Vergleichende in vivo Untersuchung verschiedener Konservierungsverfahren von allogenen Herzklappen

同种异体心脏瓣膜不同保存方法的体内比较研究

• 批准号: 51176423
• 负责人: Professor Dr. Ulrich A. Stock
• 金额: --
• 依托单位: Universitätsklinik für Thorax-, Herz- und Gefäßchirurgie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2007
• 资助国家: 德国
• 起止时间: 2006-12-31 至 2012-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/51176423?language=en
• 关键词: Vergleichende; vivo; Untersuchung; verschiedener; Konservierungsverfahren

Human heart valves or allografts represent nearly perfect heart valve substitutes. They have optimal hemodynamic characteristics and are resistant to infections. Disadvantages particularly in pediatric patients are limited availability, an inability to grow, degeneration and long-term failure. Potential culprits for the latter include immunological responses and an increased calcium metabolism. The first clinical use of allograft heart valves was as homovitals that were transplanted after antibiotic incubation without any preservation. Since 1968 standard frozen cryopreservation (SFC) has been employed including storage in vapour phase liquid nitrogen. Application of multiphoton imaging analysis (MIA) enables three dimensional visualization of elastin and collagen by induction of autofluorescence without fixation, embedding and staining in fresh tissue. MIA allowed for the first time detection of partial destruction of elastic and collagenous matrix in SFC porcine valves ex situ. As the overall amount of collagen and elastin remains unchanged the etiology of the above described destruction is postulated as freezing induced extracellular matrix (ECM) damage due to ice crystals. Disruptive interstitial ice damage that occurs during cryopreservation can be avoided by promoting vitrification, glass formation, instead of ice formation below the cryopreservation solutions glass transition temperature. As the preservation and warming is accomplished without any tissue destruction, vitrification enabled for the first time complete preservation of extracellular matrix in heart valves. In order to simplify the required infrastructure, we have further developed ice free cryopreservation (IFC), which permits storage above the solutions glass transition temperature in a 80C freezer. We applied IFC in a recent sheep study and were able to demonstrate that maintaining ECM results in better valve function and diminished degeneration. Furthermore, IFC is antiinflammatory preventing CD3 positive T cell mediated responses. The exact mechanism of this attenuated adaptive immune response remains unknown. Recent studies in a porcine tissue to human responder peripheral blood leukocyte combination have shown that treatment of valve tissue with IFC preservation medium reduces leaflet immunogenicity and induces responder monocyte differentiation towards endothelial like cells. Accordingly application of IFC for xenogeneic heart valves might overcome organ scarcity. Simplification by omitting controlled rate freezing and liquid nitrogen storage will improve cost effectiveness and allow application in third world and developing countries with limited financial resources.

人类心脏瓣膜或同种异体移植物代表了近乎完美的心脏瓣膜替代品。它们具有最佳的血流动力学特征并且具有抗感染能力。特别是对于儿科患者来说，其缺点是可用性有限、无法生长、退化和长期失败。后者的潜在罪魁祸首包括免疫反应和钙代谢增加。同种异体移植心脏瓣膜的首次临床应用是作为同种生命体，在抗生素孵育后移植，无需任何保存。自 1968 年以来，已采用标准冷冻冷冻保存 (SFC)，包括在气相液氮中储存。多光子成像分析 (MIA) 的应用通过诱导自发荧光实现弹性蛋白和胶原蛋白的三维可视化，无需在新鲜组织中固定、包埋和染色。 MIA 首次检测到 SFC 猪瓣膜异位弹性和胶原基质的部分破坏。由于胶原蛋白和弹性蛋白的总量保持不变，上述破坏的病因被推测为由于冰晶导致的冷冻诱导的细胞外基质（ECM）损伤。通过促进玻璃化、玻璃形成，而不是在低温保存溶液玻璃化转变温度以下形成冰，可以避免低温保存过程中发生的破坏性间质冰损伤。由于保存和加温是在没有任何组织破坏的情况下完成的，玻璃化冷冻首次实现了心脏瓣膜中细胞外基质的完整保存。为了简化所需的基础设施，我们进一步开发了无冰冷冻保存 (IFC)，它允许在高于溶液玻璃化转变温度的温度下储存在 80°C 的冰箱中。我们在最近的一项绵羊研究中应用了 IFC，并能够证明维持 ECM 可以改善瓣膜功能并减少退化。此外，IFC 具有抗炎作用，可防止 CD3 阳性 T 细胞介导的反应。这种减弱的适应性免疫反应的确切机制仍然未知。最近对猪组织对人类应答者外周血白细胞组合的研究表明，用 IFC 保存培养基处理瓣膜组织可降低小叶的免疫原性，并诱导应答者单核细胞向内皮样细胞分化。因此，将IFC应用于异种心脏瓣膜可能会克服器官稀缺的问题。通过省略受控速率冷冻和液氮储存进行简化将提高成本效益，并允许在财政资源有限的第三世界和发展中国家应用。