Vasodilator Effect of Glucagon: Receptorial Crosstalk Among Glucagon, GLP-1, and Receptor for Glucagon and GLP-1.

Glucagon is known for its insulin-antagonist effect in the blood glucose homeostasis, while it also reduces vascular resistance. The mechanism of the vasoactive effect of glucagon has not been studied before; thereby we aimed to investigate the mediators involved in the vasodilatation induced by glucagon. The vasoactive effect of glucagon, insulin, and glucagon-like peptide-1 was studied on isolated rat thoracic aortic rings using a wire myograph. To investigate the mechanism of the vasodilatation caused by glucagon, we determined the role of the receptor for glucagon and the receptor for GLP-1, and studied also the effect of various inhibitors of gasotransmitters, inhibitors of reactive oxygen species formation, NADPH oxidase, prostaglandin synthesis, protein kinases, potassium channels, and an inhibitor of the Na(+)/Ca(2+)-exchanger. Glucagon causes dose-dependent relaxation in the rat thoracic aorta, which is as potent as that of insulin but greater than that of GLP-1 (7-36) amide. Vasodilatation by GLP-1 is partially mediated by the glucagon receptor. The vasodilatation due to glucagon evokes via the glucagon-receptor, but also via the receptor for GLP-1, and it is endothelium-independent. Contribution of gasotransmitters, prostaglandins, the NADPH oxidase enzyme, free radicals, potassium channels, and the Na(+)/Ca(2+)-exchanger is also significant. Glucagon causes dose-dependent relaxation of rat thoracic aorta in vitro, via the receptor for glucagon and the receptor for GLP-1, while the vasodilatation evoked by GLP-1 also evolves partially via the receptor for glucagon, thereby, a possible crosstalk between the 2 hormones and receptors could occur.

Elevated vascular level of ortho-tyrosine contributes to the impairment of insulin-induced arterial relaxation.

Previous studies have shown that in diabetes mellitus, insulin-induced relaxation of arteries is impaired and the level of ortho-tyrosine (o-Tyr), an oxidized amino acid is increased. Thus, we hypothesized that elevated vascular level of o-Tyr contributes to the impairment of insulin-induced vascular relaxation. Rats were fed with o-Tyr for 4 weeks. Insulin-induced vasomotor responses of isolated femoral artery were studied using wire myography. Vascular o-Tyr content was measured by HPLC, whereas immunoblot analyses were preformed to detect eNOS phosphorylation. Sustained oral supplementation of rats with o-Tyr increased the content of o-Tyr in the arterial wall and significantly reduced the relaxations to insulin. Sustained supplementation of cultured endothelial cells with o-Tyr increased the incorporation of o-Tyr and mitigated eNOS Ser (1 177) phosphorylation to insulin. Increasing arterial wall o-Tyr level attenuates insulin-induced relaxation - at least in part - by decreasing eNOS activation. Elevated level of o-Tyr could be an underlying mechanism for vasomotor dysfunction in diabetes mellitus.

Increase in insulin-induced relaxation of consecutive arterial segments toward the periphery: Role of vascular oxidative state.

RATIONALE: The oxidative state has been implicated in the signaling of various vasomotor functions, yet its role regarding the vasomotor action of insulin is less known. OBJECTIVE: To investigate the insulin-evoked relaxations of consecutive arterial segments of different oxidative state and the role of extracellular signal-regulated kinase (ERK) pathway. METHODS AND RESULTS: The oxidative state, as assessed by the level of ortho-tyrosine, was higher in the thoracic aorta of rats than in the abdominal aorta, and was the lowest in the femoral artery. The vasomotor function of vessels of same origin was studied using a small-vessel myograph. Insulin-induced relaxations increased toward the periphery (i.e., thoracic < abdominal < femoral). Aortic banding and hydrogen peroxide/aminotriazole increased the oxidative state of the thoracic aorta that was accompanied by ERK activation and decreased relaxation to insulin, and vice versa, acutely lowered oxidative state by superoxide dismutase/catalase improved relaxation. In contrast, insulin-induced relaxation of the femoral artery could be enhanced with a higher oxidative state, and reduced with a lower state. CONCLUSIONS: Oxidative state of vessels modulates the magnitude of vasomotor responses to insulin, which appears to be mediated via the ERK signaling pathway.

Effects of erythropoietin on glucose metabolism.

We purposed to determine the impact of erythropoietin on altering glucose metabolism in the settings of in vitro and in vivo experiments. The acute effect of erythropoietin on lowering blood glucose levels was studied in animal experiments. In [³H]-deoxy-D-glucose isotope studies we measured glucose uptake with insulin and erythropoietin using 3T3-L1 cells cultured under normal or high glucose conditions. Altered activation of Akt and ERK pathways was evaluated in immunoblot analyses. Immunocytochemistry was conducted to determine the glucose transporter 4 translocation to the plasma membrane. Addition of erythropoietin significantly lowered blood glucose levels in vivo in rats. The glucose uptake was markedly increased by erythropoietin treatment (at concentrations 0.15, 0.3, and 0.625 ng/ml) in adipocytes grown in high glucose medium (p<0.05), but it remained unaltered in cells under normal glucose conditions. Significant increase of phosphorylation of ERK and Akt was detected due to erythropoietin (p<0.05). Co-administration of erythropoietin and insulin resulted in higher phosphorylation of Akt and [³H]-deoxy-D-glucose uptake in adipocytes than insulin treatment alone. We found that erythropoietin induced the trafficking of glucose transporter 4 to the plasma membrane. Our data showed that erythropoietin significantly decreased blood glucose levels both in vivo and in vitro, in part, by increasing glucose uptake via the activation of Akt pathway. Preliminary data revealed that adipocytes most likely exhibit a specific receptor for erythropoietin.

A polymorphism within the fructosamine-3-kinase gene is associated with HbA1c Levels and the onset of type 2 diabetes mellitus.

BACKGROUND: Non-enzymatic glycation is a process, which leads to the formation of advanced glycation endproducts. These compounds are involved in the development of diabetic microvascular complications. Fructosamine-3-kinase (FN3K) is an intracellular enzyme that phosphorylates fructosamines resulting in fructosamine-3-phosphate, which subsequently decomposes to inorganic phosphate, 3-deoxyglucasone and the unmodified amine. Recently, the G900C (rs1056534) single nucleotide polymorpism (SNP) of the FN3K gene was found to be associated with the enzyme activity. OBJECTIVE/DESIGN: The aim of the study was to investigate the impact of the SNP on clinical and biochemical features and microvascular complications of type 2 diabetes. PATIENTS: A total of 859 type 2 diabetic subjects and 265 healthy controls were enrolled in the study and were genotyped with PCR-RFLP method. RESULTS: Genotype frequencies were as follows, CC: 5%, GC: 54%, GG: 41% in subjects with type 2 diabetes and CC: 6%, GC: 51%, GG: 43% in the controls. Diabetic subjects with the CC variant had lower HbA (1c) levels compared with the others (CC: 6.48+/-0.05%; GC: 7.66+/-0.09%; GG: 7.68+/-0.09%; p<0.001). Furthermore, in case of the CC allelic variant type 2 diabetes was diagnosed at a later age than in case of GC or GG variants (CC: 56.0+/-1.90 years; GC: 52.0+/-0.62 years; GG: 50.1+/-0.71 years; p<0.05). Logistic regression analysis did not reveal association between CC genotype and diabetic complications, such as diabetic nephropathy, neuropathy and retinopathy (OR=1.036, CI 95% 0.652-1.647, p=0.880; OR=0.985, CI 95% 0.564-1.721 p=0.958; OR=1.213, CI 95% 0.470-3.132, p=0.690, respectively). CONCLUSION: We conclude that the G900C polymorphism associates with the level of HbA (1c) and the onset of the disease, but not with either of the diabetic microvascular complications.

Morphology of glomerular hematuria is reproduced in vitro by carbonyl stress.

BACKGROUND: In glomerulonephritides, dysmorphic red blood cells (RBCs) with membrane blebs can be found in the urine; this is referred to as glomerular hematuria. Glomerulonephritides are characterized by increased carbonyl stress and elevated methylglyoxal (MGO) levels. MGO causes oxidative stress and intracellular calcium accumulation. In the present study, we investigated whether the effect of MGO-induced calcium accumulation in RBCs develops through increased oxidative stress. Furthermore, we studied whether MGO can lead to RBC membrane blebbing. METHODS: RBC suspensions from healthy volunteers were incubated with different concentrations of MGO at 37 degrees C. We measured oxidative stress and intracellular calcium level using fluorescent indicators. We determined the frequency of dysmorphic RBCs, and also performed scanning electron microscopy. RESULTS: MGO increased oxidative stress and caused accumulation of calcium in isolated RBCs. These effects could be prevented using antioxidants. In the presence of MGO, RBC membrane blebbing developed. CONCLUSION: According to our findings, MGO causes calcium accumulation through oxidative stress. Carbonyl and oxidative stress may play an important role in the formation of dysmorphic RBCs in glomerular hematuria.