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B Activation in Monocyte Chemoattractant Protein-1 Expression by Mesangial Cells
*Hyonam Kidney Laboratory, Soon Chun Hyang University, Seoul, Korea; and University College London Medical School, The Rayne Institute, London, United Kingdom.
Correspondence to: Dr. Hi Bahl Lee, Hyonam Kidney Laboratory, Soon Chun Hyang University, 657 Hannam Dong, Yongsan Ku, Seoul 140-743, Korea. Phone: 82-2-709-9171, 82-2-792-6657; Fax: 82-2-792-5812; E-mail: hblee{at}seoul.com
ABSTRACT. Although high glucose (HG) has been shown to induce nuclear factor-
B (NF-B) activation in vascular cells, the upstream regulation and the biologic significance of NF-B activation in diabetic renal injury are not clear. It was, therefore, examined if HG-induced generation of reactive oxygen species (ROS) and protein kinase C (PKC) activation are involved in NF-B activation in mesangial cells (MC), and the role of NF-B activation in HG-induced monocyte chemoattractant protein-1 (MCP-1) expression by MC was further investigated. Recent observations suggest that MCP-1 may play a role in the development and progression of diabetic nephropathy. HG rapidly induced NF-B activation in MC as estimated by electrophoretic mobility shift assay. Supershift assay suggests that most of the binding activity arose from p50/p50 and p50/p65 dimers. Antioxidants, pyrrolidine dithiocarbamate, N-acetyl-L-cystein, and trolox effectively inhibited HG-induced NF-B activation in MC. HG rapidly generated dichlorofluorescin-sensitive intracellular ROS in MC as measured by laser-scanning confocal microscopy. HG also activated PKC rapidly in MC. Inhibition of PKC effectively blocked HG-induced intracellular ROS generation and NF-B activation in MC. HG increased MCP-1 mRNA expression by 1.9-fold and protein secretion by 1.6-fold that of control glucose in MC transfected with control vector but not in MC transfected with dominant negative mutant inhibitor of NF-B (IB
M). Inhibition of either PKC or ROS effectively blocked HG-induced, but not basal, MCP-1 protein secretion by MC transfected with control vector. Thus this study demonstrates that HG rapidly activates NF-B in MC through PKC and ROS and suggests that HG-induced NF-B activation in MC may play a role in diabetic renal injury through upregulation of MCP-1 mRNA and protein expression.
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