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. 2021 Mar 26;12:632709. doi: 10.3389/fimmu.2021.632709

Figure 3.

Figure 3

sMD-2 in plasma mediates IL-8 secretion by HUVEC. (A) Diagram shows two approaches to study sMD-2’s contribution to HUVEC IL-8 secretion. In approach 1, sMD-2 is depleted from SS plasma and in approach 2, sMD-2 is added to AA plasma. Plasma (2%) from both approaches were incubated with HUVEC for 18 hours and IL-8 secretion into the media was measured by ELISA. (B) Representative western blot of SS plasma after sMD-2 depletion. The IgG western blot was shown as loading controls. (C) IL-8 secretion by HUVEC after incubation with MD-2-depleted SS plasma. (D) Western blot of recombinant sMD-2 in conditioned medium from CHO cells transfected with MD-2-containing plasmids or non-transfected control CHO media. The total protein stain was shown as loading controls (E) sMD-2 CHO-conditioned medium or control CHO medium (10%) were added to HUVEC culture media along with 2% AA plasma in the presence or absence of TLR4 inhibitor, TAK-242 (1 µM), and incubated with HUVEC for 18 hours. The secreted IL-8 was measured by ELISA. Bars in C and E are means ± SD (n=3). *p<0.05, **p<0.01.