Development of Complex Liver Organoids Using Cell-Specific Patterned Biomaterials

使用细胞特异性图案化生物材料开发复杂的肝脏类器官

基本信息

批准号：
10654156
负责人：
Muhammad Rizwan
金额：
$ 44.34万
依托单位：
MICHIGAN TECHNOLOGICAL UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-09-01 至 2026-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10654156
关键词：
3-Dimensional Adult Affect Alagille Syndrome Area Bile Duct Diseases Bile fluid Biocompatible Materials Biological Assay Biomechanics Biomedical Research Cells Childhood Coculture Techniques Complex Cues Culture Techniques Data Development Differentiation Antigens Disease model Ductal Epithelial Cell Engineering Foundations Generations Genes Growth Hepatocyte Human Ink Ligands Liver Liver Cirrhosis Liver diseases Modeling Monitor Morphogenesis Mus Organoids Pattern Pharmaceutical Preparations Printing Public Health Relaxation Reporter Research Research Project Grants Signal Transduction Stress Structure System Technology Therapeutic Time Tissue Engineering Tissue Model Training Transplantation Tubular formation Work bile duct bioink bioprinting cancer cell cell type cholangiocyte density drug testing experience improved liver development liver function liver transplantation mechanotransduction new technology notch protein photoactivation response spatiotemporal stem cells three dimensional cell culture undergraduate student viscoelasticity

项目摘要

Liver bile-duct diseases are one of the main causes of liver transplantation, and often result in liver cirrhosis, affecting millions of US citizens. Current human liver organoids lack integrated bile ducts, which makes it difficult to accurately model liver diseases due to a lack of bile-transport system. Liver organoids containing bile ducts have been difficult to create because the hepatocytes and bile-duct cells in the liver have vastly different needs for both Notch signaling and biomechanical cues, which the current 3D-culture techniques are not capable of providing simultaneously. Thus, there is a critical need for a technology that can simultaneously deliver targeted Notch signaling and biomechanical cues to both types of liver cells in co-culture, in order to maintain their maturation. Exciting preliminary studies indicate that the dual-bioink-based bioprinted constructs can provide cell-type-specific signals within a co-culture matrix. Moreover, building on their previous work, the PI has developed new engineered matrix to precisely tune the bio-mechanical cues (stiffness and viscoelasticity) in a cell-specific manner, which supports the growth of 3D human bile-duct network. Accordingly, the objective of this proposal is to evaluate and optimize the effect of patterned Notch signaling and bio-mechanical cues in co-cultured liver cells to develop liver organoids with integrated bile-flow system. The rationale is that liver organoids with integrated bile-flow system will mimic liver function, thus will improve disease modelling and drug testing. The proposed research will pursue two specific aims: (1) Determine the effect of spatio-temporal Notch activation on liver organoid functions, and (2) Optimize the maturation of liver organoid via cell-type-specific biomechanical cues. In the first aim, bioprinted constructs will be developed using two distinct, cell-laden bioinks with and without photo- activatable Notch ligands, to achieve targeted Notch activation in co-culture. The organoids will be analyzed using Notch target genes and a range of liver functional assays. In the second aim, an 18-condition matrix screen will be developed to systematically evaluate and optimize the effect of patterned biomechanical cues on liver organoids in a bioprinted co-culture construct. Finally, the mechano-sensing mechanism will be evaluated in co-culture construct. The proposed research is expected to be significant because it will leverage targeted Notch signaling and biomechanical cues to inform the development of liver organoids with integrated bile ducts for therapeutic applications, and will train a diverse group of undergraduate students in the area of bioprinting, biomaterials, and liver tissue engineering.

肝胆管疾病是肝移植的主要原因之一，常常导致肝硬化，影响数百万美国公民。目前的人类肝脏类器官缺乏完整的胆管，由于缺乏胆汁运输系统，因此很难准确地模拟肝脏疾病。含有胆管的肝脏类器官很难制造，因为肝脏中的肝细胞和胆管细胞对 Notch 信号传导和生物力学线索的需求截然不同，而当前的 3D 培养技术无法同时提供这些需求。因此，迫切需要一种技术能够同时向共培养的两种类型的肝细胞传递靶向Notch信号传导和生物力学线索，以维持它们的成熟。令人兴奋的初步研究表明，基于双生物墨水的生物打印结构可以在共培养基质内提供细胞类型特异性信号。此外，在之前工作的基础上，PI 开发了新的工程矩阵，以细胞特定的方式精确调整生物力学线索（硬度和粘弹性），支持 3D 人类胆管网络的生长。因此，本提案的目的是评估和优化共培养肝细胞中模式化 Notch 信号传导和生物力学线索的效果，以开发具有集成胆流系统的肝脏类器官。其基本原理是，具有集成胆汁流动系统的肝脏类器官将模拟肝功能，从而改善疾病模型和药物测试。拟议的研究将追求两个具体目标：（1）确定时空Notch激活对肝脏类器官功能的影响，（2）通过细胞类型特异性生物力学线索优化肝脏类器官的成熟。在第一个目标中，将使用两种不同的、带有或不带有可光激活Notch配体的充满细胞的生物墨水来开发生物打印结构，以在共培养中实现靶向Notch激活。将使用 Notch 靶基因和一系列肝功能测定来分析类器官。第二个目标是开发 18 条件矩阵筛选，以系统地评估和优化生物打印共培养结构中图案化生物力学线索对肝脏类器官的影响。最后，将在共培养构建中评估机械传感机制。拟议的研究预计将具有重要意义，因为它将利用有针对性的 Notch 信号传导和生物力学线索来为治疗应用中具有集成胆管的肝脏类器官的开发提供信息，并将在生物打印、生物材料、和肝脏组织工程。