清除牛心包组织内细胞的方法

METHOD OF DECELLULARIZATION OF BOVINE PERICARDIUM

置换生物心脏瓣膜的病人一般不需抗凝治疗。但由于生物心脏瓣膜的逐渐损坏,术后5年至10年需行再次换瓣手术。根据生物心脏瓣膜的病理检查发现,瓣叶的钙化变性是瓣膜原发性失功的主要形式,而瓣膜内的残存细胞和细胞碎片则起了重要的初始钙化源的作用。我们清除细胞的方法是分步从新鲜的牛心包清除组织内原有的细胞。在清除细胞的过程中,为了调整牛心包内自溶体酶的活性,各片5×10nm^2的新鲜牛心包被浸在温度分别为15℃、20℃和25℃的pH7.4、0.05MHEPES的新鲜缓冲液内。实验结果表明,经过三阶段总共72小时对牛心包的处理,牛心包组织内的细胞被减少了大约70％,而组织内的胶原纤维和弹性纤维仍保持结构完整。

Bioprosthetic heart valves generally need no anticoagulant therapy, but they are progressively destroyed in patients and require reimplantation 5-10 years after their placement. Along with pathologic findings, cusp calcification and degeneration are the principal modes of primary tissue failure of bioprostheses. In bioprostheses, residual cells and cellular debris provide nucleation sites for progressive calcification. Our decellularization method includes the steps of removing native cells from fresh bovine pericardium. During the decellularization process, the fresh pericardial patches were incubated in a 0. 05 M HEPES buffered solution (pH7. 4) at a temperature of 15℃, 20℃ and 25 ℃,. respectively, in order to adjust activity of the autolytic enzymes in the pericardium. The results showed that after three stages to totaling a period of 72 hours for treatment of bovine pericardia, the number of cells was reduced by about 70% from pericardium, and the collagen bundles and elastic fibers were well preserved.