Ion Channel Fingerprinting of Tracheal and Articular Cartilage
Authors: Kadir Lina Abdul , Sharif Umar , O'Brien Fiona , Whysall Kasia , Barrett‐Jolley Richard 
Published: June 2021

Cartilage is a tough and flexible tissue that protects and lubricates joints and provides structural support in the nose, ribs and trachea. Cartilage is made up of a single cell type, chondrocytes, which synthesise the extracellular matrix (ECM) in which they reside in. Although chondrocytes are non‐excitable cells, their plasma membrane is rich in ion channels. It was recently shown that ANO1 (TMEM16A) and ANO2 (TMEM16B), which are calcium‐activated chloride channels (CaCC), are vital for normal tracheal development in mice. Deletion of these ion channels causes abnormal development of the cartilage rings and results in tracheomalacia (TM) [1], which is an increase in the collapsibility of the trachea due to structural abnormalities in the cartilage rings. It has been shown that certain microRNAs, such as miR‐125b and miR‐30a/c lead to the pathogenesis of TM. This occurs through either the targeting of the fibroblast growth factor (FGF) signalling pathway, or the TMEM16A protein, and results in abnormal cartilage formation. Thus far, numerous ion channels have already been identified in the articular chondrocyte ‘channelome’, but there are comparatively fewer existing tracheal chondrocyte data. In this project, we have combined electrophysiological and molecular biology techniques to investigate whether tracheal chondrocytes express similar ion channel sub‐types to articular chondrocytes. In particular, we are interested in the expression of chloride‐selective channels in tracheal chondrocytes that are thought to play a role in TM.Rat tracheal and articular chondrocytes were isolated with overnight digestion with collagenase II and the expression of select chloride ion channel genes was identified using qPCR. Unbiased detection of ion channel genes was achieved through RNA‐seq whilst analysis of functional ion channel expression was made with patch‐clamp electrophysiology.Current data show several significant differences between the tracheal and articular chondrocyte ion channel expression; for example the ANO1 and ANO2 genes, had significantly higher expression in tracheal chondrocytes in comparison to articular chondrocytes (qPCR: n=3, 3 respectively p<0.01; RNAseq: n=5, 3 respectively p<0.00005). These findings were validated with patch‐clamp experiments, where we identified and characterised the single‐channel gating of several ion channel subtypes.Whilst tracheal and articular chondrocytes share several common ion channels, there were distinct differences observed. For example, chloride channels and a few potassium channels were found to be significantly higher in tracheal chondrocytes compared to articular chondrocytes. Future experiments will further optimise our electrophysiological conditions to correlate the ion channel gating observed to the molecular signatures we have identified. The interplay of these ion channels with miRNAs will also be explored to try to understand the mechanisms associated with TM. Understanding how these two cartilage sub‐types differ in terms of ion channel expression will not only broaden our knowledge of tracheal physiology, but could also aid in the development of therapies for airway diseases such as TM.Support or Funding InformationWork funded by The University of Liverpool, Crossley Barnes Bequest.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.