Background Airway remodeling may explain lung function decline among asthmatic children. Extracellular matrix (ECM) deposition by human lung fibroblasts (HLFs) is implicated in airway remodeling. Airway epithelial cell (AEC) signaling may regulate HLF ECM expression. Objectives Determine whether AECs from asthmatic children differentially regulate HLF expression of ECM constituents. Methods Primary AECs were obtained from well-characterized atopic-asthmatic (N=10) and healthy children (N=10) intubated under anesthesia for an elective surgical procedure. AECs were differentiated at an air-liquid interface (ALI) for 3 weeks, then co-cultured with HLFs from a healthy child for 96 hours. Collagen I (COL1A1), collagen III (COL3A1), hyaluronan synthase 2 (HAS2), and fibronectin (FNDC) expression by HLFs and prostaglandin E2 synthase (PGE2S) expression by AECs was assessed by RT-PCR. TGFb1&2 concentrations in media were measured by ELISA. Results COL1A1 and COL3A1 expression by HLFs co-cultured with asthmatic AECs was greater than HLFs co-cultured with healthy AECs (2.2 fold, p<0.02; 10.8 fold, p<0.02). HAS2 expression by HLFs co-cultured with asthmatic AECs was 2.5-fold higher than by HLFs co-cultured with healthy AECs (p<0.002). FNDC expression by HLFs co-cultured with asthmatic AECs was significantly greater than by HLFs alone. TGFb2 activity was elevated in asthmatic AEC-HLF co-cultures (p<0.05) while PGES2 was down regulated in AEC-HLF co-cultures (2.2 fold, p<0.006). Conclusions HLFs co-cultured with asthmatic AECs showed differential expression of ECM constituents COL1A1 & COL3A1, and HAS2 compared to HLFs co-cultured with healthy AECs. These findings support a role for altered ECM production in asthmatic airway remodeling, possibly regulated by unbalanced AEC signaling.

BackgroundAirway remodeling is a proposed mechanism that underlies the persistent loss of lung function associated with childhood asthma. Previous studies have demonstrated that human lung fibroblasts (HLFs) co-cultured with primary human bronchial epithelial cells (BECs) from asthmatic children exhibit greater expression of extracellular matrix (ECM) components compared to co-culture with BECs derived from healthy children. Myofibroblasts represent a population of differentiated fibroblasts that have greater synthetic activity. We hypothesized co-culture with asthmatic BECs would lead to greater fibroblast to myofibroblast transition (FMT) compared to co-culture with healthy BECs.MethodsBECs were obtained from well-characterized asthmatic and healthy children and were proliferated and differentiated at an air-liquid interface (ALI). BEC-ALI cultures were co-cultured with HLFs for 96 hours. RT-PCR was performed in HLFs for alpha smooth muscle actin (α-SMA) and flow cytometry was used to assay for α-SMA antibody labeling of HLFs. RT-PCR was also preformed for the expression of tropomyosin-I as an additional marker of myofibroblast phenotype. In separate experiments, we investigated the role of TGFβ2 in BEC-HLF co-cultures using monoclonal antibody inhibition.ResultsExpression of α-SMA by HLFs alone was greater than by HLFs co-cultured with healthy BECs, but not different than α-SMA expression by HLFs co-cultured with asthmatic BECs. Flow cytometry also revealed significantly less α-SMA expression by healthy co-co-cultures compared to asthmatic co-cultures or HLF alone. Monoclonal antibody inhibition of TGFβ2 led to similar expression of α-SMA between healthy and asthmatic BEC-HLF co-cultures. Expression of topomyosin-I was also significantly increased in HLF co-cultured with asthmatic BECs compared to healthy BEC-HLF co-cultures or HLF cultured alone.ConclusionThese findings suggest dysregulation of FMT in HLF co-cultured with asthmatic as compared to healthy BECs. Our results suggest TGFβ2 may be involved in the differential regulation of FMT by asthmatic BECs. These findings further illustrate the importance of BEC-HLF cross-talk in asthmatic airway remodeling.