Optimisation of scaffold matrix for epithelial regeneration on tissue engineered airway implants.

Patients who have sustained trauma or required surgery to the voice box and windpipe (larynx or trachea) can develop irreversible scarring leading to severe breathing difficulties that can be life threatening. Whilst conventional treatments are able to reduce scarring and improve breathing it can leave a patient unable to talk or swallow. Furthermore, in a subset of patients reconstruction fails, resulting in severe breathing difficulties and/or the need for a permanent breathing tube to be surgically placed within the neck. Breathing tubes require the patient to nebulise daily and need daily maintenance which can significantly impact on a patient's quality of life. In addition, tracheostomy tubes can be very stigmatising leading to psychological and social problems.

The use of new laboratory techniques has made it possible to engineer new organs within the laboratory and transplant them into sufferers to replace the damaged area. Our group has demonstrated the viability of this technology by transplanting the world's first stem-cell grown windpipe in both an adult and a child to replace an irreversible stricture. Despite the clinical success, a number of hurdles exist before this technology can be translated to every day clinical practice, namely there is a pressing need to improve the regeneration of the cells that line the inside of the graft to prevent scarring and help with cough, speech and swallowing.

Cells are best grown on protein scaffolds that act as a map to direct the cells where to attach and divide. My project intends to take cells from the lung and grow them on a number of different types of scaffold in order to determine which is the best. This new knowledge will then be applied to regenerate a more robust inner lining within the laboratory. By producing a new lining that can be used as part of a stem cell based windpipe or voice box transplant we hope to greatly improve the safety and outcome of this type of surgery and thus provide new hope to hundreds of desperate sufferers across the UK.

This project will take place at University College London (UCL). UCL is amongst the world's premier centres for biomedical research, as confirmed by the results of the UK's latest research assessment exercise (RAE 2008): the numbers of our researchers shown in RAE 2008 to be carrying out world-leading research places UCL among the top universities in the world. In particularly, I will work as part of an established multidisciplinary team already tasked with providing novel therapies for diseased or damaged head and neck tissue which will allow me to draw on a range of expertise and resources unrivalled in any other part of the world.

Aim - Optimisation of scaffold matrix for epithelial regeneration on tissue engineered airway implants

Objectives
1. Determine the key extracellular matrix protein(s) important in the regeneration of respiratory epithelium from a selection of candidate proteins already identified
2. Examine if key ECM protein(s) can be used to enhance respiratory epithelial regeneration on synthetic and collagen scaffolds.
3. To investigate the role of the beta1 and alpha6 integrin subunits in cell adherence and survival of seeded airway epithelium and the downstream signalling pathways responsible

Methodology
i) To define the role of key ECM proteins in cell adhesion, survival, proliferation and differentiation using 2D and 3D organoid culture systems.
ii) To delineate the effects of protein coating of target ECM proteins on epithelialisation of POSS-PCU using air liquid interface culture systems.
iii) Examine the use of compressed collagen sheets in epithelial cell preparation using air liquid interface culture systems.
iv) Examine integrin ECM protein interactions using blocking antibodies and retroviral transduction to overexpress beta one and alpha six integrin receptors.

Scientific and medical opportunities
a) Improved understanding of respiratory epithelial homeostasis and repair
b) Improved respiratory epithelial regeneration in-vitro on POSS-PCU to improve the safety and effectiveness of tissue engineered airway implants
c) Production of respiratory epithelial collagen sheets that would open up new therapeutic options for those with chronic upper airway disease.

Respiratory Regenerative Scientists
This project will examine the role of extracellular membrane (ECM) proteins in respiratory epithelial regeneration and reveal the pathways important in this. In particularly the effect of candidate ECM proteins on integrin mediated signalling pathways will be studied in detail. These findings will greatly improve current understanding of respiratory epithelial homeostatic and repair mechanisms by demonstrating which ECM proteins are important. These proteins could then be traced to areas of potential stem and progenitor cell activity such as the terminal bronchioalveolar junction and compared to other parts of the airway. By furthering our understanding of airway homeostasis and repair it is hoped that therapies can be developed that target these processes to treat chronic lung conditions such as COPD.

Epithelial Regenerative Scientists
All epithelial cells depend on the support of an ECM and contain similar proteomic profiles. By studying the relationship between respiratory epithelium and the surrounding ECM the outcome of this project is transferrable to other epithelial scientists tasked with regenerating epithelium for no-respiratory organs. This project could consequently help drive forward other regenerative medicine strategies such as the delivery of tissue engineered bladders.

Regenerative Medicine / Patients
By improving our understanding of the relationship between respiratory epithelium and the surrounding extracellular environment we hope to enhance the regeneration of respiratory epithelium in-vitro. The combination of this with new technology such as compressed collagen has the potential to deliver a fully functioning epithelial layer in sheet form or as part of an integrated tissue engineered airway transplant. The delivery of a fully functioning and robust respiratory epithelial layer will greatly improve the safety and effectiveness of tissue engineered airway treatments and has the potential to benefit hundreds of desperate patients across the UK annually. In addition the delivery of functioning respiratory epithelial sheet has the potential to open up new therapeutic possibilities such as epithelial covered stents to treat upper airway fistulas with the potential benefit of thousands of patients globally each year.

Pharmaceutical Industry
The delivery of a functioning respiratory epithelial sheet has great potential for use as a model for the pharmaceutical industry for testing the effect of new drugs on the respiratory tract and would also have the added benefit of reducing the need for animal testing. Tissue engineered skin and buccal mucosa are already widely used for a similar purpose.