Molecular Regulation of Monocyte Chemoattractant Protein-1 Expression in Pancreatic β-Cells

Abstract

Pancreatic β-cells are selectively destroyed during the course of type 1 diabetes. In the early stages of the disease, inflammatory infiltrates of mononuclear cells, containing predominantly monocytes and T-cells, are present in the islets (insulitis). Chemokines, such as monocyte chemoattractant protein-1 (MCP-1), play a key role in the recruitment and activation of these immunocytes. We have previously described cytokine-induced MCP-1 gene expression in human and rat pancreatic islets. In the present study, the transcriptional regulation by cytokines of the rat MCP-1 gene in fluorescence-activated cell sorting-purified rat β-cells, insulin-producing INS-1E cells, and RINm5F cells was investigated. Transient transfections with luciferase-reporter constructs identified an interleukin (IL)-1β-responsive enhancer region between -2,180 bp and −2,478 bp. Mutation of either of the two nuclear factor (NF)-κB sites present in this region abrogated IL-1β-induced MCP-1 promoter activity. Binding of NF-κB to the two sites was shown in vitro by gel shift assays, while supershift assays revealed the presence of p65/p50 heterodimers and p65 homodimers. In vivo binding of NF-κB was confirmed by chromatin immunoprecipitation assay. Blocking of NF-κB activation in cytokine-exposed primary β-cells by an adenovirus overexpressing a nondegradable form of IκBα or by pyrrolidine dithiocarbamate decreased IL-1β-induced MCP-1 mRNA expression. We conclude that NF-κB plays an important role for MCP-1 expression in β-cells. This transcription factor may be an interesting target for ex vivo gene therapy before islet transplantation.