Farnebo Simon, Woon Colin Y L, Schmitt Taliah, Joubert Lydia-Marie, Kim Maxwell, Pham Hung, Chang James
1 Division of Plastic and Reconstructive Surgery, Stanford University Medical Center , Palo Alto, California.
Tissue Eng Part A. 2014 May;20(9-10):1550-61. doi: 10.1089/ten.TEA.2013.0207.
A biocompatible hydrogel consisting of extracellular matrix (ECM) from human tendons is described as a potential scaffold for guided tissue regeneration and tissue engineering purposes. Lyophilized decellularized tendons were milled and enzymatically digested to form an ECM solution. The ECM solution properties are assessed by proteome analysis with mass spectrometry, and the material's rheological properties are determined as a function of frequency, temperature, and time. In vivo application of the gel in a rat model is assessed for remodeling and host cell repopulation. Histology for macrophage invasion, fibroblast repopulation, and nanoscale properties of the gel is assessed. Gel interaction with multipotent adipoderived stem cells (ASCs) is also addressed in vitro to assess possible cytotoxicity and its ability to act as a delivery vehicle for cells. Proteome analysis of the ECM-solution and gel mass spectroscopy identified the most abundant 150 proteins, of which two isoforms of collagen I represented more than 55% of the sample. Rheology showed that storage (G') and loss (G″) of the ECM solution were stable at room temperature but displayed sigmoidal increases after ∼15 min at 37°C, matching macroscopic observations of its thermo responsiveness. G' and G″ of the gel at 1 rad/s were 213.1±19.9 and 27.1±2.4 Pa, respectively. Electron microscopy revealed fiber alignment and good structural porosity in the gel, as well as invasion of cells in vivo. Histology also showed early CD68(+) macrophage invasion throughout the gel, followed by increasing numbers of fibroblast cells. ASCs mixed with the gel in vitro proliferated, indicating good biocompatibility. This ECM solution can be delivered percutaneously into a zone of tendon injury. After injection, the thermoresponsive behavior of the ECM solution allows it to polymerize and form a porous gel at body temperature. A supportive nanostructure of collagen fibers is established that conforms to the three-dimensional space of the defect. This hydrogel holds the distinctive composition specific for tendon ECM, where tissue-specific cues facilitate host cell infiltration and remodeling. The results presented indicate that injectable ECM materials from tendon may offer a promising alternative in the treatment of tendinopathies and acute tendon injuries.
一种由人肌腱细胞外基质（ECM）组成的生物相容性水凝胶被描述为用于引导组织再生和组织工程目的的潜在支架。冻干的脱细胞肌腱经研磨和酶消化形成ECM溶液。通过质谱蛋白质组分析评估ECM溶液的性质，并确定该材料的流变性质作为频率、温度和时间的函数。在大鼠模型中评估凝胶的体内应用，以观察其重塑和宿主细胞再填充情况。评估巨噬细胞浸润、成纤维细胞再填充以及凝胶的纳米级性质的组织学情况。还在体外研究凝胶与多能脂肪来源干细胞（ASC）的相互作用，以评估可能的细胞毒性及其作为细胞递送载体的能力。对ECM溶液和凝胶进行质谱蛋白质组分析，鉴定出最丰富的150种蛋白质，其中I型胶原的两种异构体占样品的55%以上。流变学表明，ECM溶液的储能模量（G'）和损耗模量（G″）在室温下稳定，但在37°C约15分钟后呈S形增加，与其实时响应的宏观观察结果相符。凝胶在1 rad/s时的G'和G″分别为213.1±19.9和27.1±2.4 Pa。电子显微镜显示凝胶中纤维排列和良好的结构孔隙率，以及体内细胞浸润。组织学还显示，整个凝胶中早期有CD68（+）巨噬细胞浸润，随后成纤维细胞数量增加。ASC与凝胶在体外混合后增殖，表明具有良好的生物相容性。这种ECM溶液可以经皮递送至肌腱损伤区域。注射后，ECM溶液的热响应行为使其在体温下聚合形成多孔凝胶。建立了符合缺损三维空间的支持性胶原纤维纳米结构。这种水凝胶具有肌腱ECM特有的独特组成，其中组织特异性信号促进宿主细胞浸润和重塑。所呈现的结果表明，来自肌腱的可注射ECM材料可能为肌腱病和急性肌腱损伤的治疗提供一种有前景的替代方法。
