Covalent binding of aminopropanehydroxydiphosphonate to glutaraldehyde residues in pericardial bioprosthetic tissue: Stability and calcification inhibition studies

Calcification has limited the clinical utility of bioprosthetic heart valves fabricated from either glutaraldehyde-pretreated bovine pericardium or porcine aortic valves. Aminopropanehydroxydiphosphonate (APDP), covalently bound to residual aldehyde groups in glutaraldehyde-treated pericardial bioprosthetic tissue, has been shown to inhibit cardiovascular calcification in the rat subdermal model. Using 3H-labeled glutaraldehyde (GLUT) at a concentration of 0.02 M and 0.14 M 14C-labeled APDP, we assessed the effects of GLUT incubation temperature (4[deg] or 25[deg]C) and pH of the GLUT incubation solution (pH 4.0, 7.4, or 10.0) on the GLUT incorporation step and subsequent APDP binding (24 hr 25[deg]C) into bioprosthetic valve (BPV) tissue (bovine pericardium). Increased incorporation of GLUT and APDP occurred at lower GLUT incubation temperature (GLUT, 346.05 +/- 1.9 nM/mg, 4[deg]C vs 259.76 +/- 1.39 nM/mg, 25[deg]C; APDP, 57.56 +/- 4.43 nM/mg, 4[deg]C vs 36.36 +/- 0.46 nM/mg, mean +/- standard error, at 25[deg]C). There also was a greater incorporation of GLUT but not APDP at the higher glutaraldehyde pretreatment pH (GLUT, pH 10.0, 213.73 +/- 73 nM/mg vs pH 4, 132.08 +/- 43 nM/mg; APDP, pH 10.0, 51.41 +/- 12 nM/mg vs pH 4.0, 49.97 +/- 6 nM/mg). In vivo studies revealed that all groups with treated BPV implanted for 21 days in male 3-week-old CD rats demonstrated a loss of both GLUT (12%-50%) and APDP (48%-64%) compared to preimplant content. BPV implant calcification was significantly inhibited in all groups treated with APDP compared to control Ca2+ (5.54 +/- 2.1-9.64 + 1.2 [mu]g/mg, APDP pretreated, vs 93.64 +/- 11.65 [mu]g/mg, control; P [les] 0.001) despite the progressive loss of both GLUT and APDP with time. It is concluded that preincubation of BPV tissue in GLUT at lower temperature (4[deg]C) and higher pH (10.0) enhanced BPV GLUT uptake and subsequent APDP covalent binding. In addition, in the rat subdermal model, BPV tissue calcification was markedly inhibited by APDP, despite a significant loss of bound drug.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/27918/1/0000341.pd