The activity of antioxidant enzymes and the content of uncoupling protein-1 in the brown adipose tissue of hypothyroid rats: comparison with effects of iopanoic acid

The activity of antioxidant enzymes, copper-zinc superoxide dismutase (CuZnSOD), manganese superoxide dismutase (MnSOD) and catalase (CAT), as well as that of the mitochondrial FAD-dependent alpha-glycerophosphate dehydrogenase (alpha-GPD) in the rat interscapular brown adipose tissue (IBAT) were studied after the treatment with methimazole (MMI) for three weeks or with iopanoic acid (IOP) for five days. Besides, the mitochondrial concentration of uncoupling protein-1 (UCP-1) and the activity of catecholamine degrading enzyme monoamine oxidase (MAO) in the IBAT as well as the activity of the catecholamine synthesizing enzyme, dopamine beta-hydroxylase (DBH) in rat serum were examined. Judging by the significantly enhanced level of serum DBH, which is an index of sympathetic activity, and that of IBAT MAO, the increase in MnSOD and CAT activities in the IBAT of hypothyroid (MMI-treated) rats seems to be due to elevated activity of sympathetic nervous system (SNS). However, CuZnSOD activity is not affected by SNS. On the contrary, IOP, which is a potent inhibitor of T4 deiodination into T3 producing "local" hypothyroidism, did not change either SNS activity or activities of IBAT antioxidant enzyme. However, both treatments significantly decreased IBAT UCP-1 content and alpha-GPD activity suggesting that the optimal T3 concentration in the IBAT is necessary for maintaining basal levels of these key mitochondrial parameters.     

Effect of fasting and refeeding on the activities of monoamine oxidase and antioxidant enzymes in rat hypothalamus and brown adipose tissue

Fasting for 48 h and the same period of recovery induced by 48 h refeeding increased rat hypothalamic monoamine oxidase (MAO) activity. However, in the interscapular brown adipose tissue (IBAT), only refeeding induced a significant elevation of the enzyme activity. As far as hypothalamic antioxidative enzymes are concerned, the copper zinc superoxide dismutase (CuZnSOD) activity was decreased in refed rats only. However, in the IBAT both food deprivation and refeeding induced a significant decrease in catalase (CAT) activity. Under the influence of fasting the adrenal glands were strongly activated as judged by the increased dopamine-beta-hydroxylase (DBH) activity and decreased cholesterol concentration. Refeeding brought both parameters to control levels indicating full recovery of these glands. As expected, fasting for 48 h induced a significant decrease in serum glucose but an increase in FFA concentrations. Thus, it can be concluded that both fasting and refeeding resulted in increased activation of hypothalamic MAO, whereas CuZnSOD activity was decreased only by refeeding. However, in the IBAT only refeeding increased MAO activity whereas both fasting and refeeding decreased that of CAT. In conclusion, it may be assumed that food deprivation for 48 h and the same duration of refeeding influenced MAO and antioxidative enzymes activities in the rat hypothalamus and IBAT in a tissue specific manner.     

Sympathetic and hyperthermic reactions by orexin A: role of cerebral catecholaminergic neurons

This experiment tested the effect of a lesion of cerebral catecholaminergic neurons on the sympathetic and thermogenic effects induced by an intracerebroventicular (icv) injection of orexin A. The firing rates of the sympathetic nerves to the interscapular brown adipose tissue (IBAT), along with IBAT, colonic temperatures and heart rate were monitored in urethane-anesthetized male Sprague-Dawley rats before an injection of orexin A (1.5 nmol) into the lateral cerebral ventricle and over a period of 150 min after the injection. Three days before the experiment, the rats were pre-treated with an icv injection of 6-hydroxydopamine (6-OHDA) or 6-OHDA plus desipramine or saline. The results show that orexin A increases the sympathetic firing rate, IBAT, colonic temperatures and heart rate in the rats pre-treated with saline. This increase is blocked by the pre-treatment with 6-OHDA alone or 6-OHDA plus desipramine. These findings indicate that cerebral catecholaminergic neurons (particularly the dopaminergic pathway) play a fundamental role in the complex reactions related to activation of the orexinergic system.     

BRAIN DOPAMINE-β-HYDROXYLASE: REGIONAL DISTRIBUTION AND EFFECTS OF LESIONS AND 6-HYDROXY-DOPAMINE ON ACTIVITY

Abstract— A modification of a specific and sensitive radioassay was used to measure dopamine-β-hydroxylase (DBH) (EC 1.14.2.1) in various regions of the rat CNS. Highest activity was found in the hypothalamus. Relative to activity in the hypothalamus (= 100 per cent), activity in brainstem was 80 per cent, in sensory motor cortex 55 per cent, in caudate nucleus 32 per cent, and in cervical spinal cord 30 per cent. Two to three weeks after a unilateral electrolytic lesion of the lateral hypothalamus, activity of DBH in the ipsilateral cerebral cortex fell to 17 per cent of control values without changes in activity ipsi- or contra-laterally in the brainstem. Thalamic lesions did not affect DBH activity. In cerebral cortex contralateral to the hypothalamic lesion, enzymic activity rose 30 per cent. After intracisternal administration of 6-hydroxy-dopamine (6-OH-DA), cortical DBH activity fell to 20 per cent of control values. Reserpine (3 mg/kg subcutaneously for 3 days) did not increase the activity of DBH in brain regions but did increase the activity of DBH in adrenal gland 200 per cent. Our results suggest that: (a) DBH is widely distributed in neurons in CNS with a regional pattern of activities that appears to parallel the Jevels of norepinephrine; (b) DBH activity in the cerebral cortex depends on the integrity of structures (e.g. medial forebrain bundle) in lateral hypothalamus; (c) DBH in brain areas lacking cell bodies of nore- pinephrine-neurons (e.g. cerebral cortex) is contained in norepinephrine-containing axon terminals and (d) the activity of DBH in brain is not increased by reserpine under conditions that provoke marked increase of DBH activity in the adrenal gland.     

A study of selected catecholamine metabolizing enzymes: a comparison of depressive suicides and alcoholic suicides with controls

—The regional distributions of monamine oxidase (MAO) (EC 1.4.3.4), catechol-O-methyltransferase (COMT) (EC 2.1.1.6), tyrosine hydroxylase (TH) (EC 1.14.3.2), and dopamine-β-hydroxylase (DBH) (EC 1.14.2.1) have been examined in human brains obtained at autopsy from persons who died of natural causes (controls), and from persons who committed suicide and were further categorized as suffering from affective disorder (depression) or from alcoholism. Post mortem animal studies showed no changes in MAO or COMT activities in rabbit brain or in DBH activity in rat brain when the intact bodies were left at room temperature up to 24 h. TH activity in rabbit brains, however, began to decline immediately after death and after 24 h at room temperature it was approximately 48 per cent of the fresh brain level. There was no significant variation in activity of COMT, TH and DBH in human brain attributable to age or sex. MAO activities in the 60–70 yr decade were 34 per cent higher than in the 30–40 yr decade. MAO activities were highest in the hypothalamus and substantia nigra, TH activities were highest in substantia nigra, putamen and head of caudate, and DBH activities were greatest in tegmentum of pons and hypothalamus. Only minimal regional differences in COMT activities were observed. No significant differences were found between enzyme activities in brain areas of controls and suicides with the possible exception of TH in the substantia nigra, where the depressive suicides (but not the alcoholics) showed greater activity (P < 0·02). These findings appear not to support the catecholamine hypothesis of affective disorder.     

Involvement of nitric oxide in noradrenaline-induced changes in the activity of antioxidant enzymes and lipid peroxidation in rat brown adipose tissue and heart

The effect of exogenous noradrenaline (NA) (1.6 mg.kg(-1) i.p., 35 min prior sacrifice) on the activity of antioxidant enzymes (AOE) copper zinc superoxide dismutase (CuZnSOD), manganese superoxide dismutase (MnSOD) and catalase (CAT), as well as lipid peroxides (LP) concentration were studied in the rat interscapular brown adipose tissue (IBAT) and heart of saline (controls) and N(omega)-nitro-L-arginine methyl ester (L-NAME) treated rats (10 mg.kg(-1), i.p., during 3 days and 20 min before NA). NA differently affects both AOE activities and LP production in the IBAT and heart. Thus, NA inhibited the activity of all IBAT AOE and LP production while in the heart it markedly increased CAT activity only, but had no effect on any of SODs activities and LP concentration. L-NAME, a nitric oxide synthase blocker, completely abolished the NA-induced inhibition of the IBAT AOE and LP production, whereas in the heart it was without effect. In conclusion, these results indicate that both NA and L-NAME effects on AOE activity and LP production are tissue specific and also suggest that nitric oxide mediates the NA-induced inhibition of AOE activity and LP production in the IBAT only.     

DOPAMINE-β-HYDROXYLASE IN THE RAT BRAIN: DEVELOPMENTAL CHARACTERISTICS

Abstract— A sensitive and specific assay for dopamine-8-hydroxylase (DBH) in the rat brain has been developed. The enzyme in the brain has requirements for cofactors and affinity for substrate similar to DBH in the adrenal medulla. DBH activity was demonstrable in the brain of the fetal rat at 15 days of gestation; there was an increase in DBH activity with maturation that preceded and paralleled the rise in levels of endogenous norepinephrine until 3 weeks after birth. There was a shift in the distribution of total DBH activity from the caudal to the rostral regions of the brain during development. In the adult brain, DBH was highly localized in the nerve terminals. Between 17 days of gestation and adult-hood, there was 2300-fold increase in the DBH activity that sedimented with sheared-off nerve terminals.     

Great adaptability of brown adipose tissue mitochondria to extreme ambient temperatures in control and cold-acclimated gerbils as compared with mice

1. The gerbil (Gerbillus campestris) is a desert rodent able to tolerate high (38 degrees C) and low (-20 degrees C) ambient temperatures, probably due to both its low resting metabolic rate in hot environment and its high peak metabolic rate in cold. 2. Measurement of mitochondrial state IV respiration and cytochrome-oxidase activity (COX) were made in interscapular brown adipose tissue (IBAT), liver and hind limb muscles of gerbils and mice of nearly equal body mass, acclimated for 4 weeks at cold ambient temperature (CA) or reared at thermoneutrality (TN). 3. The most striking difference between these two animal species appears to be in IBAT mitochondria: in TN animals, the level of state IV respiration and COX activity was lower in gerbils than in mice, but the cold acclimation-induced increase in these parameters was greater in gerbils than in mice. 4. Alternatively, in gerbils as in mice, cold acclimation induced a reduction in muscle mitochondrial COX activity. No important change due to cold acclimation was observed in liver mitochondria, either in gerbils or in mice. 5. As compared with mice, the lower state IV respiration in IBAT mitochondria from TN gerbils may explain their low RMR, whereas the higher COX activity of IBAT mitochondria from CA gerbils may explain their higher PMR. 6. As a result of this great adaptability of BAT mitochondria, the gerbil seemed to be able to live in a wide range of ambient temperatures in its natural habitat.     

CHARACTERIZATION OF THE BASIS FOR DIFFERENCES IN SERUM DBH ACTIVITY IN IMMATURE AND ADULT RATS BY USE OF HOMOLOGOUS ANTIBODY

Abstract– Rat serum dopamine-β-hydroxylase (DBH) activity decreased 5-7-fold between 15 and 60 days of age. Immunoprecipitation performed with homologous antibody (guinea-pig anti-rat adrenal DBH) showed that during this time period the quantity of antibody necessary to precipitate 50% of the enzymatic activity (AD₅₀) decreased 5-fold from 0.25 to 0.05 μl/ml. The biochemical properties of rat serum DBH at 15 and 60 days of age were compared to test the hypothesis that there might be different biochemical forms of the enzyme in the blood of immature and adult rats. Thermal stability, apparent K_m for tyramine, electrophoretic mobility, pH optima and elution profile on gel filtratioh chromatography were all found to be similar for rat serum DBH at both ages. On the basis of homospecific activity and multiple similarities in biochemical characteristics, it appears that differences in serum activity at the two ages reflect differences in the steady-state levels of enzyme. To determine the turnover of serum DBH in the two age groups, the recovery of enzyme activity was monitored after acute clearance of the circulating pool of DBH by treatment with the homologous antiserum. Immunotitration of DBH activity in vivo indicated that the total pool of serum enzyme was 4-fold greater in the mature rat than in 4-day-olds. After treatment of adult rats with 2μl of homologous antiserum, serum DBH activity was reduced by 85% with a half-life of recovery of 3.0 ± 0.6 days; the estimated fractional rate of degradation was 0.23 ± 0.06 day^?1 and the rate of entrance was 2.3 ± 0.2 units/ml/day. After treatment of 4-day-old rats with 1 μl of homologous antiserum, serum DBH activity was reduced by 95% with a half-life of recovery of 3.3 ± 0.5 days: the estimated average fractional rate of degradation was 0.22 ± 0.06 day^?1 and the average rate of entrance was 10.7 ± 1.6 units/ml/day. Thus, the several-fold difference in steady-state levels of serum DBH in rat pups as compared to adult rats appears to be due to greatly increased rates of entrance of the enzyme in the immature rats.     

Blood dopamine beta-hydroxylase activity and noradrenaline levels in the developing white rat.

Dopamine beta-hydroxylase (DBH) (3,4-dihydroxyphenylethylamine, ascorbate:oxygen oxidoreductase (beta-hydroxylating) (EC 1.14.17.1) activity in serum of blood obtained by decapitation of white rats at 19, 20, and 21 days in utero, immediately after birth, and postnatally to 70 days, was measured. Noradrenaline (NA) and DBH in plasma from undisturbed, cannulated, postweaning rats were also assayed. During the last few days in utero and the first 2 postnatal days serum DBH activity tripled and then remained elevated during the suckling period. Upon weaning, serum DBH activity declined at first precipitously and then more slowly, until the adult level was reached around 70 days of age. This postweaning decrease in DBH activity was also observed with the cannulated animals. In contrast, plasma NA levels remained low and constant throughout the postweaning period. In suckling rats treated with 6-hydroxydopamine from 2 to 12 days of age, serum DBH activity decreased to less than half its initial value by day 8. It is suggested that the observed changes in serum DBH activity in fetal and postnatal rats reflect ontogenetic changes in sympathetic nerve terminals and that they are probably not correlated with release of NA.     

Study on the mechanism of the appearance of noise effects

This study was undertaken to investigate the dose-response relationship between the biological effect and noise exposure, and to consider the mechanism of the appearance of noise effects. Rats were exposed to noise at intensities of 60 dB (A), 80 dB (A) and 100 dB (A) for 240 min and examined for the change of activities of dopamine-beta-hydroxylase (DBH) in serum and adrenal glands. Plasma cyclic adenosine 3',5'-monophosphate (c-AMP) levels were also measured. Some rats were given 6-hydroxydopamine (6-OHDA) as a chemical sympathectomyzing agent 20 h before noise exposure in order to consider the mechanism of the appearance of noise effects. By noise exposure, serum DBH activity was significantly (P less than 0.01) increased at each intensity compared with the control group, but there were no remarkable changes in adrenal DBH activity. Plasma c-AMP level was also significantly elevated in response to the noise stress. When the rats, which had been pretreated with 6-OHDA, were exposed to noise with an intensity of 100 dB (A), the response of serum DBH activity was no longer observed. Therefore it is suggested that the effect due to noise exposure appears through the post-ganglionic sympathetic nerve fiber.     

CORRELATION OF DNA ACCUMULATION RATE WITH THYMIDYLATE SYNTHETASE AND THYMIDINE KINASE ACTIVITIES IN DEVELOPING RAT CEREBELLUM: EFFECT OF THYROXINE1

Abstract— Using the method of least squares, a logistic curve was fitted to the data points for DNA content in neonatal rat cerebellum versus postnatal age (day 0 is the day of birth). The resultant equation was differentiated to give an expression for the rate of cerebellar DNA accumulation in units of ng/h per mg wet cerebellum. The DNA accumulation rate in control rats increased from 77.0 at 2 days of age to a maximum of 108 at 7 days of age and declined thereafter to a minimum of 16.3 on day 15. Thyroxine treatment significantly (P < 0.05) increased the rate to 89.8 (117% of control) at 2 days of age, and a significant elevation was maintained to 6 days of age at which time a maximum rate of 115 (114% of control) was attained. The rate was significantly decreased below control at 9 and 12 days of age, and reached a minimum of 9.22 on day 15. The developmental pattern for the activity of cerebellar thymidylate synthetase (EC 2.1.1.6), in units of pmol/h per mg wet cerebellum, closely paralleled the pattern for DNA accumulation rate in both control and thyroxine-treated animals. In controls, thymidylate synthetase activity increased from 98.6 at 2 days of age to a maximum of 125 at 7 days of age and declined thereafter to a minimum of 30.0 at 15 days of age. In thyroxine-treated animals, the activity was significantly increased to 118 (122% of control) at 4 days of age and remained significantly elevated through 6 days of age at which time a maximum activity of 154 (115% of control) was attained; thereafter, the activity was significantly decreased below control and reached a minimum of 16.9 (56.3% of control) on day 15. The developmental pattern for the activity of cerebellar thymidine kinase (EC 2.7.1.21) did not parallel the DNA accumulation rate quite so closely, in neither treated nor control animals, as did the pattern for thymidylate synthetase activity. These data suggest that thymidylate synthetase activity in the developing rat cerebellum may be more important for maintenance of replicative DNA synthesis than is thymidine kinase activity. In addition, the thyroxine-induced acceleration of the increase and subsequent decline in rate of DNA accumulation and in the activities of thymidylate synthetase and thymidine kinase in developing rat cerebella is probably the result of alterations in the number of external granular cells undergoing replicative DNA synthesis.     

Dopamine beta-hydroxylase in rat serum and lymph: changes with age and effect of cold exposure.

The pH optimum for rat serum dopamine beta-hydroxylase (DBH) (3,4-dihydroxyphenylethylamine, ascorbate:oxygen oxidoreductase (beta-hydroxylating)(EC 1.14.17.1) was 4.0 in acetate buffer; other requirements were as reported by others. DBH activity in serum of 20-day-old fetuses is slightly higher than in that of their mothers. Levels of the enzyme in blood a few hours after birth are almost five times greater than in the adult, remain high during the suckling period, then drop rapidly during the 4th week after birth to about three times the adult level, which is then slowly reached over the next few weeks. These fluctuations in serum DBH activity coincide with the period of intense development and maturation of the sympathetic nervous system. There was not significant effect of cold exposure on blood DBH activity when newborn, suckling, weanling or adult warm- and cold-acclimated rats were exposed to cold. Similarly, exposure to cold that elicited two- to three-fold increases in O2 consumption failed to increase DBH activity in thoracic duct lymph. Therefore serum and lymph DBH activities are not sensitive indices of sympathetic secretory activity in the intact rat.     

Factors involved in changes in hepatic lipogenesis during development of the rat

1. Changes in the activities of acetyl-CoA carboxylase (EC 6.4.1.2), phosphofructokinase (EC 2.7.1.11), aldolase (EC 4.1.2.13), extramitochondrial aconitate hydratase (EC 4.2.1.3) and NADP-dependent isocitrate dehydrogenase (EC 1.1.1.42) have been measured in the livers of developing rats from late foetal life to maturity. 2. The effect of altering the weaning time on some enzymes associated with lipogenesis has been studied. Weaning rats at 15 days of age instead of 21 days results in an immediate increase in the activity of ;malic' enzyme (EC 1.1.1.40) whereas the activities of glucose 6-phosphate dehydrogenase (EC 1.1.1.49) and ATP citrate lyase (EC 4.1.3.8) did not increase until 4-5 days and acetyl-CoA carboxylase 2-3 days after early weaning. Weaning rats on to an artificial-milk diet led to complete repression of the rise in activity of hepatic enzymes associated with lipogenesis normally found on weaning, except for ;malic' enzyme, which increased in activity after 20 days of age. 3. The effect of intraperitoneal injections of glucagon, cortisol, growth hormone and thyroxine on the same hepatic enzymes has been investigated. Only thyroxine had any effect on enzyme activities and caused a 20-fold increase in ;malic' enzyme activity and a twofold increase in ATP citrate lyase activity. 4. The activities of hepatic glucose 6-phosphate dehydrogenase and ;malic' enzyme are higher in adult female than in adult male rats and it has been shown that this sex difference in enzyme activities is due to both male and female sex hormones. 5. Hepatic malate, citrate, pyruvate, glucose 6-phosphate and phosphoenolpyruvate concentrations have been measured throughout development. 6. The results are discussed in relation to the dietary and hormonal control of hepatic enzyme activities during development.     

The influence of chemical sympathectomy on oxygen uptake during development of rat brain cortical tissue

In this study the influence of 6-hydroxydopamine (6-OHDA), in vitro and in vivo, on the oxygen consumption in the rat brain cortical slices was examined. The treatment with 6-OHDA increased the oxygen uptake of brain cortical tissue of young rats. The maximum increase was observed 7 and 14 days after treatment with 6-OHDA. On the contrary, 6-OHDA added in vitro produced very marked depression of oxygen uptake in slices of brain cortical tissue of the tested animals. The addition of isoprenaline in vitro stimulated the respiratory activity in the cerebral tissue of control young rats in all the periods of examination. Thus, 18 days after the birth, the isoprenaline-stimulation of oxygen uptake in brain tissue was 44.3% as compared to the control values. The same degree of stimulation was noted in the cerebral tissue of older animals (25, 32 and 45 days after birth). However, addition of isoprenaline did not influence the respiration of cerebral tissue stimulated by 6-OHDA.     

Hypothalamic dopamine-β-hydroxylase activity: Fluctuations with time of day and their modifications by intracranial surgery,adrenalectomy, and pinealectomy

Hypothalamic and plasma dopamine--hydroxylase (DBH, EC 1.14.2.1) activities were measured by a coupled radioenzymatic method. Animals representing five experimental groups (intact controls, adrenalectomized, pinealectomized, adrenalectomized + pinealectomized, doubly sham-operated) were killed and sampled at 8 times through the 24-hr daily cycle, 15 days postoperation, and at 50–52 days of age. Hypothalamic DBH in intact control animals had statistically significant fluctuations in relation to time of day. These changes were lost or dampened in groups that had had intracranial surgery and were characteristically shifted by adrenalectomy, either alone or with pinealectomy. Plasma DBH fluctuations in the same animals resembled those in hypothalamus in some features (e.g., peak near mid-dark; shift in daily maxima and minima after adrenalectomy) and differed in others (e.g., no effect of intracranial surgery or of sham operation; adrenalectomized + pinealectomized animals resembled the solely pinealectomized). Although temporal patterns in hypothalamic DBH activity thus differed in the experimental animal groups, the daily means of hypothalamic DBH activity were similar.     

Plasma norepinephrine and dopamine-β-hydroxylase in genetic hypertensive rats

The relationship between blood pressure, plasma norepinephrine (NE), dopamine-β-hydroxylase (DBH) activity and age was investigated in spontaneously hypertensive rat (SHR), a stroke-prone substrain of the SHR and control Wistar-Kyoto rat (WKR). Blood pressure of both SHR strains increased with age and was significantly higher than that of the WKR at all ages tested (3 15 weeks). The blood pressure of stroke-prone SHR was significantly higher than that of the regular SHR after 6 weeks of age. Plasma DBH activity decreased with age in each strain, although the SHR, and especially the stroke-prone SHR, had significantly higher DBH than the controls at an early age. Plasma NE in the WKR did not change with age. Increased plasma NE was observed only in the young SHRs. The highest values were found in the 6 week old stroke-prone SHR. These data suggest that plasma DBH activity is not correlated directly with plasma NE or blood pressure, but that increased sympathetic nerve activity may occur during the development of hypertension in the SHR and the stroke-prone SHR.     

Oxidases and oxygenases in regulation of neutrophil redox pathways in Behçet's disease patients

This study aimed to determine plasma and neutrophil oxidase activities that may contribute to vascular inflammation in Beh?et's disease (BD) patients. Cyclooxygenase (COX), NADPH oxidase and myeloperoxidase (MPO) activity was determined in neutrophils isolated from BD patients and healthy controls. Functional assay of NADPH oxidase was significantly increased in BD patients, both at basal conditions and in response to fMLP stimulation. There was a significant increase in plasma MPO activity in the disease group as compared to controls. Total COX activity was significantly increased in BD neutrophils. The increase in total COX activity was accompanied with enhanced activity of COX-2, differentiated by using the COX-1 isoform-specific inhibitor SC-560. Neutrophil nitrate/nitrite levels showed no significant difference in BD; however, plasma nitrate/nitrite contents in BD patients were significantly greater compared to controls. In conclusion, increased plasma MPO, neutrophil NADPH and COX activities may contribute to intravascular inflammation documented in BD patients.     

The role of nitric oxide in remodeling of capillary network in rat interscapular brown adipose tissue after long-term cold acclimation

Cold exposure has been shown to increase blood flow in interscapular brown adipose tissue (IBAT). The aim of the present study was to evaluate the role of the L-arginine-nitric oxide (*NO) pathway on IBAT capillary network remodeling and its possible correlation with superoxide anion radical (O2(*-)). In the rats that received L-arginine (2.25%) or NG-nitro-L-arginine methyl ester (L-NAME, 0.01%) as a drinking liquid and maintained at room (22+/-1 degrees C) or low (4+/-1 degrees C) temperature for 45 days, IBAT capillaries were analyzed by stereology and observed by light and electron microscopy. Additionally, endothelial *NO synthase (eNOS) expression, nitrotyrosine immunoreactivity and both copper zinc superoxide dismutase (CuZnSOD) enzyme activity and immunohistochemical localization were examined. Stereological analyses of IBAT show that the capillary volume density, as well as capillary-to-brown adipocytes ratio, are increased in cold. L-arginine treatment increases, while L-NAME decreases both parameters, compared to respective controls. Those changes were accompanied by capillary dilatation observed by light and electron microscopy. The activity of CuZnSOD is lower in control cold-acclimated rats, as well as in both L-arginine-treated groups, when compared to control animals acclimated to room temperature. L-NAME treatment attenuates the effects both of cold and L-arginine on CuZnSOD and increases immunopositivity for CuZnSOD in room temperature-acclimated rats. Our results show that *NO induces remodeling of the IBAT capillary network by angiogenesis, and presumably that interaction with O2(*-) has a role in that modulation. The increased eNOS expression accompanied by an increased nitrotyrosine immunoreaction observed in both L-arginine-treated groups compared to corresponding controls strengthens this hypothesis.     

Oxidases and oxygenases in regulation of neutrophil redox pathways in Behçet’s disease patients

This study aimed to determine plasma and neutrophil oxidase activities that may contribute to vascular inflammation in Behçet’s disease (BD) patients. Cyclooxygenase (COX), NADPH oxidase and myeloperoxidase (MPO) activity was determined in neutrophils isolated from BD patients and healthy controls. Functional assay of NADPH oxidase was significantly increased in BD patients, both at basal conditions and in response to fMLP stimulation. There was a significant increase in plasma MPO activity in the disease group as compared to controls. Total COX activity was significantly increased in BD neutrophils. The increase in total COX activity was accompanied with enhanced activity of COX-2, differentiated by using the COX-1 isoform-specific inhibitor SC-560. Neutrophil nitrate/nitrite levels showed no significant difference in BD; however, plasma nitrate/nitrite contents in BD patients were significantly greater compared to controls. In conclusion, increased plasma MPO, neutrophil NADPH and COX activities may contribute to intravascular inflammation documented in BD patients.