The long pentraxin-3 is a useful marker for diagnosis of ovarian torsion: An experimental rat model.

Pentraxin-3 (PTX3) plays an important role in the primary inflammatory response. We aim to evaluate PTX3 as a diagnostic marker of ovarian torsion in an experimental rat model. In this study, 16 female Sprague Dawley albino mature rats were randomly allocated to Group 1 (control, sham operated) and Group 2 (experimental ovarian torsion model). A torsion model was set up using atraumatic vascular clips just above and below the right ovary for a 3-h ischaemia. Blood samples were collected before and three hours after ovarian torsion. Three hours after ovarian torsion, right ovary was surgically removed for histopathological examination in both groups. There was no significant difference in preoperative PTX3 level in both groups (1.05 ± 0.20 ng/mL vs 1.09 ± 0.28 ng/mL, p > 0.05). Three hours after the operation, mean plasma level of PTX3 was significantly higher in ovarian torsion group than the control group (2.13 ± 0.49 ng/mL vs 1.07 ± 0.22 ng/mL, p = 0.001). Also, the mean total histopathological score was significantly increased in the torsion group. PTX3 can be used in clinical practice as a useful marker for early diagnosis of ovarian torsion.

Nephro-protective effect of granulocyte colony-stimulating factor in streptozotocin induced diabetic rats.

Diabetic nephropathy is one of the most serious complications of diabetes and the major cause of end-stage renal failure. Consequences of diabetic nephropathy include increased kidney size and glomerular volume, thickening of basement membranes and progressive accumulation of extracellular matrix. Reports in the literature support an association between increased secretion of inflammatory molecules, such as cytokines, growth factors and metalloproteinases, and development of diabetic nephropathy. We investigated the potential of granulocyte colony- stimulating factor (G-CSF) as a therapeutic candidate for preventing diabetic nephropathy. We used 21 8-week-old male rats; 14 were administered a single dose of 60 mg/kg streptozotocin (STZ) to induce diabetes. The rats were divided into three groups of seven: group 1, control; group 2, diabetic; group 3, diabetic plus G-CSF treatment. After 4 weeks, immunoexpressions of transforming growth factor ß1 (TGF-ß1), Akt and CD34 levels were measured in the kidney tissue. Blood glucose, urine protein and the glomerular area also were measured for each group. We found that G-CSF treatment decreased TGF-ß1 immunoexpression, urine protein and glomerular area in kidneys of diabetic rats, and increased CD 34 and Akt immunoexpression in kidneys of diabetic rats. The effects of G-CSF were independent of blood glucose levels. G-CSF may be a useful therapeutic agent for preventing diabetic nephropathy.

Montelukast prevents ischaemia/reperfusion-induced ovarian damage in rats.

OBJECTIVE: To investigate the efficacy of montelukast for prevention of ischaemia/reperfusion (I/R) injury in rat ovary. STUDY DESIGN: Twenty-four female adult rats were included in the study. I/R injury was induced by CO2 pneumoperitoneum in a laparoscopic rat model. The rats were divided at random into three groups: the sham group was subjected to catheter insertion but was not subjected to pneumoperitoneum; the saline group was subjected to 60 min of pneumoperitoneum and 30 min of reperfusion, with 1 mg/kg physiological saline administered 10 min before pneumoperitoneum; and the montelukast group was subjected to 60 min of pneumoperitoneum and 30 min of reperfusion, with 20mg/kg montelukast administered 10 min before pneumoperitoneum. Damage to ovarian tissue was scored by histopathological evaluation. Caspase-3 expression was determined immunohistochemically. Ovarian tissue levels of malondialdehyde and glutathione, and plasma total antioxidant capacity were measured biochemically. RESULTS: In comparison with the sham group, ovarian sections in the montelukast group had higher scores for follicular degeneration and oedema (p<0.001). Montelukast treatment prevented tissue damage in ovaries, and this result was significant. Caspase-3 expression was only observed in ovarian surface epithelium in the saline and montelukast groups. However, the mean caspase-3 expression score was higher in the saline group than the montelukast group (p<0.001). Tissue levels of malondialdehyde were higher in the montelukast group than the sham group, but plasma total antioxidant capacity and tissue levels of glutathione were significantly lower. Pretreatment with montelukast reduced lipid peroxidation (p<0.005) and improved antioxidant status in rats (p<0.001). CONCLUSION: Montelukast is effective for the prevention of I/R-induced damage in rat ovary.

Mitochondrial oxidative stress in female and male rat brain after ex vivo carbon monoxide treatment.

Carbon monoxide (CO) is the most common cause of fatal poisoning all over the world. At the cellular level, a combination of tissue hypoxia and direct cellular damage underlie the pathophysiology of CO toxicity. The purpose of this study was to determine the effect of CO treatment on oxidative stress parameters in mitochondria isolated from male and female rat brains. Mitochondria prepared from frontal cortex, hippocampus and corpus striatum were treated with 0.1% CO at 37 degrees C for 30 minutes; control samples were not exposed to CO. Cytochrome c oxidase activity (COX), lipid peroxidation (thiobarbituric acid reactive species = TBARS), protein oxidation (protein carbonyls) and glutathione (GSH) levels were measured in CO treated and control samples. Our results confirmed previous studies reporting the inhibition of cytochrome c oxidase activity by CO in rat brain. Additionally, protein carbonyl levels in the hippocampus and striatum significantly increased after CO treatment in male rats. While CO treatment caused a significant decrease in GSH levels in the cortex and striatum in male rats, reduced GSH levels were observed in the cortex and hippocampus in female rats following CO exposure. Taken together, our data suggest a role for mitochondrial oxidative stress in CO toxicity at the cellular level during CO poisoning.

Serum nitric oxide metabolites in patients with multiple sclerosis.

Multiple sclerosis (MS) is an autoimmune disease of the central nervous system characterized by myelin breakdown. The free radical nitric oxide (NO), which is considered to be a major metabolite in immune function and in autoimmune disorders, is among the possible mediators causing the inflammatory reactions in MS. Consequently, NO has been implicated in the pathogenesis of MS and its animal model experimental allergic encephalomyelitis (EAE). In this study, stable metabolites of NO (NO(2-)+NO(3-)) levels were determined in sera of MS patients (n=23) and control subjects (n=16). NO(2-)+NO(3-) levels were higher in MS patients when compared to control subjects. However, there was not any correlation with serum NO(2-)+NO(3-) values and clinical features of the disease such as duration of sickness, the time elapsed from the last attack and EDSS values. Our results imply that nitric oxide may be involved in the pathogenesis of MS although further studies are required to elucidate underlying mechanisms.

Serum and cerebrospinal fluid nitrite and nitrate levels in relapsing-remitting and secondary progressive multiple sclerosis patients.

Nitric oxide (NO) has been implicated in immune mediated cellular cytotoxicity and inflammatory processes including multiple sclerosis (MS). We aimed to assess NO production in MS patients and to delineate its involvement in different stages. The stable end-products of NO; nitrite(NO2-) and nitrate(NO3-) were analysed both in serum and CSF (cerebrospinal fluid) of patients with MS and non-inflammatory neurological diseases. Nitrite levels were quantified by calorimetric assay based on the Griess reaction. Nitrate levels were examined spectrophotometrically. MS patients exhibited significantly increased serum and CSF levels of NO2-+NO3- compared with the control subjects. CSF NO2-+NO3- levels were raised significantly in MS patients with both relapsing remitting (RR) and secondary progressive (SP) course. There was no significant difference between RR and SP MS patients with regard to NO metabolites. No significant correlation was found between NO metabolites and disability score, disease progression index, MRI (magnetic resonance imaging) activity and development of cortical atrophy on MRI. This study provides further evidence for excessive NO production both in CSF and peripheral blood of MS patients. Excessive CSF NO2-+NO3- levels being more increased than the levels in sera supports pathological inflammatory process within CNS (central nervous system) in both stages of MS. Another implication for the role of NO and INOS inhibitors in the treatment of MS patients with both RR and SP courses was also suggested.

Nicotine modulates nitric oxide in rat brain.

Nicotine exerts its central actions by regulating cationic fluxes through nicotinic acetylcholine receptors (nAChRs). By this effect, the drug likely also modifies events occurring beyond the nAChR, including the regulation of nitric oxide (NO) synthesis. The present study was undertaken to assess the effects of acute and chronic nicotine administration (0.4 mg/kg, s.c.) on levels of NO(-)(2)+NO(-)(3), stable metabolites of NO, in brain regions of male and female rats. Nicotine increased levels of the metabolites, and therefore presumably of NO, with sex differences in the degree of stimulation, the brain regions affected, and the variance between the effects of acute and chronic administration. Prior inhibition of NO synthase eliminated the effect of nicotine in all regions studied. While nicotine appeared to increase NO indirectly via glutamate receptors in the cortex and hippocampus, this was not true of the corpus striatum, where blocking NMDA-type glutamate receptors with MK-801 had no effect. The findings support the view that NO is likely involved in some of the central effects of nicotine.

The effects of melatonin on the antioxidant systems in experimental spinal injury.

Melatonin has been recently shown by various in-vivo and in-vitro studies to exert potent neutralising effects on hydroxyl radicals, stimulate glutathione peroxidase (GSH-Px) activity, and protect catalase (CAT) from the destructive activity of hydroxyl radicals in neural tissue. We aimed to investigate the possible effects of pharmacological dose of melatonin on some of the antioxidant defence systems in an in-vivo study of experimental spinal injury. Seven groups of adult male Sprague Dawley rats were used in the following scheme: Group I: Naive (n = 6), Group II: Lesion (n = 8), Group III: Melatonin (n = 5), Group IV: Melatonin + Lesion (n = 8), Group V: Placebo + Lesion (n = 5), Group VI: Sham operation (n = 5), and Group VII: Placebo (n = 5). Experimental spinal injury was induced at level T7-T8 by 5 sec compression of the total cord with an aneurysm clip on anaesthetised and laminectomized animals. The total 10 mg/kg dose of melatonin (Sigma) dissolved in alcohol-water was administered i.p. four times in 2.5 mg/kg doses, at 20 min pre-, at the time of and at 1 h and 2h post-compression. At 24 +/- 2h post-injury, the rats were euthanized and the lesioned segments of cord were dissected and homogenised with special care taken to distribute equal amount of injured tissue in each sample for analysis of reduced glutathione (GSH), oxidised glutathione (GSSG), superoxide dismutase (SOD), and CAT activity. Compression injury decreased GSH/GSSG ratio significantly (p < .0001). Melatonin, by itself, significantly decreased GSSG content (p < .05) and increased CAT activity (p < .05) in the naïve rats. Melatonin treatment decreased GSSG activity, thus elevating GSH/GSSG ratio, and also increased SOD and CAT activity without reaching statistical significance in the lesioned animals. In conclusion, pharmacological dose of systemically applied melatonin seemed to support some features of the antioxidant defence systems in our hands.

The effects of chronic hepatitis C and B virus infections on liver reduced and oxidized glutathione concentrations.

The aim of this study was to evaluate the effects of hepatitis B and C virus infections on liver glutathione status. Reduced and oxidized glutathione levels were determined in liver biopsy specimens obtained from patients with chronic liver disease including chronic active hepatitis and cirrhosis. In patients with hepatitis B virus infections, GSH and GSH/GSSG levels were significantly low compared with those in controls (P<0.01). There was a significant negative correlation between histological activity indices (HAI) and hepatic GSSG levels only in patients with chronic HCV infection (P<0.01; r=-0.895). In addition to this, we also found a positive correlation between indices (HAI) and GSH/GSSG of the same group (r=0.915; P<0.05). These observations suggest that HBV and HCV infections have different effects on liver glutathione status based on diverse mechanisms.

Sexually dimorphic cognitive style in rats emerges after puberty.

In a water maze (WM), rats employ different and sexually dimorphic behavioral strategies to solve a place-learning task, a test of cognitive/propositional ability. Puberty is the last step in brain development and marks an important phase with regard to sexually dimorphic cognitive performance and behavior. The present study assessed possible sex differences in cognitive style before and after puberty in a WM place-learning task. Since nitric oxide (NO) is implicated in spatial learning and hippocampal function, and since brain NO(-)(2) + NO(-)(3) levels (stable metabolites of NO) display region-specific sex differences in rat brain, NO(-)(2) + NO(-)(3) levels were determined after behavioral testing. The sex-related style difference emerged very clearly but only in the adult rats, which suggests that the female behavioral strategy in the WM place-learning task requires the presence of female sex hormones at puberty. Although NO(-)(2) + NO(-)(3) levels were higher in the adult rats and males compared to prepubertal and female rats, respectively, no significant correlations emerged between brain NO and behavior. The fact that the behavioral sexually dimorphic cognitive-style effect observed here and in previous studies appears to emerge only after puberty suggests that awareness of such postpubertal sex differences may also be important in human educational and therapeutic contexts.

Increased cerebrospinal fluid and serum nitrite and nitrate levels in amyotrophic lateral sclerosis.

Abnormal glutamate metabolism is implied in the pathogenesis of Amyotrophic Lateral Sclerosis (ALS) and cerebrospinal fluid (CSF) glutamate levels appear to be elevated. Since nitric oxide (NO) inhibits glutamate transport, excessive amounts of nitric oxide could underlie the glutamate induced neurotoxicity in ALS. Stable metabolites of NO (NO2- + NO3-) levels were determined in serum and CSF of sporadic ALS patients and control subjects. NO2- + NO3- levels were higher in ALS, in males and in serum samples compared to controls, females and CSF, respectively. Furthermore, while the difference between serum and CSF NO2- + NO3- levels was significant in males (higher in serum) no such difference was observed in females. Our results suggest that nitric oxide may be involved in the pathogenesis of ALS directly or indirectly and in a sexually dimorphic manner.

Sexually dimorphic cognitive style, female sex hormones, and cortical nitric oxide.

Recent studies using the water maze (WM) found marked sex differences in behavioral strategy employed in place learning tasks in adult rats. When a change in the platform position is introduced following learning the place of a platform (visible or hidden) in a different position, female rats escape to the newly positioned visible platform faster than males. Nitric oxide (NO) is implicated in place learning, and there are regional sex differences in its stable metabolites, NO(2)(-)+NO(3)(-), in rat brain. Furthermore, NO(2)(-)+NO(3)(-) levels are sensitive to ovariectomy in female rats. The effect of sex hormones on brain development and function is well documented. The present study was undertaken to study the effects of ovariectomy and hormonal manipulations on cognitive performance in a WM task designed to test differences in behavioral strategy in Sprague-Dawley rats (n=48) of both sexes. Some of the females rats were ovariectomised and received either hormone replacement (estrogen or progesterone alone or in combination) or the vehicle. Cortical and hippocampal NO(2)(-)+NO(3)(-) levels were determined after behavioral testing. There were no group differences in cognitive ability or non-cognitive factors such as motivation or swim speed. Males and intact females differed in their cognitive style, but hormonal manipulations in female rats did not affect this relative use of behavioral strategy. There was a correlation between performance on the trial where sex differences were most prominent and NO(2)(-)+NO(3)(-) levels in the cortex. Our results suggest that the activational effects of circulating gonadal hormones do not play a major role in sexually dimorphic cognitive styles.

Effect of nitric oxide on postoperative adhesion formation.

Peritoneal adhesions continue to be a significant cause of postoperative complications. The purpose of the present study was to investigate the effect of nitric oxide in preventing postoperative adhesion formation in rats. Three randomized groups of Sprague-Dawley rats were subjected to a standardized lesion by cecal abrasion and parietal peritoneal defect. 0.9% NaCl (control, group 1), L-arginine (300 mg/kg, group 2) and Nomega-nitro arginine methyl ester (L-NAME; 25 mg/kg, group 3) were administered intraperitoneally before abdominal closure and during 3 consecutive days after surgery. Two weeks after surgery, a relaparotomy was performed and the extent of adhesion formation was determined. In groups 1 and 3 heavy adhesions were detected. In the L-arginine group, adhesion formation was significantly less than in the other groups (p < 0.05). This study showed that L-arginine reduced adhesion formation.

Nicotine interacts with sex in affecting rat choice between "look-out" and "navigational" cognitive styles in the Morris water maze place learning task.

The effect of sex and nicotine on cognitive style was examined in rats using a water maze task that allows differentiation between cognitive ability and style. During the 12-day acquisition period with the platform in the same location (either visible or hidden) there were no effects or interactions attributable to nicotine and sex, either in terms of learning rate or asymptotic latency. On the final test day the platform was visible and shifted in its location, and on the first trial the new location was proximal to the rats starting position, in contrast to the more distal location of the platform during the previous acquisition days. This platform relocation presented the rats with a choice between two competing cognitive styles: using local visual (look-out) cues vs. navigational cues. Performance on the test day yielded a nicotine x sex interaction, such that only saline-treated female rats showed a clear preference for the perceptual-proximal look-out cognitive style by swimming straight to the newly-relocated visible platform with mean escape latency that approximated the limits of swimming speed. The other three groups did not differ from each other, and preferred navigational cues. The results show that male and female rats use different strategies in problem solving, and that nicotine shifts the female pattern to that of the male.

Effect of N-dicyclopropylmethyl-amino-2-oxazoline (S-3341) on antioxidant status and nitric oxide in hypertensive patients.

Defective endothelium-dependent vascular relaxation has been found in animal models of hypertension and in hypertensive patients. An imbalance due to reduced production of nitric oxide (NO) or increased production of free radicals, mainly the superoxide anion, may facilitate the development of an arterial functional spasm. Although it has been shown that many antihypertensive drugs can normalise both the antioxidant activity and NO, the antioxidant effect of N-dicyclopropylmethyl-amino-2-oxazoline (S-3341), an alpha-adrenoreceptor agonist, has not been investigated. In this study we investigated the antioxidant and NO status in hypertensive patients and whether there was any effect of S-3341 on these parameters. Eleven patients with mild hypertension (mean systolic blood pressure 159.5 +/- 2.5 mmHg) were administered S-3341 (1 mg/day) for 4 weeks. Plasma vitamin E, nitrite-nitrate and MDA levels, and catalase activity, were measured both before and after treatment with S-3341. There was significant reduction in both mean systolic and diastolic blood pressure during the treatment. We found an increase in catalase activity (p < 0.05), a decrease in malondialdehyde (MDA) levels (p < 0.01) and an insignificant increase in vitamin E levels in hypertensive patients following the S-3341 treatment. We propose that S-3341 may prevent oxidant stress in hypertensive patients by inhibiting free-radical formation.

Articular cartilage homeostasis after anterior cruciate ligament reconstruction.

Several factors may be responsible for osteoarthritis after anterior cruciate ligament (ACL) reconstruction. The detrimental effect of the surgical technique may explain part of the progression of the pathologic process. In this study, the effect of ACL reconstruction on articular cartilage was investigated by measuring proteoglycan fragments (PF) in synovial fluid collected from patients who had undergone this operation. Synovial samples were obtained from 44 patients with chronic ACL deficiency aged 26 +/- 7 years preoperatively, and from some of them, samples were collected for monitoring at 1 month (n = 22), 3 months (n = 17), 6 months (n = 18) and 12s month (n = 18). Synovial fluid taken from 12 contralateral asymptomatic knees of 12 patients (not necessarily opposite knees of ACL-deficient cases) served as controls. Preoperative values were significantly larger than controls (P < 0.05). PF level reached its maximum value in the 1st month (P < 0.05) and then gradually decreased. It was significantly lower than preoperative values at 6 and 12 months but still greater than controls (P < 0.05). It seems that surgical trauma affects cartilage metabolism for the first 3 months postoperatively. Although reconstruction of the ACL contributes to articular cartilage homeostasis, a complete return to normal values cannot be achieved in 1st postoperative year at least in knees with chronic ACL deficiency. Long-term monitoring is needed to see whether these findings are early signs of osteoarthritis. Further studies may more clearly demonstrate the effect of the surgical procedure on knees with acute and subacute ACL deficiency.

Intersession interval affects performance in the Morris Water Maze.

We have recently reported an effect that shows a sexually dimorphic difference in cognitive style rather than ability. The preparation for potentially producing this proximal perceptual style effect is one where rats are first given 4-trial daily acquisition sessions for 12 days with the platform always in the same position, but sometimes visible (perceptual, "look-out" condition) and sometimes hidden (conceptual, "navigational" condition). On the first, probe trial of the 13th day, the platform's position is shifted to a point very close (proximal) to the rat's starting position, and made visible. The proximal perceptual style (PPS) effect has emerged sexually dimorphically in that only females swam straight to the newly positioned proximal platform. Other studies have shown that the PPS effect is eliminated (with females behaving like males) by nicotine and prepubertal ovariectomy, and does not occur in prepubertal females. Also, as no sex-related effects emerged during acquisition during these studies, the PPS effect appears to be a function of cognitive style rather than ability. The present study varied age, and, in an effort to economize on time, shortened acquisition to 6 days by having morning and afternoon sessions each day. To our surprise, this relatively subtle psychological manipulation eliminated the PPS effect, and also yielded some sex- and age-related effects during acquisition: A male advantage was observed and prepubertal rats had longer escape latencies; there was no significant interaction between sex and age.

Sex differences in nitrite/nitrate levels and antioxidant defense in rat brain.

This study assessed sex differences in stable metabolites of nitric oxide and major enzymes involved in antioxidant defense in various regions of rat brain. Nitrite/nitrate levels and activities of superoxide dismutase and catalase were determined in cortex, hippocampus, corpus striatum, midbrain and cerebellum of adult male and female Sprague-Dawley rats. Nitrite/nitrate levels were significantly higher in the cortex and the hippocampus of male than female rats, while catalase activity was higher in the cortex of females than in males. These sex differences may have significant effects on brain function in health and disease.

The role of nitric oxide in proteoglycan turnover by bovine articular cartilage organ cultures.

Monolayer cultures of articular chondrocytes synthesize large amounts of nitric oxide (NO) following exposure to IL-1. The latter has antianabolic and procatabolic activities on these cells, but little is known about the role, if any, of NO in the integrated metabolic pathways of the chondrocyte. In the present study, the role of endogenously produced NO in both the synthesis and degradation of proteoglycans was investigated for the first time. Bovine articular cartilage slices exposed to 20 U/ml human rIL-1 beta (hrIL-1 beta) synthesized large amounts of NO for 1 to 2 days, after which production fell to a steady state level approximately 20% of the peak value for the remainder of the 14-day incubation. The NO synthase inhibitor, N-monomethyl L-arginine (L-NMA, 1 mM), blocked NO production and enhanced the acute catabolic effects of hrIL-1 beta in cartilage derived from both cartilage derived from both calves and adult animals. However, in late cultures, release of proteoglycans was reduced in the presence of L-NMA. The proteolytic activity in conditioned medium of these cultures (measured as caseinolytic activity) was enhanced by L-NMA; however, this inhibitor did not affect the rates of synthesis of proteoglycans. Although NO is widely assumed to be a mediator of cartilage catabolism, our data suggest that it may instead have an acute protective effect. Whether this effect is maintained chronically is less clear.