Cell Biology & Molecular Medicine

Role

The Group consists of cell and molecular biologists who investigate six different respiratory diseases:

1. Asthma

2. Chronic Obstructive Pulmonary Disease

3. Idiopathic fibrosis

4. Bronchiolitis obliterans

5. Asbestos induced mesothelioma

6. Lymphangioleiomyomatosis (LAM)

In all studies we try to characterize the malfunction of cellular signalling pathways and the corresponding impact on cell proliferation and inflammation.

Research

Cell Biology

Respiratory virus studies
Brian Oliver has extended his studies on viral infection and asthma, looking at macrophages taken from patients with asthma before and after steroid treatment to see if steroids modified the levels of cytokines released from these cells in response to viral infection. Steroids in fact decreased levels of TNF-α, released from macrophages after exposure to virus, which provides evidence for their potential efficacy in viral induced infectious exacerbations. The effect of virus infection on extracellular matrix production
will be studied in 2007-9 with the aid of a recently awarded NHMRC grant.

Signal Transduction in airway smooth muscle cells
Lyn Moir is supervising a project on PI3Kinase, a step in the pathways leading to proliferation, growth factor and cytokine production in airway smooth muscle. We need to know why airway smooth muscle from people with asthma grows more rapidly and why we see excess deposition of extracellular matrix proteins. Evidence is accumulating that this pathway may be upregulated in airway diseases such as asthma and may constitute a potential target for therapeutic intervention to prevent or reverse airway smooth muscle cell proliferation and remodelling.

Contribution of the epithelial cell to airway remodelling
Qi Ge has found that profibrotic cytokines such as transforming growth factor beta, levels of which are increased in asthmatic airways, can promote or inhibit release of interleukins 6 and 8 respectively. She is exploring the way in which these can contribute to airway wall remodelling.

Angiogenesis in lung diseases
Increased new blood vessel formation is a common occurrence in lung diseases and Janette Burgess with PhD student Sarah Boustany is examining the way in which components of the extracellular matrix may contribute to angiogenesis and what determines the balance between pro and anti angiogenic factors in the airways.

Differential gene expression in airway smooth muscle in asthma
Justine Lau, PhD student in a CRC supported project is looking
for genes upregulated in airway smooth muscle. These genes
will be potential targets for new therapeutic interventions for airway remodelling.

COPD
We are continuing our research collaboration with AltanaAG (Germany) to evaluate the efficacy of a phosphodiesterase inhibitor and its active metabolite in preventing changes associated with airway wall remodelling in COPD.

Airway smooth muscle contraction in asthma
Dr Hisako Matsumoto from Kyoto University spent 18 months in the group working with Brent McParland to compare contractility of airway smooth muscle cells from asthmatic and non-asthmatic volunteers using cells obtained from biopsies embedded in collagen gels. She found that cells from asthmatic biopsies contracted more than those from non-asthmatic biopsies.

LAM
Lymphangioleiomyomatosis (LAM) is rare and affects only young women but it is potentially fatal and so far there is no treatment apart from lung transplantation. Working with Drs Deborah Yates and Allan Glanville at St Vincent’s Hospital and with the help of an anonymous philanthropic organization we are conducting clinical and laboratory studies on patients with LAM in the hope of halting or reversing the progress of the disease.

Molecular Medicine

CCAAT/Enhancer Binding Proteins (C/EBPs) control cells differentiation
Previously the lack of C/EBP-alpha in bronchial smooth muscle cells (BSMC) of asthmatics correlated with increased proliferation. We assessed disease-specific expression of C/EBP-alpha-beta-delta and -epsilon and the effects of glucocorticoids and long-acting beta2-agonists by determining the expression and function of C/EBP-alpha-beta-delta and -epsilon in BSMC of controls, asthma and COPD patients. We found that control cells expressed C/EBP-alpha-beta-delta-and -epsilon, which were up-regulated by serum.

Budesonide inhibited C/EBP-alpha and -beta expression; formoterol increased C/EBP-alpha (2-fold). C/EBP-delta and -epsilon were not affected by the drugs. sthma BSMC inappropriately expressed C/EBP-alpha, and C/EBP-beta -delta and -epsilon were up-regulated by serum. Budesonide and formoterol increased C/EBP-beta (3.4-fold & 2.5-fold), leaving C/EBP-alpha -delta and -epsilon unaffected. COPD cells expressed C/EBP-alpha -beta and -epsilon, which were up-regulated by serum, while C/EBP-delta was down-regulated. In 7/10 cell lines.

Budesonide inhibited C/EBP-alpha and -beta, up-regulated C/EBP-delta (3.2-fold). Formoterol up-regulated only C/EBP-alpha (3-fold). Protein analysis and EMSA confirmed the disease specific expression pattern of C/EBP-alpha in asthma and C/EBP-delta in COPD patients. The expression of C/EBPs in BSMC of asthma and COPD patients is disease-specific and is regulated in a drug- and disease-specific pattern.

Asthma and COPD are characterized by increased extracellular matrix (ECM) deposition
We investigated the effect of corticosteroids and LABA, alone or in combination, on total ECM and collagen deposition, gene expression, cell proliferation, and IL-6, IL-8 and TGF-beta1 levels by primary human lung fibroblasts. In our model fibroblasts in 0.3% albumin represented a non-inflammatory condition and stimulation with 5% FCS and/or TGF-beta1 mimicked an inflammatory environment with activation of tissue repair. 5% FCS increased total ECM, collagen deposition, cell proliferation, and IL-6, IL-8 and TGF-beta1 levels.

In 0.3% albumin corticosteroids reduced total ECM and collagen deposition, involving the glucocorticoid receptor (GR) and down-regulation of collagen, Hsp47 and Fli1 mRNA expression. In 5% FCS corticosteroids increased ECM deposition, involving up-regulation of COL4A1 and CTGF mRNA expression. LABA reduced total ECM and collagen deposition under all conditions partly via the beta2-adrenergic receptor. In combination, the drugs had an additive effect in the presence or absence of TGF-beta1 further decreasing ECM deposition in 0.3% albumin whereas counteracting each other in 5% FCS.

These data suggest that the effect of corticosteroids, but not of LABA, on ECM deposition by fibroblasts is altered by serum. These findings imply that as soon as airway inflammation is resolved, long-term treatment with combined drugs may beneficially reduce pathological tissue remodelling.

Mesothelioma
Pleural malignant mesothelioma is a locally aggressive tumour of epithelial cell origin. PDGF-BB induced, while TGF-beta1 inhibited cell proliferation. PDGF-BB was a chemoattractant and its effect was increased in the presence of TGF-beta1 through a collagen type-I gel in mesothelial cells only. TGF-beta1 but not PDGF-BB stimulated the de novo synthesis of pro-MMP-2 and low levels of TIMP-2 in both cell types. Only mesothelial cells expressed MT1-MMP. Pro-MMP-2 synthesis involved p38 MAP kinase.

Migration of mesothelioma cells was induced in the presence of MT1-MMP. The MMP inhibition by oleoyl-N-hydroxylamide and TIMP-2 inhibited their migration, but not proliferation. Our data suggest that proliferation of mesothelioma cells is not linked to MMP-2, MT1-MMP and low TIMP-2 expression but is involved in cell migration. However, pro-MMP-2 activation and migration of mesothelioma cells needs the presence of mesothelial cells.