Limits...
Role of lipid peroxidation and PPAR-δ in amplifying glucose-stimulated insulin secretion.

Cohen G, Riahi Y, Shamni O, Guichardant M, Chatgilialoglu C, Ferreri C, Kaiser N, Sasson S - Diabetes (2011)

Bottom Line: The latter mimicked the GSIS-amplifying effect of high glucose preexposure and of the PPAR-δ agonist GW501516 in INS-1E cells and isolated rat islets.Cytotoxic effects of 4-HNE were observed only above the physiologically effective concentration range.This molecule is an endogenous ligand for PPAR-δ, which amplifies insulin secretion in β-cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, School of Pharmacy, Faculty of Medicine, Institute for Drug Research, Hebrew University, Jerusalem, Israel.

ABSTRACT

Objective: Previous studies show that polyunsaturated fatty acids (PUFAs) increase the insulin secretory capacity of pancreatic β-cells. We aimed at identifying PUFA-derived mediators and their cellular targets that are involved in the amplification of insulin release from β-cells preexposed to high glucose levels.

Research design and methods: The content of fatty acids in phospholipids of INS-1E β-cells was determined by lipidomics analysis. High-performance liquid chromatography was used to identify peroxidation products in β-cell cultures. Static and dynamic glucose-stimulated insulin secretion (GSIS) assays were performed on isolated rat islets and/or INS-1E cells. The function of peroxisome proliferator-activated receptor-δ (PPAR-δ) in regulating insulin secretion was investigated using pharmacological agents and gene expression manipulations.

Results: High glucose activated cPLA(2) and, subsequently, the hydrolysis of arachidonic and linoleic acid (AA and LA, respectively) from phospholipids in INS-1E cells. Glucose also increased the level of reactive oxygen species, which promoted the peroxidation of these PUFAs to generate 4-hydroxy-2E-nonenal (4-HNE). The latter mimicked the GSIS-amplifying effect of high glucose preexposure and of the PPAR-δ agonist GW501516 in INS-1E cells and isolated rat islets. These effects were blocked with GSK0660, a selective PPAR-δ antagonist, and the antioxidant N-acetylcysteine or by silencing PPAR-δ expression. High glucose, 4-HNE, and GW501516 also induced luciferase expression in a PPAR-δ-mediated transactivation assay. Cytotoxic effects of 4-HNE were observed only above the physiologically effective concentration range.

Conclusions: Elevated glucose levels augment the release of AA and LA from phospholipids and their peroxidation to 4-HNE in β-cells. This molecule is an endogenous ligand for PPAR-δ, which amplifies insulin secretion in β-cells.

Show MeSH

Related in: MedlinePlus

High 4-HNE concentrations compromise β-cell survival. A: Rat islets and INS-1E cells were incubated for 48 or 24 h, respectively, with increasing concentrations of 4-HNE. Islets were pooled from three animals and divided into 10–12 islets per group. After incubation, islet cells were dispersed by mild trypsin digestion. Cell viability was determined at the end of the incubation by the trypan blue exclusion test. Cell viability in the absence of 4-HNE was >95% in both preparations. Results are mean ± SEM, n = 3. *P < 0.05 for the difference from untreated cells. B: INS-1E cells were taken for the FLICA apoptosis assay, as described in research design and methods. Results are mean ± SEM of the relative apoptotic levels in three different slides. C: Western blot analysis of 4-HNE–protein adducts in lysates prepared from INS-1E cells incubated for 48 h at 11 mmol/L glucose and the indicated concentrations of 4-HNE.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3198069&req=5

Figure 7: High 4-HNE concentrations compromise β-cell survival. A: Rat islets and INS-1E cells were incubated for 48 or 24 h, respectively, with increasing concentrations of 4-HNE. Islets were pooled from three animals and divided into 10–12 islets per group. After incubation, islet cells were dispersed by mild trypsin digestion. Cell viability was determined at the end of the incubation by the trypan blue exclusion test. Cell viability in the absence of 4-HNE was >95% in both preparations. Results are mean ± SEM, n = 3. *P < 0.05 for the difference from untreated cells. B: INS-1E cells were taken for the FLICA apoptosis assay, as described in research design and methods. Results are mean ± SEM of the relative apoptotic levels in three different slides. C: Western blot analysis of 4-HNE–protein adducts in lysates prepared from INS-1E cells incubated for 48 h at 11 mmol/L glucose and the indicated concentrations of 4-HNE.

Mentions: The effect of exogenously added 4-HNE on GSIS was then investigated; isolated rat islets and INS-1E cells were maintained for 48 or 24 h, respectively, in standard RPMI-1640 medium (11 mmol/L glucose) without or with increasing 4-HNE concentrations, followed by a static GSIS assay (Fig. 3B). The addition of 4-HNE amplified GSIS in a concentration-dependent manner; maximal effects were observed with 25 and 1 μmol/L 4-HNE in rat islets and in INS-1E cells, respectively. Above these levels, GSIS was reduced, possibly as a result of cytotoxic effects of 4-HNE (see Fig. 7).


Role of lipid peroxidation and PPAR-δ in amplifying glucose-stimulated insulin secretion.

Cohen G, Riahi Y, Shamni O, Guichardant M, Chatgilialoglu C, Ferreri C, Kaiser N, Sasson S - Diabetes (2011)

High 4-HNE concentrations compromise β-cell survival. A: Rat islets and INS-1E cells were incubated for 48 or 24 h, respectively, with increasing concentrations of 4-HNE. Islets were pooled from three animals and divided into 10–12 islets per group. After incubation, islet cells were dispersed by mild trypsin digestion. Cell viability was determined at the end of the incubation by the trypan blue exclusion test. Cell viability in the absence of 4-HNE was >95% in both preparations. Results are mean ± SEM, n = 3. *P < 0.05 for the difference from untreated cells. B: INS-1E cells were taken for the FLICA apoptosis assay, as described in research design and methods. Results are mean ± SEM of the relative apoptotic levels in three different slides. C: Western blot analysis of 4-HNE–protein adducts in lysates prepared from INS-1E cells incubated for 48 h at 11 mmol/L glucose and the indicated concentrations of 4-HNE.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3198069&req=5

Figure 7: High 4-HNE concentrations compromise β-cell survival. A: Rat islets and INS-1E cells were incubated for 48 or 24 h, respectively, with increasing concentrations of 4-HNE. Islets were pooled from three animals and divided into 10–12 islets per group. After incubation, islet cells were dispersed by mild trypsin digestion. Cell viability was determined at the end of the incubation by the trypan blue exclusion test. Cell viability in the absence of 4-HNE was >95% in both preparations. Results are mean ± SEM, n = 3. *P < 0.05 for the difference from untreated cells. B: INS-1E cells were taken for the FLICA apoptosis assay, as described in research design and methods. Results are mean ± SEM of the relative apoptotic levels in three different slides. C: Western blot analysis of 4-HNE–protein adducts in lysates prepared from INS-1E cells incubated for 48 h at 11 mmol/L glucose and the indicated concentrations of 4-HNE.
Mentions: The effect of exogenously added 4-HNE on GSIS was then investigated; isolated rat islets and INS-1E cells were maintained for 48 or 24 h, respectively, in standard RPMI-1640 medium (11 mmol/L glucose) without or with increasing 4-HNE concentrations, followed by a static GSIS assay (Fig. 3B). The addition of 4-HNE amplified GSIS in a concentration-dependent manner; maximal effects were observed with 25 and 1 μmol/L 4-HNE in rat islets and in INS-1E cells, respectively. Above these levels, GSIS was reduced, possibly as a result of cytotoxic effects of 4-HNE (see Fig. 7).

Bottom Line: The latter mimicked the GSIS-amplifying effect of high glucose preexposure and of the PPAR-δ agonist GW501516 in INS-1E cells and isolated rat islets.Cytotoxic effects of 4-HNE were observed only above the physiologically effective concentration range.This molecule is an endogenous ligand for PPAR-δ, which amplifies insulin secretion in β-cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, School of Pharmacy, Faculty of Medicine, Institute for Drug Research, Hebrew University, Jerusalem, Israel.

ABSTRACT

Objective: Previous studies show that polyunsaturated fatty acids (PUFAs) increase the insulin secretory capacity of pancreatic β-cells. We aimed at identifying PUFA-derived mediators and their cellular targets that are involved in the amplification of insulin release from β-cells preexposed to high glucose levels.

Research design and methods: The content of fatty acids in phospholipids of INS-1E β-cells was determined by lipidomics analysis. High-performance liquid chromatography was used to identify peroxidation products in β-cell cultures. Static and dynamic glucose-stimulated insulin secretion (GSIS) assays were performed on isolated rat islets and/or INS-1E cells. The function of peroxisome proliferator-activated receptor-δ (PPAR-δ) in regulating insulin secretion was investigated using pharmacological agents and gene expression manipulations.

Results: High glucose activated cPLA(2) and, subsequently, the hydrolysis of arachidonic and linoleic acid (AA and LA, respectively) from phospholipids in INS-1E cells. Glucose also increased the level of reactive oxygen species, which promoted the peroxidation of these PUFAs to generate 4-hydroxy-2E-nonenal (4-HNE). The latter mimicked the GSIS-amplifying effect of high glucose preexposure and of the PPAR-δ agonist GW501516 in INS-1E cells and isolated rat islets. These effects were blocked with GSK0660, a selective PPAR-δ antagonist, and the antioxidant N-acetylcysteine or by silencing PPAR-δ expression. High glucose, 4-HNE, and GW501516 also induced luciferase expression in a PPAR-δ-mediated transactivation assay. Cytotoxic effects of 4-HNE were observed only above the physiologically effective concentration range.

Conclusions: Elevated glucose levels augment the release of AA and LA from phospholipids and their peroxidation to 4-HNE in β-cells. This molecule is an endogenous ligand for PPAR-δ, which amplifies insulin secretion in β-cells.

Show MeSH
Related in: MedlinePlus