Tan Richard P, Lee Bob S L, Chan Alex H P, Yuen Sui Ching G, Hung Juichien, Wise Steven G, Ng Martin K C
The Heart Research Institute, Sydney, NSW 2042, Australia; Sydney Medical School, University of Sydney, NSW 2006, Australia.
The Heart Research Institute, Sydney, NSW 2042, Australia.
Acta Biomater. 2017 Apr 15;53:378-388. doi: 10.1016/j.actbio.2017.02.002. Epub 2017 Feb 3.
Biomaterial scaffolds enhancing the engraftment of transplanted bone-marrow mononuclear cells (BM-MNC) have enormous potential for tissue regeneration applications. However, development of appropriate materials is challenging given the precise microenvironments required to support BM-MNC engraftment and function. In this study, we have developed a non-invasive, real-time tracking model of injected BM-MNC engraftment to wounds and implanted biomaterial scaffolds. BM-MNCs, encoded with firefly luciferase and enhanced GFP reporter genes, were tail vein injected into subcutaneously wounded mice. Luciferase-dependent cell bioluminescence curves revealed our injected BM-MNCs homed to and engrafted within subcutaneous wound sites over the course of 21days. Further immunohistochemical characterization showed that these engrafted cells drove functional changes by increasing the number of immune cells present at early time points and remodelling cell phenotypes at later time points. Using this model, we subcutaneously implanted electrospun polycaprolactone (PCL) and PCL/Collagen scaffolds, to determine differences in exogenous BM-MNC response to these materials. Following BM-MNC injection, immunohistochemical analysis revealed a high exogenous BM-MNC density around the periphery of PCL scaffolds consistent with a classical foreign body response. In contrast, transplanted BM-MNCs engrafted throughout PCL/Collagen scaffolds indicating an improved biological response. Importantly, these differences were closely correlated with the real-time bioluminescence curves, with PCL/Collagen scaffolds exhibiting a∼2-fold increase in maximum bioluminescence compared with PCL scaffolds. Collectively, these results demonstrate a new longitudinal cell tracking model that can non-invasively determine transplanted BM-MNC homing and engraftment to biomaterials, providing a valuable tool to inform the design scaffolds that help augment current BM-MNC tissue engineering strategies.
Tracking the dynamic behaviour of transplanted bone-marrow mononuclear cells (BM-MNCs) is a long-standing research goal. Conventional methods involving contrast and tracer agents interfere with cellular function while also yielding false signals. The use of bioluminescence addresses these shortcomings while allowing for real-time non-invasive tracking in vivo. Given the failures of transplanted BM-MNCs to engraft into injured tissue, biomaterial scaffolds capable of attracting and enhancing BM-MNC engraftment at sites of injury are highly sought in numerous tissue engineering applications. To this end, the results from this study demonstrate a new longitudinal tracking model that can non-invasively determine exogenous BM-MNC homing and engraftment to biomaterials, providing a valuable tool to inform the design of scaffolds with implications for countless tissue engineering applications.
生物材料支架可增强移植骨髓单个核细胞（BM-MNC）的植入，在组织再生应用中具有巨大潜力。然而，鉴于支持BM-MNC植入和功能所需的精确微环境，开发合适的材料具有挑战性。在本研究中，我们建立了一种非侵入性的实时追踪模型，用于追踪注射的BM-MNC在伤口和植入生物材料支架中的植入情况。将编码萤火虫荧光素酶和增强型绿色荧光蛋白报告基因的BM-MNC经尾静脉注射到皮下受伤的小鼠体内。依赖荧光素酶的细胞生物发光曲线显示，在21天的过程中，我们注射的BM-MNC归巢并植入皮下伤口部位。进一步的免疫组织化学表征表明，这些植入的细胞通过在早期增加免疫细胞数量以及在后期重塑细胞表型来驱动功能变化。利用该模型，我们皮下植入了电纺聚己内酯（PCL）和PCL/胶原蛋白支架，以确定外源性BM-MNC对这些材料反应的差异。在注射BM-MNC后，免疫组织化学分析显示PCL支架周边外源性BM-MNC密度高，这与典型的异物反应一致。相比之下，移植的BM-MNC在整个PCL/胶原蛋白支架中植入，表明生物学反应有所改善。重要的是，这些差异与实时生物发光曲线密切相关，与PCL支架相比，PCL/胶原蛋白支架的最大生物发光增加了约2倍。总体而言，这些结果证明了一种新的纵向细胞追踪模型，该模型可以非侵入性地确定移植的BM-MNC对生物材料的归巢和植入情况，为设计有助于增强当前BM-MNC组织工程策略的支架提供了有价值的工具。
追踪移植骨髓单个核细胞（BM-MNC）的动态行为是一个长期的研究目标。涉及造影剂和示踪剂的传统方法会干扰细胞功能，同时还会产生假信号。生物发光的使用解决了这些缺点，同时允许在体内进行实时非侵入性追踪。鉴于移植的BM-MNC未能植入受伤组织，在众多组织工程应用中，能够在损伤部位吸引并增强BM-MNC植入的生物材料支架备受关注。为此，本研究结果证明了一种新的纵向追踪模型，该模型可以非侵入性地确定外源性BM-MNC对生物材料的归巢和植入情况，为设计具有无数组织工程应用意义的支架提供了有价值的工具。
