Xenogeneic Scaffolds for Heart Valve Tissue Engineering

描述（由申请人提供）：美国心脏协会估计，瓣膜性心脏病在美国的患病率为2.5%，每年导致20，000例死亡。虽然用异种戊二醛固定的瓣膜置换患病的瓣膜（例如，牛心包（BP））显著改善短期结果，长期免疫介导的攻击导致钙化和最终瓣膜失效（约10年寿命）。当前瓣膜置换术的缺陷导致国家心肺和血液研究所（NHLBI）心脏外科工作组建议支持心脏瓣膜假体生物材料研究。利用未固定的BP作为支架的组织工程心脏瓣膜，患者的细胞在其上生长（再细胞化），具有产生潜在理想的心脏瓣膜的潜力。然而，正如NHLBI异种移植工作组所指出的，刺激受体免疫反应的组分（异种抗原）是扩大异种生物材料临床应用的关键障碍。我们提出了一种组合的方法（我们的新的抗原去除方法结合间充质干细胞（MSC）介导的免疫调节），用于生产免疫学上可接受的异种生物材料的心脏瓣膜组织工程。抗原去除旨在去除生物材料异种抗原，同时保持细胞外基质（ECM）完整并与再细胞化相容。我们假设异种抗原的溶解对于促进其从生物材料中去除至关重要。因此，我们建议逐步应用蛋白质化学原理来溶解并因此去除BP异种抗原。目的1：从完整BP中去除水溶性异种抗原，同时保持天然ECM结构/功能关系和再细胞化潜力。目标1将分两个阶段进行。第1阶段将评估两种新因素对从BP中去除水溶性抗原的影响。第2阶段将确保所得支架保留与心脏瓣膜组织工程相容的ECM特性。目标二：在最初去除水溶性异种抗原后，以逐步的方式从完整BP中去除脂溶性异种抗原，同时保持天然ECM结构/功能关系和再细胞化潜力。目标2将使用相同的2阶段方法进行，不同之处在于目标2中使用的因子旨在去除脂溶性抗原。所得的BP支架（有或没有MSC再细胞化）在植入免疫活性受体后必须是免疫学上可接受的。因此，我们提出：目的3：评估在逐步抗原去除水溶性和脂溶性异种抗原后对BP的体内免疫应答。评估抗原去除后BP的同种异体MSC再细胞化（BP-AR）对生物材料的体内免疫应答的影响。目的3利用免疫活性兔模型评估对BP-AR和MSC再细胞化BP-AR的免疫应答。完成该提案将产生结构完整、机械可靠、免疫学上可接受的异种支架，其与心脏瓣膜组织工程的再细胞化相容。

DESCRIPTION (provided by applicant): American Heart Association estimates that valvular heart disease has a US prevalence of 2.5% and accounts for 20,000 deaths annually. Although replacement of the diseased valve with a xenogeneic glutaraldehyde- fixed valve (e.g., bovine pericardium (BP)) dramatically improves short term outcome, long term immune mediated attack results in calcification and ultimately valve failure (~10 yr lifespan). Deficiencies of curent valve replacements led the National Heart Lung and Blood Institute (NHLBI) cardiac surgery working group to recommend support for heart valve prosthesis biomaterial research. A tissue engineered heart valve utilizing un-fixed BP as a scaffold onto which patients' cells are grown (recellularization), has the potential to produce a potentially ideal heart valve. However, as noted by the NHLBI xenotransplantation working group, components which stimulate a recipient immune response (xenoantigens) represent the critical barrier to expanding clinical use of xenogeneic biomaterials. We propose a combinatorial approach (our novel antigen removal methodology combined with mesenchymal stem cell (MSC) mediated immunomodulation) for production of an immunologically-acceptable xenogeneic biomaterial for heart valve tissue engineering. Antigen removal aims to remove biomaterial xenoantigens, while leaving the extracellular matrix (ECM) intact and compatible with recellularization. We hypothesize that solubilization of xenoantigens is critical to facilitate their removal from the biomaterial. We therefore propose stepwise application of protein chemistry principles to solubilize and consequently removal BP xenoantigens. Aim 1: Remove water-soluble xenoantigens from intact BP while maintaining native ECM structure/function relationships and recellularization potential. Aim 1 will be conducted in two phases. Phase 1 will assess the effect of two novel factors on removal of water-soluble antigens from BP. Phase 2 will ensure that the resulting scaffold retains ECM properties compatible with heart valve tissue engineering. Aim 2: Remove lipid-soluble xenoantigens from intact BP in a stepwise manner after initial removal of water-soluble xenoantigens, while maintaining native ECM structure/function relationships and recellularization potential. Aim 2 will be conducted using the same 2 phased approach, with the exception that factors used in aim 2 are designed to remove lipid-soluble antigens. The resulting BP scaffold (with or without MSC recellularization) must be immunologically-acceptable following implantation in an immunocompetent recipient. We therefore propose: Aim 3: Assess the in vivo immune response to BP following stepwise antigen removal of water- and lipid-soluble xenoantigens. Assess effect of allogeneic MSC recellularization of BP following antigen removal (BP-AR) on in vivo immune response towards the biomaterial. Aim 3 utilizes an immunocompetent rabbit model for assessment of immune response to both BP-AR and MSC recellularized BP-AR. Completion of this proposal will result in a structurally integral, mechanically sound, immunologically- acceptable xenogeneic scaffold compatible with recellularization for heart valve tissue engineering.