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Background: The catalytic activity of semicarbazide-sensitive amine oxidase (SSAO) is increased in diabetes, as well as in other disorders of cardiovascular origin. Our hypothesis is that SSAO is involved in the synthesis or maturation of elastin in vascular tissue. An increased SSAO activity can thereby be involved in the development of vascular damage., Methods: Elastin quantification was performed in aorta of transgenic mice overexpressing the human form of SSAO, using electron microscopy. Furthermore, lung capacity was measured using a spirometry-mimicking method, developed for mice. The effect of vasoactive substances was estimated by measuring mean arterial pressure and pulse pressure under anesthesia., Results: No differences in elastin quantity or lung capacity could be observed between transgenic or nontransgenic littermates. Pulse pressure was higher in transgenic mice, and electron microscopy of aorta showed elastin fibers parallel with the aorta wall (ie, straight fibers instead of folded compared with control mice). No difference in the response to adrenaline or sodium chloride was observed between the transgenic and control mice. The control mice had a clear decrease in blood pressure (BP) with a longer duration as a response to injection of a nitric oxide (NO) donor, sodium nitroprusside, compared with transgenic mice where only a minor response was observed. The SSAO activity in serum of control mice was elevated in response to injection of the NO donor, but not in response to a ganglion blocker., Conclusions: An elevated pulse pressure, together with an abnormal elastin structure in the aorta, suggests a rigidity of large arteries as a result of an elevated SSAO activity as well as a physiologic role for SSAO in elastin maturation.

We have previously reported that carp (Cyprinus carpio) tissue mitochondria contain a novel form of monoamine oxidase (MAO), which belongs neither to MAO-A nor to MAO-B of the mammalian enzyme. This conclusion results from the findings that the carp MAO was equally sensitive to a selective MAO-A inhibitor clorgyline and to the MAO-B selective inhibitor l-deprenyl, when tyramine, a substrate for both forms, serotonin or beta-phenylethylamine, a substrate for either A or B-form of mammalian MAO, was used. In the present study, we tried to detect another amine oxidase, termed tissue-bound semicarbazide-sensitive amine oxidase (SSAO), activity in carp tissues. As definition of SSAO was used, such as insensitivity to inhibition of the kynuramine oxidizing activity by an MAO inhibitor pargyline and high sensitivity to the SSAO inhibitor semicarbazide. The results indicated that the oxidizing activity was selectively and almost completely inhibited by 0.1 mM pargyline alone or a combination of 0.1 mM pargyline plus 0.1 mM semicarbazide, but not by 0.1 mM semicarbazide alone. We also tried to detect any SSAO activity by changing experimental conditions, such as lower incubation temperature, higher enzyme protein concentration, a lower substrate concentration and different pH's in the reaction, as the enzyme source. However, still no SSAO activity could be detected in the tissues. These results conclusively indicate that carp tissues so far examined do not contain SSAO activity.

Semicarbazide-sensitive amine oxidase (SSAO) activity in plasma is increased in diabetes, and in particular, in diabetic patients with vascular complications. It has been speculated that SSAO is involved in the development of such complications due to the production of cytotoxic compounds. In this work, we have induced diabetes in a previously described mouse-model, overexpressing SSAO in smooth muscle cells. SSAO activity was estimated as well as expression of the endogenous mouse gene and human transgene using real-time PCR. Diabetes induced an increase in SSAO activity in serum, kidney, and adipose tissue of transgenic animals. An inverse correlation between SSAO activity and mouse SSAO mRNA levels was observed in transgenic animals with diabetes. These results further support the suggestion of a negative feedback control of the SSAO gene expression. The increased SSAO activity in diabetes is most likely dependent on post-transcriptional modifications or activation of existing inactive enzyme molecules.

Semicarbazide-sensitive amine oxidase (SSAO) is an enzyme associated with vascular systems in mammals. SSAO catalyzes the deamination of primary monoamines and has been suggested to be a risk factor in vascular disorders, e.g., diabetic vascular complications. The primary aim of the present study was to investigate if serum SSAO activity is associated with clinical parameters in non-small cell lung cancer (NSCLC) patients. Secondary aims were to investigate if there is a correlation between SSAO activity and the angiogenic factors vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF).Thirty-three patients donated 231 serum samples. Detectable levels of bFGF, VEGF, and SSAO were observed in all patients. Serum SSAO activity was not statistically associated with survival (p = 0.35). A highly significant statistical correlation was found between SSAO activity and VEGF (p < 0.0001). No significant correlation between SSAO and bFGF was observed. We conclude that SSAO was not associated with survival in patients with NSCLC. However, a strong correlation between serum SSAO activity and the angiogenic factor VEGF was found that might implicate new aspects of the mechanisms controlling angiogenesis.

The inhibitory effects of 2-bromoethylamine (2-BEA), a derivative of ethylamine, on guinea pig lung semicarbazide-sensitive amine oxidase (SAO) have been studied. Preincubation with 2-BEA time-dependently inhibited SSAO activity. The mode of the initial phase of inhibition was competitive, with a Ki value of 52 microM. After preincubation at 37 degrees C for 2 h, the inhibition was noncompetitive and irreversible, as there was no recovery of SSAO activity by dilution of the inhibited samples. Kinetic analyses confirmed previous results with rat lung SSAO that 2-BEA is a suicide SSAO inactivator with a dissociation constant of 42 microM. This latter value is similar to that of the Ki value (52 microM) for the reversible phase of inhibition by 2-BEA. Addition of the nucleophilic compound 2-mercaptoethanol could not reduce the SSAO inhibition, indicating that inactivation could not be prevented by trapping the enzymatic reaction product from 2-BEA. This finding clearly indicates that the reaction product should not diffuse away from its site of genesis and agrees with one of the characteristics of suicide inhibitors. This conclusively excludes the possibility of an affinity-labeling mechanism., ((c) 2003 Prous Science)

Elevated semicarbazide-sensitive amine oxidase (SSAO) activity has been observed in several human conditions, eg, diabetes, and it has been speculated that SSAO contributes to the development of vasculopathies associated with this disease. To investigate in vivo consequences of elevated expression of SSAO in vascular tissues, we have developed a transgenic model for overexpression of human SSAO in mice. A smooth muscle-specific promoter, smooth muscle alpha-actin promoter 8 (SMP8) was used. Transgenic expression of human SSAO in tissues with a high content of smooth muscle cells was confirmed by Northern blot analysis. Enzymatic analysis of homogenates from transgenic tissues showed elevated levels of SSAO activity compared to non-transgenic littermates. Furthermore, when plasma SSAO activity was analyzed, much higher activity was detected compared to plasma from control mice, indicating that plasma SSAO may originate from smooth muscle cells. Histopathological evaluation of aorta and renal artery from transgenic mice revealed an abnormal structure of the elastin tissue. Instead of the regularly folded elastic laminae normally found in tunica media of sacrificed mice, the elastic laminae were straight and unfolded with irregularly arranged elastic fibers, forming tangled webs, between the intercalating elastic laminae. These alterations of the elastin structures suggest that overexpression of SSAO has led to a reduced elasticity of the arteries. Moreover, the mean femoral arterial pressure of the SMP8 SSAO transgenic mice was significantly lower in comparison to non-transgenic littermates. This suggests that the transgenic mice have a defect in their ability to regulate blood pressure.

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.

Background: Plasma activity of semicarbazide-sensitive amine oxidase (SSAO) has been reported to be significantly higher in diabetic patients compared to healthy controls. Due to the production of highly angiotoxic substances in SSAO-catalyzed reactions, it has been speculated that this could be a cause for the vascular complications frequently associated with diabetes. Little is known about how the enzyme activity is regulated, and why it is high in these patients. In the present study, we assessed the possibility of transcriptional regulation by analyzing SSAO activity and SSAO-mRNA levels in mice with alloxan-induced diabetes., Materials and Methods: Diabetes was induced in NMRI mice by a single intravenous injection of alloxan. The enzyme activity was analyzed by a radiometric assay using (14) C-benzylamine as a substrate, and the mRNA-levels were analyzed by real-time PCR., Results: We found that the enzyme activity was increased in lung and adipose tissue 1 day after induction, as the glucose levels start to rise. Seven days after the injection of alloxan, the activity in serum was increased, and this activity was positively correlated with blood glucose levels in the alloxan-treated animals. Although the enzyme activity was increased in adipose tissue as a result of the treatment, SSAO-mRNA levels in this tissue were decreased, possibly suggesting a negative feedback on the gene expression., Conclusions: The main conclusion from this study is that the increased enzyme activity observed in diabetes is not a result of increased SSAO gene transcription. We speculate that the enzyme activity is controlled by posttranslational modifications of the protein, and that the catalytic activity controls the gene expression.

Plasma activity of the enzyme semicarbazide-sensitive amine oxidase (SSAO) is high in diabetes. Production of angiotoxic substances (an aldehyde, hydrogen peroxide, and ammonia) in vessel walls is catalysed by SSAO, suggesting a role for SSAO in the development of complications of diabetes. The objective of the present study was to follow up plasma SSAO activity (measured radiometrically), HbA(1c) (using ion exchange chromatography), and retinopathy (by fundus photography) after 2.8 years, in 34 patients with Type 2 diabetes. We also measured urinary levels of an SSAO substrate, methylamine, by fluorometric high-performance liquid chromatography (HPLC). As at baseline, plasma SSAO activity was now higher in subjects with retinopathy (mean 19.5) than in subjects without retinopathy (mean 16.0), 95% confidence interval (CI) for difference 0.6-6.3 nmol benzylamine ml(-1) plasma h(-1). SSAO activity had not changed significantly since baseline, mean difference -1.65 and 95% CI for difference -3.76 to 0.46 nmol benzylamine ml(-1) plasma h(-1). Mean HbA(1c) level remained higher for patients with retinopathy (now 7.9%) compared to those without retinopathy (6.1%), 95% CI for difference 0.6-3.0%. Comparing baseline and the present study, retinopathy was nonproliferative; level had worsened for five and improved for two patients. Urinary methylamine/creatinine ratio was lower in the group of patients with retinopathy (mean 0.99) than in those without retinopathy (mean 1.78), 95% CI for difference 0.1-1.5 microg mg(-1). The results of the present study are compatible with a role for SSAO in the development of diabetic retinopathy.

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.

Various mammalian tissues contain plasma membrane-bound amine oxidase, termed semicarbazide-sensitive amine oxidase (SSAO). In the present study, 2-bromoethylamine has been studied with regard to inhibitory properties towards tissue-bound SSAO in rat lung. Without preincubation, 2-bromoethylamine was a competitive and reversible SSAO inhibitor with a Ki value of 2.5 microM. After preincubation, it time-dependently and non-competitively inhibited SSAO activity, probably by forming the covalently-bound enzyme-inhibitor adduct. The data presented suggest that 2-bromoethylamine may act as a suicide inhibitor of SSAO.

Under the influence of semicarbazide-sensitive amine oxidase (SSAO), methylamine is deaminated to formaldehyde, which can react with various macromolecules and form irreversible adducts. We hereby present an autoradiographic method of visualising SSAO activity by measuring the in vivo formation of such adducts from 14C-methylamine. Our results revealed high concentrations of radioactive deposits in the intestinal wall, brown adipose tissue, spleen and bone marrow. Hydralazine is a potent SSAO inhibitor and pretreatment with this irreversible inactivator resulted in a nearly complete loss of radioactive deposits in the tissues. By giving 14C-methylamine at different time-points after irreversible inhibition of SSAO, it was also possible to determine the resynthesis rate of SSAO. Interestingly, the recovery rate of SSAO after such inactivation was tissue-specific. The possible therapeutic value of a specific SSAO inhibitory drug has been discussed.

Semicarbazide-sensitive amine oxidases (SSAOs) are widely expressed copper-containing enzymes. One enzyme of this family have high specific activity towards benzylamine and is present in human blood plasma. This enzyme is altered in several diseases, for instance in diabetes. Presently it is unclear where the plasma SSAO is synthesized. Previous autoradiographic studies have suggested that SSAO may be expressed in bone tissue. In the current study we have analyzed levels of SSAO in serum from cases with 'skeletal disease', i.e. patients with severe skeletal metastases of prostate cancer and subjects having recent fractures. Interestingly, subjects with metastases showed significantly elevated levels of SSAO in serum compared to individuals having prostate cancer without skeletal metastases. It is speculated that, at least in part, SSAO in the blood stream may be derived from bone tissue.

The semicarbazide-sensitive amine oxidases constitute a group of copper-containing enzymes whose physiological function is unclear. The enzymes are present in various tissues, including blood plasma. At present, the source of the plasma enzyme in humans is not known. Results of a recent study suggested that semicarbazide-sensitive amine oxidase is expressed in the skeleton, e.g. in the spine. Using an indirect autoradiographic method in mice, we provide evidence that semicarbazide-sensitive amine oxidase is present in high abundance in bone tissue. Specific activities of semicarbazide-sensitive amine oxidase were estimated in blood samples from subjects with femoral bone fractures. Moreover, enzyme activities were also measured in patients suffering from prostate cancer with skeletal metastases. The level of specific semicarbazide-sensitive amine oxidase activity in serum was significantly elevated in patients with skeletal metastases compared with both healthy controls and patients having prostate cancer without signs of skeletal metastases. Based on the results of the present study, we propose that semicarbazide-sensitive amine oxidase in blood plasma may originate, at least in part, from the skeleton.

Aims: To measure plasma semicarbazide-sensitive amine oxidase (SSAO) activities and detect retinopathy in Type 2 diabetes mellitus (DM)., Methods: Cross-sectional, population-based study of 65 diabetes patients (61 diagnosed from the age of 30 years) with or without retinopathy as determined by fundus photography in primary care. HbA1c was analysed by ion exchange chromatography on a Mono S for HbA1c column. SSAO activities were assayed radiometrically and formaldehyde-albumin adducts by ELISA in plasma samples from patients and 136 healthy controls., Results: Subjects with diabetes had higher plasma SSAO activity, measured as nmol benzylamine x mlplasma(-11) x h(-1)(mean 20.6), than controls (mean 14.3), P<0.0001; 95% confidence interval (CI) for difference 4.9-7.7. SSAO activity was higher in patients with retinopathy (mean 23.2) than in those without (mean 18.9), P=0.012; 95% CI for difference 1.0-7.5, and related to the HbA1c value. No statistically significant relationship between diabetes duration and SSAO activity was found. With HbA1c values and insulin treatment entered into a multiple logistic regression model, SSAO activity no longer predicted retinopathy, P increasing from 0.025 to 0.17. SSAO activity and the presence of any retinopathy were unrelated to titres of antibodies against formaldehyde-treated human serum albumin., Conclusions: SSAO activity, earlier found to be elevated in Type 1 DM, is also elevated in Type 2 DM. The SSAO family of enzymes may be involved in the development of diabetic retinopathy, possibly by catalysing the formation of toxic metabolites. A potent and specific inhibitor of human SSAO might help prevent retinopathy in Type 1 and Type 2 DM.

Semicarbazide-sensitive amine oxidase (SSAO) has been suggested to be involved in the development of vascular endothelial damage. The source of the soluble form of SSAO found in the blood serum is unknown. However, it has been speculated that it is secreted from cells within the vascular wall where high activity of the enzyme is found. Altered SSAO activity has been reported in atherosclerotic plaques of the human aorta. Stroke is a manifestation of long-term atherosclerotic disease, and in this study, plasma SSAO activities were estimated in 42 patients with cerebral thrombosis and 26 patients with cerebral embolism, and compared to two control groups of 45 individuals in total. No statistically significant differences were found between the patient groups and controls regarding plasma SSAO activity, suggesting that changes in the soluble form of SSAO found in the circulation do not play a major role in this type of acute cerebrovascular event. Furthermore, it does not seem likely that the involvement of vascular tissue occurring in stroke results in release of the enzyme into the circulation. Nevertheless, further studies on tissue-bound SSAO in cerebral vessels would be of great interest.

The activity of semicarbazide-sensitive amine oxidase (SSAO) has been reported to be elevated in blood from diabetic patients. It has been suggested that the enzyme is involved in the development of complications such as retinopathies, nephropathies and neuropathies, which are associated with advanced diabetes, possibly by the formation of toxic metabolites. Under the influence of SSAO, methylamine is deaminated to formaldehyde which is known to react with various macromolecules. It has therefore been proposed that specific inhibition of SSAO could be of therapeutic value for treatment of diabetic patients. The present results provide evidence that treatment with an SSAO inhibitor potently reduces the levels of irreversible adducts. In this study, 14C-methylamine was given intraperitoneally to NMRI mice, and the tissue distribution of irreversibly bound methylamine metabolites was estimated by an autoradiographic method. Such radioactive residues occurred in high concentrations in the intestinal wall, brown adipose tissue, spleen and bone marrow. By inhibiting SSAO irreversibly with hydralazine before giving 14C-methylamine to the mice, it was possible to determine the resynthesis rate of SSAO in different tissues. A complete recovery of SSAO activity was seen in the intestinal wall after 6 days, whereas only about 60% was recovered in adipose tissue after 14 days. This suggests that factors controlling the synthesis of SSAO differ in these tissues, or that these tissues express different forms of enzymes.

The deamination of 5-hydroxytryptamine, tryptamine and benzylamine by porcine dental pulp membrane preparations is brought about not only by monoamine oxidase, but also by a clorgyline (and deprenyl) resistant), semicarbazide sensitive enzyme. The semicarbazide sensitive enzyme was also inhibited by aminoguanidine, hydroxylamine and phenylhydrazine, but was not affected to any significant extent by incubation at 50 degrees for up to 100 min. There was, on the other hand, considerable inhibition of monoamine oxidase activity after incubation at this temperature. The semicarbazide sensitive enzyme neither metabolised, nor was inhibited by putrescine or cadaverine. Mixed substrate experiments indicated that 5-hydroxytryptamine and tryptamine interacted at the same catalytic centre on the semicarbazide sensitive enzyme.