Role of pluripotent stem cells (PSCs) isolated from adult tissue in bone tissue regeneration

虽然轻微的骨损伤会自行愈合和/或再生，但由于先天性畸形、创伤和手术切除病变组织而导致的重大缺陷，会使骨骼再生能力不堪重负，导致多次且昂贵的干预措施，并延长患者的痛苦。目前的治疗方法已经取得了不同程度的成功，但也伴随着一些缺点。组织工程(TE)技术使用不同的支架、生长因子和骨祖细胞的组合，直接应用于骨缺损；在消除许多缺点的同时，为实现最佳愈合提供了巨大的希望。利用这些技术，我们和其他人纯化和扩增了MSC、EPC和BM-MNC，并在不同的组合中将它们种植到支架上，在动物模型中治疗了大型骨缺损，并证明了改善了骨愈合。虽然大多数组合的纯化细胞促进了愈合，但我们惊讶地发现，愈合的速度和质量并不比传统的自体骨移植治疗好。这些意想不到的发现促使我们重新检查BM-MNC部分中自体移植物的成分，试图确定哪些是/是促进所观察到的增强的骨愈合的主要因素(S)。这些治疗中使用的BM-MNC由含有间充质干细胞、造血祖细胞、内皮祖细胞和多能干细胞的单个核细胞组成。后者已被几个小组描述，并根据分离方案的不同被赋予不同的名称：孢子样干细胞、多能成体干细胞(MASCs)、多系分化应激耐受(MUSE)细胞、多能成体祖细胞(MAP)、极小胚胎样干细胞(VSEL)。在大多数成人组织中都发现了它们，在活体模型中被证明是多能的，并被证明能刺激新骨形成。在此基础上，我们提出了一个假设，即在BM-MNC中的VSEL细胞在骨损伤后的再生反应中发挥关键作用。为了验证这一假设，我们将测量和比较BM-MNC，有和没有VSEL细胞，以及单独VSEL细胞，在大鼠股骨临界大小骨缺损模型中对骨愈合的影响。我们将组织这些实验，以回答以下问题：VSEL细胞或其后代是否存在于大鼠股骨的愈合骨中？如果是这样的话，VSEL细胞是否有助于在基于BM-MNC的治疗中观察到的促愈合反应？VSEL细胞对骨愈合有哪些方面的贡献？

While minor bone injuries heal and/or regenerate on their own, major defects caused by congenital anomalies, trauma and surgical excision of diseased tissues, overwhelm bones regenerative capacity, resulting in multiple and costly interventions, and prolonged patient suffering. Current treatments have met with varying degrees of success, however are associated with several drawbacks. Tissue engineering (TE) techniques, that use different combinations of, scaffolds, growth factors, and osteoprogenitor cells, applied directly into the bone defect; hold great promise for achieving optimal healing, while eliminating many of the drawbacks. Using these techniques we and others have purified and expanded MSC, EPC and BM-MNC, and in different combinations have seeded them onto scaffolds and, in animal models, treated large bone defects, and have demonstrated improved bone healing.While most combinations of purified cells promote healing, we were surprised to find that the rate and quality of healing was not significantly better than, conventional treatment with autologous bone grafts. These unexpected findings led us to reexamine the constituents of autologous grafts, in the BM-MNC fraction, in an attempt to identify which is/are the primary actor(s) promoting the observed enhanced bone healing. The BM-MNC used in these treatments consists of a heterogeneous mix of mononuclear cells containing mesenchymal stem cells, hematopoietic progenitor cells, endothelial progenitor cells, and pluripotent stem cells. The latter have been described by several groups, and depending on the isolation protocol used, have been assigned different names; spore-like stem cells, multipotent adult stem cells (MASCs), multilineage-differentiating stress enduring (Muse) cells, multipotent adult progenitor cells (MAPs), very small embryonic-like stem cells (VSEL). They have been found in most adult tissues, have been shown to be pluripotent in an in vivo models and have been shown to stimulate new bone formation.Based on the above we formed a hypothesis that VSEL cells, within BM-MNC, play a key role in the regenerative response in bone following injury. To test this hypothesis, we will measure and compare the effects of BM-MNC, with and without VSEL cells, and VSEL cells alone, on bone healing in a rat femur critical size bone defect model. We will structure the experiments in order to answer the following questions: Are VSEL cells, or their descendants, present in healing bone in the rat femur? If so do VSEL cells contribute to the pro-healing response observed in BM-MNC based treatments? And what aspect of bone healing do VSEL cells contribute to?