İyisan Nergishan, Hausdörfer Oliver, Wang Chen, Hiendlmeier Lukas, Harder Philipp, Wolfrum Bernhard, Özkale Berna
Microrobotic Bioengineering Lab (MRBL), School of Computation, Information, and Technology, Department of Electrical Engineering, Technical University of Munich (TUM), Hans-Piloty-Straße 1, 85748, Garching, Germany.
Munich Institute of Robotics and Machine Intelligence, Technical University of Munich, Georg-Brauchle-Ring 60, 80992, München, Germany.
Small Methods. 2024 Dec;8(12):e2400272. doi: 10.1002/smtd.202400272. Epub 2024 Jul 16.
In this study, the novel 3D-printed pressure chamber for encapsulated single-cell stimulation (3D-PRESS) platform is introduced for the mechanical stimulation of single stem cells in 3D microgels. The custom-designed 3D-PRESS, allows precise pressure application up to 400 kPa at the single-cell level. Microfluidics is employed to encapsulate single mesenchymal stem cells within ionically cross-linked alginate microgels with cell adhesion RGD peptides. Rigorous testing affirms the leak-proof performance of the 3D-PRESS device up to 400 kPa, which is fully biocompatible. 3D-PRESS is implemented on mesenchymal stem cells for mechanotransduction studies, by specifically targeting intracellular calcium signaling and the nuclear translocation of a mechanically sensitive transcription factor. Applying 200 kPa pressure on individually encapsulated stem cells reveals heightened calcium signaling in 3D microgels compared to conventional 2D culture. Similarly, Yes-associated protein (YAP) translocation into the nucleus occurs at 200 kPa in 3D microgels with cell-binding RGD peptides unveiling the involvement of integrin-mediated mechanotransduction in singly encapsulated stem cells in 3D microgels. Combining live-cell imaging with precise mechanical control, the 3D-PRESS platform emerges as a versatile tool for exploring cellular responses to pressure stimuli, applicable to various cell types, providing novel insights into single-cell mechanobiology.
在本研究中，我们介绍了一种用于3D微凝胶中单个干细胞机械刺激的新型3D打印压力室（3D-PRESS）平台。定制设计的3D-PRESS能够在单细胞水平上精确施加高达400 kPa的压力。利用微流控技术将单个间充质干细胞包裹在含有细胞黏附RGD肽的离子交联海藻酸盐微凝胶中。严格测试证实了3D-PRESS装置在高达400 kPa时的防漏性能，且该装置具有完全的生物相容性。通过特异性靶向细胞内钙信号和机械敏感转录因子的核转位，将3D-PRESS应用于间充质干细胞进行机械转导研究。与传统的二维培养相比，对单独包裹的干细胞施加200 kPa的压力会使3D微凝胶中的钙信号增强。同样，在含有细胞结合RGD肽的3D微凝胶中，Yes相关蛋白（YAP）在200 kPa时发生核转位，揭示了整合素介导的机械转导在3D微凝胶中单个包裹的干细胞中的作用。将活细胞成像与精确的机械控制相结合，3D-PRESS平台成为探索细胞对压力刺激反应的通用工具，适用于各种细胞类型，为单细胞机械生物学提供了新的见解。
