Cellular and molecular control of human embryonic alveolar development: towards lung regeneration

During human embryonic development the lung is one of the last organs to become fully formed and ready for birth. When a baby is born prematurely normal embryonic lung development is interrupted and many premature infants require breathing support. Some of these children, particularly the most premature, develop a long-term lung condition called bronchopulmonary dysplasia (BPD). However, even premature infants who do not develop BPD frequently have decreased lung function throughout life. These facts illustrate that interruptions in embryonic lung development are not naturally caught-up during childhood growth. The Wold Health Organisation estimates that every year 15 million babies are born prematurely and that this figure is steadily rising. The aim of this proposal is to study the normal mechanisms of human embryonic lung development in order to identify new strategies to improve the lung health of premature infants.

Surprisingly little is known about the cell types which work together to build a lung in the human embryo and even less is known about the cell-cell communication mechanisms that coordinate the process. Instead of working with animal models of lung development, this project will focus on studying human embryonic lung development using human embryonic lungs. This is now possible due to advances in molecular genetics and new techniques for growing human organs in the laboratory as mini-organs, or organoids.

We will identify the different cells that are involved in building the lung and determine the signals that these cells use to communicate with each other. This study will provide a base-line for normal human lung development, allowing us to identify points for which therapies could be developed to promote lung maturation. In addition, during this project we will improve our techniques for growing human mini-lungs in the laboratory. These mini-lungs will provide a system for us to investigate the genetic causes of some premature lung conditions like BPD. These genetic experiments will assist with the development of new markers for the disease helping doctors to determine which premature babies are at risk. In addition, they will provide a system in which treatments to reduce the effects of BPD, and other lung problems associated with prematurity, can be tested.

Interruptions in normal embryonic lung development caused by premature birth lead to a life-long decrease in respiratory capacity. We will study the cellular and molecular mechanisms underlying human embryonic alveolar development, and develop improved in vitro alveolar models, as the first steps to improving outcomes for premature neonates.

Our preliminary data show that there are numerous differences in the molecular regulation of mouse and human embryonic lungs. We will therefore focus our research directly on human embryonic lungs and a human embryonic lung organoid system that my lab has developed.

We will reconstruct the alveolar epithelial cell lineage in human embryos, and our organoid system, using single cell RNA-seq. A major focus of the bioinformatics approaches will also be analysis of epithelial cell signalling pathways.

The human embryonic lung epithelial organoid system that we have developed will be used to screen for combinations of signalling molecules that are sufficient to promote alveolar epithelial cell differentiation.

We will use wholemount antibody staining to identify the mesenchymal cell populations participating in human alveolar development. Lentiviral-mediated lineage-labelling in multiple single cells in lung slice cultures and kidney capsule grafts, followed by quantitative analysis, will be used to determine the mesenchymal cellular hierarchy. The organoid system will be combined with specific populations of mesenchyme and endothelial cells to determine cell-cell interaction nodes that control specific aspects of differentiation and morphogenesis.

The organoid system will be further developed into a genetically-manipulable model for studying human alveolar disease. We will focus on using this to study contributing genetic factors to BPD and will also investigate its utility for other lung conditions.

1) Benefits to patients, the health service and the public
Ultimately the main beneficiaries of this work will be patients with decreased lung capacity due to interruption of in utero lung development, or degenerative lung disease. In the long-term our work on the cellular and molecular mechanisms of normal lung development, and the development of new model systems, will lead to improved diagnoses and new therapeutic options for these patients. Although it needs to be recognised that we are laying the ground-work for a long-term programme that will ultimately require the efforts of many academic and industrial labs over a time-scale of more than 10 years.

2) Commercialisation
This project has several potential commercial applications which could be important for the economic competitiveness of the UK. Although therapeutic benefits to patients will take many years to realise, commercial application of any advances could begin within the lifetime of the award. In particular, our work on the development of culture conditions for the growth of populations of lung epithelial cells. These media could potentially be transferred to pluripotent cell differentiation and maintenance of adult lung epithelial cells. Such growth media could make the possibility of industrial drug toxicology testing/screening on large numbers of mature lung cells a realistic possibility. With the assistance of Cambridge Enterprise we will investigate possibilities for commercialisation, including patenting, with a view to developing media with pharmaceutical companies through collaboration or licensing. We are already discussing the commercialisation potential of our current organoid growth medium with Cambridge Enterprise.

3) Staff Development
During this project we will develop professional skills that we will be able to use in our future careers. We will develop our research skills and expand our area of expertise. In particular, the adaptation of our organoid culture conditions to high-throughput screening techniques and the use of CRISPR-Cas9 for gene editing will be techniques that are transferable to multiple scientific roles in academic and industrial settings. In addition, the University of Cambridge offers numerous formal training opportunities in leadership, mentoring, science communication that we will take advantage of. These will all be important impacts for myself and the staff involved which will be achieved within the life-time of the award.