Biomaterial processing for organ-on-a-chip engineering

组织工程可以为旁分泌信号、组织功能和化合物发现的研究提供微生理学的芯片平台。干细胞生物学的最新进展使得通过将多能干细胞(PSC)分化为专门的谱系，如心肌细胞(CM)、肝细胞等，可以获得几乎所有类型的人类细胞。然而，整个领域面临着两个关键限制：1)将几种细胞类型集成到高保真的芯片上组织模型中的复杂性仍然存在。2)缺乏对体外构建功能器官的结构-功能关系和设计规则的了解。利用生物材料和聚合物科学、材料加工、微制造和化学工程的方法，这项研究计划将克服这两个限制。目前大多数单芯片器件使用聚二甲基硅氧烷(PDMS)，这是臭名昭著的小分子吸收和浸出。片上设备具有封闭的配置，通常将组织密封在PDMS层和玻璃基板之间，需要泵驱动的流动。这使得直接接触组织变得困难，各种隔室的整合呈指数级复杂，需要笨重的外部设备。在体内，代谢物通过连接不同器官的血管系统对流输送，穿过内皮进入实质空间。在项目1中，为了设计一个坚固的组织然后是芯片上器官的平台，我们将首先建立一个通用的和功能强大的血管床来支撑并连接不同的组织。为此，我们将开发一种新的3D微冲压技术，以创建基于聚酯的可灌流的分支血管系统，该系统满足两个相反的标准：机械稳定的壁，但可渗透到小分子、蛋白质和最终迁移的细胞。血管床将在无PDMS的生物反应器中种植实质细胞，具有孔板的占地面积和开放的液体分配通道。我们的目标是将心脏、肝脏和乳腺肿瘤组织整合到一个单一平台上。在项目2中，我们假设可以通过生物打印多层3D结构来设计具有功能的心脏左室(LV)，该多层3D结构概括了天然LV的复杂纤维方向，方法是使用微流控生物打印机在水凝胶片中指定CM附着的粘附区和非粘附区。我们预计，具有适当纤维取向的LV能够像自然心室一样同步扭曲和收缩，而各向同性或周向排列的纤维则不能。项目1和项目2作为项目1的分支血管相互连接，可用于项目2的放大研究。这两个项目都将使用从人类诱导的PSC分化而来的细胞。三名博士、两名硕士和五名本科生将通过拟议的学习得到培训。

Tissue engineering may provide microhpysiological on-a-chip platforms for studies of paracrine signaling, tissue function and compound discovery. Recent advances in stem cell biology enable procurement of virtually all human cell types by differentiation of pluripotent stem cells (PSC) into specialized lineages e.g. cardiomyocytes (CM), hepatocytes, etc. However, the entire field is faced with two critical limitations: 1) Complexities related to integrating several cell types into high-fidelity tissue-on-a-chip models still remain. 2) Lack of understanding of structure-function relationships and design rules for building functional organs in vitro. Using approaches from biomaterial and polymer sciences, materials processing, microfabrication and chemical engineering this research program will overcome both limitations. Most current on-a-chip devices utilize poly(dimethylsiloxane), PDMS, which is notorious for absorption and leaching of small molecules. The on-a-chip devices have a closed configuration, usually with the tissue sealed between the PDMS layer and a glass substrate, requiring a pump driven flow. This makes direct access to the tissue difficult and integration of various compartments exponentially complex, requiring bulky external equipment. In vivo, metabolites are delivered by convection through the vasculature connecting different organs, passing through the endothelium into the parenchymal space. In Project 1, to engineer a robust tissue then organ-on-a-chip platform, we will first build a generic and functional vascular bed to support and then connect different tissues. For this purpose, we will develop a new 3D micro-stamping technique to create polyester-based perfusable branching vasculature that accommodates two opposing criteria: mechanically stable walls yet permeable to small molecules, proteins and ultimately migrating cells. The vascular bed will be seeded with parenchymal cells in a PDMS-free bioreactor, with a footprint of a well plate and open access for liquid dispensing. We aim to integrate cardiac, liver and breast tumor tissue onto a single platform. In Project 2 we hypothesize that a functional heart left ventricle (LV) can be engineered by bioprinting a multilayer 3D construct that recapitulates complex fiber orientation of the native LV, by specifying adhesive and non-adhesive regions for CM attachment in a hydrogel sheet using a microfluidic bioprinter. We expect the LV with appropriate fiber orientation to be capable of synchronously twisting and contracting as the native ventricle, while isotropic or circumferentially aligned fibers will not. Project 1 and 2 are interconnected as branching vasculature from Project 1  could be used in scale-up studies for Project 2. Both Projects will use cells differentiated from human induced PSC. Three PhD, 2 MASc and 5 undergraduate students will be trained through the proposed studies.