双面纳米管器件的构建及其作用下细胞间蛋白交换的机制研究

Exploring the intercellular proteins transferring mechanisms and techniques involving direct transferring of proteins between cells, which plays essential roles in understanding the biological mechanism of cells and pathogenesis of diseases. Nowadays the technology of transferring intracellular proteins between cells requires multi-step protein extraction and delivery. Inspired by natural intercellular protein transfer through tunneling intercellular tubular connections, in this project, a double-sided nanostraws array is proposed to fabricate and integrate with a microfluidic device to transfer intracellular proteins between different cells directly. Nanostraws is fabricated on the top and bottom surfaces of substrate by MEMES/NEMS and how to be integrated with a microfluidic device, allowing cells cultured on both top and bottom substrates. This project explores that, with optimization of nanostarws size and parameters of applied electrical field, how to apply the double-sided nanostraws coupled with electrical filed to penetrate both top and bottom cell membrane without harming cell viability. Then intracellular proteins can be extracted from the donor cells on the bottom surface and immediately driven by the electric field through the nanostraw channel and into the living cells on the top surface. This work holds great promise on improving the efficiency of directly transferring of proteins between cells by MEMES/NEMS and facilitating the development of cell to cell communication.

了解细胞间蛋白交换的机制并人工实现不同细胞间蛋白交换，对于了解细胞生物机制和疾病发作机制意义重大。现有技术对细胞导入外来蛋白需要多步骤法蛋白质提取、纯化和释放。受到自然界不同的细胞之间通过隧道纳米管转移蛋白质的启发，本项目原创性地提出利用双面纳米管-电穿孔器件的作用，实现在不同细胞间直接传递交换细胞内蛋白质。探索利用微纳加工工艺制备上下表面都具有纳米管的平面阵列，并探索其与电穿孔器件的整合。细胞分别培养在上下层纳米管表面。在不影响细胞活性的前提下，探索纳米管尺寸和外加电场条件，使得纳米管在电场的协助下能够同时穿透上下表面的细胞的细胞膜，当细胞膜破裂之后，细胞内的细胞液能够沿着纳米管内腔扩散到对面的细胞内部，从而实现上下表面的细胞间互相传递各自细胞内的蛋白分子。此技术有望通过微纳加工技术实现高效的细胞间蛋白交换，为细胞信号通讯问题提供重要支持。

细胞之间良好地通讯是细胞间各项活动协调有序进行的基础，探索细胞间物质交换机制的研究以及提高物质转运的效率，对于了解细胞的生物机制和疾病的发作机制意义重大。克服传统的多步法蛋白质提取释放以达到在细胞内导入外来蛋白的缺陷，本项目模拟自然界细胞间通过天然纳米隧道进行通信的方法，原创性地利用双面纳米管-电穿孔器件的作用，实现在不同细胞间直接传递交换细胞内蛋白质。本项目成功探索如何将双面纳米管阵列与电穿孔器件整合，探索细胞培养在纳米管表面的活性和行为是否被影响。探索纳米管尺寸和外加电场条件，使得纳米管在电场的协助下能够同时穿透上下表面的细胞的细胞膜，但不影响细胞的活性。探索当细胞膜破裂之后，细胞内的细胞液能够沿着纳米管内腔扩散到对面的细胞内部，从而实现上下表面的细胞间互相传递各自细胞内的蛋白分子。此技术有望实现不同细胞之间直接转移传输细胞内蛋白分子的高效方法。

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