Plasmatic ADAMTS-13 metalloprotease and von Willebrand factor in children with cyanotic congenital heart disease

Abstract

Changes in plasma von Willebrand factor concentration (VWF:Ag) and ADAMTS-13 activity (the metalloprotease that cleaves VWF physiologically) have been reported in several cardiovascular disorders with prognostic implications. We therefore determined the level of these proteins in the plasma of children with cyanotic congenital heart disease (CCHD) undergoing surgical treatment. Forty-eight children were enrolled (age 0.83 to 7.58 years). Measurements were performed at baseline and 48 h after surgery. ELISA, collagen-binding assays and Western blotting were used to estimate antigenic and biological activities, and proteolysis of VWF multimers. Preoperatively, VWF:Ag and ADAMTS-13 activity were decreased (65 and 71% of normal levels considered as 113 (105-129) U/dL and 91 ± 24% respectively, P < 0.003) and correlated (r = 0.39, P = 0.0064). High molecular weight VWF multimers were not related, suggesting an interaction of VWF with cell membranes, followed by proteolytic cleavage. A low preoperative ADAMTS-13 activity, a longer activated partial thromboplastin time and the need for cardiopulmonary bypass correlated with postoperative bleeding (P < 0.05). Postoperatively, ADAMTS-13 activity increased but less extensively than VWF:Ag (respectively, 2.23 and 2.83 times baseline, P < 0.0001), resulting in an increased VWF:Ag/ADAMTS-13 activity ratio (1.20 to 1.54, respectively, pre- and postoperative median values, P = 0.0029). ADAMTS-13 consumption was further confirmed by decreased ADAMTS-13 antigenic concentration (0.91 ± 0.30 to 0.70 ± 0.25 µg/mL, P < 0.0001) and persistent proteolysis of VWF multimers. We conclude that, in pediatric CCHD, changes in circulating ADAMTS-13 suggest enzyme consumption, associated with abnormal structure and function of VWF.

Figure 1. Representative Western immunoblot showing plasma von Willebrand factor multimeric structure (pattern observed in 36 of 48 children with cyanotic congenital heart disease). At baseline, reduction of high molecular weight multimers (HMW), with increased density of low molecular weight fractions (LMW, five bands migrating just above IgM) compared with control plasma (relative optical density of LMW multimers 0.44 (0.11-0.56) and 0.36 (0.25-0.55), respective median value and range in patients and controls, P = 0.0081). Postoperatively, the density of LMW fractions remained high [0.45 (0.17-0.63), P = 0.5360 versus baseline]. The migration of immunoglobulin M (IgM, 950 kDa) is indicated.

Figure 2. Preoperative (filled squares) and postoperative (open squares) correlations of plasma ADAMTS-13 activity (A) and von Willebrand factor activity (B, VWF:CB) with VWF antigenic concentration (VWF:Ag) in children with cyanotic congenital heart disease. Preoperative and postoperative mean values of ADAMTS-13 activity adjusted for VWF:Ag were 84 and 122%, respectively (P = 0.0066, ANCOVA). In B, the slopes are different (P = 0.0477), indicating lowered VWF:CB relative to VWF:Ag. Preoperative and postoperative adjusted mean values of VWF:CB were 62 and 79%, respectively (P = 0.0395, ANCOVA).