Use of a three dimensional cell culture model to study airway smooth muscle - mast cell interactions in asthma

Asthma is a disease that still causes a significant amount of morbidity and mortality in the UK. Airway smooth muscle hypertrophy and hyperplasia, with a corresponding infiltration by mast cells, are key features of the inflammatory process that results in airway remodelling and fixed airflow obstruction. A 3 dimensional collagen I model was developed to examine differences in airway smooth muscle cell morphology, phenotype and MMP 2 expression, when cultured in a 30 environment in comparison to a 20 monolayer. Using :: this technique, airway smooth muscle cells and HMC-1s, a mast cell line, ,; were co-cultured to assess the effects of cell to cell interaction on airway I smooth muscle proliferation rates and MMP2 production. A novel method for assessing HMC-1 migration towards airway smooth cells was also established. To further study mast cell migration in asthma, a new technique of patient recruitment and endobronchial biopsying of individuals had to be established. Airway smooth muscle cells when cultured in 30 were observed to adopt a spindle like morphology. They also appeared to express lower levels of a smooth muscle actin and vimentin. In the model, basal rates of airway smooth muscle cell proliferation, when examined using Ki67, are reduced by over 50% compared to the 20 controls (p<0.05). Rates of airway smooth muscle cell proliferation were significantly increased in the model when they were co-cultured with the HMC-1s, and activated HMC-1s; the rate doubling in the latter from sole culture alone (p<0.05). 6 --. ---------- - Culturing airway smooth muscle cells in 3D also resulted in increased and sustained MMP2 production with a further increase when co-cultured with HMC-1s. This also resulted in increased expression of the activated form. Co-culturing of the cells resulted in increased rates of gel contraction, by 22%. This contraction could be reduced by using a broad spectrum MMP inhibitor, 1I0mastat. HMC-1 migration towards airway smooth muscle is partially MMP dependent. The rate of HMC-1 s migrating when treated with 1I0mastat was reduced by 45% (p