Development of Human Intestinal Simulacra

 描述（由申请人提供）：在本申请中，提出了一个合作的多学科研究项目，该项目有可能为健康和疾病中的人类生理学研究创造一个新的范式。将开发最先进的微制造平台，以创建人结肠上皮及其相关微生物组的功能性体外复制品，即模拟物。这项新技术将用于进行新的研究和假设检验肠道生理学，目前不能进行。此外，这里开发的技术和方案将为从正常和患病的原代人体组织中创建器官模拟物奠定基础，这将改变组织功能，微生物组影响和药物作用的研究方式。结肠上皮干细胞器官型培养的最新进展使得在凝胶状基质中产生长寿命的球状体成为可能。然而，不存在化学梯度以及图案化的物理支撑导致无序和混乱的构象，其很难模拟结肠的结构和功能，并且此外不适合微生物组共培养。我们建议开发一种新型的微制造系统，该系统将使人类结肠上皮细胞和覆盖的微生物群的离体培养成为可能，该微生物群在非极化细胞培养系统和完整人体有机体的复杂性之间的设置中再现了结肠粘膜的3D结构和环境。我们实验室在结肠上皮干细胞单层持续培养方面的最新技术进展将与微加工支架和设备相结合，以创建结肠模拟物。许多创新将被纳入微工程系统，目标是重现结肠干细胞生态位，分化的肠粘膜，微生物群以及这些隔间之间的动态信息流。该平台将允许通过提供独立的流体和气体进入这些隔室来严格控制腔和基底隐窝环境。该平台将支持有丝分裂原，形态发生，分化因子，饮食化合物和气体梯度的形成，以实现对结肠生理学的前所未有的研究。将测试与饮食因素、微生物组和结肠上皮相互作用相关的一些假设，以证明这种变革性技术的力量和广泛适用性。

 DESCRIPTION (provided by applicant): In the current application, a collaborative, multidisciplinary research project is proposed that has the potential to create a new paradigm for the study of human physiology in health and disease. A state-of-the-art microfabricated platform will be developed to create a functional, in vitro replica, i.e. simulacrum, of the human colonic epithelium and its associated microbiome. This new technology will be used to perform novel studies and hypothesis testing of intestinal physiology that cannot currently be performed. Furthermore, the technology and protocols developed here will establish the basis for creating organ simulacra from normal and diseased primary human tissues that will change the manner in which studies of tissue function, microbiome influence, and drug effect are performed. Recent progress in organotypic culture of colonic epithelial stem cells has made it possible to create long-lived spheroids in a gelatinous matrix. However, the absence of chemical gradients as well as patterned physical support results in a disorganized and chaotic conformation that poorly mimics the structure and function of the colon, and furthermore is not amenable to microbiome co-culture. We propose to develop a novel microfabricated system that will enable ex vivo culture of human colonic epithelium and overlying microbiota that recapitulates the 3D structure and environment of the colonic mucosa in a setting which lies between an unpolarized cell culture system and the complexity of the intact human organism. Recent technical advances from our labs in sustained monolayer culture of colonic epithelial stem cells will be integrated with microfabricated scaffoldings and devices to create the colonic simulacrum. A number of innovations will be incorporated into the microengineered system with the goal of recapitulating the colonic stem-cell niche, the differentiated intestinal mucosa, the microbiota, and the dynamic information flow between these compartments. The platform will permit tight control of the luminal and basal crypt environments by providing independent fluidic and gaseous access to these compartments. The platform will support formation of mitogen, morphogen, differentiation-factor, dietary-compound and gaseous gradients to enable unprecedented investigations into colonic physiology. A number of hypotheses related to the interplay of dietary factors, the microbiome and the colonic epithelium will be tested to demonstrate the power and broad applicability of this transformative technology.