Asthma
Dr Stuart J Hirst, Dr Vladimir A Snetkov & Prof Tak H Lee
King’s College London
2005 – 2008 Research Grant
Pathobiology of airway smooth muscle from asthmatics.
Dr Stuart Hirst is a Reader in Cell Pharmacology in the Department of Asthma, Allergy and Respiratory Science, King’s College London.
Dr Vladimir A Snetkov is a Senior Research Fellow in the Division of Asthma Allergy and Respiratory Science, King’s College London.
Prof Tak H Lee is Head of the Division of Asthma, Allergy and Respiratory Science, King’s College London.
Asthma rates are soaring worldwide, especially in children, although the reasons for this remain unclear. Asthma affects an estimated eight million people in the UK – one in 13 adults and one in eight children.
Animals are widely used in asthma research, but important species differences between the lungs and airways of humans, rats, mice and other animals, mean that there are marked variations between findings in animal experiments and humans with asthma. Moreover, none of the animals commonly used in asthma research (guineapigs, rabbits, rats and mice) naturally exhibit an asthma-like syndrome.
Pathologically, asthma is characterised by excessive airways narrowing due to bronchospasm, hyper-responsiveness, inflammation. In more chronic cases, alterations in normal airway dimensions (remodelling) involve marked increases in the amount of smooth muscle surrounding the airways.
This project will study the most important aspect of the remodeling process – alterations in the function of airway smooth muscle (ASM). Confounding findings from animal ‘models’ combined with variable reports from post-mortem human lung tissue have fuelled controversy in this area. However, new information is coming to light in studies of ASM tissue obtained by fibreoptic endobronchoscopy in volunteers with and without asthma.
Human ASM cells cultured from bronchial biopsies from asthmatic patients show significantly higher basal levels of free cytosolic calcium, indicating a change in calcium homeostatic mechanisms in asthma. Hyperreactivity of ASM to inflammatory mediators in asthma may be due to increased levels of free cytosolic calcium in asthmatic cells.
The goal of this Dr Hadwen Trust-funded research project is to elucidate the specific features of calcium handling in ASM cells that underlie airway hyper-responsiveness in asthmatic patients; and to establish the systematic use of these cells as a research tool to supersede animal and lung resection work, in the field of airway remodelling.
Asthma and the role of calcium
A work in progress report by Dr Katharina Mahn
Asthma is a chronic disease of the lung. Common complications in asthma include wheezing, coughing and chest tightness. During an asthma attack, the bronchial tubes narrow excessively in response to irritants such as allergens, cold air or occupational triggers. Airway narrowing arises mainly from contraction of the muscle cells that surround the bronchial tubes [1] and the contraction relies heavily on increases in calcium inside these cells [2] .
It has been suggested that the control of calcium is somehow impaired in asthma and that this contributes to the airway narrowing that gives rise to symptoms. However, due to limited availability of lung tissue from patients with asthma there is insufficient information either to support or disregard this possibility. A large proportion of asthma research involves animal models, including mice sensitised to allergen and/or having single genes deleted or overexpressed. Mice, however, do not develop asthma and so far have been of limited benefit in understanding the critical mechanisms behind this uniquely human disease [3] .
We have developed techniques which allow muscle samples to be obtained using fibreoptic bronchoscopy from the airways of volunteers with and without mild or moderate asthma. The muscle cells can then be grown in cell culture and studied in the laboratory over periods of many weeks, so that differences can be compared in muscle from patients with varying degrees of asthma severity and from healthy volunteers. Funding from the Dr Hadwen Trust has greatly enhanced our ability to recruit and screen sufficient numbers of suitable patients and volunteers for this research.
Many of our studies involve measuring levels of calcium inside muscle cells using a calcium-sensing fluorescent dye, recorded with a microscope provided by funds from the Dr Hadwen Trust. By studying RNA from these cells we can also detect if there are differences in the genes that control the production of specific proteins involved in the release and storage of calcium that can be linked to muscle contraction in asthma.
We have discovered that the control or handling of calcium is different inside the muscle cells from asthmatics as compared to healthy individuals. Calcium inside the cell is regulated by proteins in the membrane, which transport calcium in and out of the cell, and by a key storage organelle inside the cell – the sarcoplasmic reticulum. The sarcoplasmic reticulum acts as an internal reservoir for calcium and can be emptied, leading to an increase in available calcium inside the cell and initiation of a contraction. Furthermore, this reservoir can ‘mop up’ free calcium allowing relaxation to occur and the opening of the airways in asthma to help relieve symptoms. One of the molecules important for the emptying and refilling function of this calcium store is called SERCA or the sarco(endo)plasmic reticular calcium ATPase. SERCA sequesters available calcium by pumping it into the sarcoplasmic reticulum reservoir.
We have shown that the muscle cells from patients with asthma have less SERCA than cells from healthy controls. The reduction is by almost 50% in muscle cells from moderate asthmatics. These important findings suggest that in asthma once a contraction has begun it takes longer for the calcium to be sequestered, possibly strengthening and thereby drawing out each contraction of the muscle, keeping the airways closed for longer.
We have now started to modify these muscle cells by changing the expression of key genes involved in the storage of calcium. We are performing gene intervention studies to reduce the SERCA content of healthy cells to induce an asthma phenotype and conversely increasing SERCA in muscle from asthmatics. Our goal is to show that muscle cells from asthmatics can once again function normally.
Our results may provide an explanation for why the airways narrow excessively in asthma, and allow development of new forms of treatment, as well as reducing the reliance on animal models of asthma.
Dr Katharina Mahn, who wrote this report, is a postdoctoral fellow in Dr Hirst’s research group at King’s College London.
References
1. Hirst S (2000). Airway smooth muscle as a target in asthma. Clin Exp Allergy 30:54-59.
PubMed Link
2. Sanders KM (2001). Mechanisms of calcium handling in smooth muscles. J Appl Physiol 91:1438-1449.
PubMed Link
3. Wenzel SE & Holgate ST (2006). The mouse trap: It still yields few answers in asthma. Am J Respir Crit Care Med 174:1173-1176.