Enhancement of Myoblast Chemotactic Migration

描述(由申请人提供)： 一些退行性肌肉疾病如Duchenne肌营养不良症(DMD)的遗传缺陷是已知的，但还没有找到这些疾病的有效治疗方法。一种方法是基于细胞的治疗，将正常的成肌细胞或转基因的患者成肌细胞注射到患病肌肉中，目的是植入足以弥补蛋白质缺乏的肌肉。 然而，由于移植物存活率低和大量肌肉注射的不切实际要求等问题，这种方法几乎没有取得什么成功。最近，通过尾静脉或动脉注射在小鼠体内全身输送肌肉前体细胞，导致低水平的供体细胞植入再生肌肉组织。因此，前体细胞发生血管迁移和外渗，为改进基于细胞的治疗这些破坏性疾病提供了一条有用的途径。这项工作的具体目的是1)确定成肌细胞中表达的与激活的内皮细胞附着有关的分子，并促进内皮细胞的跨内皮细胞迁移；2)如果可能，通过细胞因子诱导免疫系统细胞的附着和外渗调节分子的表达，提高成肌细胞跨内皮细胞迁移的效率。方法：对小鼠骨骼肌成肌细胞进行研究，以确定已知在白细胞外渗中起作用的蛋白质的表达水平。炎性细胞因子将被用来诱导成肌细胞表达与趋化运动相关的蛋白质。体外跨内皮细胞迁移实验将被用来评估特定的趋化因子、受体和细胞黏附分子在这一过程中的作用以及炎性细胞因子刺激对成肌细胞迁移的影响。将正常的成肌细胞和由细胞因子诱导的成肌细胞注射到MDX小鼠(DMD模型)的尾静脉中，接受肌肉再生和渗入组织的评估。这一结果将进一步加深我们对促进供体成肌细胞系统植入病变肌肉的机制的理解，从而显着促进成肌前体细胞用于肌营养不良的治疗。

DESCRIPTION (provided by applicant): Genetic defects underlying several degenerative muscle diseases such as Duchenne muscular dystrophy (DMD) are known, yet effective therapies for these disorders have not been found. One approach has been cell-based therapy in which normal myoblasts or genetically modified patient myoblasts are injected into diseased muscle with the intent that engraftment would be sufficient to compensate for protein deficiencies. Little success has been achieved with this approach however due to problems such as poor graft survival and impractical requirements for numerous muscle injections. Recently, systemic delivery of muscle precursor cells via tail vein or arterial injection in mice has been demonstrated resulting in low-level donor cell engraftment of regenerating muscle tissue. Vascular migration and extravasation of precursor cells thus occurs and could provide a useful route for improved cell-based therapy for these devastating diseases. The specific aims of the proposed work are to 1) Identify molecules expressed in myoblasts that are involved in the attachment to activated endothelial cells and promote trans-endothelial cell migration, 2) Improve the efficiency of myoblast trans-endothelial migration, if possible, by cytokine-induced expression of molecules known to regulate attachment and extravasation of immune system cells. Methods: Murine skeletal muscle myoblasts will be studied to determine expression levels of proteins known to function in leukocyte extravasation. Inflammatory cytokines will be used to induce myoblast expression of proteins relevant to chemotactic movement. In vitro trans-endothelial cell migration assays will be used to assess the role of specific chemokines, receptors and cell adhesion molecules in this process and the influence of inflammatory cytokine stimulation on myoblast migration. Normal myoblasts and those induced by cytokines will be injected into tail veins of mdx mice (model for DMD) undergoing muscle regeneration and extravasation into tissues assessed. Results will further our understanding of the mechanisms that promote systemic engraftment of donor myoblasts into diseased muscle could significantly advance the therapeutic use of myogenic precursor cells for the treatment of muscular dystrophy.