CCAAT/enhancer-binding protein-delta is induced in mesangial area during the early stages of anti-Thy1.1 glomerulonephritis and regulates cell proliferation and inflammatory gene expression in cultured rat mesangial cells.
BACKGROUND
Interleukin (IL)-6, cyclooxygenase (COX)-2, and monocyte chemoattractant protein (MCP)-1 contribute to renal injury. The promoter regions of these genes contain CCAAT/enhancer-binding protein (C/EBP)-binding sites. In this study, we investigated the role of C/EBP-delta in mesangial cells (MCs).


METHODS
In an in vivo study, anti-Thy 1.1 glomerulonephritis rats were generated and C/EBP-delta, IL-6, COX-2, and MCP-1 expressions were assessed by immunohistochemistry. In an in vitro study, cultured MCs were transfected with non-silencing (NS) short interfering RNA (siRNA) or C/EBP-delta siRNA. Subsequently, after stimulation with IL-1beta, C/EBP-delta, IL-6, COX-2, and MCP-1 mRNA expression levels were evaluated using real-time polymerase chain reaction (PCR). IL-6 concentration in the culture medium was determined by enzyme-linked immunosorbent assay. In addition, cell proliferative activity against IL-1beta or platelet-derived growth factor-BB was assessed by bromodeoxyuridine incorporation.


RESULTS
In the in vivo study, C/EBP-delta, IL-6, COX-2, and MCP-1 were expressed in the mesangial region of anti-Thy 1.1 glomerulonephritis rats on day 1. In the in vitro study, IL-1beta increased C/EBP-delta mRNA levels in NS siRNA-transfected MCs (7.3-fold), but no increase was evident in C/EBP-delta siRNA-transfected MCs. IL-6, COX-2, and MCP-1 mRNA levels in C/EBP-delta siRNA-transfected MCs were all lower than those in NS siRNA-transfected MCs (decreases of 57.7%, 85.7%, and 69.3%, respectively). The IL-6 concentration in the culture medium from C/EBP-delta siRNA transfected MCs (7.37 +/- 4.3 pg/ml) was also lower than that in the culture medium from NS siRNA-transfected MCs (25.2 +/- 3.4 pg/ml). Cell proliferative activity in C/EBP-delta siRNA-transfected MCs was lower than that in NS siRNA transfected MCs.


CONCLUSIONS
C/EBP-delta was induced in the mesangial region during the early stages of anti-Thy1.1 glomerulonephritis. C/EBP-delta contributes to inflammatory gene expression and MC proliferation.
Miyoshi
Ken-ichi
K
Department of Integrated Medicine and Informatics, Ehime University Graduate School of Medicine, Shitsukawa, Toon, Ehime 791-0295, Japan.
Okura
Takafumi
T
Fukuoka
Tomikazu
T
Higaki
Jitsuo
J
D000818:Animals; D000077214:Becaplermin; D051743:CCAAT-Enhancer-Binding Protein-delta / Q000378:metabolism*; D049109:Cell Proliferation; D002478:Cells, Cultured; D018932:Chemokine CCL2 / Q000378:metabolism*; D051546:Cyclooxygenase 2 / Q000378:metabolism*; D015870:Gene Expression; D005921:Glomerulonephritis / Q000276:immunology / Q000378:metabolism*; D053583:Interleukin-1beta / Q000494:pharmacology; D015850:Interleukin-6 / Q000378:metabolism*; D007518:Isoantibodies; D050527:Mesangial Cells / Q000378:metabolism*; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D020574:Proto-Oncogene Proteins c-sis; D051381:Rats; D017207:Rats, Sprague-Dawley
C491766:Ccl2 protein, rat; D018932:Chemokine CCL2; D053583:Interleukin-1beta; D015850:Interleukin-6; D007518:Isoantibodies; D010982:Platelet-Derived Growth Factor; D020574:Proto-Oncogene Proteins c-sis; C035678:anti-Thy antibody; D051743:CCAAT-Enhancer-Binding Protein-delta; D000077214:Becaplermin; D051546:Cyclooxygenase 2