Signal transduction and the regulation of ion transport in airway epithelia
It has recently been shown that a critical component of airways innate defense is the thin (7 µm) liquid layer lining airway surfaces, the periciliary liquid (PCL), that provides a low viscosity solution for ciliary beating and acts a lubricant layer for mucus transport. Normal airways appear to be able to sense the PCL volume and adjust ion channel activity accordingly by unknown mechanisms. A proposed model for ASL volume regulation is shown in Figure 1.
Apical membrane ion channel activity controls the amount of salt (and water) on airway surfaces and hence, PCL volume and mucus hydration levels. It has recently been proposed that the initiating event in CF lung disease is depletion of the PCL due to abnormal ion channel activity (i.e. a lack of CFTR), which causes dehydrated mucus to adhere to airway surfaces, preventing it from being cleared (Figure 2), causing increased bacterial infections.
The long term goal of this laboratory is to understand how homeostasis of PCL volume occurs in airway epithelia under normal and pathophysiological conditions. Currently, research in the Tarran lab is focused on three main areas, listed below, and we utilize cell biological and biochemical techniques coupled with in vivo translational approaches to address these questions:
- Regulation of epithelial cell function by the extracellular environment: We have hypothesized that nucleotides, proteases and other molecules contained in the ASL (ATP for example) can regulate airway ion transport. Fig. 3 depicts cystic fibrosis airway cells that have lost the ability to regulate ASL volume following infection with GFP-labeled viruses that inhibit intracellular Ca2+ signaling by depleting extracellular ATP.
Gender differences in cystic fibrosis lung disease: Females suffer more severely with cystic fibrosis than their male counterparts, resulting in a significantly shorter lifespan. This gender difference becomes apparent after puberty. Accordingly, we are currently investigating whether estrogens affect Ca2+signaling and ASL homeostasis. Figure 4 shows an example of simultaneous Fura-2 imaging (as a marker of intracellular Ca2+) and a fusion protein of Estrogen Receptor α (ER α) conjugated to orange fluorescent protein (mOr).
Figure 3. Figure 4. Figure 5.
- The effects of cigarette smoke on epithelial airway ion transport: Similar to CF, long term tobacco exposure also results in chronic mucus accumulation. We have found that acute tobacco exposure causes a loss of CFTR from the plasma membrane (Figure 5) and are currently investigating this phenomenon.
Techniques used in our lab:
- Ca2+ imaging
- Confocal microscopy
- Electrophysiology (In vivo nasal potential difference measurements, microelectrodes & Ussing chambers)
- Fluorescence resonance energy transfer (FRET)
- Mass Spectrometry
- Molecular Biology & Real Time (q)PCR
- Tissue culture
- Western Blot