ErbB2 activity is required for airway epithelial repair following neutrophil elastase exposure.
In cystic fibrosis and chronic bronchitis, airways are chronically injured by exposure to neutrophil elastase (NE). We sought to identify factors required for epithelial repair following NE exposure. Normal human bronchial epithelial cells were treated with NE (50 nM, 22 h) or control vehicle. Following NE treatment, we found a marked and sustained decrease in epithelial proliferation as detected by Ki67 immunostaining. 3H-thymidine incorporation was also initially depressed but increased over 72 h in NE-treated cells, which suggests that DNA synthesis constitutes an early repair process following NE exposure. We hypothesized that ErbB2 receptor tyrosine kinase, a regulator of cancer cell proliferation, was required for epithelial DNA synthesis following NE exposure. Immediately following NE treatment, by flow cytometry analysis, we found a decrease in ErbB2 surface expression. Protein levels of the full-length 185 kD ErbB2 receptor significantly decreased following NE treatment and smaller ErbB2-positive bands, ranging in size from 23 to 40 kD, appeared, which suggests that NE caused ErbB2 degradation. By real-time RT-PCR analysis, we found no change in ErbB2 mRNA expression following NE treatment, which suggests that changes in ErbB2 protein levels were regulated at the post-translational level. Following NE treatment, full-length 185 kD ErbB2 levels increased to pretreatment levels, correlating with the increase in thymidine incorporation during the same time period. Importantly, inhibition of ErbB2 activity with AG825 (5 microM) or Herceptin (3.1 microM), an ErbB2-neutralizing antibody, blocked thymidine incorporation only in NE-treated cells. These results suggest ErbB2 is a critical factor for epithelial recovery following NE exposure.
Fischer
Bernard M
BM
Department of Pediatrics, Duke University Medical Center, Durham, North Carolina 27710, USA.
Cuellar
Jacob G
JG
Byrd
Angela S
AS
Rice
Annette B
AB
Bonner
James C
JC
Martin
Linda D
LD
Voynow
Judith A
JA
D052160:Benzothiazoles; D001980:Bronchi / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D004247:DNA / Q000096:biosynthesis; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects; D066246:ErbB Receptors / Q000378:metabolism; D006801:Humans; D019272:Leukocyte Elastase / Q000633:toxicity*; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000502:physiology*; D020032:Tyrphostins / Q000494:pharmacology
D052160:Benzothiazoles; D004247:DNA; D066246:ErbB Receptors; D019272:Leukocyte Elastase; D018719:Erb-b2 Receptor Tyrosine Kinases; D020032:Tyrphostins; C098214:tyrphostin AG825