Regeneration of Nucleus Pulposus by Tissue Engineering

描述(由申请人提供)： 背痛是我们社会发病率的一个重要来源，与椎间盘的病理直接相关。目前，退行性腰椎间盘疾病的治疗包括以缓解症状为目的的保守和侵入性治疗。在了解组织修复和干细胞生物学方面的最新进展有助于确定重建病变/退化组织的可能的新方法。因此，这项建议中描述的研究的目标是开发一种基于“组织工程学”的方法来进行椎间盘治疗。本研究的具体目的是：1.研究骨髓间充质干细胞向髓核样细胞的分化；评价这些工程化细胞在体外细胞和器官培养系统中的定位和命运。骨髓间充质干细胞将在促进分化为髓核表型的条件下保持。在选定的时间间隔，将评估椎间盘细胞向与髓核一致的表型分化。我们将验证这一假设，即椎间盘独特的微环境有助于促进MSC向表现为髓核样表型的细胞分化。2.从髓核分离脊索细胞群；检测这些细胞对MSC与髓核表型结合的影响。我们将研究从幼年大鼠髓核分离的脊索细胞对MSC分化和形成髓核样组织的影响。我们将在将预分化的MSC植入髓核腔之前将其与脊索细胞共培养。我们将检验脊索细胞刺激髓核表型表达的假设。3.评价移植的MSC能否在体内再生大鼠髓核。悬浮在可注射的水凝胶中，我们将在体内将预分化的MSC移植到正常大鼠的髓核内。我们将评估细胞的活力以及它们创造出的髓核样组织。这项研究的结果将验证在特定目标1和2中进行的体外研究，并允许我们测试MSC可以在体内分化为髓核样细胞的假设。从长远来看，如果成功，这种治疗方法将需要使用患者自己的细胞进行椎间盘再生。

DESCRIPTION (provided by applicant): Back pain, a significant source of morbidity in our society, is directly related to the pathology of the intervertebral disc. At present, the treatment of degenerative disc disease consists of therapies, both conservative and invasive, that are aimed at symptomatic relief. Recent advances in the understanding of tissue repair and stem cell biology have served to identify possible new approaches to rebuild diseased/degenerate tissues. Accordingly, the goal of the investigations described in this proposal is to develop a "tissue engineering" based approach to disc therapy. The Specific Aims of the investigation are: 1. To examine the differentiation of MSC into a nucleus pulposus-like cell; to evaluate the commitment and fate of these engineered cells in an in vitro cell and organ culture system. MSC will be maintained under conditions that enhance differentiation into a nucleus pulposus phenotype. At selected time intervals, differentiation of the disc cells towards a phenotype consistent with that of the nucleus pulposus will be assessed. We will test the hypothesis that the unique micro-environment of the intervertebral disc serves to enhance the differentiation of MSC to cells displaying a nucleus pulposus-like phenotype. 2. To isolate a notochordal population of cells from the nucleus pulposus; to examine the impact of these cells on the commitment of MSC to the nucleus pulposus phenotype. We will examine the impact of notochordal cells, isolated from the nucleus pulposus of the young rat, on the differentiation of MSC and the formation of a nucleus pulposus-like tissue. We will co-culture notochordal with the pre-differentiated MSC prior to their insertion into the nucleus pulposus cavity. We will test the hypothesis that cells of notochordal origin stimulate the expression of the nucleus pulposus phenotype. 3. To assess whether transplanted MSC can be used in vivo to repopulate the rat nucleus pulposus. Suspended in an injectable hydrogel, we will surgically transplant pre-differentiated MSC into the nucleus pulposus cavity of normal rat discs in vivo. We will evaluate the vitality of the cells and their creation of a nucleus pulposus-like tissue. Outcomes from this study will validate the in vitro studies performed in Specific Aims 1 and 2 and permit us to test the hypothesis that MSC could differentiate into nucleus pulposus-like cells in vivo. In the long term, if successful, this approach to therapy will entail using patients' own cells for disc regeneration.