E1A has no effect on LPS-induced IL-6 secretion in rat alveolar epithelial cells.

BACKGROUND

The adenoviral protein E1A has been suggested to play a role in the pathophysiological development of chronic obstructive pulmonary disease (COPD) by inducing expression of inflammatory factors. It is well known that glucocorticoids are important inhibitors of inflammation. In the treatment of COPD corticosteroid therapy commonly has little or no anti-inflammatory effect. We hypothesized that the anti-inflammatory effect of glucocorticoids may be decreased or abolished by E1A expression, which aggravates the airway inflammation and results in the late stage of 'corticosteroid resistance' in COPD development. Corticosteroid therapy is a widely used method to treat respiratory inflammation. However, loss of anti-inflammatory effect is a common and serious complication during the therapy for COPD.

OBJECTIVES

To test whether the E1A gene, which is 1 of the viral genes responsible for enhanced inflammatory responses, is involved in corticosteroid resistance during clinical therapy for COPD.

METHODS

Rat alveolar epithelial cells were transfected with an E1A gene vector. Stably transfected cells were screened by PCR amplification of the E1A gene fragment. Transcription of the E1A gene was assayed by QT-RT-PCR. Protein expression was measured by immunohistochemistry and Western blot. E1A induced interleukin (IL)-6 secretion. mRNA levels were tested by ELISA and QT-RT-PCR. E1A-positive cells and E1A-negative cells were treated with lipopolysaccharide (LPS), physiological concentrations of hydrocortisone and trichostatin A (TSA), an inhibitor of histone deacetylase (HDAC). The activity of HDAC was confirmed by colorimetric HDAC activity assay kit.

RESULTS

In all 4 E1A-expressing transfectants, expression of the E1A 243R protein was much higher than that of the 289R protein. LPS significantly increased IL-6 secretion in both E1A-positive and E1A-negative cells and no significant differences were found. Treatment with hydrocortisone at a physiological concentration clearly decreased LPS-induced IL-6 secretion, and this down-regulation effect could be abolished in the presence of TSA. LPS significantly decreased HDAC activity, which was completely abolished by application of hydrocortisone. Hydrocortisone alone had no effect on HDAC activity in cells without LPS stimulation. There were no differences in HDAC activity or IL-6 secretion under these treatments between E1A-positive and E1A-negative cells.

CONCLUSIONS

Our results indicated that a physiological concentration of glucocorticoid suppresses LPS-mediated IL-6 secretion, through the participation of HDAC. Expression of adenoviral E1A had no effect on HDAC activity and hydrocortisones induced an anti-inflammatory effect.