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A2869 - Hyaluronan Production by Human Lung Fibroblasts Co-Cultured with Asthmatic Bronchial Epithelial Cells Is Enhanced by Epithelial Infection with Respiratory Syncytial Virus
Author Block: S. R. Reeves1, C. K. Chan2, K. A. Barrow1, I. Kang2, M. P. White1, T. N. Wight2, J. S. Debley1; 1Center for Immunity and Immunotherapies, Seattle Children's, Seattle, WA, United States, 2Matrix Biology Program, Benaroya Research Institute, Seattle, WA, United States.
Rationale: Viruses such as respiratory syncytial virus (RSV) significantly contribute to asthma exacerbations. Bronchial epithelial cells (BECs) represent the interface between the host and the environment and can modulate airway remodeling. Recent work from our group has shown that BECs obtained from asthmatic children drive human lung fibroblasts (HLFs) to upregulate several extracellular matrix (ECM) components such as hyaluronan (HA). The effect of RSV infection of BECs on HLF production of HA is unknown.
Aim: To determine if HLFs co-cultured with BECs derived from children with asthma produce an ECM that is more enriched with HA compared to co-culture with BECs derived from healthy children following RSV infection.
Methods: BECs were obtained via bronchial brushings from well-characterized asthmatic (n=6) and healthy children (n=6) ages 6-18 years. Cells were proliferated and differentiated at an air-liquid interface (ALI) for 21 days. HLFs from a healthy child were obtained from a commercial vendor and used for all experiments. HLFs were co-cultured with BECs ± RSV infection (MOI=1) for 96 h following which samples were harvested for analysis of HA content. Cultures were radiolabeled with tritiated acetate 24 h before harvesting. The hyaluronidase-sensitive material was assayed via slot blot.
Results: HLFs co-cultured with asthmatic BECs displayed greater total production of HA under control conditions compared to HLFs co-cultured with healthy BECs (0.073±0.008 vs. 0.043±0.008 dpm, p=0.02). HLFs co-cultures with RSV-infected BECs displayed increased total HA production that was greater in the asthmatic BEC co-culture group compared to co-culture with healthy BECs (0.26±0.05 vs. 0.14±0.03 dpm, p=0.04). Increased HA production in the RSV-infected healthy BEC/HLF co-cultures were the result of increases of HA secreted into the culture media (0.080±0.02 vs. 0.012±0.002 dpm, p=0.004). Increased HA production in the RSV-infected asthmatic BEC/HLF co-cultures compared to uninfected were the result of increases in HA content of both the cell layer (0.090±0.01 vs. 0.051±0.007 dpm, p=0.02) and culture media (0.16±0.04 vs. 0.021±0.005 dpm, p=0.02).
Conclusions: HLFs co-cultured with BECs derived from asthmatic children produce more HA than HLFs co-cultured with BECs derived from healthy children. RSV infection of BECs from both groups significantly increased HA production in our BEC/HLF co-culture model; however, this effect was significantly greater in asthmatic co-cultures. Enrichment of the ECM with HA and/or HA degradation products may contribute to the airway inflammation observed in asthma and may be enhanced by altered epithelial signaling in response to viral infections.