IL-1β INDUCES SERINE PROTEASE INHIBITOR 3 (SPI-3) GENE EXPRESSION IN RAT PANCREATIC β-CELLS. DETECTION BY DIFFERENTIAL DISPLAY OF MESSENGER RNA

Abstract

Immune-mediated β-cell damage induces diverse intracellular signals, leading to transcription of different genes which may either contribute to β-cell repair and/or defence or lead to cell death. The cytokine interleukin-1β (IL-1) is a potential mediator of β-cell dysfunction and damage in type 1 diabetes mellitus. To understand the molecular actions of this cytokine upon β-cells, this study aimed at the cloning of genes induced in FACS-purified rat pancreatic β-cells by a 6- or 24-h exposure to IL-1 by using differential display of mRNA with reverse transcription-polymerase chain reaction (DDRT-PCR). Among these cytokine-induced genes, a gene encoding for rat serine protease inhibitor (SPI-3) was isolated. SPI-3 may be involved in cellular defence responses against inflammatory stress. RT-PCR analysis confirmed that SPI-3 mRNA expression in rat β-cells is increased by IL-1 at an early stage (2h), with maximal accumulation during 6–12h and decline after 24h. Similar observations were made in mouse pancreatic islets and in the rat insulinoma cell line RINm5F. IFN-γ neither increased SPI-3 gene expression nor potentiated its induction by IL-1 in rat β-cells. The stimulatory effects of IL-1 on SPI-3 mRNA expression were decreased by co-incubation with an inhibitor of gene transcription (actinomycin D), an inhibitor of protein synthesis (cycloheximide) or an inhibitor of NF-κB activation (PDTC). On the other hand, a blocker of inducible nitric oxide synthase (iNOS) activity (NG-methyl-L-arginine) did not prevent IL-1-induced SPI-3 expression. Thus, SPI-3 mRNA expression following IL-1 exposure depends on gene transcription, protein synthesis and activation of the nuclear transcription factor NF-κB, but it is independent of NO formation.