or MCT4 siRNA and subjected to Hypoxia/Reoxygenation and IL-1β (E) THP1 monocyte adhesion to inflamed ECs. Labeled THP1 cells were introduced on a monolayer of HPAECs that had been subjected to the indicated experimental conditions. Following incubation, adhered THP1 cells were visualized quantified as in C. Representative images of fluorescent THP1 cells attached to ECs in different experimental conditions are presented. Scale bar, 200 μm. Data are represented as mean ± SEM. Statistics were determined by one-way ANOVA with Sidak correction for multiple comparisons. n=3–4; *, P < 0.05; **, P <0.01; ***, P <0.001; ****, P < 0.0001; ns, not statistically significant. Nx, Normoxia; Hx, Hypoxia/Reoxygenation; Veh, vehicle; Syro, syrosingopine; C, negative control siRNA; MCT4si, MCT4siRNA.