Rhythms in Disease- Determining Circadian Clock Mechanism in Asthma

Asthma is a common disease affecting the airways by causing narrowing, leading to symptoms of wheeze and breathlessness. Everyday in the UK, 3 people die from asthma, mostly during the night-time. Those with severe asthma are difficult to treat and there is an urgent need for new therapeutic options. Asthma is a very rhythmic disease; symptoms go up and down during the day, often peaking during the night. My research has shown that many aspects of lung function and disease processes (e.g. airway narrowing) fluctuate over the day, and this fluctuation impacts not only symptoms of asthma but also diagnosis (since patients are seen in the day when symptoms are typically reduced). Critically, we do not know why asthma is so rhythmic. Understanding what causes this rhythmicity may lead us to new treatment options and how to use existing drugs at the best time of the day.

My recent work, and that of my collaborators have shown that the daily variation in lung function in asthma is strongly influenced by innate timing mechanisms present in our bodies, known as the circadian clock. The circadian system allows us to track the external environment (e.g. light/dark, mealtimes) to ensure that biological processes in the body happen at the correct time of day. There is a master clock, centrally, within the brain, which dictates major rhythms in the body such as body temperature and sleep/wake cycles; however we now know that virtually every organ in the body (including the lung) has its own internal clock and that these peripheral clocks are essential to tissue function. My research has shown that the circadian clock in asthma may not be working as it should and this leads to more pronounced rhythmicity in asthma compared to healthy individuals. Furthermore, my research has found that night shift workers (who have an imbalance between their circadian clock and their environment) have an increased risk of having asthma and having more severe disease. This fellowship proposal will examine how clocks in the brain and lung contribute to the rhythmicity observed in asthma, and define processes within the disease which are under the control of these clocks to allow us to identify novel ways to treat the disease.

Firstly, I will determine how the peripheral clock in the lung controls airway narrowing, to do this I will genetically alter the clock in the lung and in the nerves controlling the airway and measure what happens to airway narrowing under normal conditions and under inflamed conditions (reminiscent of asthma). Next. I will find out if the lung clock itself is affected by allergic inflammation (reminiscent of asthma). I will also determine if it is possible to treat airway narrowing by using a drug that affects the clock. Then, I will determine how much influence the central clock in the brain has in controlling airway narrowing. Here, I will disrupt the synchronisation between the central clock and the peripheral lung clock by mimicking a night shift, or by changing eating patterns. This research will be carried out in mice, as this allows me to genetically manipulate the circadian clock in order to tease out a mechanism; this is not possible in humans. During my fellowship I will work with my collaborators to design a clinical circadian study to determine whether night shifts affect airway narrowing in individuals with asthma. The results will be helpful in working out how best to treat asthma in night shift workers in the future. I plan to apply for funding for this study during year 3-4 of my fellowship.

The proposed research will provide the mechanism underlying the rhythmic variation in airway narrowing in asthma. This will lead to better management and treatment of patients with asthma, by revealing new drug targets, and providing the timing information needed to use existing asthma treatments at the best time of the day. My research will also help in developing guidelines for managing asthma in night shift workers.

Asthma is highly rhythmic; asthma deaths occur at night; symptoms, airway narrowing and inflammation also peak overnight. Evidence suggests the circadian clock is important in mediating rhythmicity. Here, I will determine the mechanism controlling rhythmicity in asthma.

Aim 1. Using transgenic mice, I will target the molecular clock selectively in bronchiolar epithelial cells to define how local clock function in the lung impacts severity and rhythmicity in allergic airways inflammation. To determine the role of the autonomic nervous system , pharmacological and genetic interventions (removing clock function in cholinergic neurons) in vivo, will be combined with precision cut lung slice (PCLS) cultures. Assessment of disease will involve non-invasive and invasive determination of lung function, and monitoring of physiological rhythms (e.g. temperature, activity).

Aim 2. I will examine how clock function itself is impacted by lung inflammation, and identify pathways affected by time-of-day, disease state, and local loss of clock function. Using a Reverba reporter mouse, I will track Reverba expression in bronchiolar epithelial cells during airway inflammation both in vivo and in PCLS. I will analyse RNAseq profiles of laser captured microdissected airway smooth muscle cells from targeted mice at 2 times of day under normal/ disease state.

Aim 3. To understand the role of the central clock in the control of lung function I will manipulate the environment to decouple the central clock from the peripheral lung clock by employing a restricted feeding schedule or an acute shift in lighting schedule. This will allow me to determine the effect of misalignment on airway function (akin to the effects of night shift work).

This research provides an opportunity to discover novel therapeutic targets, use existing drugs at more efficacious times of day and inform guidelines to manage asthma in night shift workers.