Coating with fibronectin and stromal cell-derived factor-1α of decellularized homografts used for right ventricular outflow tract reconstruction eliminates immune response-related degeneration.

OBJECTIVE: This study assesses the performance and cellular features of decellularized ovine aortic homografts coated with stromal cell-derived factor-1α (SDF-1α) and its natural linker, fibronectin (FN), after implantation in the right ventricular outflow tract of adolescent sheep. METHODS: Right ventricular outflow tract reconstructions using cryopreserved (n = 7), decellularized (n = 8), and decellularized FN/SDF-1α-coated aortic ovine homografts (n = 6) were performed. Echocardiographic, morphologic, radiographic, histologic, and immunohistochemical examinations were performed 5 months after implantation. RESULTS: There were no hemodynamic differences between groups, except for the decellularized homografts' tendency to develop more valve regurgitation (3 of 8 grafts had regurgitation >2/4). All decellularized, but coated, grafts had normal hemodynamics. Decellularized valve conduits were less calcified than cryopreserved conduits (P < .05), but coated valve conduits were free of calcification (P < .05). The same was found for pannus in the outflow parts. Immune response (CD45(+), CD45R(+), or CD11b(+) cells) was decreased in decellularized valves compared with cryopreserved grafts, but was virtually absent (P < .05) in coated grafts. Collagen organization and density in the leaflets and walls were decreased in cryopreserved and decellularized valves, but not in coated valves (P < .05). Coating improved re-endothelialization (P < .05). CONCLUSIONS: Coating of decellularized allografts with FN/SDF-1α prevents cryopreserved heart valve-mediated immune response, conduit calcification, and pannus formation and stimulates re-endothelialization.

Calcification of allograft and stentless xenograft valves for right ventricular outflow tract reconstruction: an experimental study in adolescent sheep.

OBJECTIVE: Aortic homografts were compared with pulmonary homografts in the setting of right ventricular outflow tract reconstruction in adolescent sheep. Furthermore, clinically available stentless porcine and bovine xenografts were studied as an alternative to homografts. METHODS: In 51 adolescent sheep cryopreserved aortic and pulmonary (ovine) homografts, as well as 6 different types of clinically available stentless bioprostheses (Prima Plus, Toronto SPV, Toronto BiLinx, Freestyle, Pericarbon Stentless, and Contegra) were implanted in the pulmonary position. After 5 to 6 months, the valves were explanted and studied for structural valve degeneration by means of radiographic analysis, histology, and calcium content determination. RESULTS: Pulmonary homografts calcified significantly less than aortic homografts in the wall portion. Leaflet calcification was mild, hardly detectable on radiographic analysis, and comparable between aortic and pulmonary homografts. Stentless porcine xenografts showed severe calcification in the aortic wall portion, irrespective of the antimineralization treatment. Leaflet calcification was mild and in the range of that seen in homografts. Pannus formation was present but never induced leaflet retraction or cusp immobilization. Calcification was absent in the stentless Pericarbon valve implants, but all valves showed extensive pannus overgrowth, leaflet retraction, and cusp immobilization. The Contegra valves showed wall calcification, but the leaflets were completely free of calcification and pannus. CONCLUSIONS: For right ventricular outflow tract reconstruction, the pulmonary homograft remains the first choice. All xenografts result in either calcific degeneration or cusp immobilization.

Gene expression study of monocytes/macrophages during early foreign body reaction and identification of potential precursors of myofibroblasts.

Foreign body reaction (FBR), initiated by adherence of macrophages to biomaterials, is associated with several complications. Searching for mechanisms potentially useful to overcome these complications, we have established the signaling role of monocytes/macrophages in the development of FBR and the presence of CD34(+) cells that potentially differentiate into myofibroblasts. Therefore, CD68(+) cells were in vitro activated with fibrinogen and also purified from the FBR after 3 days of implantation in rats. Gene expression profiles showed a switch from monocytes and macrophages attracted by fibrinogen to activated macrophages and eventually wound-healing macrophages. The immature FBR also contained a subpopulation of CD34(+) cells, which could be differentiated into myofibroblasts. This study showed that macrophages are the clear driving force of FBR, dependent on milieu, and myofibroblast deposition and differentiation.

Selection of an immunohistochemical panel for cardiovascular research in sheep.

Large animal research, often required as a final phase before commencing clinical trials for devices, has generally been hampered by the lack of appropriate tools to compare it with either initial small animal tests or to later evaluation in humans. Setting out to tissue engineer heart valves, we were particularly struck by the limited availability of immunohistochemical markers for sheep tissue, despite sheep being the FDA-approved animal for heart valve testing. This paper, therefore, aims to compile the available knowledge and extend the marker list with antibodies cross-reacting with sheep tissue. Thirty-seven antibodies attributed to 1 of these classes were found to be useful: (1) endothelium, (2) mesenchymal cells, myofibroblasts, and smooth muscle cells, (3) immune response, (4) primitive cells, (5) extracellular matrix, and (6) miscellaneous. Twelve had already been used in sheep tissue, but to our knowledge, the remaining 25 have not been described for use in sheep. From this result, we can conclude that the immunohistochemical panel for sheep has been extensively expanded with respect to cardiovascular research.

The remodeling of cardiovascular bioprostheses under influence of stem cell homing signal pathways.

Optimizing current heart valve replacement strategies by creating living prostheses is a necessity to alleviate complications with current bioprosthetic devices such as calcification and degeneration. Regenerative medicine, mostly in vitro tissue engineering, is the forerunner of this optimization search, yet here we show the functionality of an in vivo alternative making use of 2 homing axes for stem cells. In rats we studied the signaling pathways of stem cells on implanted bioprosthetic tissue (photooxidized bovine pericardium (POP)), by gene and protein expression analysis. We found that SDF-1alpha/CXCR4 and FN/VLA4 homing axes play a role. When we implanted vascular grafts impregnated with SDF-1alpha and/or FN as carotid artery interpositions, primitive cells were attracted from the circulation. Next, bioprosthetic heart valves, constructed from POP impregnated with SDF-1alpha and/or FN, were implanted in pulmonary position. As shown by CD90, CD34 and CD117 immunofluorescent staining they became completely recellularized after 5 months, had a normal function and biomechanical properties and specifically the combination of SDF-1alpha and FN had an optimal valve-cell phenotype.