Anatomical distribution of primary amine oxidase activity in four adipose depots and plasma of severely obese women with or without a dysmetabolic profile

Semicarbazide-sensitive amine oxidase (SSAO), identical to primary amine oxidase or vascular adhesion protein-1, is a membrane enzyme that generates hydrogen peroxide. SSAO is highly expressed at the adipocyte surface, and its plasma levels increase with type 2 diabetes. Since visceral adipose tissue (AT) is more tightly associated with obesity complications than subcutaneous (SC) abdominal fat, we compared SSAO activity in plasma and 4 distinct AT locations in 48 severely obese women (body mass index (BMI), averaging 54 ± 11 kg/m²), with or without a dysmetabolic profile. Higher glucose and triacylglycerol levels vs lower high-density lipoprotein (HDL)-cholesterol characterized dysmetabolic women (DYS; n = 25) from non-dysmetabolic (NDYS; n = 23), age- and weight-matched subjects. SC, mesenteric (ME), omental (OM), and round ligament (RL) fat locations were collected during bariatric surgery. SSAO capacity to oxidize up to 1 mM benzylamine was determined in AT and plasma with radiometric and fluorimetric methods. Plasma SSAO was higher in the DYS group. SSAO activity was higher in fat than in plasma, when expressed as radiolabeled benzaldehyde per milligram of protein. In ATs from DYS women, protein content was 10 % higher, and basal hydrogen peroxide release lower than in NDYS subjects, except for RL location. The SSAO affinity towards benzylamine did not exhibit regional variation and was not altered by a dysmetabolic profile (K _m averaging 184 ± 7 μM; n = 183). Although radiometric and fluorimetric methods gave different estimates of oxidase activity, both indicated that AT SSAO activity did not vary according to anatomical location and/or metabolic status in severely obese women.     

Alteration of Amine Oxidase Activity in the Adipose Tissue of Obese Subjects

Objective: To explore the activity of monoamine oxidases (MAOs) and semicarbazide‐sensitive amine oxidases (SSAOs) in adipose tissue and blood of lean and moderately obese subjects and to study whether there is a link between these hydrogen peroxide‐generating enzymes and blood markers of oxidative stress. Research Methods and Procedures: Nine obese male subjects (BMI 32.6 ± 0.4 kg/m²) and nine controls (BMI 23.4 ± 0.5) of 24‐ to 40‐year‐old subjects were included in the study. MAO and SSAO activities were measured on microbiopsies of abdominal subcutaneous adipose tissue by quantifying ¹⁴C‐tyramine and ¹⁴C‐benzylamine oxidation. Levels of soluble SSAO, lipid peroxidation products, and antioxidant agents were measured in plasma, whereas cytoprotective enzymes were determined in blood lysates. Results: The high MAO activity found in adipose tissue was diminished by one‐half in obese subjects (maximum initial velocity of 1.2 vs. 2.3 nmol tyramine oxidized/mg protein/min). There was no change in SSAO activity, either under its adipose tissue‐bound or plasma‐soluble form. Plasma levels of lipid peroxidation products and antioxidant vitamins remained unmodified, as well as erythrocyte antioxidant enzymes, whereas circulating triglycerides, insulin, and leptin were increased. Discussion: Although they already exhibited several signs of endocrino‐metabolic disorders, the obese men did not exhibit the increase in blood markers of oxidative stress or the decrease in antioxidant defenses reported to occur in very obese or diabetic subjects. The reduced MAO and the unchanged SSAO activities found in obesity suggest that these hydrogen peroxide‐generating enzymes expressed in adipocytes are probably not involved in the onset of the oxidative stress found in severe obesity and/or in its complications.     

Variation in semicarbazide-sensitive amine oxidase activity in plasma and tissues of mammals

Semicarbazide-sensitive amine oxidase (SSAO) (E.C. 1.4.3.6) is a group of enzymes with as yet poorly understood function which is widely present in nature. The variation in methodology for determination of activity, differences in substrates used and in nomenclature have made it difficult to compare SSAO in different species and tissues. Since SSAO is implicated in the pathophysiology of diabetes mellitus and congestive heart failure, our aim was to analyse the importance and abundance of SSAO in human plasma and tissues compared to other mammals. In plasma of ten different mammals, Vmax values were found to vary more than 10,000-fold, while KM differed much less; in human plasma SSAO activity is relatively low. In some species more than one SSAO entity was present in plasma. SSAO activity was ubiquitous in tissues of human, rat and pig, but varied considerably, both between species and between tissues. In human tissues, SSAO activity is higher than in tissues from rat and pig. Relative to monoamine oxidase-B there is also wide variation in SSAO, with much higher relative activities in human than in rat and pig tissues. We conclude that in plasma, SSAO activity is highest in ruminants, while in tissues, SSAO activity is more prominently present in human than in rat and pig.     

Blood-brain barrier dysfunction underlying Alzheimer's disease is induced by an SSAO/VAP-1-dependent cerebrovascular activation with enhanced Aβ deposition

Dysfunctions of the vascular system directly contribute to the onset and progression of Alzheimer's disease (AD). The blood-brain barrier (BBB) shows signs of malfunction at early stages of the disease. When Abeta peptide (Aβ) is deposited on brain vessels, it induces vascular degeneration by producing reactive oxygen species and promoting inflammation. These molecular processes are also related to an excessive SSAO/VAP-1 (semicarbazide-sensitive amine oxidase) enzymatic activity, observed in plasma and in cerebrovascular tissue of AD patients. We studied the contribution of vascular SSAO/VAP-1 to the BBB dysfunction in AD using in vitro BBB models. Our results show that SSAO/VAP-1 expression is associated to endothelial activation by altering the release of pro-inflammatory and pro-angiogenic angioneurins, most highly IL-6, IL-8 and VEGF. It is also related to a BBB structure alteration, with a decrease in tight-junction proteins such as zona occludens or claudin-5. Moreover, the BBB function reveals increased permeability and leukocyte adhesion in cells expressing SSAO/VAP-1, as well as an enhancement of the vascular Aβ deposition induced by mechanisms both dependent and independent of the enzymatic activity of SSAO/VAP-1. These results reveal an interesting role of vascular SSAO/VAP-1 in BBB dysfunction related to AD progression, opening a new window in the search of alternative therapeutic targets for fighting AD.     

Inactivation of Semicarbazide-Sensitive Amine Oxidase Stabilizes the Established Atherosclerotic Lesions via Inducing the Phenotypic Switch of Smooth Muscle Cells

Given that the elevated serum semicarbazide-sensitive amine oxidase (SSAO) activity is associated with the severity of carotid atherosclerosis in clinic, the current study aims to investigate whether SSAO inactivation by semicarbazide is beneficial for established atherosclerotic lesions in LDLr knockout mice on a high-fat/high- cholesterol Western-type diet or after dietary lipid lowering. Despite no impact on plasma total cholesterol levels, the infiltration of circulating monocytes into peripheral tissues, and the size of atherosclerotic lesions, abrogation of SSAO activity resulted in the stabilization of established lesions as evidenced by the increased collagen contents under both conditions. Moreover, SSAO inactivation decreased Ly6C^high monocytosis and lesion macrophage contents in hypercholesterolemic mice, while no effect was observed in mice after normalization of hypercholesterolemia by dietary lipid lowering. Strikingly, abrogation of SSAO activity significantly increased not only the absolute numbers of smooth muscle cells (SMCs), but also the percent of SMCs with a synthetic phenotype in established lesions of mice regardless of plasma cholesterol levels. Overall, our data indicate that SSAO inactivation in vivo stabilizes the established plaques mainly via inducing the switch of SMCs from a contractile to a synthetic phenotype. Targeting SSAO activity thus may represent a potential treatment for patients with atherosclerosis.     

Abnormality in Translational Regulation of Catalase Expression in Disorders of Peroxisomal Biogenesis

Abstract: Peroxisomal disorders are a newly described group of inherited neurological diseases. In disorders of peroxisomal biogenesis, e.g., Zellweger syndrome, owing to the lack of peroxisomes, catalase, a peroxisomal enzyme, is found to be present in the cytoplasm instead. We observed higher catalase activity (7.59 ± 0.41 mU/mg of protein) in cultured skin fibroblasts from Zellweger patients than in control fibroblasts (4.45 ± 0.29 mU/mg of protein). Moreover, we also found that the majority of the catalase in Zellweger cells was present in the inactive form. The specific activities following reactivation in Zellweger and control cells were 12.1 and 4.9 mU/mg of protein, respectively. To understand the molecular basis of higher levels of catalase in Zellweger than control cells, we examined the rate of synthesis and turnover of catalase and levels of catalase mRNA and protein levels in Zellweger cells as compared with control cells. The initial rates of synthesis of catalase in Zellweger (1.68 ± 0.15 mU/mg of protein) and control (1.51 ± 0.14 mU/mg of protein) cells were similar. The rates of turnover of catalase in Zellweger (t_1/2 = 47 ± 8 h) and control (t_1/2 = 49 ± 7 h) were also similar. Consistent with the enzyme activity, the levels of catalase protein were higher in Zellweger cells as compared with control cells. On the other hand, there was no difference in the level of catalase mRNA between control and Zellweger cells. Although the rate of synthesis in Zellweger and control cells were initially similar, it was down-regulated to a lower level at ～72 h of culture in control fibroblasts as compared with Zellweger cells, which continued to synthesize catalase at the same rate up to 5 days in culture. The presence of similar levels of mRNA in control and Zellweger cells and continued synthesis of catalase in Zellweger cells at a higher level as compared with control cells suggest a loss of regulation at the translational level.     

Plasma semicarbazide-sensitive amine oxidase in human (patho)physiology

Semicarbazide-sensitive amine oxidases (SSAO) are widely distributed enzymes, with as yet not fully elucidated functions and roles, present in many tissues but also circulating in plasma. The enzyme also functions as an adhesion molecule, the vascular adhesion protein-1. In healthy humans, plasma SSAO activity is constant from birth until 16 years of age, when it drops to lower values, gradually increasing again at advanced ages. When measuring SSAO activity, care should be taken to ensure proper preparation and storage conditions, and it should be realized that quite a few drugs unintentionally are good inhibitors, and sometimes even substrates, of SSAO. Under normal conditions SSAO activity is constant and inter-individual variation is small. In various pathophysiological conditions plasma SSAO activities are increased, most notably in diabetes mellitus (both type I and type II), in congestive heart failure and in cirrhotic liver inflammation. In patients with other vascular and inflammatory diseases plasma SSAO is normal, while it is low in children with congenital lung diseases. Interpretation of these changes is speculative, since source and regulation of plasma SSAO are as yet unknown. However, in two situations where the disease-causing process was ended (transplantation, delivery), plasma SSAO returned to normal. Many questions remain to be answered.     

Salivary gland apyrase determines probing time in anopheline mosquitoes

Salivary gland homogenates of adult female anopheline mosquitoes, of three different species, hydrolysed ATP and ADP, thereby demonstrating an apyrase activity. Total enzyme activity was greatest in the vector species A. freeborni (20.7 ± 2.4 mU/pair of glands) and least in the autogenous mosquito A. sp. nr. salbaii (3.0 ± 0.4 mU/pair of glands); another vector species, A. stephensi, produced intermediate levels of the enzyme (7.8 ± 0.7 mU/pair of glands). In all cases, the reaction was activated by divalent cations and maximal at pH 9.0 and in the presence of 2-mercaptoethanol. Apyrase activity in each salivary gland correlated with the degree of inhibition of ADP-induced platelet aggregation in vitro. Duration of probing correlated inversely with salivary apyrase content. We conclude that salivary apyrase largely determines a mosquito's ability to locate blood. Differential selective pressures for facility of blood location would have influenced the level of salivary apyrase in these mosquitoes.     

A colorimetric assay method to measure acetyl-CoA synthetase activity: Application to woodchuck model of hepatitis virus-induced hepatocellular carcinoma

A new spectrophotometric method for quantitation of acetyl-CoA synthetase (ACAS) activity is developed. It has been applied for ACAS assay in the liver tissues of a woodchuck model of hepatitis virus-induced hepatocellular carcinoma (HCC). The assay is based on the established pyrophosphate (PPi) detection system. ACAS activity is indexed by the amount of PPi, the product of ACAS reaction system of activated form of acetate (acetyl-CoA) with ACAS catalysis. PPi is determined quantitatively as the amount of chromophore formed with molybdate reagent, 1-amino-2-naphthol-4-sulfonic acid in bisulfite and 2-mercaptoethanol. PPi reacts with molybdate reagent to produce phosphomolybdate and PPi-molybdate complexes. 2-mercaptoethanol is responsible for color formation which has the peak absorbance at 580 nm. This method was sensitive from 1 to 20 nmol of PPi in a 380-μl sample (1-cm cuvette). A ten-fold excess of Pi did not interfere with the determination of PPi. To study the major metabolic pathways of imaging tracer [1-¹¹C]-acetate in tumors for detection of HCC by Positron Emission Tomography (PET), the activity of one of the key enzymes involved in acetate or [1-¹¹C]-acetate metabolism, ACAS was assayed by this newly developed assay in the tissue samples of woodchuck HCCs. A significant increase of ACAS activity was observed in the liver tissues of woodchuck HCCs as compared with neighboring regions surrounding the tumors (P < 0.05). The respective ACAS activities in the subcellular locations were also significantly higher in HCCs than in the surrounding tissues (P < 0.05) (total soluble fraction: 876.61 ± 34.64 vs. 361.62 ± 49.97 mU/g tissue; cytoplasmic fraction: 1122.02 ± 112.39 vs. 732.32 ± 84.44 mU/g tissue; organelle content: 815.79 ± 100.77 vs. 547.91 ± 97.05 mU/ g tissue; sedimentable fragment: 251.92 ± 51.56 vs. 90.94 ± 18.98 mU/ g tissue). The finding suggests an increase in ACAS activity in the liver cancer of woodchuck models of HCC as compared to that in the normal woodchuck liver. The developed assay is rapid, simple and accurate and is suitable for the investigation of ACAS activity under physiologic and pathophysiologic conditions.     

Inhibition of tissue-bound semicarbazide-sensitive amine oxidase by two haloamines, 2-bromoethylamine and 3-bromopropylamine

Various mammalian tissues contain membrane-bound amine oxidase termed semicarbazide-sensitive amine oxidase (SSAO). A variety of compounds has been identified as relatively selective SSAO inhibitors, but those inhibitors currently available also inhibit monoamine oxidase (MAO). In the present study, inhibitory properties of 2-bromoethylamine (2-BEA) and 3-bromopropylamine (3-BPA) toward rat lung-bound SSAO have been studied. Regardless of preincubation, 2-BEA could not appreciably inhibit MAO-A and MAO-B activity, but 3-BPA at relatively high concentrations inhibited only MAO-B activity. 3-BPA was a competitive and reversible SSAO inhibitor with a Ki value of 17 microM regardless of preincubation. In contrast, without preincubation, 2-BEA competitively inhibited SSAO activity with the Ki value of 2.5 microM and after preincubation, the mode of inhibition changed to be noncompetitive, indicating irreversible inhibition after the preincubation. Dialysis experiments with 2-BEA-pretreated homogenate resulted in no recovery of SSAO activity even after overnight dialysis. A decreased rate of SSAO inhibition under N2 atmosphere to that obtained under O2 was produced upon preincubation of enzyme with 2-BEA, suggesting that oxidized intermediate was necessary for its inhibitory activity. Thus, 2-BEA first interacts with SSAO to form a reversible complex with a subsequent reaction, leading this complex to the covalently bound enzyme-inhibitor adduct. The data analyzed by the plot of 1/k' vs 1/2-BEA concentrations intersected on the y-axis indicate that the inhibition by 2-BEA is not mediated by a bimolecular reaction; thus it is not an affinity-labeling agent, but a suicide SSAO inhibitor. 2-BEA may be employed as a useful compound in the studying SSAO.     

Synthetic liver X receptor agonist T0901317 inhibits semicarbazide-sensitive amine oxidase gene expression and activity in apolipoprotein E knockout mice

Semicarbazide-sensitive amine oxidase (SSAO) catalyzes oxidative deamination of primary aromatic and aliphatic amines. Increased SSAO activity has been found in atherosclerosis and diabetes mellitus. We hypothesize that the anti-atherogenic effect of liver X receptors (LXRs) might be related to the inhibition of SSAO gene expression and its activity. In this study, we investigated the effect of LXRagonist T0901317 on SSAO gene expression and its activity in apolipoprotein E knockout (apoE^−/−) mice. Male apoE^−/− mice (8 weeks old) were randomly divided into four groups: basal control group; vehicle group; prevention group; and treatment group. SSAO gene expression was analyzed by real-time quantitative polymerase chain reaction and its activity was determined. The activity of superoxide dismutase and content of malondialdehyde in the aorta and liver were also determined. In T0901317-treated mice, SSAO gene expression was significantly decreased in the aorta, liver, small intestine, and brain. SSAO activities in serum and in these tissues were also inhibited. The amount of superoxide dismutase in the aorta and liver of the prevention group and treatment group was significantly higher compared with the vehicle group ( P < 0.05). Malondialdehyde in the tissues of these two groups was significantly lower compared with the vehicle group ( P < 0.05). Our results showed that T0901317 inhibits SSAO gene expression and its activity in atherogenic apoE^−/− mice. The atheroprotective effect of LXR agonist T0901317 is related to the inhibition of SSAO gene expression and its activity.     

Semicarbazide-sensitive amine oxidase activity exerts insulin-like effects on glucose metabolism and insulin-signaling pathways in adipose cells

Semicarbazide-sensitive amine oxidase (SSAO) is very abundant at the plasma membrane in adipocytes. The combination of SSAO substrates and low concentrations of vanadate markedly stimulates glucose transport and GLUT4 glucose transporter recruitment to the cell surface in rat adipocytes by a mechanism that requires SSAO activity and hydrogen peroxide formation. Substrates of SSAO such as benzylamine or tyramine in combination with vanadate potently stimulate tyrosine phosphorylation of both insulin-receptor substrates 1 (IRS-1) and 3 (IRS-3) and phosphatidylinositol 3-kinase (PI 3-kinase) activity in adipose cells, which occurs in the presence of a weak stimulation of insulin-receptor kinase. Moreover, the acute administration of benzylamine and vanadate in vivo enhances glucose tolerance in non-diabetic and streptozotocin-induced diabetic rats and reduces hyperglycemia after chronic treatment in streptozotocin-diabetic rats. Based on these observations, we propose that SSAO activity and vanadate potently mimic insulin effects in adipose cells and exert an anti-diabetic action in an animal model of type 1 diabetes mellitus.     

Production and regulation of extracellular chitinase from the entomopathogenic fungus Isaria fumosorosea

Extracellular chitinase production by the entomopathogenic fungus, Isaria fumosorosea IF28.2 was studied by using submerged fermentation. Maximum chitinase production (178.34±3.91 mU/mL) was obtained when fermentation was carried out at 25°C for 120 h using 72-h-old mycelium in a medium. The effect of inoculum size on chitinase activity was also observed and maximum chitinase activity (159.41±2.91 mU/mL) was obtained with an inoculum size of 3 discs while an incubation period of 96 h proved the most active inducer of chitinase production yielding a chitinase activity of 186.14±3.81 mU/mL. Colloidal chitin (1.5%, w/v) proved to be the best concentration. The optimum pH for chitinase production was 5.7 while 25°C proved to be the best temperature for chitinase production. Supplementation of additional carbon source like 1.5% N-acetylglucosamine (GlcNAc) showed further enhancement in chitinase production. The divalent metal salts, CaCl₂, MgCl₂ and ZnSO₄, inhibited chitinase activity at 10 and 100 mM concentration, whereas inhibition of chitinase activity by KCl, FeSO₄ and EDTA was observed only at higher concentrations. The results presented in this study increase the knowledge on chitinase production in I. fumosoroseus opening new avenues for the study of the role of this enzyme in virulence against different insect pests during the infection process.     

Semicarbazide-Sensitive Amine Oxidase (SSAO) in the Brain

Semicarbazide-sensitive amine oxidase (SSAO) is widely distributed in almost tissues. However, its presence in brain microvessels is still controversial. The affinity of SSAO towards benzylamine (Bz) is considerably higher than that of monoamine oxidase (MAO). SSAO plays a role in the toxicity of several environmental and endogenous amines. SSAO-mediated production of toxic aldehydes has been proposed to be related to pathophysiological conditions. The most potent of inhibition of SSAO in monkey brain was observed by tricyclic antidepressant drug imipramine, as compared to tetracyclic drug maprotiline or non-cyclic drug nomifensine. An endogenous SSAO modulator in rat brain cytosol after immobilization stress (IMMO) was found and that this inhibitor could be induced by IMMO. SSAO activity in rat brain might be regulated by the level of this inhibitor. Semicarbazide, a SSAO inhibitor, enhances the formation of OH products of efflux/oxidation due to 1-methyl-4-phenylpyridinium ion (MPP⁺). The precise physiological functions of SSAO could play an important role in the control of energy balance in adipose tissue. SSAO could play an important role in the regulation of adipocyte homeostasis.     

Plasma methylglyoxal and glyoxal are elevated and related to early membrane alteration in young, complication-free patients with Type 1 diabetes

The reactive aldehydes methylglyoxal and glyoxal, arise from enzymatic and non-enzymatic degradation of glucose, lipid and protein catabolism, and lipid peroxidation. In Type 1 diabetes mellitus (T1DM) where hyperglycemia, oxidative stress, and lipid peroxidation are common, these aldehydes may be elevated. These aldehydes form advanced glycation end products (AGEs) with proteins that are implicated in diabetic complications. We measured plasma methylglyoxal and glyoxal in young, complication-free T1DM patients and assessed activity of the ubiquitous membrane enzyme, Na⁺/K⁺ ATPase. A total of 56 patients with TIDM (DM group), 6–22 years, and 18 non-diabetics (ND group), 6–21 years, were enrolled. Mean plasma A1C (%) was higher in the DM group (8.5 ± 1.3) as compared to the ND group (5.0 ± 0.3). Using a novel liquid chromatography-mass spectrophotometry method, we found that mean plasma methylglyoxal (nmol/l) and glyoxal levels (nmol/l), respectively, were higher in the DM group (841.7 ± 237.7, 1051.8 ± 515.2) versus the ND group (439.2 ± 90.1, 328.2 ± 207.5). Erythrocyte membrane Na⁺/K⁺ ATPase activity (nmol NADH oxidized/min/mg protein) was elevated in the DM group (4.47 ± 0.98) compared to the ND group (2.16 ± 0.59). A1C correlated with plasma methylglyoxal and glyoxal, and both aldehydes correlated with each other. A high correlation of A1C with Na⁺/K⁺ ATPase activity, and a regression analysis showing A1C as a good predictor of activity of this enzyme, point to a role for glucose in membrane alteration. In complication-free patients, increased plasma methylglyoxal, plasma glyoxal, and erythrocyte Na⁺/K⁺ ATPase activity may foretell future diabetic complications, and emphasize a need for aggressive management.     

Studies on semicarbazide-sensitive amine oxidase in patients with diabetes mellitus and in transgenic mice

Patients with diabetes mellitus and with vascular complications in particular, exhibit higher plasma activities of semicarbazide-sensitive amine oxidase (SSAO) compared to control subjects. It has been speculated that production of cytotoxic products of SSAO may cause endothelial damage and thus contribute to the development of diabetic vascular complications such as retino-, nephro-, and neuropathies as a result of SSAO activity.In order to explore the possibility that high SSAO activity contributes to the development of vascular complications in diabetes, we have performed two studies in patients with Type-2 diabetes quantifying plasma SSAO activity, HbA(1c), and urinary levels of the SSAO substrate, methylamine. We also examined the prevalence of retinopathy in these patients. Additionally, we have studied a model of transgenic mice expressing human SSAO in smooth muscle cells. The transgenic mice have an increased SSAO activity as well as mRNA expression. Histological studies revealed a specific aorta phenotype with a condensed and rigid vessel wall in some of the transgenic mice. No wild-type animals displayed this phenotype.In conclusion, we suggest that this transgenic mouse model may be of great value for increasing the knowledge about to what extent human SSAO contributes to vascular complications in diabetes, and also to which extent inhibition of SSAO can prevent the development of such complications.     

Synthetic polyamines as potential amine oxidase inhibitors: a preliminary study

In the last few decades, medicinal chemists have carried out extensive research on synthetic polyamines for use as anticancer drugs and multitarget-directed ligands in neurodegenerative diseases. The aim of this study was to evaluate the effect of some synthetic polyamines as inhibitors of two new potential targets, human semicarbazide-sensitive amine oxidase/vascular adhesion protein-1 (SSAO/VAP-1) and monoamine oxidases B (MAO B), enzymes involved in various multi-factorial diseases such as Alzheimer's disease. N,N'-Dibenzyl-dodecane-1,12-diamine (Bis-Bza-Diado), a newly synthesised compound, and ELP 12, a muscarinic cholinergic M(2) receptor antagonist, were found to behave as reversible and mixed non-competitive inhibitors of both amine oxidases (dissociation constants of about 100 μM). ELP 12 was found to be more selective for SSAO/VAP-1. Combining kinetic and structural approaches, the binding mode of ELP 12 to SSAO/VAP-1 was investigated. ELP 12 may bind at the entrance of the active site channel by ionic interactions with ASP446 and/or ASP180; one end of the polyamine may be accommodated inside the channel, reaching the TPQ cofactor area. The binding of ELP 12 induces rearrangement of the secondary structure of the enzyme and impedes substrate entry and/or product release and catalysis. These structural data reveal that the entrance and the first part of the SSAO/VAP-1 channel may be considered as a new target area, or a "secondary binding site", for modulators of human SSAO/VAP-1 activity. These results indicate ELP 12 and Bis-Bza-Diado as new "skeletons" for the development of novel SSAO/VAP-1 and MAO B inhibitors.     

Diabetes and semicarbazide-sensitive amine oxidase (SSAO) activity: a review

The enzyme of semicarbazide-sensitive amine oxidase (SSAO) activity has been reported to be elevated in blood from diabetic patients. SSAO are widely distributed in plasma membranes of various tissues and blood plasma. SSAO-mediated production of toxic aldehydes has been proposed to be related to pathophysiological conditions. Cytotoxic metabolites by SSAO may cause endothelial injury and subsequently induce atherosclerosis. The precise physiological functions of SSAO could play an important role in the control of energy balance in adipose tissue. It is possible that the increased SSAO activity in diabetes may be a result of up-regulation due to increase of SSAO substrates, such as methylamine or aminoacetone. SSAO could play an important role in the regulation of adipocyte homeostasis. Inhibition of SSAO could be of therapeutic value for treatment of diabetic patient.     

Reduction of Visceral Adipose Tissue and Improvement of Metabolic Indices: Effect of Dexfenfluramine in NIDDM

Increased visceral adipose tissue is thought to contribute to impaired glucose tolerance. We studied 10 men with non-insulin dependent diabetes (NIDDM) before and after a 12-week intervention study using dexfenfluramine. Subjects had a mean body mass index (BMI) of 26.4 ± 1.7 kg\m² and had an abdominal distribution of body fatness (waist-to hip ratio >0.9). Anthropometric indices, biochemistry, macronutrient intake from 7-day food records as well as a euglycaemic glucose clamp and magnetic resonance imaging (MRI) were performed at week 0 and week 12. Abdominal adipose tissue area measured by MRI was reduced from 854 ± 270 cm² to 666 ± 231 cm² (p=0.003) due mainly to a selective 32% reduction in visceral fat area from 484 ± 230 cm² to 333 ± 72 cm² (p=0.002). Insulin sensitivity improved from 0.29 ± 0.13 [min^?1 (mU/L)] to 0.54 ± 0.21 [min^?1 (mU/L)] (p=0.01) and C-peptide levels reduced from 0.77 ± 0.24 μmol/L to 0.58 ± 0.15 μmol/L (p=0.002). The reductions in fasting glucose and glycated haemoglobin failed to achieve significance. Fasting total cholesterol and triglyceride levels significantly reduced (p=<0.001 and p=0.021 respectively). There was a reduction in total energy intake (p=0.005) due to a significant reduction in calories obtained from fat (p<0.001). Thus dexfenfluramine was shown to be a useful adjunct therapy for the reduction of visceral fat in abdominally-obese men with NIDDM with an associated improvement in insulin sensitivity.