Proteomic Discovery of Biomaterial Antigens

描述（由申请人提供）：组织工程假体，如使用异种生物材料作为支架的置换心脏瓣膜，提供了一种可以解决耐久性问题的活体植入物的可能性。在该应用中，异种生物材料经历“去细胞化”以去除细胞相关抗原，从而防止组织排斥。这种方法的假设是：1）重要的异种抗原仅与组织的细胞成分相关，2）组织切片中细胞的消失等同于抗原从组织中完全去除。这些有缺陷的假设的一个可能和不幸的后果是最近报告的Synergraft-decellulized非固定猪心脏瓣膜在儿童中的灾难性失败。通过更好地了解异种生物材料的免疫原性，可以加强异种生物材料在戊二醛固定和组织工程生物假体制造中的应用。该方法的关键是更好地理解异种组织中抗原的数量和分布（例如细胞膜，细胞外基质）。因此，本研究的具体目标是：1.通过二维凝胶电泳提取和分离候选异种生物材料的细胞膜、细胞质和细胞外基质蛋白组分。2.通过将分离的蛋白质暴露于针对候选组织产生的天然抗体和获得性抗体，测定异种生物材料的抗原性。3.通过MALDI-TOF-TOF质谱鉴定激发抗原反应的蛋白质。本申请中描述的蛋白质组学方法提供了一种强大且特异的方法，用于筛选大量蛋白质的跨物种抗原性的关键重要性质。这些信息将为异种组织工程组织的可行性和异种移植提供重要的见解。异种生物材料是可用于制造人类替代组织（即组织工程）的动物组织。该应用提供了关于生物材料的免疫排斥的信息，在动物组织可用于组织工程之前必须更好地理解这些信息。

DESCRIPTION (provided by applicant): Tissue-engineered prostheses such as replacement heart valves that utilize xenogeneic biomaterials as scaffold offer the possibility of a living implant that could resolve durability issues. In this application, xenogeneic biomaterials undergo "decellularization" to remove cell-associated antigens, thereby preventing tissue rejection. The assumptions of this approach are: 1) that important xenogeneic antigens are associated solely with the cellular component of the tissue, and 2) that disappearance of cells from histologic section equates to complete removal of antigens from the tissue. A likely and unfortunate consequence of these flawed assumptions was the recently reported catastrophic failure of Synergraft-decellularized non-fixed porcine heart valves in children. The application of xenogeneic biomaterials to fabrication of both glutaraldehyde-fixed and tissue-engineered bioprostheses could be enhanced by a better understanding of the immunogenicity of these materials. Critical to that approach is a better understanding of both the number and distribution (e.g. cell-membrane, extracellular matrix) of antigens within xenogeneic tissues. Thus, the specific aims of this proposal are: 1.To extract and separate by 2-D gel electrophoresis the cell membrane, cytoplasmic and extracellular matrix protein fractions of candidate xenogeneic biomaterials. 2. To determine antigenicity of xenogeneic biomaterials by exposing separated proteins to naturally-occurring and acquired antibodies generated against the candidate tissues. 3. To identify by MALDI-TOF-TOF mass spectrometry proteins that elicits an antigenic response. The proteomic approach described in this application offers a powerful and specific method for screening a large number of proteins for the critically important property of antigenicity across species. This information will provide important insight about the feasibility of xenogeneic tissue-engineered tissues and about xenotransplantation in general. Xenogeneic biomaterials are tissues from animals that could be used in the fabrication of replacement tissues for humans (i.e. tissue engineering). This application provides information about immune rejection of biomaterials that must be better understood before animal tissues can be used in tissue engineering.