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SP600125, an inhibitor of c-Jun NH2-terminal kinase, blocks expression of angiotensin II-induced monocyte chemoattractant protein-1 in human mesangial cells

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Abstract

Background

We investigated the role of c-Jun NH2-terminal kinase (JNK), a member of the mitogen-activated protein kinase family, in the expression of angiotensin II (Ang II)-induced monocyte chemoattractant protein-1 (MCP-1) and transforming growth factor-1 (TGF-1), and in the production of fi bronectin (FN), by human mesangial cells (HMCs).

Methods

JNK activation in cultured human mesangial cells was determined by Western blotting with an antibody against the phosphorylated Ser63 residue of c-Jun. Binding of the activator protein (AP-1) to the MCP-1 AP-1 motif was detected via the electrophoretic mobility shift assay (EMSA). The transient luciferase reporter was used to examine MCP-1 promoter activity; an RNase protection assay and ELISA were used respectively to detect the expression of MCP-1 mRNA and production of MCP-1, TGF-β and FN.

Results

Anthra (1,9-cd) pyrazol-6(2H)-one (SP600125), a pharmacological inhibitor of JNK, almost completely abolished Ang II-induced Ser63 phosphorylation of c-Jun at concentrations of 5–20 μmol/L: JNK activity was reduced by 75% with 10 μmol/L SP600125, and by 90% with 20 μmol/L. Ang II increased AP-1 binding to the MCP-1 AP-1 motif in a time-dependent manner, as detected by EMSA, while SP600125 effectively blocked this increased AP-1 binding in a concentration-dependent manner. Treatment with 100 nmol/L Ang II led to a steady increase in MCP-1 mRNA expression, and to an enhanced production of MCP-1, TGF-β and FN. These effects were blocked by SP60025 in a dose-dependent manner. SP600125 also reduced MCP-1 mRNA stability: the halflife of MCP-1 mRNA was approximately 5 hours in cells treated with Ang II only, but was reduced to 2 hours when treated with a combination of Ang II and SP600125.

Conclusions

These results show that the JNK/AP-1 pathway is involved in the expression of MCP-1 and TGF-β, and in extracellular matrix production. JNK is an important therapeutic target for glomerulonephritis and glomerulosclerosis.

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Authors and Affiliations

  1. Department of Nephrology, Nanjing Children’s Hospital, Affiliated to Nanjing Medical University, Nanjing, China

    Gui-Xia Ding, Ai-Hua Zhang & Song-Ming Huang

  2. Institute of Pediatrics, Nanjing Medical University, Nanjing, China

    Gui-Xia Ding, Ai-Hua Zhang, Song-Ming Huang, Xiao-Qin Pan & Rong-Hua Chen

  3. Department of Nephrology, Nanjing Children’s Hospital, Affiliated to Nanjing Medical University, Nanjing, 210008, Jiangsu, China

    Ai-Hua Zhang

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  1. Gui-Xia Ding
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Correspondence to Ai-Hua Zhang.

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Ding, GX., Zhang, AH., Huang, SM. et al. SP600125, an inhibitor of c-Jun NH2-terminal kinase, blocks expression of angiotensin II-induced monocyte chemoattractant protein-1 in human mesangial cells. World J Pediatr 6, 169–176 (2010). https://doi.org/10.1007/s12519-010-0033-2

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  • DOI: https://doi.org/10.1007/s12519-010-0033-2

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