In our system, metalloproteases seem not to play a role in GC metaplasia because responses were not inhibited by the metalloprotease inhibitor GM6001. cells is usually associated with HA depolymerization and EGF processing by TK followed by EGFR signaling, suggesting that increases in TK activity could contribute to GC metaplasia and mucus hypersecretion in diseases such as asthma and chronic bronchitis. The data also suggest that increases in GC populace could be sustained by the associated upregulation of Bcl-2 in airway epithelial cells. studies. The antiapoptotic factor Bcl-2 appears to be crucial to sustain the increase BI-78D3 in GC numbers (20), and its expression is usually upregulated in GC in the airways of patients with asthma (21) or cystic fibrosis compared with healthy individuals (20). Bcl-2 is also increased in lipopolysaccharide-induced GC metaplasia in rat airways (22). Until recently, mucin production was considered Rabbit Polyclonal to RFX2 the only function of GC necessary to provide a defense shield between the epithelium and the environment. BI-78D3 However, GCs have been shown to synthesize additional products important for host defense, such as lactoperoxidase (LPO) (23C25) in human and sheep and surfactant protein D (26) in rats. Epidermal growth factor receptor (EGFR) activation (27) with subsequent signaling via EGFR tyrosine kinase and mitogen-activated protein kinase (MAPK) seems to be a convergent pathway involved in GC hyperplasia and increased MUC5AC gene expression (12, 15) induced by oxidative stress and T helper-2 cytokines. EGFR activation can be achieved by multiple EGFR ligands: EGF, transforming growth factor (TGF)-, heparin-binding EGF-like growth factor (HB-EGF), amphiregulin, epiregulin, and betacellulin (28C31). These ligands are synthesized as transmembrane precursors and are cleaved BI-78D3 to their active form by matrix metalloproteases of the ADAM family (32C35), with the exception of EGF, which is usually cleaved by the serine protease tissue kallikrein (TK) in kidney (36), in salivary and mammary glands (37, 38), and in airways submucosal gland cells (39). TK is present in the airways (40, 41), BI-78D3 and its activity is usually elevated in the bronchoalveolar lavage of patients with asthma (42, 43) and in patients with chronic bronchitis (44, 45); exposure to air pollutants can also induce TK upregulation (46). Studies from this laboratory have shown that this glycosaminoglycan hyaluronan (HA) immobilizes TK at BI-78D3 the apical pole of airway epithelium and inhibits its catalytic activity (41, 47); consequently, HA degradation results in a dramatic increase in TK activity (41). Because HA is usually depolymerized by reactive oxygen species (ROS) (48), increased airway TK activity is usually expected in diseases associated with increased airway ROS production. TK activity is usually increased in patients with chronic bronchitis and other diseases associated with oxidative stress and mucus hypersecretion when compared with normal subjects (42, 44) possibly due to continued degradation of HA by ROS (49C51). We have also shown that ROS-induced EGFR signaling and MUC5AC hypersecretion is usually associated with HA depolymerization followed by pro-EGF processing by TK in primary cultures of human submucosal gland (SMG) cells (39). The goal of the present study was to test whether ROSCinduced, EGFR-dependent GC metaplasia is initiated by HA depolymerization in polarized human airway epithelial cells. To test this hypothesis, we uncovered normal human bronchial epithelial (NHBE) cells produced and differentiated at the airCliquid interface (ALI) to oxidative stress and assessed HA depolymerization, TK activation, pro-EGF processing, EGFR and 44/42MAPK phosphorylation, gene and protein expression of specific goblet cell products (MUC5AC, LPO and Bcl-2), and goblet cell.