Manipulation of stem cell differentiation by noninvasive electrical stimulus

描述（由申请人提供）：使用干细胞的潜在治疗性治疗需要阐明控制干细胞更新和分化的机制。必须开发和测试诱导干细胞分化为组织特异性谱系的操作技术。然后可以设想基于干细胞的组织工程技术。然而，实现这些目标首先需要工程原理的知识和应用，以整合生物和物理信号并放大分子信号传导机制，以促进和增强选择性干细胞增殖和分化。我们最近首次证明了使用非侵入性电刺激可以应用于操纵间充质干细胞（MSC）分化。尽管电刺激在过去已被有益地用于诱导不同的细胞和分子反应，但电刺激的使用既未被优化，也未探索调节人MSC分化的电偶联机制。因此，我们建议结合联合收割机独特的，新颖的物理和光学技术，（1）优化电刺激参数促进hMSC成骨分化和（2）阐明整合素介导的信号通路，包括丝裂原活化蛋白激酶信号机制的作用。我们建议使用量子点结合的整合素，以确定在人MSC表面上的整合素动力学的变化，并将它们与成骨分化的不同阶段的整合素下调。基于整合素在电刺激下的重新分布和聚集，提出了假设和至少2种替代的电耦合机制。电刺激的最佳应用和电耦合机制的阐明将为一种新的生物技术方法来操纵干细胞分化奠定基础，为建立一个新的范式组织工程方法，整合物理和分子技术铺平了道路。长期研究目标将包括通过最佳使用物理刺激来操纵和控制干细胞增殖和分化，从而调节干细胞衍生的工程化组织构建体的完整性和功能性。

DESCRIPTION (provided by applicant): Potential therapeutic treatment using stem cells requires elucidation of mechanisms that control stem cell renewal and differentiation. Manipulation techniques to induce stem cell differentiation into tissue-specific lineages will have to be developed and tested. Stem cell-based technologies for tissue engineering can then be envisioned. However, achievement of such goals would first require the knowledge and application of engineering principles to integrate biological and physical signals and amplify the molecular signaling mechanism(s) in order to promote and enhance selective stem cell proliferation and differentiation. We have recently demonstrated for the first time that use of non-invasive electrical stimulus can be applied to manipulate mesenchymal stem cell (MSC) differentiation. Although electrical stimulus has been used beneficially in the past to induce diverse cellular and molecular responses, neither the use of electrical stimulus has been optimized nor have the electrocoupling mechanisms regulating human MSC differentiation have been explored. We therefore propose to combine unique, novel physical and optical techniques to (1) optimize the electrical stimulus parameters for facilitated hMSC osteogenic differentiation and (2) elucidate the role of integrin-mediated signaling pathways, including the mitogen-activated protein kinase signaling mechanisms. We propose to use quantum dot-conjugated integrins to determine changes in the integrin dynamics on the human MSC surface and correlate them with integrin down-regulation at the different stages of osteogenic differentiation. Hypotheses and at least 2 alternate electrocoupling mechanisms are proposed based on integrin redistribution and clustering in response to electrical stimulus. Optimal application of electrical stimulus and elucidation of electrocoupling mechanisms will lay the foundation for a novel biotechnology approach to manipulate stem cell differentiation, paving the way to establish a new paradigm for tissue engineering methodologies that integrates physical and molecular techniques. The long- term research objectives would include manipulation and control of stem cell proliferation and differentiation by the optimal use of physical stimuli and, thereby, regulate the integrity and functionality of stem cell-derived engineered tissue constructs.