Interleukin-1 reduces the glycolytic utilization of glucose by pancreatic islets and reduces glucokinase mRNA content and protein synthesis by a nitric oxide-dependent mechanism.

Michael Landt,Zhongmin Ma,John Turk,Sasanka Ramanadham,David M. Kipnis,Alan Bohrer

doi:10.1074/jbc.272.28.17827

Michael Landt, Zhongmin Ma + Show 4 more

Open Access

https://doi.org/10.1074/jbc.272.28.17827

Copy DOI

Abstract

Culture of rat pancreatic islets with interleukin-1 (IL-1) results in up-regulation of the inducible isoform of nitric oxide synthase and overproduction of nitric oxide (NO). This is associated with reversible inhibition of both glucose-induced insulin secretion and islet glucose oxidation, and these effects are prevented by the inducible nitric oxide synthase inhibitor NG-monomethylarginine. IL-1 also induces accumulation of nonesterified arachidonic acid in islets by an NO-dependent mechanism, and one potential explanation for that effect would involve an IL-1-induced enhancement of islet glycolytic flux. We have therefore examined effects of IL-1 on islet glycolytic utilization of glucose and find that culture of islets with IL-1 in medium containing 5.5 mM glucose results in suppression of islet glucose utilization subsequently measured at glucose concentrations between 6 and 18 mM. The IL-1-induced suppression of islet glucose utilization is associated with a decline in islet glucokinase mRNA content, as determined by competitive reverse transcriptase-polymerase chain reaction, and in glucokinase protein synthesis, as determined by immuoprecipitation experiments, and all of these effects are prevented by NG-monomethylarginine. These findings suggest that IL-1 can down-regulate islet glucokinase, which is the primary component of the islet glucose-sensor apparatus, by an NO-dependent mechanism. Because reductions in islet glucokinase levels are known to cause a form of type II diabetes mellitus, these observations raise the possibility that factors which increase islet NO levels might contribute to development of glucose intolerance.

Highlights

Culture of rat pancreatic islets with interleukin-1 (IL-1)1 induces islet expression of the inducible isoform of nitric oxide synthase and overproduction of nitric oxide (NO) [1,2,3,4,5,6]
1 The abbreviations used are: IL-1, interleukin-1; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; KRB, Krebs-Ringer bicarbonate buffer; MEM, modified Eagle’s medium; MODY, maturity-onset diabetes of the young; NMMA, NG-monomethylarginine; NO, nitric oxide; PAGE, polyacrylamide gel electrophoresis; PBS, phosphate-buffered saline; Polymerase chain reactions (PCR), polymerase chain reaction; RT, reverse transcriptase; bp, base pair(s). Of these effects are prevented by the inducible nitric oxide synthase inhibitor NG-monomethylarginine (NMMA) [1, 2, 4], indicating that they occur through NO-dependent mechanisms
The islets were placed in fresh medium and incubated with various concentrations of [5-3H]glucose. Glycolytic utilization of this substrate was determined by measurement of [3H]OH production, which is generated at the triose-phosphate isomerase and enolase reactions reactions (20 –28) and provides a quantitative measure of islet glycolytic flux [27]

Summary

EXPERIMENTAL PROCEDURES

Materials—Male Sprague-Dawley rats (180 –220 g body weight) were purchased from Sasco (O’Fallon, MO); collagenase from Boehringer Mannheim; tissue culture medium (CMRL-1066), penicillin, streptomycin, Hanks’ balanced salt solution, heat-inactivated fetal bovine serum, and L-glutamine from Life Technologies, Inc. (Grand Island, NY); Pentex bovine serum albumin (fatty acid free, fraction V) from Miles Laboratories (Elkhart, IN); Rodent Chow 5001 from Ralston Purina Materials—Male Sprague-Dawley rats (180 –220 g body weight) were purchased from Sasco (O’Fallon, MO); collagenase from Boehringer Mannheim; tissue culture medium (CMRL-1066), penicillin, streptomycin, Hanks’ balanced salt solution, heat-inactivated fetal bovine serum, and L-glutamine from Life Technologies, Inc. Media—Media included KRB (Krebs-Ringer bicarbonate buffer: 25 mM HEPES, pH 7.4, 115 mM NaCl, 24 mM NaHCO3, 5 mM KCl, 2.5 mM CaCl2, 1 mM MgCl2), nKRB (KRB supplemented with 3 mM D-glucose), cCMRL-1066 (CMRL-1066 supplemented with 10% heat-inactivated fetal bovine serum, 1% L-glutamine and 1% (w/v) each of penicillin and streptomycin), and Hank’s balanced salt solution supplemented with 0.5% penicillin-streptomycin. Incubation of Islets with IL-1 and Measurement of Insulin Secretion—Islets (400 per condition) were placed in Petri dishes (10 ϫ 35 mm); suspended in cCMRL medium (1 ml) containing no additives, IL-1 (5 units/ml) alone, or both IL-1 and NMMA (0.5 mM); and incubated (2– 48 h, 37 °C). Islets were removed from the incubation medium, and their secretion of insulin, oxidation of

NO Reduces Islet Glucokinase mRNA

RESULTS

DISCUSSION