Faculty Collaboration Database

Evidence for involvement of the proteasome complex (26S) and NFkappaB in IL-1beta-induced nitric oxide and prostaglandin production by rat islets and RINm5F cells. Diabetes 1998 Apr;47(4):583-91

Date

05/06/1998

Pubmed ID

9568691

DOI

10.2337/diabetes.47.4.583

Scopus ID

2-s2.0-0031899972 (requires institutional sign-in at Scopus site)   76 Citations

Abstract

Interleukin-1beta (IL-1beta) has been implicated as an effector molecule of beta-cell destruction in autoimmune diabetes. IL-1beta inhibits insulin secretion from pancreatic beta-cells by stimulating the expression of inducible nitric oxide synthase (iNOS) that generates the free radical nitric oxide. IL-1beta 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-1beta-induced iNOS and COX-2 expression by rat islets and transformed rat pancreatic beta-cells. Because of the limitations of cell numbers of purified primary beta-cells obtained from rat islets, biochemical and molecular studies were performed using the rat insulinoma beta-cell line RINm5F. A serine protease inhibitor, Nalpha-P-tosyl-L-lysine chloromethyl ketone (TLCK), and a proteasome complex (26S) inhibitor, MG 132, inhibited IL-1beta-induced nitrite formation, an oxidation product of nitric oxide produced by iNOS, in a concentration-dependent manner, with complete inhibition observed at 100 micromol/l and 10 micromol/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 micromol/l) and MG 132 (10 micromol/l) inhibited IL-1beta-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 NFkappaB is essential for activation of a number of cytokine-inducible enzymes and was evaluated as a possible site of protease action necessary for IL-1beta-induced coexpression of iNOS and COX-2. TLCK and MG 132 inhibited both IL-1beta-induced activation of NFkappaB and degradation of IkappaBalpha by islets and RINm5F cells. These results implicate protease activation as an early signaling event in IL-1beta-induced inhibition of beta-cell function. This study also suggests that IL-1beta-induced iNOS and COX-2 coexpression by pancreatic beta-cells share a common signaling pathway in utilizing the proteasome complex (26S) and the transcription factor NFkappaB, and it identifies sites of intervention to prevent the overproduction of their inflammatory products.

Author List

Kwon G, Corbett JA, Hauser S, Hill JR, Turk J, McDaniel ML

Author

John A. Corbett PhD Chair, Professor in the Biochemistry department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

Animals
Cell Line, Transformed
Cyclooxygenase 2
Cysteine Proteinase Inhibitors
Dinoprostone
Enzyme Induction
Gene Expression Regulation, Enzymologic
Humans
In Vitro Techniques
Interleukin-1
Islets of Langerhans
Isoenzymes
Leupeptins
Male
Membrane Proteins
NF-kappa B
Nitric Oxide
Nitric Oxide Synthase
Nitric Oxide Synthase Type II
Peptide Hydrolases
Phosphorylation
Prostaglandin-Endoperoxide Synthases
Proteasome Endopeptidase Complex
RNA, Messenger
Rats
Rats, Sprague-Dawley
Signal Transduction
Tosyllysine Chloromethyl Ketone
Tumor Cells, Cultured