E1A expression dysregulates IL-8 production and suppresses IL-6 production by lung epithelial cells.
BACKGROUND
The adenoviral protein E1A has been proposed to play a role in the pathophysiology of COPD, in particular by increasing IL-8 gene transcription of lung epithelial cells in response to cigarette smoke-constituents such as LPS. As IL-8 production is also under tight post-transcriptional control, we planned to study whether E1A affected IL-8 production post-transcriptionally. The production of IL-6 by E1A-positive cells had not been addressed and was studied in parallel. Based on our previous work into the regulation of IL-8 and IL-6 production in airway epithelial cells, we used the lung epithelial-like cell line NCI-H292 to generate stable transfectants expressing either E1A and/or E1B, which is known to frequently co-integrate with E1A. We analyzed IL-8 and IL-6 production and the underlying regulatory processes in response to LPS and TNF-alpha.


METHODS
Stable transfectants were generated and characterized with immunohistochemistry, western blot and flow cytometry. IL-8 and IL-6 protein production was measured by ELISA. Levels of IL-8 and IL-6 mRNA were measured using specific radiolabeled probes. EMSA was used to assess transcriptional activation of relevant transcription factors. Post-transcriptional regulation of mRNA half-life was measured by Actinomycin D chase experiments.


RESULTS
Most of the sixteen E1A-expressing transfectants showed suppression of IL-6 production, indicative of biologically active E1A. Significant but no uniform effects on IL-8 production, nor on transcriptional and post-transcriptional regulation of IL-8 production, were observed in the panel of E1A-expressing transfectants. E1B expression exerted similar effects as E1A on IL-8 production.


CONCLUSIONS
Our results indicate that integration of adenoviral DNA and expression of E1A and E1B can either increase or decrease IL-8 production. Furthermore, we conclude that expression of E1A suppresses IL-6 production. These findings question the unique role of E1A protein in the pathophysiology of COPD, but do not exclude a role for adenoviral E1A/E1B DNA in modulating inflammatory responses nor in the pathogenesis of COPD.
van den Berg
Arjen
A
Department of Pulmonology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands. a.vandenberg@amc.uva.nl
Snoek
Mieke
M
Jansen
Henk M
HM
Lutter
René
R
D017500:Adenovirus E1A Proteins / Q000235:genetics / Q000378:metabolism*; D017501:Adenovirus E1B Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D003001:Cloning, Molecular; D004847:Epithelial Cells / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000502:physiology; D006801:Humans; D015850:Interleukin-6 / Q000378:metabolism*; D016209:Interleukin-8 / Q000378:metabolism*; D008168:Lung / Q000166:cytology / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D020545:Respiratory Mucosa / Q000166:cytology / Q000378:metabolism*
D017500:Adenovirus E1A Proteins; D017501:Adenovirus E1B Proteins; D015850:Interleukin-6; D016209:Interleukin-8; D011994:Recombinant Proteins