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

虽然轻微的骨损伤可以自行愈合和/或再生，但由先天性异常、创伤和病变组织的手术切除造成的重大缺陷会压倒骨骼的再生能力，导致多次昂贵的干预措施，并延长患者的痛苦。目前的治疗方法取得了不同程度的成功，但也存在一些缺陷。组织工程（TE）技术，使用支架、生长因子和骨祖细胞的不同组合，直接应用于骨缺损；在消除许多缺点的同时，对实现最佳治疗有很大的希望。使用这些技术，我们和其他人纯化和扩展了MSC、EPC和BM-MNC，并以不同的组合将它们植入支架，并在动物模型中治疗了大骨缺损，并证明了骨愈合的改善。虽然大多数纯化细胞的组合促进愈合，但我们惊讶地发现愈合的速度和质量并不明显优于传统的自体骨移植治疗。这些意想不到的发现促使我们重新检查自体移植物的成分，在BM-MNC部分中，试图确定哪些是促进观察到的骨愈合增强的主要因素。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核细胞组成。后者已被几个小组描述过，并根据所使用的隔离协议被分配了不同的名称；孢子样干细胞、多能成体干细胞（MASCs）、多系分化耐压细胞（Muse）、多能成体祖细胞（MAPs）、极小胚胎样干细胞（VSEL）。它们在大多数成人组织中被发现，在体内模型中被证明是多能的，并被证明能刺激新骨的形成。基于以上，我们提出了一个假设，即BM-MNC内的VSEL细胞在骨损伤后的再生反应中起关键作用。为了验证这一假设，我们将在大鼠股骨临界尺寸骨缺损模型中测量和比较BM-MNC（含和不含VSEL细胞）和单独VSEL细胞对骨愈合的影响。我们将组织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，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细胞在骨愈合的哪个方面有贡献？

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?