Interleukin-10 activates the transcription factor C/EBP and the interleukin-6 gene promoter in human intestinal epithelial cells.
BACKGROUND
Interleukin (IL)-6 is produced by enterocytes in response to sepsis and after treatment with IL-1beta. The IL-6 promoter contains binding sites for multiple transcription factors, including nuclear factor-kappaB and C/EBP. The anti-inflammatory cytokine IL-10 downregulates nuclear factor-kappaB activity, but its effects on C/EBP activation and IL-6 production in the enterocyte are not known.


METHODS
Caco-2 cells were treated with IL-1beta, IL-10, or a combination of the cytokines. C/EBP DNA binding activity was determined by electrophoretic mobility shift assay and IL-6 levels by enzyme-linked immunosorbent assay. IL-6 promoter activation was assessed by luciferase assay.


RESULTS
IL-10 treatment of cultured Caco-2 cells resulted in increased C/EBP DNA binding activity. Supershift analysis revealed upregulated DNA binding activity of C/EBP-beta but not C/EBP-delta. To examine if the increased DNA binding reflected gene activation, cells were transfected with a wild-type IL-6 promoter luciferase construct or with a mutated C/EBP binding site. IL-10 potentiated IL-1 beta-induced IL-6 promoter activity. Replacing the wild-type promoter with the promoter containing a mutated C/EBP DNA binding sequence blocked the effect of IL-10. When cells were treated with 0.5 ng/mL of IL-1 beta for 24 hours, IL-6 production increased, and this response to IL-1 beta was potentiated several-fold by IL-10.


CONCLUSIONS
IL-10 may activate the IL-6 gene in stimulated enterocytes by upregulating the expression and activity of C/EBP.
Robb
Bruce W
BW
Department of Surgery, University of Cincinnati, Ohio, USA.
Hershko
Dan D
DD
Paxton
James H
JH
Luo
Guang-Ju
GJ
Hasselgren
Per-Olof
PO
D022762:CCAAT-Enhancer-Binding Proteins / Q000378:metabolism*; D018938:Caco-2 Cells; D020895:Enterocytes / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects / Q000276:immunology; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology; D016753:Interleukin-10 / Q000494:pharmacology*; D015847:Interleukin-4 / Q000494:pharmacology; D015850:Interleukin-6 / Q000235:genetics*; D007413:Intestinal Mucosa / Q000166:cytology / Q000276:immunology / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000502:physiology*; D015533:Transcriptional Activation
D022762:CCAAT-Enhancer-Binding Proteins; D007375:Interleukin-1; D015850:Interleukin-6; D016753:Interleukin-10; D015847:Interleukin-4