Figure 2. Neutralizing antibody against toll-like receptor 4 (TLR4) reduces retinol-binding protein 4 (RBP4)-induced endothelial inflammation. A-E: Human retinal microvascular endothelial cells (HRECs) were exposed to TLR4 neutralizing antibody (mAb) or control immunoglobulin G (IgG) at 10 µg/ml for 2 h, followed by RBP4 treatment for 24 h. Cell media were collected for enzyme-linked immunosorbent assay (ELISA)-based quantification of soluble extracellular levels of (A) vascular cell adhesion molecule 1 (VCAM-1), (B) intracellular adhesion molecule 1 (ICAM-1), (C) endothelial cell selectin (E-selectin), (D) monocyte chemoattractant protein (MCP-1), and (E) interleukin 6 (IL-6). F: Representative images of monocyte adherence to HRECs after the indicated treatment. Data are representative of three separate experiments. Magnification: 20×. G: Adherent monocytes were counted in four randomly selected visual fields (20× objective) for each treatment group. ***, p<0.001 versus bovine serum albumin (BSA); #, p<0.05; ##, p<0.01; ###, p<0.001 versus RBP4 by one-way analysis of variance (ANOVA) with Tukey’s post hoc test.