Figure 4

Effects of α ₂ AP and derivatives on plasma clot formation and lysis. (A) Clot formation and lysis was followed by monitoring turbidity (absorbance at 340 nm) every 30 seconds for 4 hours using a plate reader, of clots formed using diluted α₂AP-deficient plasma containing both 5 nM thrombin and 0.125 nM tPA, and taking the area under the turbidity versus time curve (AUC). Reactions were supplemented with increasing concentrations of purified plasma-derived α₂AP, and the AUC relative to that of reactions lacking tPA reported as a percentage. Results of a single experiment are shown. (B) shows turbidity plots for reactions similar to those shown in A, under 7 conditions described in the + or - table in panel C; for instance, (1) shows stable clot formation in the absence of tPA, (3) shows clot formation and rapid lysis in the presence of tPA, (4) shows attenuation of clot lysis in the presence of 1.0 μM α₂AP and (6) and (7) show competition of the α₂AP effect by 14 μM α₂AP(13-42) synthetic peptide (Pep) or α₂AP(13-42)-HSA fusion protein (FP), respectively. C shows the results of quantification of the experiment shown in B and repeated 4 times (n = 3 to 11 ± SD), under the conditions summarized below the graph, as indicated below the lanes. A control peptide unrelated to α₂AP corresponding to residues 54–75 of heparin cofactor II was used at 14 μM (HCII Pep). Asterisks indicate significant differences between groups compared between the horizontal lines (p < 0.05).