Department of Mechanical and Industrial Engineering, University of Toronto, Toronto, Canada.
Institute of Biomaterials and Biomedical Engineering, University of Toronto, Toronto, Canada.
Tissue Eng Part C Methods. 2019 Oct;25(10):581-592. doi: 10.1089/ten.TEC.2019.0121. Epub 2019 Sep 12.
Biomechanical stimulation is a common strategy to improve the growth, maturation, and function of a variety of engineered tissues. However, identifying optimized biomechanical conditioning protocols is challenging, as cell responses to mechanical stimuli are modulated by other multifactorial microenvironmental cues, including soluble factors and biomaterial properties. Traditional bioreactors lack the throughput necessary for combinatorial testing of cell activity in mechanically stimulated engineered tissues. Microfabricated systems can improve experimental throughput, but often do not provide uniform mechanical loading, are challenging to use, lack robustness, and offer limited amounts of cells and tissue for analysis. To address the need for higher-throughput, combinatorial testing of cell activity in a tissue engineering context, we developed a hybrid approach, in which flexible polydimethylsiloxane microfabricated inserts were designed to simultaneously generate multiple tensile strains when stretched cyclically in a standard dynamic bioreactor. In the embodiment presented in this study, each insert contained an array of 35 dog bone-shaped wells in which cell-seeded microscale hydrogels can be polymerized, with up to eight inserts stretched simultaneously in the bioreactor. Uniformity of the applied strains, both along the length of a microtissue and across multiple microtissues at the same strain level, was confirmed experimentally. In proof-of-principle experiments, the combinatorial effects of dynamic strain, biomaterial stiffness, and transforming growth factor (TGF)-β1 stimulation on myofibroblast differentiation were tested, revealing both known and novel interaction effects and suggesting tissue engineering strategies to regulate myofibroblast activation. This platform is expected to have wide applicability in systematically probing combinations of mechanobiological tissue engineering parameters for desired effects on cell fate and tissue function. Impact Statement In this study, we introduce a dynamic bioreactor system incorporating microfabricated inserts to enable systematic probing of the effects of combinations of mechanobiological parameters on engineered tissues. This novel platform offers the ease of use, robustness, and well-defined mechanical strain stimuli inherent in traditional dynamic bioreactors, but significantly improves throughput (up to 280 microtissues can be tested simultaneously in the embodiment presented in this study). This platform has wide applicability to systematically probe combinations of dynamic mechanical strain, biomaterial properties, biochemical stimulation, and other parameters for desired effects on cell fate and engineered tissue development.
生物力学刺激是一种提高多种工程组织生长、成熟和功能的常用策略。然而，确定优化的生物力学条件方案具有挑战性，因为细胞对机械刺激的反应受到其他多因素微环境线索的调节，包括可溶性因子和生物材料特性。传统的生物反应器缺乏组合测试工程组织中细胞活性所需的高通量。微制造系统可以提高实验通量，但通常不能提供均匀的机械加载，使用起来具有挑战性，缺乏稳健性，并且为分析提供的细胞和组织数量有限。为了解决在组织工程背景下对细胞活性进行高通量、组合测试的需求，我们开发了一种混合方法，其中柔性聚二甲基硅氧烷微制造插入物被设计为在标准动态生物反应器中周期性拉伸时同时产生多种拉伸应变。在本研究中提出的实施方案中，每个插入物包含一个数组的 35 个狗骨形状的井，其中可以聚合细胞接种的微尺度水凝胶，最多可以同时在生物反应器中拉伸八个插入物。通过实验证实了施加应变的均匀性，无论是在微组织的长度上还是在同一应变水平的多个微组织上。在原理验证实验中，测试了动态应变、生物材料刚度和转化生长因子（TGF）-β1刺激对肌成纤维细胞分化的组合效应，揭示了已知和新的相互作用效应，并提出了调节肌成纤维细胞激活的组织工程策略。该平台有望在系统地探测机械生物学组织工程参数的组合对细胞命运和组织功能的期望影响方面具有广泛的适用性。 影响陈述 在本研究中，我们引入了一种动态生物反应器系统，该系统结合了微制造插入物，以能够系统地探测机械生物学参数组合对工程组织的影响。这种新平台提供了传统动态生物反应器固有的易用性、稳健性和明确定义的机械应变刺激，但大大提高了通量（在本研究中提出的实施方案中，最多可以同时测试 280 个微组织）。该平台具有广泛的适用性，可以系统地探测动态机械应变、生物材料特性、生化刺激和其他参数的组合，以实现对细胞命运和工程组织发育的预期影响。
