MECHANISMS OF GOBLET CELL HYPERPLASIA INDUCED BY SIMULATED VIRAL EXPOSURE OR TH2 CYTOKINES.
Introduction: Goblet cell hyperplasia is a common feature of respiratory epithelial response to numerous environmental challenges (e.g. cigarette smoke, air pollution, viral infection) and is also commonly observed in respiratory diseases, such as chronic obstructive pulmonary disease (COPD) and asthma (1-4). In the current work we sought to reproduce goblet cell hyperplasia in an organotypic in vitro model of the human airway epithelium, and to gain mechanistic insights by determining epithelial gene expression changes associated with goblet cell induction. Methods: The EpiAirway-FT™ in vitro human airway model was exposed for up to six days to Poly(I:C) to simulate viral exposure, or to the Th2 cytokine IL-13 to simulate Th2 imbalance. Induction of goblet cell hyperplasia was determined by histological evaluation of H&E stained paraffin sections and by mucin staining with alcian blue (AB)/periodic acid Schiff (PAS). Gene expression changes associated with the goblet cell hyperplasia were determined by qPCR utilizing a custom qPCR array containing 45 mucin related genes. IL-8 production by the EpiAirway-FT tissues was monitored by ELISA. The effect of a prototypic in vivo inhibitor of goblet cell hyperplasia (azithromycin, Az) on in vitro goblet cell hyperplasia and gene expression was also evaluated. Results: Treatment with the Th2 cytokine IL-13 caused a dramatic induction of goblet cell hyperplasia after 6 days of exposure. Quantitative PCR experiments identified several genes that were associated with Th2 cytokine-induced goblet cell hyperplasia, including MUC5AC, ALOX15, CLCA1, SPDEF and TFF3. Az at 30 mM did not diminish the histological appearance of IL-13-induced goblet cells or AB/PAS staining. However, higher doses of Az were more effective for reduction of goblet cell metaplasia and PAS staining. Az also significantly reduced IL-13- induced expression of CLCA1, MUC5AC and TFF3 mRNA. Treatment of EpiAirway-FT™ with Poly(I:C) induced modest goblet cell hyperplasia. Numerous genes including MUC5AC, MUC5B, STAT1 and TFF3 were upregulated more than 3-fold compared to untreated controls. Exposure to Poly(I:C), but not IL-13, produced a 5-fold increase in IL-8 secretion into the culture medium. Az caused a dose-dependent decrease in Poly(I:C) induced goblet cell number and size, and also diminished Poly(I:C) induced changes in mucin gene expression. Conclusions: Taken together, the results show that the Th2 cytokine IL-13 and Poly(I:C) are each capable of inducing goblet cell hyperplasia in organotypic in vitro human airway epithelial models. However, the mechanisms of induction appear to involve substantially different molecular pathways.
15-Lox, Alcian blue (AB), ALOX15, Asthma, Chronic obstructive pulmonary disease (COPD), CLCA1, Goblet cell hyperplasia, MUC RNA stabilization, MUC transcription, MUC5AC, MUC5B, Mucin content, Mucus hyperproduction, PAS staining, Periodic acid Schiff (PAS), SPDEF, STAT1, TFF3, Th2 cytokine, Th2 imbalance, Viral exposure
Azithromycin, EpiAirway-FT, IL-13, Poly(I:C)
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