Development of a novel biomaterial model of collagen mediated genetic disease for study of collagen organisation and drug development

开发胶原蛋白介导的遗传病的新型生物材料模型，用于研究胶原蛋白组织和药物开发

• 批准号: 2897511
• 金额: --
• 依托单位: University of Sheffield
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2023
• 资助国家: 英国
• 起止时间: 2023 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2897511
• 关键词: Development; novel; biomaterial; model; collagen

Osteogenesis imperfecta (OI) is a group of 17 inherited connective tissue disorders mediated by mutations in collagen genes that lead to bone deformation and fragility. Currently, in vivo testing is the standard practice for treatments, with over 20 animal models representing OI. These models are technically limited, but also with regards to accessibility and reproducibility of collagen alignment and organisation. A validated in vitro method would allow for accurate preclinical modelling.Mesenchymal stem cells (MSC) can differentiate into bone forming cells and create a bone-like collagen extracellular matrix. Work in Prof. Reilly's group has demonstrated that the organisation and quality of the collagen can be modelled in a tissue specific manner. We propose that the genetic mutations that cause OI will also affect the ability of cells to organise collagen correctly, and that using this scaffold-cell system will provide important insight into OI pathogenesis. The aim of this project is to develop characterised and validated in vitro model of OI collagen pathology. This will be achieved by using electo-spinning to produce polycaprolactone scaffolds on which cells will be grown following established protocols. The relationship between scaffold fibre characteristics and collagen structure will be mapped using light microscopy and SEM for scaffold characteristics, and histology and second harmonic generation microscopy for cellular and extracellular factors. Mechanical testing will be used to determine the role of collagen organisation in tissue structure. OI mutations identified from patient populations will be introduced to MSC cells using CRISPR endogenous gene editing enabling comparison between healthy vs OI collagen. This will allow for investigation of the molecular pathways and testing of therapeutic agents. A high throughput image analysis protocol using ImageJ will be developed for systematic analysis of large data sets.

成骨不完美（OI）是一组17组遗传性结缔组织疾病，该疾病是由胶原基因突变介导的，导致骨变形和脆弱性。当前，体内测试是治疗的标准实践，代表20多个动物模型。这些模型在技术上受到限制，但在胶原蛋白对齐和组织的可及性和可重复性方面也有限。经过验证的体外方法将允许准确的临床前模量。间质干细胞（MSC）可以区分骨形成细胞并产生类似骨状胶原蛋白的细胞外基质。 Reilly教授小组的工作表明，胶原蛋白的组织和质量可以以特定于组织的方式进行建模。我们建议引起OI的遗传突变也会影响细胞正确组织胶原蛋白的能力，并且使用这种支架细胞系统将为OI发病机理提供重要的见解。该项目的目的是开发经过胶原蛋白病理学的特征和验证的体外模型。这将通过使用选举旋转产生多碳酸酯支架来实现，在建立方案之后将在其上种植细胞。脚手架纤维特征与胶原蛋白结构之间的关系将使用光学显微镜和SEM进行绘制，以进行脚手架特征，以及细胞和细胞外因子的组织学和第二次谐波生成显微镜。机械测试将用于确定胶原蛋白组织在组织结构中的作用。使用CRISPR内源性基因编辑从患者种群中鉴定出的OI突变将被引入MSC细胞，从而可以比较健康与OI胶原蛋白。这将允许研究分子途径和治疗剂的测试。使用ImageJ的高吞吐量分析协议将开发用于对大数据集的系统分析。