No abstract

Background: Surgical reconstruction of congenital heart defects is often limited by the nonresorbable material used to approximate normal anatomy. In contrast, biologic scaffold materials composed of resorbable non-cross-linked extracellular matrix (ECM) have been used for tissue reconstruction of multiple organs and are replaced by host tissue. Preparation of whole organ ECM by decellularization through vascular perfusion can maintain much of the native three-dimensional (3D) structure, strength, and tissue-specific composition. A 3D cardiac ECM (C-ECM) biologic scaffold material would logically have structural and functional advantages over materials such as Dacron™ for myocardial repair, but the in vivo remodeling characteristics of C-ECM have not been investigated to date. Methods and Results: A porcine C-ECM patch or Dacron patch was used to reconstruct a full-thickness right ventricular outflow tract (RVOT) defect in a rat model with end points of structural remodeling function at 16 weeks. The Dacron patch was encapsulated by dense fibrous tissue and showed little cellular infiltration. Echocardiographic analysis showed that the right ventricle of the hearts patched with Dacron were dilated at 16 weeks compared to presurgery baseline values. The C-ECM patch remodeled into dense, cellular connective tissue with scattered small islands of cardiomyocytes. The hearts patched with C-ECM showed no difference in the size or function of the ventricles as compared to baseline values at both 4 and 16 weeks. Conclusions: The C-ECM patch was associated with better functional and histomorphological outcomes compared to the Dacron patch in this rat model of RVOT reconstruction.

Perfusion-based whole organ decellularization has recently gained interest in the field of tissue engineering as a means to create site-specific extracellular matrix scaffolds, while largely preserving the native architecture of the scaffold. To date, this approach has been utilized in a variety of organ systems, including the heart, lung, and liver [1][2][3][4][5] . Previous decellularization methods for tissues without an easily accessible vascular network have relied upon prolonged exposure of tissue to solutions of detergents, acids, or enzymatic treatments as a means to remove the cellular and nuclear components from the surrounding extracellular environment [6][7][8] . However, the effectiveness of these methods hinged upon the ability of the solutions to permeate the tissue via diffusion. In contrast, perfusion of organs through the natural vascular system effectively reduced the diffusion distance and facilitated transport of decellularization agents into the tissue and cellular components out of the tissue. Herein, we describe a method to fully decellularize an intact porcine heart through coronary retrograde perfusion. The protocol yielded a fully decellularized cardiac extracellular matrix (c-ECM) scaffold with the three-dimensional structure of the heart intact. Our method used a series of enzymes, detergents, and acids coupled with hypertonic and hypotonic rinses to aid in the lysis and removal of cells. The protocol used a Trypsin solution to detach cells from the matrix followed by Triton X-100 and sodium deoxycholate solutions to aid in removal of cellular material. The described protocol also uses perfusion speeds of greater than 2 L/min for extended periods of time. The high flow rate, coupled with solution changes allowed transport of agents to the tissue without contamination of cellular debris and ensured effective rinsing of the tissue. The described method removed all nuclear material from native porcine cardiac tissue, creating a site-specific cardiac ECM scaffold that can be used for a variety of applications. Video LinkThe video component of this article can be found at