Controlling cellular and organismal microenvironment using light

细胞和有机体的行为与其直接微环境的物理和化学特性密切相关。无论是在体内还是在培养中，细胞所附着的底物的硬度和粘附性都可以深刻地影响细胞的特性和行为，包括分化过程中的形状、运动性和命运选择。模式生物，如线虫，使用机械感官将地形和环境的质地整合到它们的行为中。细胞和有机体对其微环境的反应可以反过来被用来控制它们的行为。例如，将细胞置于细胞外基质(ECM)的线性微图案上可以诱导持续的定向迁移，而将它们电镀在孤立的盘状图案上可以使它们固定不动。因此，细胞外基质的微模式可以用来产生刻板的、高度重复性的细胞行为，便于在活细胞荧光显微镜研究中对潜在机制进行系统的定量分析。同样，通过将线虫放置在定义的3D微环境中，可以控制它们的行为，并确定它们对地形特征和化学线索的神经元反应。 我们建议在McGill的生命科学复合体中建立一个光调制系统，用于三个主要应用：(1)控制细胞黏附和迁移行为的ECM的微图案化，(2)用于微流体的复杂聚二甲基硅氧烷(PDMS)三维结构的微制造或研究细胞和线虫如何对复杂的三维地形做出响应，以及(3)构建水凝胶来模拟细胞遇到的复杂的三维组织环境。 为了建立这个光敏系统，我们要求一个“Primo”紫外光光敏系统(法国Alvéole)，以及能够将其集成到现有的、目前未得到充分利用的倒置光学显微镜系统中的部件。Primo系统将置于高级生物成像设备(ABIF)的管理之下，ABIF将为用户提供培训并支持该仪器，使受训人员和研究人员能够方便地进行微图形、微结构和微制造。 Primo系统对于我们由NSERC资助的不同研究领域的研究项目至关重要，这些研究领域包括单分子生物物理学、焦点黏附动力学/细胞迁移、集体细胞迁移/形态发生、免疫细胞动力学、神经电路功能和计算建模。控制细胞和组织微环境是实验设计的一个新的关键方面，使新的问题能够通过在基础科学、有机生物学、组织工程和再生医学中的应用来解决。因此，有了这个系统，将大大加强HQP培训。虽然目前的申请包括六个小组，但这些设备将通过ABIF向生命科学综合体的&gt；100实验室和更广泛的研究社区开放。

The behavior of cells and organisms is intimately linked to the physical and chemical properties of their immediate microenvironment. The stiffness and adhesiveness of the substrate to which cells adhere, both in vivo and in culture, can profoundly affect cellular properties and behavior, including shape, motility, and fate choices during differentiation. Model organisms such as the nematode C. elegans use mechanosensation to integrate the topography and the texture of their environment into their behavior. The response of cells and organisms to their microenvironment can in turn be exploited to control their behaviors. For example, putting cells on linear micropatterns of extracellular matrix (ECM) can induce persistent directional migration, whereas plating them on isolated disc-shaped patterns can render them immobile. Thus, micropatterns of ECM can be used to generate stereotypical, highly reproducible cell behaviors facilitating systematic quantitative analysis of underlying mechanisms in live-cell fluorescence microscopy studies. Similarly, by placing C. elegans in defined 3D microenvironments, their behavior can be controlled and their neuronal responses to topographical features and chemical cues be determined. We propose to establish a photopatterning system in McGill's Life Sciences Complex for three main applications: (1) micropatterning of ECM to control cell adhesion and migration behavior, (2) microfabrication of complex polydimethylsiloxane (PDMS) 3D structures for microfluidics or to study how cells and C. elegans respond to complex 3D topography and (3) structuring hydrogels to mimic complex 3D tissue environments that cells encounter. To establish this photopatterning system, we are requesting a “PRIMO” UV photopatterning system (Alvéole, France), as well as components to enable its integration into an existing, currently under-utilized inverted light microscopy system. The PRIMO system will be placed under the management of the Advanced BioImaging Facility (ABIF), who will provide training to users and support the instrument, making micropatterning, microstructuring, and microfabrication highly accessible to trainees and researchers. The PRIMO system is essential for our NSERC-funded research programs in diverse research areas including single-molecule biophysics, focal adhesion dynamics/cell migration, collective cell migration/morphogenesis, immune cell dynamics, neural circuit function and computational modeling. Controlling the cellular and organismal microenvironment is an emerging key aspect of experimental design enabling new questions to be addressed with applications in basic science, organoid biology, tissue engineering, and regenerative medicine. Thus, having this system in place will significantly enhance HQP training. While six groups are included in the current application, the equipment will be accessible to >100 labs in the Life Sciences Complex and the broader research community through the ABIF.