Additive Manufacturing of and for Organ-on-a-Chip models and analysis of their Extravesicular Secretions

细胞外囊泡（EV）是由细胞分泌的微小液滴，由一层薄薄的脂质膜（想想肥皂泡）组成，充满液体，并装有蛋白质和核酸。人们已经认识到EV在许多疾病中起着关键作用，并作为细胞和器官之间的主要通信途径，充当细胞之间发送的“包裹”，但分析仍然具有挑战性。EV，特别是直径为60至120 nm的EV，称为外泌体或小EV，由细胞分泌，并且可以从各种体液（如唾液、尿液和血液）中回收。小EV含有特定的分子货物，可用于识别其亲本细胞，描绘细胞组成和失调，特别是神经系统疾病，癌症和最近的糖尿病。EV可以通过血脑屏障迁移，而来自胰岛的EV已被证明含有糖尿病特征。已经确定癌细胞比健康细胞分泌更多的EV，并且它们可以在血液中发现，并且含有癌症特异性特征。因此，EV作为疾病和癌症的生物标志物具有很高的潜力。 芯片上器官和微型器官的生物制造是一个相对较新的、快速发展的研究课题，它使组织、芯片上器官和类器官的3D构建能够在体外重现人体生理学的相关特征。我们的目标是分析复杂的3D结构和类器官分泌的EV，并揭示EV信号在健康和疾病中的作用。在这里，我们提出了一个平台，该平台构建并结合了我们团队在微生物工程和生物医学研究方面的集体专业知识，以在类器官模型的背景下研究EV，包括脑和hPSC衍生的胰岛类器官。我们的目标是创建更先进的器官模型，并了解EV在健康和疾病期间细胞与细胞和器官通讯中的作用。所要求的仪器包括最先进的高分辨率和大构建面积3D打印机，其允许使用生物相容性墨水来创建（i）用于引导血管化的复制模制微流体结构，（ii）直接生物打印细胞以形成芯片上器官，以及（iii）脑和胰岛样类器官。所要求的仪器还包括用于表征这些细胞系统的EV分泌物（特别是尺寸分布）的纳米颗粒尺寸测量仪器。

Extracellular vesicles (EVs) are tiny droplets secreted by cells and are made up of a thin lipid membrane (think of soap bubble), filled with liquid, and packed with protein and nucleic acids. EVs have been recognized to play a critical role in many diseases and function as a major communication pathway between cells and organs, acting as “parcels” sent among cells, but remain challenging to analyze. EVs, in particular those with a diameter of 60 to 120 nm, known as exosomes or small EVs, are secreted by cells, and can be retrieved from various bodily fluids, such as saliva, urine and blood. Small EVs contain specific molecular cargos, which could be used to identify their parental cells, delineate cellular composition and dysregulation, in particular for neurological diseases, cancer, and recently diabetes. EVs can migrate through the blood-brain barrier, while EVs derived from islets have been shown to contain diabetes signatures. It is already established that cancer cells secrete more EVs than healthy cells and that they can be found in blood, and contain cancer-specific signatures. Hence EVs have a high potential as accessible biomarkers for diseases and cancer. Biomanufacturing of organs-on-a-chip and miniorgans is a relatively new, rapidly evolving research topics, which enables 3D constructs of tissue, organ-on-a-chip and organoids that recapitulate relevant features of human physiology in vitro. We aim to analyze EVs secreted by complex 3D constructs and organoids, and shed light on EV signaling in health and disease. Here, we propose a platform that builds and combines the collective expertise of our team in microbioengineering and biomedical research to study EVs in the context of organoid models, including brain and hPSC-derived islet organoids. We aim to create more advanced organ models and gain understanding of EVs' role in cell-cell and organ communication during health and disease. The requested instruments include a state-of-the-art, high resolution and large build area 3D printer that allow the use of biocompatible ink for creating (i) replica molded microfluidic structures for guiding vascularization, (ii) direct bioprinting of cells to form organ-on-a-chip, and (iii) brain and islet-like organoids. The requested instruments also include a nanoparticle sizing instrument for characterizing, particularly the size distribution, EV secretions of these cellular systems.