The Free Fatty Acid Receptor GPR40 - expression and role in islet hormone secretion

Type 2 diabetes (T2D) is a serious condition of growing proportions. Developing via an increasing imbalance between insulin sensitivity in the peripheral tissues and insulin release from pancreatic beta-cells, it ultimately renders the individual incapable of regulating the blood glucose concentration, e.g. after a meal. The increased prevalence of T2D is associated with an increase in the prevalence of obesity, with obesity being the single largest risk factor for the development of T2D. This work describes a molecule in the border zone between T2D and obesity. Our initial characterization of GPR40 identified it as a receptor for medium- to long-chain free fatty acids (FFAs). With a marked expression in pancreatic beta-cell lines, we expected GPR40 to be involved in FFA-mediated augmentation of insulin release. This was confirmed when we examined the dose-response relationship between FFA stimulation of GPR40 and both intracellular second messengers in a beta-cell line and insulin release from isolated pancreatic islets. A similarly increased glucagon secretion from alpha-cells was demonstrated after we established that these cells also express GPR40. Antisense knock-down of GPR40 abolished the effect of FFA stimulation on hormone secretion from both cell types. In the final part of this work, FFAs that activate GPR40 were shown to negatively regulate its mRNA expression, indicating a mechanism of protection from detrimental effects of sustained GPR40 stimulation. FFAs mediate effects on both alpha- and beta-cells that are potentially harmful in the development of T2D and it is possible that at least part of those occur via GPR40

Palmitate-Induced β-Cell Dysfunction Is Associated with Excessive NO Production and Is Reversed by Thiazolidinedione-Mediated Inhibition of GPR40 Transduction Mechanisms

BACKGROUND: Type 2 diabetes often displays hyperlipidemia. We examined palmitate effects on pancreatic islet function in relation to FFA receptor GPR40, NO generation, insulin release, and the PPARgamma agonistic thiazolidinedione, rosiglitazone. PRINCIPAL FINDINGS: Rosiglitazone suppressed acute palmitate-stimulated GPR40-transduced PI hydrolysis in HEK293 cells and insulin release from MIN6c cells and mouse islets. Culturing islets 24 h with palmitate at 5 mmol/l glucose induced beta-cell iNOS expression as revealed by confocal microscopy and increased the activities of ncNOS and iNOS associated with suppression of glucose-stimulated insulin response. Rosiglitazone reversed these effects. The expression of iNOS after high-glucose culturing was unaffected by rosiglitazone. Downregulation of GPR40 by antisense treatment abrogated GPR40 expression and suppressed palmitate-induced iNOS activity and insulin release. CONCLUSION: We conclude that, in addition to mediating acute FFA-stimulated insulin release, GPR40 is an important regulator of iNOS expression and dysfunctional insulin release during long-term exposure to FFA. The adverse effects of palmitate were counteracted by rosiglitazone at GPR40, suggesting that thiazolidinediones are beneficial for beta-cell function in hyperlipidemic type 2 diabetes

GPR40 is expressed in glucagon producing cells and affects glucagon secretion.

The free fatty acid receptor, GPR40, has been coupled with insulin secretion via its expression in pancreatic beta-cells. However, the role of GPR40 in the release of glucagon has not been studied and previous attempts to identify the receptor in alpha-cells have been unfruitful. Using double-staining for glucagon and GPR40 expression, we demonstrate that the two are expressed in the same cells in the periphery of mouse islets. In-R1-G9 hamster glucagonoma cells respond dose-dependently to linoleic acid stimulation by elevated phosphatidyl inositol hydrolysis and glucagon release and the cells become increasingly responsive to fatty acid stimulation when overexpressing GPR40. Isolated mouse islets also secrete glucagon in response to linoleic acid, a response that was abolished by antisense treatment against GPR40. This study demonstrates that GPR40 is present and active in pancreatic alpha-cells and puts further emphasis on the importance of this nutrient sensing receptor in islet function. (c) 2006 Elsevier Inc. All rights reserved

A human cell surface receptor activated by free fatty acids and thiazolidinedione drugs.

Fatty acids, which are essential nutritional components, are also involved in cardiovascular and metabolic diseases. Here we report a human cell surface receptor that we name free fatty acid receptor (FFAR), because it is specifically activated by medium to long-chain free fatty acids. The receptor belongs to the class of seven-transmembrane, G-protein coupled receptors (GPCRs) and also mediates responses to antidiabetic drugs of the thiazolidinedione type. It is expressed in skeletal muscle, heart, liver, and pancreatic β-cells. Stimulation of FFAR increases the intracellular calcium concentration in cells expressing the receptor in a native (pancreatic β-cell line) or in a recombinant form. In view of the nature of the activating substances, their physiological role in the body, and the tissue distribution of FFAR we suggest the term “nutrient sensing receptor” for receptors acting at the interface between dietary components and signalling molecules

Correction: Palmitate-Induced β-Cell Dysfunction Is Associated with Excessive NO Production and Is Reversed by Thiazolidinedione-Mediated Inhibition of GPR40 Transduction Mechanisms.

https://researchrepository.wvu.edu/wvu_herbwvfern/2768/thumbnail.jp

Free fatty acid receptor 1 (FFA(1)R/GPR40) and its involvement in fatty-acid-stimulated insulin secretion.

Free fatty acids (FFA) have generally been proposed to regulate pancreatic insulin release by an intracellular mechanism involving inhibition of CPT-1. The recently de-orphanized G-protein coupled receptor, FFA(1)R/GPR40, has been shown to be essential for fatty-acid-stimulated insulin release in MIN6 mouse insulinoma cells. The CPT-1 inhibitor, 2-bromo palmitate (2BrP), was investigated for its ability to interact with mouse FFA(1)R/GPR40. It was found to inhibit phosphatidyl inositol hydrolysis induced by linoleic acid (LA) (100 mu M in all experiments) in HEK293 cells transfected with FFA(1)R/GPR40 and in the MIN6 subclone, MIN6c4. 2BrP also inhibited LA-stimulated insulin release from mouse pancreatic islets. Mouse islets were subjected to antisense intervention by treatment with a FFA(1)R/GPR40-specific morpholino oligonucleotide for 48 h. Antisense treatment of islets suppressed LA-stimulated insulin release by 50% and by almost 100% when islets were pretreated with LA for 30 min before applying the antisense. Antisense treatment had no effect on tolbutamide-stimulated insulin release. Confocal microscopy using an FFA(1)R/GPR40-specific antibody revealed receptor expression largely localized to the plasma membrane of insulin-producing cells. Pretreating the islets with LA for 30 min followed by antisense oligonucleotide treatment for 48 h reduced the FFA(1)R/GPR40 immunoreactivity to background levels. The results demonstrate that FFA(1)R/GPR40 is inhibited by the CPT-1 inhibitor, 2BrP, and confirm that FFA(1)R/GPR40 is indeed necessary, at least in part, for fatty-acid-stimulated insulin release

Recent High Incidence of Fulminant Hepatitis in Samara, Russia: Molecular Analysis of Prevailing Hepatitis B and D Virus Strains

Until 1991, the Russian city of Samara was largely isolated from other parts of Russia and the rest of the world. Very recently, Samara has seen an alarming increase in the incidence of hepatitis. The proportion of fulminant cases is unusually high. We wanted to assess the roles of hepatitis B virus (HBV) and hepatitis D virus (HDV) in acute viral hepatitis in this region by analyzing the prevailing strains of both and by determining their genotypes and possible origin. Serum samples were screened for different serological markers and by PCR followed by direct sequencing. Of the 94 HBV-positive samples (80% of which were acute infections), 37 (39%) were also HDV positive. Sixty-seven percent of the patients had anti-HCV antibodies. Twenty-five percent of all patients in the study had fulminant hepatitis. Statistically significant sex differences were found among fulminant cases. For HBV, the core promoter sequences of 62 strains were determined and all but one were found to be of genotype D. None of these had any deletions. Only one strain, from a patient with fulminant fatal hepatitis, showed multiple mutations. The pre-S2 region sequences of 31 HBV strains were also compared. Phylogenetically, these fell into two distinct groups within genotype D, suggesting different origins. For HDV, part of the region encoding the δ-antigen was sequenced from four strains. All proved to be of genotype I and were similar to Far Eastern and Eastern European strains. The contribution of intravenous drug use to the sharp increase in viral hepatitis in this unique setting is discussed

Islet NOS activities, insulin secretion, and expression of GPR40 and iNOS after treatment with palmitate and GPR40 antisense.

<p>Isolated islets were pretreated for 24 h with either the M40 antisense morpholino or a non-specific random sequence morpholino (control). The islets were incubated for 30 min in absence or presence of palmitate and then cultured with palmitate±M40 for a further 24 h. (A) M40 caused a marked suppression of palmitate-induced iNOS and ncNOS activities as well as a reduced insulin release (black bars). The results from the control morpholino are indicated by white bars. Values are mean±s.e.m for 4 different experiments performed at different occasions. ** p<0.01; *** p<0.001. (B) Expression of GPR40 (green) and iNOS (red) in islets cultured with palmitate (A-C) or palmitate+M40 (D-F). A and D = GPR40; B and E = iNOS, C and F = overlay. Bar indicates 5 μm. (C) Immunostaining and confocal images of formaldehyde-fixed β-cells. The expression pattern of insulin (red), GPR40 (green) and iNOS (yellow) from dispersed β-cells cultured with palmitate is shown. A = insulin, B = GPR40, C = iNOS and D = overlay. E–H show absence of expression of GPR40 (F) and iNOS (G) after M40 treatment. E = insulin, H = overlay. Bar indicates 5 μm.</p