Manipulation of stem cell differentiation by noninvasive electrical stimulus

描述(由申请人提供)：使用干细胞进行潜在的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，基于干细胞的组织工程技术就可以被设想出来。然而，实现这些目标首先需要了解和应用工程学原理，以整合生物和物理信号并放大分子信号机制(S)，以促进和增强选择性干细胞的增殖和分化。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)的分化。虽然电刺激在过去被用来诱导不同的细胞和分子反应，但电刺激的使用既没有得到优化，也没有探索到调节人类MSC分化的电耦合机制。因此，我们建议结合独特的、新颖的物理和光学技术来(1)优化促进hMSC成骨分化的电刺激参数和(2)阐明整合素介导的信号通路的作用，包括丝裂原激活的蛋白激酶信号机制。我们建议使用量子点结合的整合素来确定人骨髓间充质干细胞表面整合素动力学的变化，并将其与成骨分化不同阶段整合素下调相关。基于整合素的重新分布和聚集对电刺激的响应，提出了假说和至少两种可选的电耦合机制。电刺激的最佳应用和电耦合机制的阐明将为操纵干细胞分化的新的生物技术方法奠定基础，为建立整合物理和分子技术的组织工程方法的新范式铺平道路。长期的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的增殖和分化，从而调节干细胞来源的工程化组织结构的完整性和功能性。

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.