Catechol estrogens stimulate insulin secretion in pancreatic β-cells via activation of the transient receptor potential A1 (TRPA1) channel

Estrogen hormones play an important role in controlling glucose homeostasis and pancreatic β-cell function. Despite the significance of estrogen hormones for regulation of glucose metabolism, little is known about the roles of endogenous estrogen metabolites in modulating pancreatic β-cell function. In this study, we evaluated the effects of major natural estrogen metabolites, catechol estrogens, on insulin secretion in pancreatic β-cells. We show that catechol estrogens, hydroxylated at positions C2 and C4 of the steroid A ring, rapidly potentiated glucose-induced insulin secretion via a nongenomic mechanism. 2-Hydroxyestrone, the most abundant endogenous estrogen metabolite, was more efficacious in stimulating insulin secretion than any other tested catechol estrogens. In insulin-secreting cells, catechol estrogens produced rapid activation of calcium influx and elevation in cytosolic free calcium. Catechol estrogens also generated sustained elevations in cytosolic free calcium and evoked inward ion current in HEK293 cells expressing the transient receptor potential A1 (TRPA1) cation channel. Calcium influx and insulin secretion stimulated by estrogen metabolites were dependent on the TRPA1 activity and inhibited with the channel-specific pharmacological antagonists or the siRNA. Our results suggest the role of estrogen metabolism in a direct regulation of TRPA1 activity with potential implications for metabolic diseases

Increase in cellular glutamate levels stimulates exocytosis in pancreatic β-cells

AbstractGlutamate has been implicated as an intracellular messenger in the regulation of insulin secretion in response to glucose. Here we demonstrate by measurements of cell capacitance in rat pancreatic β-cells that glutamate (1 mM) enhanced Ca2+-dependent exocytosis. Glutamate (1 mM) also stimulated insulin secretion from permeabilized rat β-cells. The effect was dose-dependent (half-maximum at 5.1 mM) and maximal at 10 mM glutamate. Glutamate-induced exocytosis was stronger in rat β-cells and clonal INS-1E cells compared to β-cells isolated from mice and in parental INS-1 cells, which correlated with the expressed levels of glutamate dehydrogenase. Glutamate-induced exocytosis was inhibited by the protonophores FCCP and SF6847, by the vacuolar-type H+-ATPase inhibitor bafilomycin A1 and by the glutamate transport inhibitor Evans Blue. Our data provide evidence that exocytosis in β-cells can be modulated by physiological increases in cellular glutamate levels. The results suggest that stimulation of exocytosis is associated with accumulation of glutamate in the secretory granules, a process that is dependent on the transgranular proton gradient

GPR142 Controls Tryptophan-Induced Insulin and Incretin Hormone Secretion to Improve Glucose Metabolism.

GPR142, a putative amino acid receptor, is expressed in pancreatic islets and the gastrointestinal tract, but the ligand affinity and physiological role of this receptor remain obscure. In this study, we show that in addition to L-Tryptophan, GPR142 signaling is also activated by L-Phenylalanine but not by other naturally occurring amino acids. Furthermore, we show that Tryptophan and a synthetic GPR142 agonist increase insulin and incretin hormones and improve glucose disposal in mice in a GPR142-dependent manner. In contrast, Phenylalanine improves in vivo glucose disposal independently of GPR142. Noteworthy, refeeding-induced elevations in insulin and glucose-dependent insulinotropic polypeptide are blunted in Gpr142 null mice. In conclusion, these findings demonstrate GPR142 is a Tryptophan receptor critically required for insulin and incretin hormone regulation and suggest GPR142 agonists may be effective therapies that leverage amino acid sensing pathways for the treatment of type 2 diabetes

GPR142 prompts glucagon-like Peptide-1 release from islets to improve β cell function

Objective: GPR142 agonists are being pursued as novel diabetes therapies by virtue of their insulin secretagogue effects. But it is undetermined whether GPR142's functions in pancreatic islets are limited to regulating insulin secretion. The current study expands research on its action. Methods and Results: We demonstrated by in situ hybridization and immunostaining that GPR142 is expressed not only in β cells but also in a subset of α cells. Stimulation of GPR142 by a selective agonist increased glucagon secretion in both human and mouse islets. More importantly, the GPR142 agonist also potentiated glucagon-like peptide-1 (GLP-1) production and its release from islets through a mechanism that involves upregulation of prohormone convertase 1/3 expression. Strikingly, stimulation of insulin secretion and increase in insulin content via GPR142 engagement requires intact GLP-1 receptor signaling. Furthermore, GPR142 agonist increased β cell proliferation and protected both mouse and human islets against stress-induced apoptosis. Conclusions: Collectively, we provide here evidence that local GLP-1 release from α cells defines GPR142's beneficial effects on improving β cell function and mass, and we propose that GPR142 agonism may have translatable and durable efficacy for the treatment of type 2 diabetes. Keywords: GPR142, Intra-islet GLP-1, α cell, PC1/

Selective activation of GPR142 signaling by L-Tryptophan and L-Phenylalanine and GPR142 mRNA expression in mouse and human tissues.

<p>(A) IP-1 levels in HEK293 cells expressing human GPR142 treated with amino acids and metabolites at varying concentrations. (B) IP-1 levels in HEK293-hGPR142 cells and untransfected control HEK293 cells treated with L-Tryptophan or L-Phenylalanine. Data are mean ± SD of 2 replicate wells. Data representative of 3 independent experiments are shown. (C-E) Mouse <i>Gpr142</i> mRNA expression determined by quantitative RT-PCR in (C) tissues of normal C57BL/6 male mice, (D) pancreatic islets isolated from male <i>Gpr142</i> KO mice and WT littermate controls, and (E) pancreatic islets isolated from male 8-week-old db/db mice and age-matched C57 control mice. (F, G) Human <i>GPR142</i> mRNA expression determined by quantitative RT-PCR in (F) a panel of human tissues and (G) pancreatic islets isolated from healthy non-diabetic or type 2 diabetic donors. Data are normalized against Rplp0 mRNA in each tissue sample, then normalized to the mean value of islet expression (C, F) or control group (D, E, G) set as 1, and expressed as fold expression. N = 2–4 (C-D), N = 5 per group (E), N = 1 pooled biological samples from ≥ 3 donors per tissue type (F), or N = 12–15 donors per group (G). Error bars represent SEM. ND: not detectable. **: p<0.01 control vs. db/db.</p

Effects of compound A on insulin secretion, plasma hormones and glycemia during oral glucose tolerance test.

<p>(A-B) Pancreatic islets isolated from normal C57 mice (A) or a non-diabetic human donor (B) were incubated in the presence of 11 mM glucose and varying concentrations of compound A for 1 hour. Insulin concentrations in the culture media were measured. Data representative of 2–3 independent experiments are shown. Data are mean ± SEM. N = 5 replicates per group. *,**,***: p<0.05, 0.01, 0.001 vs. control. (C-D) Vehicle (1% w/v HEC, 0.25% v/v Tween80, 0.05% v/v Antifoam in DI water) or compound A (30 mg/kg) was dosed orally to overnight fasted normal male C57 mice. 30 minutes later, glucose (2 g/kg) was orally dosed, and cardiac blood was collected at T = 0 (no glucose dosing), 3, or 15min after glucose challenge. Plasma levels of GIP (C) and total GLP-1 (D) were measured. (E-F) Vehicle or compound A (10 mg/kg) was dosed orally to 5hr daytime fasted diet-induced obese male C57 mice. 30 minutes later, glucose (2 g/kg) was orally dosed. Blood glucose was monitored for the next 120 minutes (E) and plasma insulin at 15 minutes after glucose challenge was measured (F). Data are mean ± SEM. N = 6 per group. *,**,***: p<0.05, 0.01, 0.001 compound A vs. vehicle.</p