Combination of Metformin and Inorganic Nitrate: Effects on Metabolic Disease and Lactic Acidosis

Nitric oxide (NO) is a diatomic gaseous molecule generated in our bodies by enzymes known as NO synthases. More recently another pathway for generation of NO has been delineated in which dietary or endogenous inorganic nitrate and nitrite are metabolized to form NO. Dietary supplementation with nitrate has proven beneficial for a number of disorders e.g. by improving metabolic function and lowering blood pressure, making it an attractive novel drug candidate in metabolic and cardiovascular disease. Besides utilizing direct therapeutic effects of nitrate itself, another attractive option might be to combine nitrate with another drug in order to increase the overall effect or reduce the side effects of the other drug. The aim of the proposed thesis was to determine whether simultaneous acute administration of a low dose of metformin and nitrate can act synergistically to improve metabolic functions and decrease blood glucose level in a metabolic syndrome genetic mouse model (A2BKO mice) and also if this combination can prevent lactic acidosis produced by metformin in rats. The results showed that both nitrate and metformin alone had a beneficial effect on glucose tolerance but there was no clear synergistic effect to decrease blood glucose levels. We speculate that the lack of synergy may be due to their same mechanism of action in targeting NADPH oxidase and AMP-activated protein kinase. However, the glucose disposal was better in metformin and nitrate combination group compared to metformin group. Moreover, in separate experiments we found that acute administration of nitrate had no impact on prevention of lactic acidosis induced by high dose of metformin in rats. However, future chronic studies are necessary in order to ameliorate our knowledge of the proposed combination mechanism.</p

Combination of Metformin and Inorganic Nitrate: Effects on Metabolic Disease and Lactic Acidosis

Nitric oxide (NO) is a diatomic gaseous molecule generated in our bodies by enzymes known as NO synthases. More recently another pathway for generation of NO has been delineated in which dietary or endogenous inorganic nitrate and nitrite are metabolized to form NO. Dietary supplementation with nitrate has proven beneficial for a number of disorders e.g. by improving metabolic function and lowering blood pressure, making it an attractive novel drug candidate in metabolic and cardiovascular disease. Besides utilizing direct therapeutic effects of nitrate itself, another attractive option might be to combine nitrate with another drug in order to increase the overall effect or reduce the side effects of the other drug. The aim of the proposed thesis was to determine whether simultaneous acute administration of a low dose of metformin and nitrate can act synergistically to improve metabolic functions and decrease blood glucose level in a metabolic syndrome genetic mouse model (A2BKO mice) and also if this combination can prevent lactic acidosis produced by metformin in rats. The results showed that both nitrate and metformin alone had a beneficial effect on glucose tolerance but there was no clear synergistic effect to decrease blood glucose levels. We speculate that the lack of synergy may be due to their same mechanism of action in targeting NADPH oxidase and AMP-activated protein kinase. However, the glucose disposal was better in metformin and nitrate combination group compared to metformin group. Moreover, in separate experiments we found that acute administration of nitrate had no impact on prevention of lactic acidosis induced by high dose of metformin in rats. However, future chronic studies are necessary in order to ameliorate our knowledge of the proposed combination mechanism.Siirretty Doriast

AMP-activated protein kinase activation and NADPH oxidase inhibition by inorganic nitrate and nitrite prevent liver steatosis

Advanced age and unhealthy dietary habits contribute to the increasing incidence of obesity and type 2 diabetes. These metabolic disorders, which are often accompanied by oxidative stress and compromised nitric oxide (NO) signaling, increase the risk of adverse cardiovascular complications and development of fatty liver disease. Here, we investigated the therapeutic effects of dietary nitrate, which is found in high levels in green leafy vegetables, on liver steatosis associated with metabolic syndrome. Dietary nitrate fuels a nitrate–nitrite–NO signaling pathway, which prevented many features of metabolic syndrome and liver steatosis that developed in mice fed a high-fat diet, with or without combination with an inhibitor of NOS (L-NAME). These favorable effects of nitrate were absent in germ-free mice, demonstrating the central importance of host microbiota in bioactivation of nitrate. In a human liver cell line (HepG2) and in a validated hepatic 3D model with primary human hepatocyte spheroids, nitrite treatment reduced the degree of metabolically induced steatosis (i.e., high glucose, insulin, and free fatty acids), as well as drug-induced steatosis (i.e., amiodarone). Mechanistically, the salutary metabolic effects of nitrate and nitrite can be ascribed to nitrite-derived formation of NO species and activation of soluble guanylyl cyclase, where xanthine oxidoreductase is proposed to mediate the reduction of nitrite. Boosting this nitrate–nitrite–NO pathway results in attenuation of NADPH oxidase-derived oxidative stress and stimulation of AMP-activated protein kinase and downstream signaling pathways regulating lipogenesis, fatty acid oxidation, and glucose homeostasis. These findings may have implications for novel nutrition-based preventive and therapeutic strategies against liver steatosis associated with metabolic dysfunction.</p

Folate Receptor β Targeted PET Imaging of Macrophages in Autoimmune Myocarditis

Rationale: Currently available imaging techniques have limited specificity for the detection of active myocardial inflammation. Aluminum fluoride-18-labeled 1,4,7-triazacyclononane-N,N′,N″-triacetic acid conjugated folate (18F-FOL) is a positron emission tomography (PET) tracer targeting folate receptor β (FR-β) that is expressed on activated macrophages at sites of inflammation. We evaluated 18F-FOL PET for the detection of myocardial inflammation in rats with autoimmune myocarditis and studied expression of FR-β in human cardiac sarcoidosis specimens. Methods: Myocarditis was induced by immunizing rats (n = 18) with porcine cardiac myosin in complete Freund’s adjuvant. Control rats (n = 6) were injected with Freund’s adjuvant alone. 18F-FOL was intravenously injected followed by imaging with a small animal PET/computed tomography (CT) scanner and autoradiography. Contrast-enhanced high-resolution CT or 2-deoxy-2-18F-fluoro-D-glucose (18F-FDG) PET images were used for co-registration. Rat tissue sections and myocardial autopsy samples of 6 patients with cardiac sarcoidosis were studied for macrophages and FR-β. Results: The myocardium of 10 out of 18 immunized rats showed focal macrophage-rich inflammatory lesions with FR-β expression occurring mainly in M1-polarized macrophages. PET images showed focal myocardial 18F-FOL uptake co-localizing with inflammatory lesions (SUVmean, 2.1 ± 1.1), whereas uptake in the remote myocardium of immunized rats and controls was low (SUVmean, 0.4 ± 0.2 and 0.4 ± 0.1, respectively; P </p

Combination of Metformin and Inorganic Nitrate: Effects on Metabolic Disease and Lactic Acidosis

Nitric oxide (NO) is a diatomic gaseous molecule generated in our bodies by enzymes known as NO synthases. More recently another pathway for generation of NO has been delineated in which dietary or endogenous inorganic nitrate and nitrite are metabolized to form NO. Dietary supplementation with nitrate has proven beneficial for a number of disorders e.g. by improving metabolic function and lowering blood pressure, making it an attractive novel drug candidate in metabolic and cardiovascular disease. Besides utilizing direct therapeutic effects of nitrate itself, another attractive option might be to combine nitrate with another drug in order to increase the overall effect or reduce the side effects of the other drug. The aim of the proposed thesis was to determine whether simultaneous acute administration of a low dose of metformin and nitrate can act synergistically to improve metabolic functions and decrease blood glucose level in a metabolic syndrome genetic mouse model (A2BKO mice) and also if this combination can prevent lactic acidosis produced by metformin in rats. The results showed that both nitrate and metformin alone had a beneficial effect on glucose tolerance but there was no clear synergistic effect to decrease blood glucose levels. We speculate that the lack of synergy may be due to their same mechanism of action in targeting NADPH oxidase and AMP-activated protein kinase. However, the glucose disposal was better in metformin and nitrate combination group compared to metformin group. Moreover, in separate experiments we found that acute administration of nitrate had no impact on prevention of lactic acidosis induced by high dose of metformin in rats. However, future chronic studies are necessary in order to ameliorate our knowledge of the proposed combination mechanism.Siirretty Doriast

Synthesis and characterization of a novel organic nitrate NDHP: Role of xanthine oxidoreductase-mediated nitric oxide formation

In this report, we describe the synthesis and characterization of 1,3-bis(hexyloxy)propan-2-yl nitrate (NDHP), a novel organic mono nitrate. Using purified xanthine oxidoreductase (XOR), chemiluminescence and electron paramagnetic resonance (EPR) spectroscopy, we found that XOR catalyzes nitric oxide (NO) generation from NDHP under anaerobic conditions, and that thiols are not involved or required in this process. Further mechanistic studies revealed that NDHP could be reduced to NO at both the FAD and the molybdenum sites of XOR, but that the FAD site required an unoccupied molybdenum site. Conversely, the molybdenum site was able to reduce NDHP independently of an active FAD site. Moreover, using isolated vessels in a myograph, we demonstrate that NDHP dilates pre-constricted mesenteric arteries from rats and mice. These effects were diminished when XOR was blocked using the selective inhibitor febuxostat. Finally, we demonstrate that NDHP, in contrast to glyceryl trinitrate (GTN), is not subject to development of tolerance in isolated mesenteric arteries

Vascular adhesion protein-1-targeted PET imaging in autoimmune myocarditis

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