NAD(P)H oxidase participates in the palmitate‐induced superoxide production and insulin secretion by rat pancreatic islets

Abstract

Nicotinamide adenine dinucleotide phosphate [NAD(P)H] oxidase complex has been shown to be involved in the process of glucose-stimulated insulin secretion (GSIS). In this study, we examined the effect of palmitic acid on superoxide production and insulin secretion by rat pancreatic islets and the mechanism involved. Rat pancreatic islets were incubated during 1 h with 1 mM palmitate, 1% fatty acid free-albumin, 5.6 or 10 mM glucose and in the presence of inhibitors of NAD(P)H oxidase (DPI--diphenyleneiodonium), PKC (calphostin C) and carnitine palmitoyl transferase-I (CPT-I) (etomoxir). Superoxide content was determined by hydroethidine assays. Palmitate increased superoxide production in the presence of 5.6 and 10 mM glucose. This effect was dependent on activation of PKC and NAD(P)H oxidase. Palmitic acid oxidation was demonstrated to contribute for the fatty acid induction of superoxide production in the presence of 5.6 mM glucose. In fact, palmitate caused p47(PHOX) translocation to plasma membrane, as shown by immunohistochemistry. Exposure to palmitate for 1 h up-regulated the protein content of p47(PHOX) and the mRNA levels of p22(PHOX), gp91(PHOX), p47(PHOX), proinsulin and the G protein-coupled receptor 40 (GPR40). Fatty acid stimulation of insulin secretion in the presence of high glucose concentration was reduced by inhibition of NAD(P)H oxidase activity. In conclusion, NAD(P)H oxidase is an important source of superoxide in pancreatic islets and the activity of NAD(P)H oxidase is involved in the control of insulin secretion by palmitate.

Augmented Rac1 Expression and Activity are Associated with Oxidative Stress and Decline of β Cell Function in Obesity

“…The Nox holoenzyme consists of membrane and cytosolic components. The membrane-associated catalytic core consists of gp91phox and p22phox, and the cytosolic regulatory core includes p47phox, p67phox, p40phox, and Rac1 [20]. Under stimulatory conditions, the cytosolic components translocate to the membrane fraction for association with the catalytic core for Nox2 holoenzyme assembly and catalytic activation.…”

Biphasic Roles of a Small G-Protein, RAC1 in Pancreatic Β-Cell

“…Unlike first category of small G-proteins, requisite for posttranslational modifications and mechanism[s] involved in the activation of Rab GTPases under the physiological insulin secretagogues remains elusive. However, Rap1 is activated transiently by glucose 50 and undergoes carboxymethylation. 18, 51 The third group of small G-proteins consists of Rab2, Rhes and Rem2 which are under-studied, [52][53][54][55] whereas, RalA appears to draw direct regulatory effects in exocytosis.…”

“…Immunological localization and functional regulation of Nox have been described in clonal β-cells, rat and human islets. 50,75,90,92 Studies by Shen and associates in cardiac myocytes have also suggested regulatory roles for Rac1 in the activation of Nox and associated generation of ROS in animal models of diabetes. 93 In addition, significant increase in Nox-mediated oxidative stress and subsequent metabolic dysfunction has been clearly reviewed in a recent article by Kowluru.…”

“…Along these lines, Pi and Collins have overviewed the existing evidence in supporting "secondary messenger" roles of ROS in physiological insulin secretion. 50 In addition, studies have also emphasized roles for Nox in physiological insulin secretion. For example, Diphenyleneiodonium [DPI], a selective inhibitor of Nox, inhibited glucose-induced Nox activity and GSIS.…”

“…90 Graciano and co-workers demonstrated regulatory roles for Nox in palmitate-induced superoxide generation and insulin secretion in rat islets. 50 Furthermore, recent findings suggests that prenylation and activation of Rac1 are critical for glucose-and mitochondrial fuel-induced Noxdependent ROS generation in clonal pancreatic β-cells and rodent islets. 91 In summary, a tonic increase in intracellular ROS is necessary for normal physiological insulin secretion and Rac1 initiates subsequent signaling steps including Nox activation and insulin release.…”