Targeted Cellular Repair of Articular Cartilage

描述（由申请人提供）：已知成人软骨几乎没有修复能力，主要是由于缺乏活跃的软骨祖细胞。因此，治疗骨关节炎的选择有限，其中大多数针对疾病的症状，而不是治愈。从患者身上分离出的间充质干细胞和培养的软骨前细胞，如果能被运送到需要的部位，就有可能应用于软骨的修复。据推测，包被软骨基质特异性抗体的软骨前细胞可以作为再生或修复受损软骨的一种方法。本建议的重点是开发一种针对软骨前细胞的靶向策略，以便这些细胞可以在体内用于修复关节软骨缺损。为了实现这一目标，MSCs和软骨前细胞将被棕榈化蛋白G（或A [PPG/A]）包裹，然后与II型胶原蛋白或聚集蛋白分子上的表位抗体孵育。这些抗体将与PPNG结合，它们的抗原结合位点将向外定向。这些细胞包被不同的抗体，单独或联合，将测试其结合软骨基质在冷冻切片和兔关节软骨移植体的能力。软骨前细胞将用一种重要的荧光染料染色，涂上靶向抗体，加入切片或外植体，清洗，然后观察荧光细胞的存在。将对包被的细胞进行体外研究，以确认包被过程不会显著阻碍细胞增殖或分化为软骨细胞的能力。一旦证实了软骨基质的靶向和合成，这些靶向软骨前细胞将在兔体内测试其修复标准关节软骨缺损的能力。修复程度将通过关节软骨缺损的组织学检查和形态测量检查来评估。待测实验组为：(1)控制未处理缺陷，(2)靶向细胞注射，(3)PPA/g包被，(4)未处理细胞注射。这些实验将确定这种新的细胞靶向方法是否有效地将软骨前细胞导向软骨基质，以及这些靶向细胞是否有助于修复受损软骨。这些研究的长期目标是开发一种治疗骨关节炎的方法。

DESCRIPTION (provided by applicant): Adult cartilage is known to have little ability for repair, due primarily to a lack of active chondrogenic progenitor cells. As a result, there are limited alternatives for the treatment of osteoarthritis, most of which are targeted at the symptoms of the disease and not at a cure. Mesenchymal stem cells and cultured pre-chondrocytes isolated from a patient have the potential to be applied to the repair of cartilage if they can be delivered to the sites where needed. It is hypothesized that pre-chondrogenic cells coated with antibodies specific for cartilage matrix could be used as a method to regenerate or repair damaged cartilage. The focus of this proposal is to develop a targeting strategy for pre-chondrocytes so that these cells can be used to repair an articular cartilage defect in vivo. To achieve this goal, MSCs and pre-chondrocytes will be coated with palmitated protein G (or A [PPG/A]) and then incubated with antibodies to type II collagen or to epitopes on aggrecan molecules. These antibodies will bind to the PPNG and their antigen binding sites will be oriented outward. These cells coated with different antibodies, alone or in combination, will be tested for their ability to bind cartilage matrix on frozen sections and on explants of rabbit articular cartilage. Pre-chondrocytes will be stained with a vital fluorescent dye, coated with targeting antibodies, added to sections or explants, washed and then observed for the presence of fluorescent cells. In vitro studies will be carried out on coated cells to confirm that the coating procedure does not significantly impede the cells ability to proliferate or to differentiate into chondrocytes. Once targeting and synthesis of cartilage matrix have been demonstrated, these targeted pre-chondrocytes will be tested for their ability to repair a standard articular cartilage defect in rabbits, in vivo. The degree of repair will be assessed by histologic examination and morphometric examination of the articular cartilage defects. Experimental groups to be tested are: (1) control untreated defects, (2) targeted cell-injected, (3) PPA/g coated, and (4) untreated cell-injected. These experiments will determine if this novel cell targeting methodology effectively directs pre-chondrocytes to cartilage matrix and whether these targeted cells can contribute to the repair of damaged cartilage. The long-term goal of these studies is to develop a treatment for osteoarthritis.