Evidence for involvement of the proteasome complex (26S) and NFκB in IL-1β-induced nitric oxide and prostaglandin production by rat islets and RINm5F cells

Guirn Kwon, John A. Corbett, Scott Hauser, Jeanette R. Hill, John Turk, Michael L. McDaniel

Research output: Contribution to journalArticle

71 Scopus citations

Abstract

Interleukin-lβ (IL-1β) has been implicated as an effector molecule of β-cell destruction in autoimmune diabetes. IL-1β inhibits insulin secretion from pancreatic β cells by stimulating the expression of inducible nitric oxide synthase (iNOS) that generates the free radical nitric oxide. IL-1β also induces the coexpression of the inducible isoform of cyclooxygenase (COX-2) that results in the overproduction of proinflammatory prostaglandins. The current studies were designed to characterize the involvement of protease(s) in the signaling pathway of IL-1β-induced iNOS and COX-2 expression by rat islets and transformed rat pancreatic β-cells. Because of the limitations of cell numbers of purified primary β-cells obtained from rat islets, biochemical and molecular studies were performed using the rat insulinoma β-cell line RINm5F. A serine protease inhibitor, Nα-P-tosyl-L- lysine chloromethyl ketone (TLCK), and a proteasome complex (26S) inhibitor, MG 132, inhibited IL-1β-induced nitrite formation, an oxidation product of nitric oxide produced by iNOS, in a concentration-dependent manner, with complete inhibition observed at 100 μmol/l and 10 μmol/l, respectively. Both TLCK and MG 132 also inhibited iNOS gene expression at the level of mRNA and protein. In an analogous manner, TLCK (100 μmol/l) and MG 132 (10 μmol/l) inhibited IL-1β-induced COX-2 enzyme activity (PGE2 formation) and COX-2 gene expression at the level of mRNA and protein. In human islets, the proteasome inhibitor MG 132 also inhibited the formation of the products of iNOS and COX-2 enzyme activity, nitrite, and PGE2, respectively. These findings suggest that the inhibitory action of TLCK and MG 132 on iNOS and COX-2 expression precedes transcription. The transcription factor NFκB is essential for activation of a number of cytokine-inducible enzymes and was evaluated as a possible sire of protease action necessary for IL-1β-induced coexpression of iNOS and COX-2. TLCK and MG 132 inhibited both IL-1β- induced activation of NFκB and degradation of IκBα by islets and RINm5F cells. These results implicate protease activation as an early signaling event in IL-1β-induced inhibition of β-cell function. This study also suggests that IL-1β-induced iNOS and COX-2 coexpression by pancreatic β- cells share a common signaling pathway in utilizing the proteasome complex (26S) and the transcription factor NFκB, and it identifies sites of intervention to prevent the overproduction of their inflammatory products.

Original languageEnglish
Pages (from-to)583-591
Number of pages9
JournalDiabetes
Volume47
Issue number4
DOIs
StatePublished - Apr 21 1998

Fingerprint Dive into the research topics of 'Evidence for involvement of the proteasome complex (26S) and NFκB in IL-1β-induced nitric oxide and prostaglandin production by rat islets and RINm5F cells'. Together they form a unique fingerprint.

  • Cite this