Abstract

Osteocytes are the major mechanosensing cells in bone remodeling. Current in vitro bone mechanotransduction research use macroscale devices such as flow chambers; however, in vitro microfluidic devices provide an optimal tool to better understand this biological process with its flexible design, physiologically relevant dimensions and high-throughput capabilities. This project aims to design and fabricate a multi-shear stress, co-culture platform to study the interaction between osteocytes and other bone cells under varying flow conditions. Standard microfluidic design utilizing changing geometric parameters is used to induce different flow rates that are directly proportional to the levels of shear stress, with devices fabricated from standard polydimethylsiloxane (PDMS)-based softlithography processes. Each osteocyte channel (OCY) is connected to an adjacent osteoclast channel (OC) by 20-μm perfusion channels for cellular signaling molecule transport. Significant differences in RANKL levels are observed between channels with different shear stress levels, and we observed that pre-osteoclast differentiation was directly affected by adjacent flow-stimulated osteocytes. Significant decrease in the number of differentiating osteoclasts is observed in the OC channel adjacent to the 2-Pa shear stress OCY channel, while differentiation adjacent to the 0.5-Pa shear stress OCY channel is unaffected compared with no-flow controls. Addition of zoledronic acid showed a significant decrease in osteoclast differentiation, compounding to effect instigated by increasing fluid shear stress. Using this platform, we are able to mimic the interaction between osteocytes and osteoclasts in vitro under physiologically relevant bone interstitial fluid flow shear stress. Our novel microfluidic co-culture platform provides an optimal tool for bone cell mechanistic studies and provides a platform for the discovery of potential drug targets for clinical treatments of bone-related diseases.

中文翻译：

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用于骨细胞机械转导研究的新型体外微流体平台

摘要

骨细胞是骨重塑中的主要机械感应细胞。目前的体外骨机械转导研究使用大型设备，如流动室；然而，体外微流体设备以其灵活的设计、生理相关的尺寸和高通量能力提供了一种最佳工具，可以更好地了解这一生物过程。该项目旨在设计和制造一个多剪切应力共培养平台，以研究不同流动条件下骨细胞与其他骨细胞之间的相互作用。利用改变几何参数的标准微流体设计用于诱导与剪切应力水平成正比的不同流速，设备由基于标准聚二甲基硅氧烷 (PDMS) 的软光刻工艺制成。每个骨细胞通道 (OCY) 通过 20-μm 灌注通道连接到相邻的破骨细胞通道 (OC)，用于细胞信号分子传输。在具有不同剪切应力水平的通道之间观察到 RANKL 水平的显着差异，我们观察到前破骨细胞分化直接受到相邻流动刺激的骨细胞的影响。在与 2-Pa 剪切应力 OCY 通道相邻的 OC 通道中观察到分化破骨细胞的数量显着减少，而与无流量控制相比，与 0.5-Pa 剪切应力 OCY 通道相邻的分化不受影响。添加唑来膦酸显示破骨细胞分化显着降低，复合效应由增加的流体剪切应力引起。使用这个平台，我们能够模拟骨细胞和破骨细胞之间的相互作用 并且我们观察到前破骨细胞分化直接受到邻近流动刺激的骨细胞的影响。在与 2-Pa 剪切应力 OCY 通道相邻的 OC 通道中观察到分化破骨细胞的数量显着减少，而与无流量控制相比，与 0.5-Pa 剪切应力 OCY 通道相邻的分化不受影响。添加唑来膦酸显示破骨细胞分化显着降低，复合效应由增加的流体剪切应力引起。使用这个平台，我们能够模拟骨细胞和破骨细胞之间的相互作用 并且我们观察到前破骨细胞分化直接受到邻近流动刺激的骨细胞的影响。在与 2-Pa 剪切应力 OCY 通道相邻的 OC 通道中观察到分化破骨细胞的数量显着减少，而与无流量控制相比，与 0.5-Pa 剪切应力 OCY 通道相邻的分化不受影响。添加唑来膦酸显示破骨细胞分化显着降低，复合效应由增加的流体剪切应力引起。使用这个平台，我们能够模拟骨细胞和破骨细胞之间的相互作用 与无流量控制相比，5-Pa 剪切应力 OCY 通道不受影响。添加唑来膦酸显示破骨细胞分化显着降低，复合效应由增加的流体剪切应力引起。使用这个平台，我们能够模拟骨细胞和破骨细胞之间的相互作用 与无流量控制相比，5-Pa 剪切应力 OCY 通道不受影响。添加唑来膦酸显示破骨细胞分化显着降低，复合效应由增加的流体剪切应力引起。使用这个平台，我们能够模拟骨细胞和破骨细胞之间的相互作用体外在生理相关骨间质液流动剪切应力下。我们新型的微流体共培养平台为骨细胞机制研究提供了最佳工具，并为发现用于骨相关疾病临床治疗的潜在药物靶标提供了平台。