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Arachidonic acid actions on functional integrity and attenuation of the negative effects of palmitic acid in a clonal pancreatic β-cell line.

Keane DC, Takahashi HK, Dhayal S, Morgan NG, Curi R, Newsholme P - Clin. Sci. (2011)

Bottom Line: Experiments using specific COX and LOX (lipoxygenase) inhibitors demonstrated the importance of COX-1 activity for acute (20 min) stimulation of insulin secretion, suggesting that AA metabolites may be responsible for the insulinotropic effects.AA decreased the protein expression of iNOS (inducible NO synthase), the p65 subunit of NF-κB (nuclear factor κB) and the p47 subunit of NADPH oxidase in PA-treated cells.These findings indicate that AA has an important regulatory and protective β-cell action, which may be beneficial to function and survival in the 'lipotoxic' environment commonly associated with Type 2 diabetes mellitus.

View Article: PubMed Central - PubMed

Affiliation: UCD School of Biomolecular and Biomedical Science, UCD Conway Institute and UCD Institute of Sport and Health, UCD Dublin, Belfield, Dublin 4, Ireland.

ABSTRACT
Chronic exposure of pancreatic β-cells to saturated non-esterified fatty acids can lead to inhibition of insulin secretion and apoptosis. Several previous studies have demonstrated that saturated fatty acids such as PA (palmitic acid) are detrimental to β-cell function compared with unsaturated fatty acids. In the present study, we describe the effect of the polyunsaturated AA (arachidonic acid) on the function of the clonal pancreatic β-cell line BRIN-BD11 and demonstrate AA-dependent attenuation of PA effects. When added to β-cell incubations at 100 μM, AA can stimulate cell proliferation and chronic (24 h) basal insulin secretion. Microarray analysis and/or real-time PCR indicated significant AA-dependent up-regulation of genes involved in proliferation and fatty acid metabolism [e.g. Angptl (angiopoietin-like protein 4), Ech1 (peroxisomal Δ3,5,Δ2,4-dienoyl-CoA isomerase), Cox-1 (cyclo-oxygenase-1) and Cox-2, P<0.05]. Experiments using specific COX and LOX (lipoxygenase) inhibitors demonstrated the importance of COX-1 activity for acute (20 min) stimulation of insulin secretion, suggesting that AA metabolites may be responsible for the insulinotropic effects. Moreover, concomitant incubation of AA with PA dose-dependently attenuated the detrimental effects of the saturated fatty acid, so reducing apoptosis and decreasing parameters of oxidative stress [ROS (reactive oxygen species) and NO levels] while improving the GSH/GSSG ratio. AA decreased the protein expression of iNOS (inducible NO synthase), the p65 subunit of NF-κB (nuclear factor κB) and the p47 subunit of NADPH oxidase in PA-treated cells. These findings indicate that AA has an important regulatory and protective β-cell action, which may be beneficial to function and survival in the 'lipotoxic' environment commonly associated with Type 2 diabetes mellitus.

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Related in: MedlinePlus

Effect of AA on relative cell numberBRIN-BD11 β-cells were incubated in the absence or presence of various concentrations of AA for 24 h (50–150 μM AA). Relative cell number was assessed using an assay based on the reduction of WST-1, a tetrazolium salt, to formazan by cellular mitochondrial dehydrogenase activity. An increase in the number of viable cells results in an increase in the overall mitochondrial dehydrogenase activity. Results are normalized to 100% and are expressed as means±S.E.M., for six independent experiments. *P<0.05 compared with WST-1 absorbance from control cells.
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Figure 1: Effect of AA on relative cell numberBRIN-BD11 β-cells were incubated in the absence or presence of various concentrations of AA for 24 h (50–150 μM AA). Relative cell number was assessed using an assay based on the reduction of WST-1, a tetrazolium salt, to formazan by cellular mitochondrial dehydrogenase activity. An increase in the number of viable cells results in an increase in the overall mitochondrial dehydrogenase activity. Results are normalized to 100% and are expressed as means±S.E.M., for six independent experiments. *P<0.05 compared with WST-1 absorbance from control cells.

Mentions: Incubation of BRIN-BD11 cells with AA (up to 150 μM) for 24 h did not impair cell viability when compared with control, as determined by WST-1 analysis or flow cytometry. WST-1 is widely used to determine mitochondrial dehydrogenase activity, making the assumption that viable cells contain fully functional mitochondria, and therefore WST-1 absorbance is proportional to cell number. The addition of 100 μM of AA, however, stimulated cell proliferation, increasing the cell population by 19% when compared with the control. Incubation of the BRIN-BD11 cells with a relatively high level of 150 μM AA resulted in a decrease in cell viability and therefore a reduction in relative cell number when compared with cells incubated with 100 μM, but not when compared with the vehicle control (Figure 1).


Arachidonic acid actions on functional integrity and attenuation of the negative effects of palmitic acid in a clonal pancreatic β-cell line.

Keane DC, Takahashi HK, Dhayal S, Morgan NG, Curi R, Newsholme P - Clin. Sci. (2011)

Effect of AA on relative cell numberBRIN-BD11 β-cells were incubated in the absence or presence of various concentrations of AA for 24 h (50–150 μM AA). Relative cell number was assessed using an assay based on the reduction of WST-1, a tetrazolium salt, to formazan by cellular mitochondrial dehydrogenase activity. An increase in the number of viable cells results in an increase in the overall mitochondrial dehydrogenase activity. Results are normalized to 100% and are expressed as means±S.E.M., for six independent experiments. *P<0.05 compared with WST-1 absorbance from control cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2990202&req=5

Figure 1: Effect of AA on relative cell numberBRIN-BD11 β-cells were incubated in the absence or presence of various concentrations of AA for 24 h (50–150 μM AA). Relative cell number was assessed using an assay based on the reduction of WST-1, a tetrazolium salt, to formazan by cellular mitochondrial dehydrogenase activity. An increase in the number of viable cells results in an increase in the overall mitochondrial dehydrogenase activity. Results are normalized to 100% and are expressed as means±S.E.M., for six independent experiments. *P<0.05 compared with WST-1 absorbance from control cells.
Mentions: Incubation of BRIN-BD11 cells with AA (up to 150 μM) for 24 h did not impair cell viability when compared with control, as determined by WST-1 analysis or flow cytometry. WST-1 is widely used to determine mitochondrial dehydrogenase activity, making the assumption that viable cells contain fully functional mitochondria, and therefore WST-1 absorbance is proportional to cell number. The addition of 100 μM of AA, however, stimulated cell proliferation, increasing the cell population by 19% when compared with the control. Incubation of the BRIN-BD11 cells with a relatively high level of 150 μM AA resulted in a decrease in cell viability and therefore a reduction in relative cell number when compared with cells incubated with 100 μM, but not when compared with the vehicle control (Figure 1).

Bottom Line: Experiments using specific COX and LOX (lipoxygenase) inhibitors demonstrated the importance of COX-1 activity for acute (20 min) stimulation of insulin secretion, suggesting that AA metabolites may be responsible for the insulinotropic effects.AA decreased the protein expression of iNOS (inducible NO synthase), the p65 subunit of NF-κB (nuclear factor κB) and the p47 subunit of NADPH oxidase in PA-treated cells.These findings indicate that AA has an important regulatory and protective β-cell action, which may be beneficial to function and survival in the 'lipotoxic' environment commonly associated with Type 2 diabetes mellitus.

View Article: PubMed Central - PubMed

Affiliation: UCD School of Biomolecular and Biomedical Science, UCD Conway Institute and UCD Institute of Sport and Health, UCD Dublin, Belfield, Dublin 4, Ireland.

ABSTRACT
Chronic exposure of pancreatic β-cells to saturated non-esterified fatty acids can lead to inhibition of insulin secretion and apoptosis. Several previous studies have demonstrated that saturated fatty acids such as PA (palmitic acid) are detrimental to β-cell function compared with unsaturated fatty acids. In the present study, we describe the effect of the polyunsaturated AA (arachidonic acid) on the function of the clonal pancreatic β-cell line BRIN-BD11 and demonstrate AA-dependent attenuation of PA effects. When added to β-cell incubations at 100 μM, AA can stimulate cell proliferation and chronic (24 h) basal insulin secretion. Microarray analysis and/or real-time PCR indicated significant AA-dependent up-regulation of genes involved in proliferation and fatty acid metabolism [e.g. Angptl (angiopoietin-like protein 4), Ech1 (peroxisomal Δ3,5,Δ2,4-dienoyl-CoA isomerase), Cox-1 (cyclo-oxygenase-1) and Cox-2, P<0.05]. Experiments using specific COX and LOX (lipoxygenase) inhibitors demonstrated the importance of COX-1 activity for acute (20 min) stimulation of insulin secretion, suggesting that AA metabolites may be responsible for the insulinotropic effects. Moreover, concomitant incubation of AA with PA dose-dependently attenuated the detrimental effects of the saturated fatty acid, so reducing apoptosis and decreasing parameters of oxidative stress [ROS (reactive oxygen species) and NO levels] while improving the GSH/GSSG ratio. AA decreased the protein expression of iNOS (inducible NO synthase), the p65 subunit of NF-κB (nuclear factor κB) and the p47 subunit of NADPH oxidase in PA-treated cells. These findings indicate that AA has an important regulatory and protective β-cell action, which may be beneficial to function and survival in the 'lipotoxic' environment commonly associated with Type 2 diabetes mellitus.

Show MeSH
Related in: MedlinePlus