Abstract: Objective To examine the cell viability and hemodynamic functions of the stented homograft valves preserved in liquid nitrogen. Methods Cell viability of the stented homograft valve preserved in liquid nitrogen after 3 months of preservation (experimental group，n=6) was examined using flow cytometer. Fresh homografts served as control group (n=6). We prepared three sorts of stented homograft valve(21#, 23#, 25#) preserved by liquid nitrogen. In vitro pulsatile flow tests were performed on valves of two groups. Effective opening area EOA),transvalve pressure gradient and regurgitation ratio were recorded at various flow volume, and compare with Perfect bioprosthetic valve. Results The results revealed that the death ratio of endothelial cell was 10.24%±1.71% in the experimental group, and 9.09%±2.72% in the control group (P=0.441). The death ratio of smooth muscle cell was 8.76%±1.82% in the experimental group, and 7.84%±0.59% (P=0.178) in the control group. The death ratio of total cell was 8.79%±1.44% in the experimental group, and 7.40%±0.49% in the control group (P=0.072). There were no significantly differences between two groups. The transvalve pressure gradient of two groups of valve depended on the flow volume, and increased with the flow volume increasing. The transvalve pressure gradient of the stented homograft valve was higher than that of Perfect valve. Regurgitation ratio of the stented homograft valve was bigger than Perfect valve’s. EOA had an increasing character when flow volume increased. EOA of the stented homograft valve was smaller than that of Perfect valve’s. Conclusion Liquid nitrogen can offer the benefit of cell viability of the stented homograft bioprosthetic valves. The stented homograft valve has salisfactory hemodynamic functions.
ObjectiveExtracting the endothelial cells or all endothelial cells and interstitial cells from the cryopreserved homograft valves (HV), to evaluate the immunogenicity of this two kinds of decellular HV. MethodsFor extracting the endothelial cells, the leaflet and wall of the HV were decellularized by a 4-step detergent-enzymatic extraction method involving the 1% triton in combination with RNase (1μg/ml) and DNase (10μg/ml). For extracting the endothelial cells and interstitial cells, the leaflet and wall of the HV were decellularized by a 3-step detergent-enzymatic extraction method involving the 1% deoxycholic acid (DOA) in combination with RNase (20μg/ml) and DNase (200μg/ml). HLA-DR antigen expression was detected by using immunohistochemical techniques. The valve and wall of the HV were transplanted subcutaneously in the mice for 8 weeks, and the histology, calcium assay and calcium content were examined. ResultsFor the staining of the HLA-DR antigens, the immunogenic potential of the HV with extracting all endothelial cells and interstitial cells or only the endothelial cells was lower than cryopreserved HV, but it more obviously decreased for the HV with extracting all endothelial cells and interstitial cells. After 8 weeks embedded in the mice, the histological signs of the inflammatory reactions and the calcification extent to the cryopreserved HV and the HV with only extracting endothelial cells were stronger than the HV with extracting all endothelial cells and interstitial cells predominantly. And calcification extent or the inflammatory reactions to the wall of the HV were more severe than those of the leaflet. ConclusionsThe immunogenicity of the HV with extracting all endothelial cells and interstitial cells is much less than HV with only extracting endothelial cells. The histological signs of the inflammatory reactions and the calcification extent in vivo experiments is obviously decreased. For the HV with only extracting endothelial cells, though the histological signs of the inflammatory reactions slightly decrease, the calcification extent in vivo experiments is more severe, especially for the wall. The interstitial cells may be the important factor for the donor-reactive immune responses that is related to the graft calcification or destruction after implantation.