Bioactive fragments of the extracellular matrix orchestrate lung epithelial cell repair.

On a daily basis, our lungs are exposed to an array of environmental insults including viruses, bacteria and toxic particles. Specialised 'epithelial' cells run from our nose all the way down to the depths of our lungs, forming a barrier to the external environment and protecting us from these insults. However, these epithelial cells are frequently and sometimes severely damaged by these exposures. It is critical that after injury, the epithelial cells are quickly repaired to restore this barrier. If epithelial cells are not efficiently repaired then we are more prone to infection, and our ability to breathe is compromised. The lungs are able to repair themselves when damaged but this ability deteriorates as we get older and in some people the repair process doesn't work properly, leading to disease. Currently, there are no treatments able to restore damaged lung tissue, and this is clearly an urgent clinical need. A greater understanding of how the lungs repair themselves is required to promote long term lung health and to identify new treatments that can promote lung repair.

The extracellular matrix (ECM) is a three-dimensional meshwork of proteins and other factors that supports the structure of the lungs and acts as a scaffold for cells that populate the lungs. We are interested in a small fragment of this ECM called Pro-Gly-Pro (PGP), which is normally hidden but becomes released from the ECM in response to infection or injury. We have exciting data that demonstrates that PGP is potent at promoting repair responses in lung epithelial cells. Furthermore, PGP can also operate to drive the influx of cells called neutrophils into the lungs. Neutrophils are essentially the soldiers of our immune system that can kill any invading organisms that have entered the lung as a result of injury. Therefore, we believe that PGP is a fragment of the lung tissue that is released in response to injury and then subsequently acts to direct localised epithelial repair to seal the breach to the external environment, whilst simultaneously causing the influx of neutrophils to sterilise the lung tissue. We also believe that pathways governing the levels of PGP may be disrupted in disease settings. Consequently, understanding how PGP promotes repair responses could yield novel treatments to counteract lung injury. Because the ECM is a critical component of all tissues, our data is highly likely to also be relevant for repair of other organs in the body.

In this proposal, we want to understand more about how PGP drives repair in epithelial cells and ascertain the relative importance of PGP as a mediator of repair following lung injury. We will use epithelial cells isolated from the lungs of healthy individuals to probe how exactly PGP is able to drive repair responses, thus revealing potential strategies for therapeutic intervention. Subsequently, we will induce micro-injuries in slices of human and mouse lung tissue that are essentially 'mini lungs' and assess how manipulation of PGP in this more complex 3D setting modulates subsequent repair responses. The use of human lung cells and tissue is critical if we are to understand the importance of PGP to human lung injury and repair. However, to truly demonstrate the capacity of PGP to instigate lung repair and minimize pathology over a prolonged period of time, it is also necessary to assess the role of PGP in a mouse model of lung epithelial cell injury. We will determine the importance of naturally generated PGP in supporting epithelial repair and also ascertain to what extent supplementation of PGP can enhance repair. The results of this proposal could lead in future to new treatments that can promote lung repair via modulation of PGP.

The respiratory epithelium is a target of frequent and often severe injury. It is critical that a reparative programme is readily initiated to restore epithelial integrity and preserve lung function, whilst minimizing pathogen invasion. Deficient epithelial repair is a feature of aging and lung diseases. To date, research into lung repair has focused on identifying key stem/progenitor populations and growth factors regulating them. However, it is equally important to understood how microenvironmental cues orchestrate appropriate and targeted epithelial repair responses. This is critical to long term respiratory health and identification of novel pro-repair therapeutics.

The tripeptide Pro-Gly-Pro (PGP) is a collagen fragment liberated during inflammation and classically described as a neutrophil chemoattractant. The physiological importance of PGP in health is poorly understood. We challenge the central dogma, arguing that PGP primarily functions to promote airway epithelial cell (AEC) repair, whilst operating indirectly via AECs to drive neutrophilic inflammation. In response to injury, we hypothesize that AECs drive PGP generation, which then functions to repair proximal AECs to seal the breach to the external environment, whilst promoting neutrophilic inflammation to sterilise the site.

Aim (1) will use primary human AECs to establish their capacity to generate PGP in response to environmental stressors, and subsequently interrogate how PGP regulates distinct processes central to epithelial repair. Aim (2) will use established injury models in mouse and human precision cut lung slices to validate the importance of PGP in defining repair in situ in the context of complex lung architecture. Aim (3) will utilise a mouse model of naphthalene-induced lung injury to demonstrate how manipulation of PGP can modulate epithelial repair in vivo. Combined, these studies will generate novel mechanistic insight into pathways coordinating epithelial repair.