[Effects of airway epithelium injury on the transdifferentiation of sub-epithelial fibroblasts and its role in the development of airway hyperresponsiveness in asthma].
OBJECTIVE
To address the possible role of injured airway epithelium in initiating transdifferentiation of sub-epithelial fibroblasts into myofibroblasts and accelerating cell proliferation in sub-epithelial fibroblasts, which may be involved in airway hyperresponsiveness in asthma.


METHODS
Human primary cultured sub-epithelial fibroblasts were co-cultured with human bronchial epithelial cells (16HBE) which were treated with lipopolysaccharide (LPS) plus mechanical scratch prior to co-culture. The procedure was also performed in the presence or absence of endothelin (ET) receptor A inhibitor (BQ123), transforming growth factor-beta(1) (TGF-beta(1)) neutralized antibody, respectively or simultaneously, followed by immunostaining, Western blotting and bromodeoxyuridine (BrdU) incorporation respectively to detect alpha-SMA expression and cell proliferation in the co-cultured sub-epithelial fibroblasts. Using the inhibitors specific for mitogen-activated protein kinases (MAPKs) pathways, the role of MAPKs pathways in activating the expression of alpha-SMA was evaluated. In addition, the interaction between matrix metalloproteinases (MMPs) and ET-1 was investigated by cell transfection with anti-ET-1 converting enzyme (anti-ECE) mRNA expression plasmid followed by gelatin zymography analysis.


RESULTS
16HBE treated with LPS plus mechanical injury induced alpha-SMA expression in sub-epithelial fibroblasts and accelerated BrdU incorporation in the cells. BQ123, TGF-beta(1) neutralized antibody, specific inhibitors for p38 MAPK and extracellular signal-regulated kinase 1/2 (ERK1/2) were able to block the induction respectively to a certain extent. Phosphorylated p38 MAPK and ERK1/2 were detected in the sub-epithelial fibroblasts 10 min after being co-cultured with injured 16HBE. Compared to normal control (16HBE transfected with pEGFPN(2)) or those cells transfected with anti-ECE mRNA expression plasmids, ET-1 released from the 16HBE cells transfected with pEGFPN(2) into supernatants were increased significantly after the treatment described as above: 16HBE pre-transfected with pEGFP-N(2) expression plasmid before being treated with mechanical scrape plus LPS stimulation: (15.00 +/- 0.86) pg/ml; 16HBE pre-transfected with anti-ECE expression plasmid before being treated with mechanical scrape plus LPS stimulation: (7.57 +/- 0.94) pg/ml (all P < 0.01). At the same time, the activities of MMP-2 and MMP-9 were enhanced.


CONCLUSIONS
Injured airway epithelial cells induced the transdifferentiation of sub-epithelial fibroblasts into myofibroblasts, which may be mediated by ET-1 and TGF-beta(1) through MARKs pathways such as p38 MAPK and ERK 1/2.
Chen
Xing-wu
XW
Guangzhou Institute of Respiratory Diseases, First Affiliated Hospital, Guanzhou Medical College, Guangzhou 510120, China.
Xu
Jun
J
D000199:Actins / Q000378:metabolism; D001249:Asthma / Q000378:metabolism / Q000473:pathology*; D002454:Cell Differentiation; D002461:Cell Line, Transformed; D049109:Cell Proliferation; D054338:Cell Transdifferentiation; D018920:Coculture Techniques; D019332:Endothelin-1 / Q000378:metabolism; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology*; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology; D048052:Mitogen-Activated Protein Kinase 3 / Q000037:antagonists & inhibitors / Q000378:metabolism; D010456:Peptides, Cyclic / Q000494:pharmacology; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D020545:Respiratory Mucosa / Q000378:metabolism / Q000473:pathology*; D014162:Transfection; D053773:Transforming Growth Factor beta1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism
D000199:Actins; D019332:Endothelin-1; D008070:Lipopolysaccharides; D010456:Peptides, Cyclic; D047428:Protein Kinase Inhibitors; D053773:Transforming Growth Factor beta1; D048052:Mitogen-Activated Protein Kinase 3; D048051:p38 Mitogen-Activated Protein Kinases; C072247:cyclo(Trp-Asp-Pro-Val-Leu)