Bovine pericardial patch with reduced crosslinking time preserves matrix integrity and mitigates calcification in rat subcutaneous tissue.

Prosthetic valves derived from bovine pericardium (BP) are crucial for heart valve replacement, yet current crosslinking methods with glutaraldehyde can lead to immune responses and calcification. This study evaluated the effects of reducing the glutaraldehyde crosslinking time from 10 to 5 days in bovine pericardial patches for use as heart valve substitutes. In addition to examining the physical properties of the BP, the study analyzed the biocompatibility, tissue structure, and calcification of the pericardial tissue. BPs were processed using two protocols based on the fixation time with glutaraldehyde: BP10d (10 days) and BP5d (5 days). All samples were treated with glutamic acid to neutralize residual aldehyde groups from the glutaraldehyde. Subsequently, the resulting material was assessed for mechanical and thermal properties and histologically using light and scanning electron microscopy. Post-implantation histological evaluation and calcium content determination were conducted after 7, 14, 30, 60 and 120 days. The calcification was a rare occurrence. However, some samples from the BP10d group displayed positive Von Kossa staining, indicating mineral deposition. Chemical analysis using ICP-OES revealed low calcium concentrations in the explants of both groups, with higher concentrations observed in the BP10d group during the later analysis periods. Mechanical and thermal stability assessments showed no significant differences between experimental groups. Histological examination revealed more collagen and elastic fibers deformation, and inflammation in the BP10d group compared to the BP5d group. The revised manufacturing protocol, with a 5-day fixation time, showed promising anti-calcifying activity, biocompatibility, and tissue preservation.