Hydrolase activities in the rat aorta. I. Effects of diabetes mellitus and insulin treatment.

Vascular disease in diabetics could arise in part from altered vessel wall catebolism. Specific activities of hydrolases in aortic smooth muscle cells from rats with streptozotocin-induced diabetes were measured. Enyzmes included: neutral alpha-glucosidase, alpha-mannosidase, and lysosomal N-acetyl beta-glucosaminidase, beta-galactosidase, cathepsin C, acid alpha-glucosidase, and acid cholesteryl esterase. After 4,8, and 11 weeks of diabetes, activities of all enzymes studied were decreased significantly in diabetic vessels, decreases ranging from 15% for cathepsin C to 62% for alpha-mannosidase. After 3 weeks of diabetes, insulin treatment for 1 week restored enzyme levels to normal. After 7 weeks of diabetes, 1 week of insulin treatment did not restore enzyme levels fully to normal (acid cholesteryl esterase was unchanged); 4 weeks of insulin did. Acid phosphatase and N-acetyl beta-glucosaminidase activities were reduced markedly in histochemical studies of diabetic aortas at all time periods and were restored by insulin treatment. Alloxan-induced diabetes gave results similar to those with streptozotocin. Significant decreases of aortic hydrolase activities, including those of lysosomes, occur in experimental diabetes mellitus and could contribute to accumulation of substrates in vascular smooth muscle cells.     

Hydrolase activities in the rat aorta. II. Effects of hypertension alone and in combination with diabetes mellitus

Hypertension is an important risk factor for atherosclerosis and often occurs in association with diabetes mellitus. Specific activities of hydrolases in homogenates of aortas from rats with renal-clip hypertension, normotension following a period of hypertension, and hypertension combined with streptozotocin-induced diabetes mellitus were measured. Enzymes included: neutral alpha-glucosidase, and lysosomal N-acetyl-beta-glucosaminidase, beta-galactosidase, cathepsin C, acid alpha-glucosidase, and acid cholesteryl esterase. After 6 or 12 weeks of hypertension, specific activities of all enzymes measured were significantly increased, levels ranging from 24% above normal for cathepsin C to 351% above normal for N-acetyl-beta-glucosaminidase. Six weeks of normotension following 6 weeks of hypertension resulted in restoration to normal of four of the six enzyme activities; the remaining two enzymes were significantly below normal levels. Combined hypertension and diabetes mellitus showed smooth muscle cell levels of four of the five hydrolases measured to be significantly lower than those present with hypertension alone. In every instance, histochemical studies of aortas showed acid phosphatase and N-acetyl-beta-glucosaminidase activities which corresponded to the biochemical findings. These findings indicate profound and discrete effects of two clinical risk factors on vascular smooth muscle cell lysosomes.     

Influence of diabetes on enzyme activities in rat aorta

The activities of hexokinase, glucose-6-phosphate dehydrogenase, hydroxyacyl-CoA-dehydrogenase, adenylate kinase and glutathione reductase were determined in the aorta of rats made diabetics with streptozotocin for over two weeks and in noninjected controls. Adenosinetriphosphate (ATP) and total adenine nucleotide content were also measured. Glutathione reductase activity was not significantly changed in the diabetic aorta whereas the activities of glucose-6-phosphate dehydrogenase, hydroxyacyl-CoA-dehydrogenase and adenylate kinase were all increased. Hexokinase activity was significantly decreased in diabetic rat aorta. When measured after incubation in vitro for 2 h with 5.6 mmol/l glucose, the ATP-concentration was reduced in the diabetic aorta while the total concentration of adenine nucleotides was unchanged. Insulin treatment started three days after induction of diabetes with streptozotocin and continued for twelve days restored the growth rate of the rats but their glucose metabolism was not completely normalized. After insulin treatment no significant differences between diabetic and normal rats were found in the aortic activities of glucose-6-phosphate dehydrogenase, hydroxyacyl-CoA-dehydrogenase, adenylate kinase or in the ATP content.     

Demonstration of muscular activity in the human aorta. Effects of trinitrin

The human thoracic aorta is usually considered to be a purely elastic vessel. Transoesophageal echo-cardiography (TEE) provides a new approach to study the mechanical properties of the descending aorta. The aim of the study was to evaluate the reproducibility and accuracy of M mode recordings of the human descending thoracic aorta and to appreciate the changes produced by an infusion of glyceryl trinitrate (GTN). The reproducibility of M mode recordings was studied in vitro on plexiglass tubes of different calibre, and also in vivo, the inter and intra-observer error was estimated to 0.6% in vitro and 1% in vivo. The accuracy of the method was evaluated in vitro by comparing the measured values (MV) with the actual diameters of the plexiglass tubes (T): MV = 1.012T + 0.9; r = 0.99; SE = 0.8 mm. The systolic and diastolic diameters of the descending thoracic aorta were measured in 8 healthy volunteers by TEE, before and during continuous intravenous infusion of GTN at a rate of 0.9 mg/hr and then 1.35 mg/hr after a 10 minutes interval. Systolic and diastolic blood pressures were recorded automatically every minute. The results showed a very significant increase in the systolic (from 20.3 +/- 0.7 to 20.9 +/- 1.3 and 22.1 +/- 2.2 mm) and diastolic diameters of the aorta (from 18.3 +/- 0.7 to 19.1 +/- 1.4 and 20.1 +/- 2.4 mm) despite a fall in systolic blood pressure (from 121.3 +/- 7.7 to 114.5 +/- 6.6 and 108.4 +/- 5.4 mmHg). This study shows that TEE is a reliable and reproducible method of measuring the diameter of the human descending thoracic aorta.(ABSTRACT TRUNCATED AT 250 WORDS)     

老年大鼠主动脉对胰岛素敏感性的改变及其机制

目的观察大鼠主动脉对胰岛素敏感性的增龄改变并分析其可能机制。方法老年组采用老年（18月龄）SD大鼠20只,随机选取成年（15周龄）SD大鼠20只为对照组。采用离体血管灌流技术,观察胸主动脉对胰岛素反应性的变化,并同时测定两组主动脉血管一氧化氮（NO）释放量及血管一氧化氮合酶（eNOS）活性;免疫组织化学法及Western Blot法检测老年组及成年组胸主动脉eNOS的蛋白表达变化。结果胰岛素可以浓度依赖性舒张成年大鼠胸主动脉,舒血管作用具有内皮依赖性。与之相比,老年大鼠主动脉对胰岛素的舒张反应显著下降（P<0.05）。同时发现,与成年组相比,老年组NO释放量以及eNOS活性显著降低（P<0.05）,免疫组织化学染色显示老年组主动脉eNOS的蛋白表达显著降低;而Western Blot检测发现老年组血管eNOS磷酸化水平显著降低（P<0.05）。结论血管组织内源性eNOS-NO系统活性下降可能是衰老导致的血管胰岛素敏感性下降的重要机制。     

Effects of exercise training and its cessation on components of the insulin resistance syndrome in obese children.

OBJECTIVE: To determine the effect of exercise training (ET) on components of the insulin resistance syndrome (IRS) in obese children. DESIGN: Randomized, modified cross-over study, with subjects assigned to one of two conditions: (1) 4 months of ET followed by 4 months of no-ET; or (2) 4 months of no-ET followed by 4 months of ET. Measurements were made at three time points: 0, 4 and 8 months. SUBJECTS: 79 obese, but otherwise healthy children (age: 7-11 y, percent fat (%fat) 27-61%). MEASUREMENTS: Plasma lipid and lipoprotein concentrations, plasma insulin and glucose concentrations; %fat; submaximal heart rate (HR) as an index of fitness. EXERCISE TRAINING: ET was offered 5 d/week 40 min/d. For the 73 children who completed 4 months of ET, the mean attendance was 80% (that is, 4 d/week) and the average HR during ET was 157 bpm. RESULTS: Significant (P < 0.05) group x time interactions were found for plasma triglyceride (TG) and insulin concentrations and %fat. The average change for both groups, from just before ET to just after the 4 month ET was -0.24 mmol.l-1 for TG, -25.4 pmol.l-1 for insulin and -1.6 units for %fat. When Group 1 ceased ET, over the following 4 month period the average change for insulin was +26.6 pmol.l-1 and for %fat +1.3 units. CONCLUSION: Some components (plasma TG, insulin, %fat) of the IRS are improved as a result of 4 months of ET in obese children. However, the benefits of ET are lost when obese children become less active.     

Triglyceride lipase activity in bovine aorta.

The cell-free supernatant from homogenized bovine aorta hydrolyzed triglycerides, beta-naphthylesters of lauric and stearic acid and Tween 20, 40 and 60. The rate of hydrolysis decreases as the acyl chain length of the substrates increases. The activity against triglycerides of short-chain fatty acids and monoacylesters could be partially separated from that of glycerol-trioleate lipase by ammonium sulfate fractionation. The activity of glycerol-trioleate lipase remained unaffected by heating for 5 min at 60 degrees C or by addition of bile acids, whereas the activity causing hydrolysis of triglycerides with short-chain fatty acids and monoacylesters decreases up to 60% by analogous treatment.     

Effects of cigarette smoking and its cessation on body weight and plasma leptin levels.

Smokers weigh less than age-matched nonsmokers, and most smokers gain weight after smoking cessation due to an increase in food intake and a decrease in energy expenditure. Leptin is an endocrine signal thought to regulate body fat stores through hypothalamic control of energy intake and expenditure. To determine whether the "weight-reducing" effects of smoking may be mediated by leptin, we measured plasma leptin concentrations in 22 middle-aged and older male smokers (body mass index [BMI], 28 +/- 1 kg/m2, mean +/- SEM) and 22 nonsmokers matched to the smokers for age (64 +/- 1 years) and BMI (28 +/- 1 kg/m2). The body weight and leptin concentration were remeasured at 3 and 6 months in 13 of the smokers who successfully stopped smoking. The leptin concentration correlated positively with the BMI in both smokers (r = .74, P < .001) and nonsmokers (r = .76, P < .001). However, the intercept of the regression line was higher for smokers versus nonsmokers (P < .05), with no difference in the slope. Thus, male smokers have a higher leptin level for a given BMI than nonsmokers. Following 6 months of smoking cessation, body weight increased by 7% (6.0 +/- 0.1 kg, n = 13, P < .01). Despite this weight gain, the mean leptin concentration did not increase with smoking cessation. On average, leptin levels were 25% lower than would be expected for the amount of weight gained after smoking cessation. These findings suggest that cigarette smoking directly elevates circulating plasma leptin concentrations, and this increase may be one mechanism for the lower body weight of smokers compared with nonsmokers.     

Effects of hypertension on the intercellular contacts between smooth muscle cells in the rat thoracic aorta.

A qualitative and quantitative study was made of the contacts between muscle cells in the media of the thoracic aorta of hypertensive and normotensive Sprague-Dawley rats. In ultrathin sections, the type and number of contacts per 100 microns cell perimeter and per 100 cell profiles were determined using an image analysis computer. The intercellular contacts in both groups were in the form of simple appositions, interdigitations, intermediate junctions and nexus junctions. In the hypertensive group, there was a reduction in the number of simple appositions and intermediate junctions, but the nexus junctions (which provide intercellular communication) were increased in density. Interdigitations demonstrated no change. Decreased intercellular cohesion may play an important role in the mechanisms by which hypertension predisposes degeneration of the media with progression to ectasia, aneurysm formation and dissection.     

Eicosanoid-dependent and endothelium-independent oscillations of rat aorta.

We studied the role of endothelium and eicosanoids in rhythmic contractions of rat thoracic aorta. Spontaneous oscillations were observed in 35% of 385 endothelium-intact and in 46% of 22 endothelium-denuded aortic strips from normotensive Sprague-Dawley male rats. Vasoactive agents (norepinephrine, epinephrine, phenylephrine, isoproterenol, arachidonic acid, PGF2 alpha, serotonin, potassium, endothelin, atrial natriuretic factor and angiotensin II) induced rhythmic contractions in the majority of tissues. Rhythmic activity was also observed in aortic strips from adult female, pregnant and old male rats. Aortic oscillations were partially inhibited by indomethacin, ibuprofen and nordihydroguaiaretic acid (NDGA), and completely inhibited by indomethacin plus NDGA, nifedipine, low external calcium (less than 1 mM) and pretreatment with dexamethasone. Indomethacin, NDGA and arachidonic acid did not affect oscillations of the portal vein. Rhythmic contractions were observed in thoracic aortic strips from neonatal but not from adult rabbits. However, oscillations could be induced in strips of the mesenteric artery and terminal abdominal aorta of adult rabbits. Also, adult rabbit thoracic aortic strips exhibited oscillations when set up in close proximity of rat aorta. It is suggested that rhythmic contractions are physiological characteristics of many and perhaps all blood vessels and may play a role in blood flow and turbulence; the likely cause of these oscillations is the cyclic release of one or more eicosanoids.     

Static mechanical properties of the developing and mature rat aorta.

The static mechanical properties of the aorta have been examined in rats, aged between 4 weeks and 2 years. Internal radius, relative wall thickness, circumferential incremental strain, and circumferential incremental elastic modulus have been measured at pressures between 1.33 and 33.3 kPa (10 and 250 mm Hg). The results show that the value of the incremental elastic modulus does not change after 12 weeks. Changes observed in younger animals are related to alterations in relative wall thickness. The findings correlate with previously observed chemical and morphological changes. Studies of muscle function indicate that the smooth muscle of the media does not contribute significantly to the static elastic properties of the vessel wall.     

Effects of melittin on endothelium-dependent relaxation and cyclic GMP levels in rat aorta

The present study investigates the mechanism of endothelium-dependent relaxation of vascular smooth muscle. Melittin, a polypeptide found in honeybee venom and a known activator of phospholipase A2, induced transient, endothelium-dependent relaxations of rat thoracic aortae contracted with norepinephrine. Higher concentrations of melittin induced relaxations followed by contractions. Prior incubation of melittin with trypsin abolished the changes in relaxation and contraction due to melittin. Melittin (10 micrograms/ml)-induced relaxations were associated with transiently elevated levels of cyclic GMP with a peak increase of 30-fold, which occurred 30 seconds after melittin exposure. Melittin (10 micrograms/ml) elevated cyclic AMP levels less than twofold and this effect was variable. A lower concentration of melittin (1 microgram/ml) elevated cyclic GMP levels approximately twofold, while exposure to 1 microgram/ml melittin in the presence of the cyclic GMP phosphodiesterase inhibitor, M&B 22948 (1 mM), increased cyclic GMP levels fivefold. Removal of the endothelium prevented the increased levels of cyclic GMP and cyclic AMP due to melittin. Exposure to the guanylate cyclase inhibitor, methylene blue, prevented the increased levels of cyclic GMP. Methylene blue, nordihydroguaiaretic acid, and the phospholipase A2 inhibitor, parabromophenacyl bromide, inhibited melittin-induced relaxations, while the cyclo-oxygenase inhibitor, indomethacin, was without effect. Arachidonic acid increased cyclic AMP levels but had no effect on cyclic GMP levels in the presence or absence of indomethacin. Relaxations to melittin, and to the endothelium-dependent vasodilators acetylcholine, trypsin, histamine, and the Ca2+ ionophore A23187, and/or the associated increased cyclic GMP levels, were reduced following exposure to melittin. Prior exposure to polyarginine (10 micrograms/ml), which induced endothelium-dependent relaxations that were prevented by methylene blue, also inhibited relaxations to the endothelium-dependent vasodilators. In contrast, relaxations to sodium nitroprusside were potentiated in tissues previously exposed to melittin. Removal of the endothelium by rubbing the intimal surface also potentiated relaxations to sodium nitroprusside. Scanning electron micrographs of the intimal surface demonstrated that melittin and polyarginine greatly damaged the endothelial cells. The present results suggest that polycation containing peptides induce endothelium-dependent relaxation through elevation of cyclic GMP levels within the smooth muscle.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effects of high-cholesterol diet on the interendothelial clefts and the associated junctional complexes in rat aorta

The arterial endothelial intercellular cleft (AEC) and its associated junctional complex (JC) are the determinants of permeability to macromolecules. This study analyzed frequencies of AEC and JC profile types in the rat thoracic aorta at 1 and 12 months after feeding the animals with a normal or a high-cholesterol diet. Rats on either a normal diet or high-cholesterol diet for 12 months showed more of the simple 'end to end' or 'overlap' types (P < 0.01) but fewer complex 'interdigitating' type (P < 0.01) of AEC compared to the 1 month group. With regard to JC, the frequencies of gap junctions were decreased (P < 0.01) while the tight junctions and the normal junctionless complex were increased (P < 0.01) after 12 months of normal diet as compared with 1 month on the normal diet. These changes in frequencies for gap junction and tight junction were even greater for the high-cholesterol diet than for the normal diet treatment. Moreover, the incidence of open junctions was also noticeably increased after 12 months of high-cholesterol diet. These findings suggest that the proportions of the AEC and JC were highly responsive to aging whereas those of JC were more susceptible to the high-cholesterol diet treatment.     

Distribution of the activity of the angiotensin-converting enzyme in the rat aorta and changes in the activity with aging and by the action of l-NAME

The activity of the angiotensin-converting enzyme (ACE) of the inner surface (the endothelium surface) of rat aorta sections has been studied depending on their distance from the aortic arch, age of rats, and the duration of treatment of rats with the NO synthase inhibitor, N_ω-nitro-l-arginine (l-NAME). The activity of ACE of aorta sections was determined by measuring the hydrolysis of hippuryl-l-histidyl-l-leucine and was expressed as picomoles of Hip–His–Leu hydrolyzed per minute per square millimeter of the endothelium surface. It was found that the ACE activity considerably varies along the aorta of young rats. This variability decreases with increasing age of rats and by the action of l-NAME. The average ACE activity in the aorta increases with the age of rats and with increasing time of l-NAME treatment. Enalapril normalizes the distribution of the ACE activity along the aorta and decreases the average ACE activity. The changes in the distribution of the ACE activity along the aorta and in the average ACE activity in the aorta with increasing age of the rat and by the action of l-NAME may play a role in the development of atherosclerosis of vessels on aging and the inhibition of formation of nitric oxide.     

Endothelium-independent vasorelaxant activity of Trachyspermum ammi essential oil on rat aorta

Several pharmacological activities of the essential oil of Trachyspermum ammi seeds (TAEO) have been previously studied. These include antitussive, antihypertensive, and antispasmodic effects. However, its action on isolated aorta has not yet been studied. This study was aimed to investigate the vasorelaxant activity of TAEO and characterize its mechanism of action. Extraction of TAEO was performed using Clevenger-type apparatus with the final content of 4.5% (v/w). To evaluate some probable mechanisms of action of TAEO, the action isometric tension was then measured in the aortic rings from Wistar rats which were precontracted with phenylephrine (PHE) (1 µM) or KCl (60 mM). The major constituents of TAEO included Thymol (38.1%), gamma-terpinene (33.3%), and p-cymene (23.1%), as was analyzed by GC-MS. The cumulative concentrations of TAEO reduced precontraction caused by PHE and KCl (p < 0.05) significantly, which was dose dependent. The vasorelaxation caused by TAEO was not influenced in the presence of methylene blue and L-NAME in the endothelium-intact and denuded aorta ring. The inhibitory effect of TAEO on the aortic rings precontracted with KCl and PHE was considerably reduced by nifedipine. These findings hypothesized that the vasorelaxation caused by TAEO is completely endothelium independent and the extracellular Ca²⁺ influx was also inhibited by TAEO.