ADSC Isolation by MMPs

描述(申请人提供)：已知间充质干细胞(MSCs)具有自我更新和分化为间充质系细胞类型的能力，包括成骨细胞、脂肪细胞、软骨细胞、腱细胞和成肌细胞，并有助于各种间充质组织的再生。脂肪组织代表了间充质干细胞的丰富来源，并提供了丰富和可获得的成人干细胞来源，患者的不适感最小。脂肪组织源干细胞(ADSCs)的分离主要是用Liberase HI实现的，该酶由溶组织梭菌胶原酶I、II和溶血素组成。所有的Liberase制剂都含有内毒素，内毒素对细胞活力有害。I型胶原酶的不稳定性影响了Liberase酶的重组生产。基质金属蛋白酶(MMPs)是一类能够催化几乎所有ECM成分的降解的酶，包括胶原蛋白、层粘连蛋白、纤维连接蛋白和弹性蛋白。目前还没有文献报道使用基质金属蛋白酶从脂肪组织中分离干细胞。本文描述的研究计划集中于探索重组基质金属蛋白酶鸡尾酒以有效分离ADSCs。为了实现这一目标，具体目标是(1)评估基质金属蛋白酶诱导的干细胞从脂肪组织中分离的有效性，以及(2)评价基质金属蛋白酶与细菌胶原酶在干细胞分离中的效率。重组MMPs的应用研究和生产具有非常重要的意义，因为如果没有最高质量的结缔组织降解酶，几乎不可能释放出具有良好和稳定的增殖能力的活性ADSCs。干细胞移植领域拟议的研究和应用以及在治疗一系列疾病方面的进展预计将产生重大的整体影响。

DESCRIPTION (provided by applicant): Mesenchymal stem cells (MSCs) are known to have the capacity for self-renewal and differentiation into mesenchyme-lineage cell types, including osteoblasts, adipocytes, chondrocytes, tenocytes, and myoblasts, and contribute to the regeneration of a variety of mesenchymal tissues. Adipose tissue represents a rich source of mesenchymal stem cells, and provides an abundant and accessible source of adult stem cells with minimal patient discomfort. Isolation of adipose tissue-derived stem cells (ADSCs) has primarily been achieved with Liberase HI, which is composed of Clostridium histolyticum collagenases I and II and thermolysin. All Liberase preparations contain endotoxin, which is harmful for cell viability. The recombinant production of Liberase enzymes is marred by the instability of collagenase I. The matrix metalloproteinases (MMPs) are a family of enzymes capable of catalyzing the degradation of virtually all ECM components, including collagens, laminin, fibronectin, and elastin. There are no literature studies describing the use of MMPs for the isolation of stem cells from adipose tissue. The research plan described herein focuses on the exploration of recombinant MMP cocktails for the efficient isolation of ADSCs. To achieve this goal the specific aims are (1) evaluate the efficacy of MMP-induced isolation of stem cells from adipose tissue and (2) evaluate MMP versus bacterial collagenase efficiencies for stem cell isolation. The applied research and production of recombinant MMPs proposed in this application is very significant, because without the highest quality of connective tissue degrading enzymes it is virtually impossible to liberate viable ADSCs with good and stable proliferative capabilities. The overall impact of the proposed research and applications in the field of stem cell transplantation and advancements in treatment of a broad array of diseases is expected to be significant.