The use of precision cut lung slices and co-culture modelling to investigate the effect of human rhinovirus on cough and airway inflammation

Stinson, Rebecca Jane

Abstract

Human Rhinovirus (hRV) is a major cause of upper respiratory tract infections (URTIs) and is linked to lower respiratory tract infections (LRTIs) and airway diseases exacerbations. Cough, airway inflammation and hypersensation are common symptoms of hRV infections however, the mechanisms involved remain elusive. A promising hypothesis is neuromodulation, whereby stimulation of transient receptor potential channels (TRPV4) in airway epithelial cells release ATP, subsequently activating the purinoreceptor P2X3 on vagal afferent nerve fibres. Here it is hypothesised that hRV infection will alter expression of channels and receptors known to be involved in cough and airway inflammation, causing lung tissue to become hyperresponsive to cough and bronchoconstrictive agents.

A co-culture model of human airways using a lung epithelial cell line (A549 or Beas-2b) and astrocytes (1321N1 transfected with P2X3) to mimic neurones showed that activation of TRPV4 on lung epithelial cells led to ATP release which could evoke a measurable calcium influx into astrocytes. Additionally, A549 and Beas-2b cells responded differently to RV16 infection, with A549 cells exhibiting a reduced cytotoxicity, increased viability and proliferation compared to Beas-2b cells. Furthermore, RV16 infected ex vivo tissue showed limited impact on viability and cytotoxicity, reduced TEER measurements, increased bronchoconstriction and upregulated ICAM-1 protein expression. Finally, comparison of microfluidic devices to standard culture conditions demonstrated A549 cells could be cultured for 72 hours with similar rates of cell proliferation and viability, whilst PCLS could be cultured for 6 days with minimal impact on tissue viability or morphology.

Overall, findings suggest that the TRPV4–ATP–P2X3 neuromodulation theory may play a role in the mechanism of cough and supports observations indicating hRV causes URTIs more frequently than LRTIs. Furthermore, hRV infection of ex vivo tissue appears promising as a model that can replicate in vivo and in vitro observations. Whilst the microfluidic devices used to replicate elements of the aforementioned models provide a promising start to better recapitulating the in vivo environment.

Citation

Stinson, R. J. (2020). The use of precision cut lung slices and co-culture modelling to investigate the effect of human rhinovirus on cough and airway inflammation. (Thesis). Hull York Medical School, the University of Hull and the University of York. Retrieved from https://hull-repository.worktribe.com/output/4222863

• Thesis Type: Thesis
• Deposit Date: Feb 25, 2021
• Publicly Available Date: Feb 23, 2023
• Keywords: Medicine
• Public URL: https://hull-repository.worktribe.com/output/4222863
• Additional Information: Hull York Medical School, The University of Hull and the University of York
• Award Date: Feb 1, 2020