Direct and indirect contributions of perivascular stem cells to bone healing

项目总结 作为自体移植骨的替代，间充质干细胞(MSC)已经被用来加速 骨修复。然而，当前的MSC来源有显著的缺点，包括：(1)稀缺的可用性，(2) 需要培养；(3)明显的细胞异质性，成骨效率降低。我们的解决方案是 使用一种新的、纯化的和未培养的干细胞来源，称为血管周围干细胞(PSC)。 PSC是培养衍生的、传统的MSC的预期纯化的祖先，并通过它们的 出现在船只周围。与骨髓干细胞(BMSC)不同，PSC在 临床翻译，包括：(1)通过荧光激活细胞分选(FACS)而不是培养分离，(2) 均一性高；(3)临床用量大。 我们的数据显示，PSC通过直接和旁分泌机制发挥骨愈合作用，包括 对宿主骨组织的促成骨和促血管生成作用的结合。最近令人兴奋的数据表明 PSC的频率、活性、生长和分化因子(GDF)的表达与成骨分化 能抵抗骨质疏松性骨质流失的有害影响。相比之下，BMSC的数量有所减少， 骨质疏松症患者的增殖和成骨分化潜能。这一显著的差异使我们 在目前的方案中，直接系统地比较PSC和BMSC的骨愈合潜力。 总的来说，以下早期/New Investigator应用程序将解决蜂窝和信令问题 血管周围干细胞通过直接和间接作用促进骨愈合的机制 以及它们对骨质疏松条件的独特抵抗力。 目的1：评价PSC在骨愈合中的直接和旁分泌作用。通过一系列系统化的 在研究中，我们将检查PSC对骨祖细胞迁移和分化的影响，以及 骨愈合中的血管生成和新生血管。成骨细胞和内皮细胞报告动物将 用来比较我们先前描述的股骨节段性缺损(FSD)中宿主：供体细胞的贡献 模特。研究将与培养衍生的骨髓间充质干细胞应用进行面对面的研究。骨骼上的差异 治疗组之间的愈合结果将与体外GDF精化在统计学上相关。 目的2：论证宿主与供体骨质疏松在PSC骨愈合中的重要性。AS 如上所述，PSC对骨质疏松骨丢失的有害影响表现出显著的抵抗力。 然而，我们的试验数据表明，“宿主骨”而不是“供体间充质干细胞”的骨质疏松状态可能会影响 骨愈合。在这里，我们将确定移植的PSC或宿主骨内骨质疏松的程度 组织会影响骨骼的愈合。研究将与培养衍生的骨髓间充质干细胞在 我们的消防车模型。骨愈合的差异将与GDF和骨抑制信号的表达有关。

PROJECT SUMMARY As an alternative to autograft bone, mesenchymal stem cells (MSC) have been employed to accelerate bone repair. However, current MSC sources have significant drawbacks, including: (1) scarce availability, (2) need for culture, and (3) significant cell heterogeneity with decreased bone-forming efficacy. Our solution is the use of a novel, purified and uncultured stem cell source, known as Perivascular Stem Cells (PSC). PSC are prospectively purified ancestors of culture-derived, traditional MSC, and are identified by their presence around vessels. Unlike bone marrow stem cells (BMSC), PSC have significant advantages for clinical translation, including: (1) isolation by fluorescence activated cell sorting (FACS) rather than culture, (2) high homogeneity, and (3) high quantity for clinical application. Our data show that PSC exert bone healing effects via both direct and paracrine mechanisms, including a combination of pro-osteogenic and pro-vasculogenic effects on host bone tissue. Exciting recent data suggest that PSC frequency, viability, growth and differentiation factor (GDF) elaboration and osteogenic differentiation are resistant to the detrimental effects of osteoporotic bone loss. In contrast, BMSC show reduced numbers, proliferation, and osteogenic differentiation potential with osteoporosis. This marked difference has led us to directly and systematically compare the bone healing potential of PSC versus BMSC in the current proposal. Overall, the following Early Stage / New Investigator application will address the cellular and signaling mechanisms through which perivascular stem cells promote bone healing through direct and indirect effects, and their unique resistance to osteoporotic conditions. AIM 1: Evaluate the direct and paracrine effects of PSC in bone healing. Through a systematic series of studies, we will examine the effects of PSC on osteoprogenitor cell migration and differentiation, as well as angiogenesis and neovascularization in bone healing. Osteoblast and endothelial cell reporter animals will be utilized to compare host:donor cell contributions in our previously described femoral segmental defect (FSD) model. Studies will be performed head-to-head with culture-derived BMSC application. Differences in bone healing outcomes between treatment groups will be statistically correlated with in vitro GDF elaboration. AIM 2: Demonstrate the importance of host versus donor osteoporosis in PSC bone healing. As mentioned, PSC demonstrate marked resistance to the detrimental effects of osteoporotic bone loss. However, our pilot data suggest that osteoporotic status of the ‘host bone’ but not ‘donor MSC’ may impact bone healing. Here, we will determine the extent to which osteoporosis within transplanted PSC or host bone tissue affects bone healing. Studies will be performed head-to-head with culture-derived BMSC application in our FSD model. Differences in bone healing will be correlated with GDF and bone inhibitory signal expression.