Construction of an immuno-competent and self reporting human lung model using nanosensor incorporated scaffolds

The study of human lung biology has a huge impact on our understanding of the disease process in a number of lung conditions such as asthma, cystic fibrosis and chronic obstructive pulmonary disease (COPD), disorders which have significant health and socioeconomic implications worldwide. At the moment, it is difficult to carry out such research on humans, because in many cases it is not safe or procedures are too invasive, and the use of animal models is not always appropriate. For instance, mice do not develop asthma naturally which suggest that the biology of their lungs is different to that of humans. These limitations in the availability of physiologically relevant human lung models are therefore set to continue having a negative knock-on effect on the search for novel targets and molecules for therapeutic interventions. For example, despite enhanced patient care, the morbidity and mortality of asthma has remained high with one asthma related death every 19 minutes and 20 million lost working days per annum in the UK alone. This is partly due to lack of efficient therapeutic strategies and the fact that a large proportion of patients do not respond to treatment. What we want to do in this project is to develop a model of the human lung in the laboratory using cells previously isolated from donated tissue or blood. We will grow these cells on materials that contain sensors that can pick up changes in, for example, oxygen, glucose and acidity. These sensors will allow us to observe how cells respond to stimulation in real time. By growing these cell types on these materials, we can arrange them into layers such that the cells are grown in the laboratory in the same position as in the lung. Such a model would give scientists and pharmaceutical companies a better tool for investigating some aspects of human lung biology, identifying new targets for treatment and testing new drug compounds.

The study of human lung biology has a huge impact on our understanding of the pathophysiology of lung diseases. Unfortunately, such studies in human have been hampered by technical and ethical issues which have led to the use of animal models that in most cases have little relevance to human disease. These limitations in the availability of physiologically relevant human lung models are thus set to continue having a negative knock-on effect on the search for novel targets for therapy. Thus, there is a real need to develop biotechnologically-advanced human-based lung models which can be used for studying disease pathogenesis, identifying novel therapeutic targets and assessing response to new drug leads. There are at present only a few in vitro models of bronchial mucosa and none that incorporate the immune elements of this tissue, which are essential for sensing cellular and environmental changes, as well as exerting a crucial role in the pathogenesis of lung diseases. This, therefore, questions the physiological relevance of existing models in studying lung disease pathogenesis and intervention. Another equally important shortfall in the existing models is the lack of appropriate means for non-invasive, real-time and integrated monitoring of cellular responses. To circumvent these limitations, we propose to develop an immuno-competent and self-reporting human lung model comprising dendritic cells, epithelial cells and fibroblasts within a perfusable 3D environment on nanosensor-incorporated scaffolds. The presence of the key cell types and the use of nanosensor-incorporated scaffold sheets will enable us to interactively monitor the microenvironment, cell-cell interaction and cell activation without disturbing the 3D architecture of the model. The adequate cellular and structural representation of the lung tissue and being amenable to in situ monitoring make this model an invaluable tool for research in lung biology, disease modeling and drug discovery/delivery.

This proposal directly addresses the BBSRC's Research and Policy Priority areas of Bionanotechnology, Synthetic Biology and the 3Rs priority for reduction, refinement and replacement of animals, specifically for the purpose of assessment. Beneficiaries from this research include scientists with an interest in nanotechnology, lung biology, immunology biomaterials, tissue engineering and developing 3D in vitro models. Another set of main beneficiaries are the many industries involved in the translation and use of these models, policy regulators and the general public at large. The wider beneficiaries will be as follows: The research staff: The research staff employed on this grant will gain interdisciplinary expertise as a result of the project, through collaborative working with researchers from distinctly different fields of science (material science, nanotechnology, tissue engineering, immunology & imaging). It will bring opportunities to publish in new areas, present to a diverse audience, and to strengthen and develop the direction of their careers. Policy regulators: If this technology is successful and enters routine practice, the avoidance of animal use will directly address the 3Rs criteria. Since March 2009 European directives now impose a marketing ban on any cosmetic substances that involve animal usage, and as such a ban on animal use is effectively now in place. Alternative methods must be used where possible, however the scientific rigour and relevance of many alternative tests is presently lacking. Our technology addresses the need to develop more relevant models while directly addressing new regulatory guidance requiring such change. Industry: This an IPA application and is supported by Kirkstall Ltd., a British company with extensive expertise in developing high throughput screening technologies and multi-chamber bioreactors for growing different tissues. Once the methodology for creating immuno-competent lung epithelia is developed and shown to be effective, it is hoped that it would be translated via industry and become available for routine use. Kirkstall will not only be involved in developing and validating the model, it would also be an ideal vehicle for manufacturing of customised constructs on a commercial basis. This would bring a range of benefits including employment, expertise, and future research and development opportunities. A realistic timescale would be that this could be achieved within 5-7 years. This work would place the UK at the forefront of 3D immuno-competent epithelial models. Currently there are two ECVAM approved epithelial models available for testing. However, both are single cell-type models and as such do not take in to account epithelial-stromal interactions, or immune cell interactions. It is clear from existing research that 3D models populated with these cells give a much more relevant response than single cell type models, so there is an opportunity here for the UK to lead this field, bringing associated economic benefits. General public: The general public as consumers will be exposed directly to a multitude of environmental chemicals. Under European legislation all raw material chemicals must be assessed under REACH legislation. General public concern exists in the use of animal models for such assessment, and so engagement with the public using 3D in vitro models as a basis for accurate and relevant alternatives should be viewed as extremely important. Animals: The major potential beneficiaries of this study will be animals originally utilised for assessment. If our novel approach proves successful and is translated via the relevant industries a new technology could be made available to a multitude of companies worldwide. A major benefit will be the relevance and accuracy of readouts available from the model, and the potential for high-throughout operation. Thus the direct outcomes are expected to have a substantial impact in animal alternative.