Understanding Mechanisms Driving Lung Disease Caused by Environmental Particulate Matter

总的来说，呼吸系统疾病是全世界死亡的主要原因。因此，呼吸系统研究增加了500%，动物模型支持了大约20%的研究。仅在过去十年中，就有超过90万只小鼠被用于帮助了解人类呼吸系统健康，然而，人类和小鼠肺部的生理和解剖差异使得研究结果难以转化为人类治疗方法。吸入研究的重点是吸入的化学物质、颗粒、细菌、病毒、药物输送系统和污染对健康和患病肺的影响。最广泛使用的人类肺特异性模型是基于细胞的模型，主要关注肺上皮细胞，即构成肺表面的细胞。虽然这些模型可用于模拟上皮细胞对吸入物质的反应，但整个肺反应极其重要，包括复杂的细胞间相互作用和信号，免疫细胞和细胞外基质修饰，目前只有小鼠模型才能完全复制，因此它们被过度使用。我们将通过创建一个多细胞型、免疫能力强的人肺泡模型来解决模型平台上的这一空白，该模型不需要动物实验，能够拉伸，并适用于许多呼吸研究领域。为了实现这一目标，我们将使用干细胞衍生的II型肺泡上皮细胞，巨噬细胞，成纤维细胞和内皮细胞，在由我们的工业合作伙伴曼彻斯特BIOGEL开发的合成水凝胶中。我们将演示该模型的应用，以了解大气颗粒物（世界上最重要的环境污染物）如何影响肺部健康。以下目标将为“下一代”人类肺泡模型提供机会，该模型能够模拟上皮反应，纤维化以及免疫系统，并可以在许多呼吸研究领域提供重要的机制，诊断和治疗信息。1)：将hIPSC衍生的肺细胞类型合成无动物水凝胶。我们将把干细胞衍生的II型肺泡上皮细胞、巨噬细胞、成纤维细胞和内皮细胞结合在合成的水凝胶中，生成一个3D多细胞型、免疫能力强的人类肺泡类器官模型。我们将在不同的水凝胶配方中描述类器官的结构，模拟有和没有拉伸力的纤维化。然后，我们将比较和验证在同等条件下培养的人类精确切割肺切片的单个细胞行为。2)了解颗粒物对肺泡组织的影响为了进一步证明该模型的应用，我们将在实验1中确定的不同水凝胶和拉伸特性的环境颗粒物中暴露肺泡类器官。然后，我们将描述每种细胞类型的反应，并使用目标1中人类精确切割的肺切片比较和验证反应。3)教育和培训新用户开发和操作hIPSC衍生肺泡类器官。我们将举办一次会议，重点讨论当前的人类健康和疾病无动物模型，以教育当前和未来的研究领导者使用和开发无动物的人类健康模型。我们还将为希望采用和使用我们的模型进行研究的研究人员举办为期一周的培训课程。总之，这些目标将创建适用的“下一代”人类肺泡组织模型，可以减少和取代许多不同呼吸系统疾病的动物模型。重要的是，将为其他研究人员提供培训机会，使他们能够根据自己的实验需要采用和修改模型，进一步提高该平台在减少研究中动物使用方面产生积极影响的可能性。

Collectively, respiratory diseases are the leading cause of death worldwide. As a consequence respiratory research has increased by 500% and animal models have supported approximately 20% of this research. More than 900,000 mice have been used in the last decade alone to help understand human respiratory health, however, the physiological and anatomic differences between the human and mouse lungs have made it difficult to translate research findings into human therapies. Inhalation research focuses on the effect of inhaled chemicals, particles, bacteria, viruses, drug delivery systems and pollution in both healthy lung and diseased lungs. The most widely used human specific models of the human lung are cell-based models that focus on lung epithelial cells, the cells that make up the surface of the lungs. While these models are useful to model the response of epithelium to inhaled substances, the whole lung response is extremely important and encompasses complex cell-to-cell interactions and signalling, immune cells and extracellular matrix modifications that currently only mouse models can fully replicate, hence their over-use.We will address this gap in model platforms by creating a multi-cell type, immune competent model of human lung alveoli that is animal-free, capable of stretching and applicable to many areas of respiratory research. To achieve this we will use stem cell derived type II alveoli epithelial cells with macrophages, fibroblasts and endothelial cells in a synthetic hydrogel developed by our industrial partner Manchester BIOGEL. We will demonstrate application of the model to understand how atmospheric particulate matter, the most significant environmental pollutant in the world, affects lung health. The objectives below will provide an opportunity for a "next-generation" model of the human lung alveoli, capable of modelling epithelial response, fibrosis as well as that of the immune system and could provide important mechanistic, diagnostic and therapeutic information across many areas of respiratory research. 1): Combine hIPSC derived lung cell types into synthetic animal-free hydrogels.We will bring together our stem cell derived type II alveoli epithelial cells, with macrophages, fibroblasts and endothelial cells in a synthetic hydrogel to generate a 3D multi-cell-type, immune competent organoid model of the human lung alveoli. We will characterise structure of the organoid in different hydrogel formulations that mimic fibrosis both with and without stretch forces. We will then compare and validate individual cell behaviour to human precision cut lung slices cultured under equivalent conditions. 2) Understand the impact of particulate matter on lung alveoli tissueTo further demonstrate application of the model we will expose the alveoli organoids to environmental particulate matter in different hydrogel and stretch characteristics identified in Obj 1. We will then characterise the response of each cell type and compare and validate the response using human precision cut lung slices as in Obj 1. 3) Educate and train new users in development and manipulation of hIPSC derived alveoli organoids.We will host a conference focussed on current animal free models of human health and disease to educate current and future research leaders in use and development of animal-free models of human health. We will also host a 1-week training course for researchers that would like to adopt and use our model for their own research purposes.Together these objective will create a "next generation" model of human alveoli tissue applicable that could reduce and replace animal models across many different respiratory diseases. Importantly, there will be training opportunities provided to other researchers that will allow them to adopt and modify the model to their own experimental needs, further enhancing the possibility that the platform will have a positive impact in reducing animal use in research.