Lim Hong-Gook, Kim Gi Beom, Jeong Saeromi, Kim Yong Jin
Seoul National University Hospital Clinical Research Institute, Xenotransplantation Research Center, Seoul, Republic of Korea Department of Thoracic and Cardiovascular Surgery, Seoul National University Hospital, Seoul National University College of Medicine, Seoul, Republic of Korea.
Seoul National University Hospital Clinical Research Institute, Xenotransplantation Research Center, Seoul, Republic of Korea Department of Pediatrics, Seoul National University Hospital, Seoul National University College of Medicine, Seoul, Republic of Korea.
Eur J Cardiothorac Surg. 2015 Jul;48(1):104-13. doi: 10.1093/ejcts/ezu385. Epub 2014 Oct 14.
Conventional crosslinking with glutaraldehyde (GA) renders cardiac xenografts inert, non-biodegradable and non-antigenic, but is a main cause for dystrophic calcification due to phospholipids, free aldehyde groups and residual antigenicity. A significant immune reaction to the galactose-α-1,3 galactose β-1,4-N-acetylglucosamine (α-Gal) of a GA-fixed cardiac xenograft occurs, leading to calcification. We developed a next-generation α-Gal-free tissue valve with GA-fixed cardiac xenografts, treated using a novel combined anticalcification protocol including immunological modification, which was demonstrated effective in a small animal study.
Porcine aortic valves were decellularized with 1% sodium dodecyl sulphate, 1% Triton X-100 and 1% sodium lauroyl sarcosinate and immunologically modified with α-galactosidase. The valves were treated by a polyethylene glycol space filler, fixed with GA in 75% ethanol + 5% octanol and detoxified with glycine. We manufactured the tissue valve with the porcine aortic valve mounted on a Nitinol (nickel-titanium memory alloy) plate. The tissue valve was placed under in vitro mock circulation, and durability from mechanical stress was evaluated for 100 days. Ten sheep underwent mitral valve replacement with the tissue valve, and haemodynamic, radiological, immunohistopathological and biochemical results were obtained for 18 months after implantation.
The in vitro circulation experiment demonstrated that the valve functioned well with good morphology. Eight sheep survived for 1, 2, 5, 10, 14, 53, 546 and 552 days after mitral valve replacement, but two sheep did not survive. An evaluation by echocardiography and cardiac catheterization demonstrated good haemodynamic status and function of the mitral valve at 18 months after implantation. The xenografts were well preserved without a α-Gal immune reaction or calcification based on the immunological, radiographic, microscopic and biochemical examinations.
We developed a next-generation α-Gal-free tissue valve with simultaneous use of multiple anticalcification therapies and novel tissue treatments such as decellularization, immunological modification with α-galactosidase, space filler, an organic solvent and detoxification. Future investigations should evaluate α-Gal-free substitutes such as our tissue valve, and a future clinical study is warranted based on these promising preclinical results.
用戊二醛（GA）进行传统交联可使心脏异种移植物失去活性、不可生物降解且无抗原性，但由于磷脂、游离醛基和残留抗原性，这是营养不良性钙化的主要原因。对GA固定的心脏异种移植物的半乳糖-α-1,3-半乳糖β-1,4-N-乙酰葡糖胺（α-Gal）会发生显著的免疫反应，导致钙化。我们开发了一种使用GA固定的心脏异种移植物的下一代无α-Gal组织瓣膜，采用包括免疫修饰在内的新型联合抗钙化方案进行处理，在一项小动物研究中证明其有效。
用1%十二烷基硫酸钠、1% Triton X-100和1%月桂酰肌氨酸钠对猪主动脉瓣进行去细胞处理，并用α-半乳糖苷酶进行免疫修饰。瓣膜用聚乙二醇空间填充剂处理，在75%乙醇+5%辛醇中用GA固定，并用甘氨酸解毒。我们将猪主动脉瓣安装在镍钛诺（镍钛记忆合金）板上制作组织瓣膜。将组织瓣膜置于体外模拟循环下，评估其在100天内承受机械应力的耐久性。10只绵羊接受了用该组织瓣膜进行的二尖瓣置换术，并在植入后18个月获得了血流动力学、放射学、免疫组织病理学和生化结果。
体外循环实验表明瓣膜功能良好，形态正常。8只绵羊在二尖瓣置换术后存活了1、2、5、10、14、53、546和552天，但2只绵羊未存活。超声心动图和心导管检查评估显示，植入后18个月二尖瓣的血流动力学状态和功能良好。基于免疫、放射、显微镜和生化检查，异种移植物保存良好，无α-Gal免疫反应或钙化。
我们开发了一种下一代无α-Gal组织瓣膜，同时使用了多种抗钙化疗法以及去细胞、用α-半乳糖苷酶进行免疫修饰、空间填充剂、有机溶剂和解毒等新型组织处理方法。未来的研究应评估像我们的组织瓣膜这样的无α-Gal替代物，基于这些有前景的临床前结果，有必要开展未来的临床研究。
