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. Author manuscript; available in PMC: 2009 Oct 1.
Published in final edited form as: Transl Res. 2008 Sep 4;152(4):165–177. doi: 10.1016/j.trsl.2008.07.007

Figure-3. Modulation of PS-mediated erythrocyte binding to immobilized TSP by annexin V, heparin, enoxaparin, HDS, CSA, and sulfatide.

Figure-3

Panel A: Annexin V reduces PS binding sites on the red cell surface (measured as mean cell fluorescence or MCF) and inhibits erythrocyte adhesion to immobilized TSP. Erythrocytes (1×108 cells/ml containing 12% PS-positive cells) were pre-incubated in the presence or absence of the indicated concentrations of annexin-V, and then assessed for their adhesive potential (diamonds) and residual surface PS (circles) as described. Values presented are the means ± SD from 3 (annexin V between 3 and 28 μg/ml) to 5 (annexin V at 57 μg/ml) different experiments. The number of PS-positive erythrocytes adhered in the absence of annexin V was 639 ± 147 cells/mm2. *Changes noted with annexin V at 57 μg/ml were statistically significant at P<0.05.

Panel B: Heparin and enoxaparin inhibit PS-mediated erythrocyte binding to immobilized thrombospondin in a concentration-dependent manner. Red cells (2×108 cells/ml containing 12% PS-positive cells) were pre-treated with the indicated concentrations (0.1 to 50 U/ml) of heparin (circles) or Lovenox (squares), and then assessed for their adhesive potential using a TSP matrix containing 50 ng protein per cm2 as described in methods. Values presented are the means ± SD from 5 (Lovenox), or 9 (heparin) different experiments. *Inhibitory effects noted with heparin (between 0.5 and 50 U/ml) and Lovenox (between 0.5 and 50 U/ml) were statistically significant at P<0.05. The number of PS-positive erythrocytes adhered in the absence of modulators were 632 ± 241 cells/mm2.

Panel C:Effects of glycosaminoglycans (GAGs), and sulfatide on PS-mediated erythrocyte binding to immobilized thrombospondin. Effects of high molecular weight dextran sulfate, MW 500,000 (HDS, 200 μg/ml), chondroitin sulfate A (CSA, 200 μg/ml), and bovine brain sulfatide (200 μg/ml) on PS-mediated RBC-TSP binding were evaluated using erythrocytes (2×108 cells/ml) containing 12% PS and TSP matrix at 50 ng/cm2. Values presented are the means ± SD from 6 different experiments. Heparin (200 μg/ml, equivalent to 50 units/ml) tested in parallel with other GAGs is shown for paired comparison with HDS and CSA. *Red cell adhesion in the presence of heparin, HDS and CSA were different from the respective paired controls as assessed by the paired student’s t-test at P<0.01.