Induction of interleukin-1 and interleukin-8 mRNAs and proteins by TGF beta 1 in rat lung alveolar epithelial cells.
The transforming growth factor-beta (TGF-beta) has been shown to increase in lung injury and in fibrotic states of the lung. In the current study, we sought to investigate whether TGF beta 1 induced the expression of IL-1 alpha and IL-8 in rat alveolar epithelial cells. We evaluated TGF beta 1, IL-1 alpha, and IL-8 expression by immunofluorescence in silica-injured and saline-treated control rat lungs. Antibodies to IL-1 alpha, IL-8, and TGF beta 1 showed intense staining in silica-injured lungs as compared to saline-instilled lungs. Primary isolated type II cells from silica-injured lungs showed increased expression of IL-1 alpha as compared to saline-instilled lungs. To evaluate the effects of TGF beta 1, we treated an immortalized rat type II cell-derived cell line (LM5) with 100 pg/ml of TGF beta 1 in serum-free medium for 0-24 hours and analyzed the expression of IL-1 alpha and IL-8 mRNAs and proteins using semiquantitative RT-PCR, Northern blot analysis, Western blot analysis, and immunohistochemistry. Densitometric analysis of Northern blots showed modest constitutive expression of IL-1 alpha gene in untreated control LM5 cells. TGF beta 1 treatment resulted in an increase in IL-1 alpha mRNA, that reached maximum levels (4-fold) by 2 hours and remained elevated for 4-16 hours, with a subsequent decline by 24 hours. Similarly, Northern blot and RT-PCR analysis demonstrated that TGF beta 1 treatment resulted in maximum induction of IL-8 mRNA (6- 8.5-fold) within 1-4 hours. The levels remained elevated for up to 24 hours afterwards. Western blot analysis results further confirmed the expression of both IL-1 alpha and IL-8 proteins by LM5 cells. TGF beta 1 treatment resulted in increased expression of both IL-1 alpha and IL-8 proteins. Immunofluorescence studies demonstrated increased staining of IL-1 alpha by TGF beta 1 as compared to untreated cells. These results suggest that TGF beta 1 may regulate IL-1 alpha and IL-8 expression in alveolar epithelial cells and contribute to polymorphonuclear leukocyte recruitment and lung injury in clinical states with increased TGF beta 1.
Kumar
N M
NM
Department of Internal Medicine, SUNY at Buffalo School of Medicine, Buffalo General Hospital 14203, USA.
Rabadi
N H
NH
Sigurdson
L S
LS
Schünemann
H J
HJ
Lwebuga-Mukasa
J S
JS
D000818:Animals; D015152:Blotting, Northern; D005455:Fluorescent Antibody Technique; D005786:Gene Expression Regulation; D007375:Interleukin-1 / Q000235:genetics / Q000378:metabolism*; D016209:Interleukin-8 / Q000235:genetics / Q000378:metabolism*; D008168:Lung / Q000187:drug effects / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011650:Pulmonary Alveoli / Q000187:drug effects* / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012822:Silicon Dioxide / Q000494:pharmacology; D014158:Transcription, Genetic; D016212:Transforming Growth Factor beta / Q000378:metabolism / Q000494:pharmacology*
D007375:Interleukin-1; D016209:Interleukin-8; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D012822:Silicon Dioxide