Quenching the effects ofl-arginine on free radical injury in cultured cardiomyocytes

l -arginine on the free radical injury of cultured cardiomyocytes which were obtained from patients undergoing corrective surgery for tetralogy of Fallot. Free radicals were generated from hypoxanthine via xanthine oxidase, and the cellular changes were determined microscopically. All concentrations of l-arginine (0.5 to 3 mM) prolonged the myocyte survival time compared to the control group, with 0.5 mM l-arginine increasing the survival time to the greatest extent. Cellular susceptibility to free radical injury was the lowest with 0.5 mM l-arginine. Further experiments were performed with 0.5 mM l-arginine plus 100 mM or 1 000 mM of the NO synthase (NOS) inhibitor N ^G-nitro-l-arginine methylester (l-NAME) to determine whether or not the effects of l-arginine are mediated through the NO pathway. The survival time for the cells treated with a concentration of l-NAME was shorter than for the cells treated with 0.5 mM l-arginine alone. These results suggest that l-arginine acts through the NO-dependent pathway. In conclusion, our findings thus confirmed the quenching effects of NO on free radical injury in cultured cardiomyocytes. (Received for publication on Feb. 19, 1997; accepted on July 8, 1997)     

Limitations of Exogenous L-Arginine in Exerting a Cytoprotective Effect on Hepatic Ischemia/Reperfusion Injury

-arginine/nitric oxide (NO) pathway induces a protective effect, we investigated the effect of exogenous L-arginine on hepatic ischemia/reperfusion (I/R) injury, using ex vivo perfusion of the isolated rat liver. The rat liver was removed and preserved in cold saline for 60 min, followed by 120 min of reperfusion with oxygenated perfusate at 37°C. Either 600 mg/kg of L-arginine (groups 1 and 4), D-arginine (group 2), N ^G-nitro-L-arginine methyl ester (L-NAME) (group 3), or saline (group 5) were administered through the portal vein starting from 5 min before reperfusion to 5 min after reperfusion. In group 4, 600 mg/kg of L-NAME was preadministered at 10 min prior to the administration of L-arginine. The intrahepatic nitric oxide (NO) levels showed only a temporal elevation (227% ± 70% of the pre-reperfusion levels at 5 min) after reperfusion in group 1. Pretreatment with L-NAME suppressed the elevation of the NO levels immediately after reperfusion in group 4. The lactate dehydrogenase release to the effluent perfusate significantly decreased and the histological findings showed that the sinusoidal damage observed after reperfusion was mitigated in group 1 more than in the other groups. These results thus suggest that exogenous L-arginine produced a relatively small amount of NO and therefore resulted in a slight decrease of hepatic I/R injury. (Received for publication on Feb. 12, 1999; accepted on Nov. 11, 1999)     

Protective Effect of Oral <Emphasis Type="SmallCaps">L</Emphasis>-arginine Supplementation on Cyclosporine Induced Nephropathy in Rats

Background: One of the major adverse effects of long term cyclosporine A (CyA) administration is chronic nephrotoxicity. Several studies have suggested that alterations of the L-arginine (L-Arg) nitric oxide (NO) pathway may be involved in the pathogenesis of CyA-induced kidney damage. Aim: We postulated that in vivo activation of L-Arg-NO pathway might have a beneficial effect on CyA-induced renal damage. Conditions of chronic NO enhancement was established with L-Arg supplementation and chronic NO blockade with N-nitro-L-Arg methyl ester (L-NAME). We tested the hypothesis that, if CyA administration alters intrarenal NO synthesis, then exogenous L-Arg supplementation could limit renal injury, on the contrary, L-NAME, a potent competitive inhibitor of NO synthesis, could enhance CyA nephrotoxicity. Harmful effect of NO blockade indirectly supports the beneficial effect of NO in a model of CyA nephrotoxicity. Methods: Rats were administered vehicle (VH), CyA (7.5 mg/kg/day), CyA + L-Arg (2g/kg/day), CyA + L-NAME (5 mg/100 ml/day), CyA + L-Arg + L-NAME, VH + L-Arg, VH + L-NAME and were sacrificed at the end of the experiment. Body weight, serum creatinine, blood urea nitrogen (BUN) and NO levels were determined. Tubular injury and interstitial fibrosis were evaluated semiquantitatively using scoring systems on paraffin sections stained with hematoxylin/eosin (H/E), Masson’s trichromic and periodic acid-Schiff (PAS). Results: The CyA group developed marked renal injury, characterized by a significant increase in serum creatinine and BUN, and histopathological alterations including tubular dilatation, vacuolization, necrosis, interstitial cell infiltration and tubulointerstitial fibrosis. CyA reduced serum NO level. L-Arg treatment significantly enhanced NO biosynthesis and protected animals from CyA-induced kidney damage. In contrast L-NAME strikingly reduced serum NO level, and worsened biochemical and histopathological alterations. Conclusion: Chronic CyA nephrotoxicity can be aggravated by NO blockade and ameliorated by NO enhancement suggesting that L-Arg supplementation may be protective in CyA nephrotoxicity.     

Renal hemodynamic effects of L-NAME during postnatal maturation in conscious lambs

The purpose of the present study was to investigate the effects of the L-arginine analogue, N ^G -nitro-L-arginine methyl ester, L-NAME, in modulating renal hemodynamics during postnatal maturation in conscious chronically instrumented lambs. To this end, renal hemodynamic responses to intravenous injection of 10, 20, and 40 mg/kg L-NAME, as well as its inactive enantiomer D-NAME, were measured for 4 h in conscious lambs approximately 1 week (n=10), 3 weeks (n=12), and 6 weeks (n=14) of age. Administration of L-NAME was associated with an increase in renal vascular resistance (RVR) leading to a decrease in renal blood flow. One-week-old lambs were more sensitive to the effects of L-NAME, a marked increase in RVR occurring after the smallest dose administered at 1 week but not at 3 and 6 weeks of age. Renal hemodynamic effects of L-NAME as well as the duration of the responses were also age dependent, such that changes in RVR were greatest and most prolonged in 1-week-old lambs. In addition, a smaller dose of L-NAME was required to attenuate acetylcholine-induced renal vasodilation in lambs aged 1 week compared with older animals. Our data provide new evidence to support the premise that endogenously produced nitric oxide plays a predominant role in regulating renal vascular tone early in life. Received: 27 November 2000 / Revised: 30 May 2001 / Accepted: 6 June 2001     

Supplementation of l-arginine prevents glucocorticoid-induced reduction of bone growth and bone turnover abnormalities in a growing rat model

The present study was designed to evaluate the effects of glucocorticoid (GC) treatment on bone turnover and bone mineral density in the growing rat. Because of the recent evidence that nitric oxide (NO) can counteract prednisolone-induced bone loss in mature rats, we examined the effect on bone of the NO donor l-arginine in young male rats, in which bone mass is increased by the same biological mechanism as in children and adolescents. Thirty-six 10-week-old Sprague-Dawley male rats were assigned to six groups of six animals each, and treated for 4 weeks with either vehicle (once a week subcutaneous injection of 100µl of sesame oil); prednisolone sodium succinate, 5mg/kg, 5 days per week by intramuscular injection (i.m.); l-arginine, 10mg/kg intraperitoneally (i.p.) once a day; N^G-nitro-l-arginine methylester (l-NAME), 50mg/kg subcutaneously once a day; prednisolone sodium succinate 5mg/kg, 5 days per week i.m. +l-arginine 10mg/kg i.p. once a day; or prednisolone sodium succinate, 5mg/kg, 5 days per week i.m. +l-NAME 50mg/kg subcutaneously once a day. Serum calcium, alkaline phosphatase (ALP), osteocalcin, and the C-terminal telopeptides of type I collagen (RatLaps) were measured at baseline conditions and after 2 and 4 weeks. Prior to treatment, and after 2 and 4 weeks, the whole body, vertebral, pelvic, and femoral bone mineral density (BMD) was measured by dual-energy X-ray absorptiometry (DXA) scanning. Prednisolone and prednisolone + l-NAME treated rats had significantly lower ALP and osteocalcin levels than controls at 2 and 4 weeks, and significantly higher levels of Rat-Laps than controls at 4 weeks. Prednisolone, l-NAME, and prednisolone + l-NAME produced a significant inhibition of bone accumulation and bone growth at all sites measured. Supplementation with l-arginine appeared to prevent the inhibition of bone growth and increase in bone resorption induced by prednisolone. These data would suggest, for the first time, that supplementation with an NO donor could be considered as a treatment for steroid-induced osteoporosis in the developing skeleton.     

Endothelin-1 produces no renal vasoconstriction in conscious newborn lambs

Experiments were carried out to investigate the effects of endothelin-1 (ET-1) on renal vascular tone during development under physiological conditions in conscious lambs. Renal blood flow (RBF), renal vascular resistance (RVR), mean arterial pressure (MAP), and heart rate (HR) were measured in conscious, chronically instrumented lambs aged approximately 1 week and 6 weeks before and after intra-arterial (i.a.) injection of 0, 100, 200, and 400 ng/kg body weight of ET-1. In addition, plasma levels of ET-1 were measured in 39 sheep aged 5-85 days. In 6-week-old lambs, i.a. injection of ET-1 was associated with a rapid dose-dependent decrease in RBF that resulted from a dose-dependent increase in RVR. In 1-week-old lambs, there was no renal vasoconstriction observed after ET-1 administration, even at the highest dose tested. In response to i.a. injection of ET-1 to 1-week-old and 6-week-old lambs, MAP increased and there was a concomitant decrease in HR; these effects were dose dependent but not age dependent. Plasma levels of ET-1 were 10.7+/-4.2 pg/ml at 5-10 days, and remained constant throughout the first 3 months of life in conscious sheep. We conclude that ET-1 is not a renal vasoconstrictor agent in the immediate newborn period, and that the effects of ET-1 on renal vascular tone appear to be developmentally regulated.     

Is glandular formation of nitric oxide a prerequisite for muscarinic secretion of fructose in the guinea-pig seminal vesicle?

The significance of nitric oxide (NO) formation in seminal secretion was studied in guinea-pig seminal vesicles. The nitric oxide synthase (NOS) activity was estimated and reduced nicotinamide-adenine dinucleotide phosphate (NADPH)-diaphorase histochemistry was performed. Furthermore, cyclic guanosine 3,5-monophosphate (cGMP) concentration as well as fructose secretion from isolated vesicles was estimated. High Ca²⁺-dependent NOS activity as well as prominent glandular NADPH-diaphorase staining was found in the secretory epithelium. The NOS inhibitorsN ^G-nitrol-arginine methyl ester (L-NAME) andN ^G-nitrol-ar-ginine (L-NNA) inhibited carbachol-induced fructose secretion but the D-isomer to L-NAME had no effect. Whenl-arginine was administered together with L-NAME, no inhibitory effect on the carbachol-induced fructose secretion could be seen. Nerve-induced fructose secretion was also inhibited by L-NAME. The NO donor glyceryl trinitrate (GTN) increased the fructose secretion. Carbachol or GTN did not increase cGMP levels, nor was fructose secretion inhibited by a guanylate cyclase inhibitor (ODQ). Our results suggests that glandular NO production is a prerequisite for muscarinic fructose secretion in the seminal vesicle via a cGMP-independent pathway.     

Effects ofl-arginine andN-nitro-l-arginine treatment on hemodynamics, DO2, VO2, and extravascular lung water in a dog endotoxin shock model

To verify the effect of nitric oxide pathway modification during sepsis, experiments were conducted in four groups of anesthetized dogs which received lipopolysaccharide (LPS) intravenously (group 1), 300 mg·kg⁻¹ ofl-arginine plus LPS (group 2), 20 mg·kg⁻¹ ofN-nitro-l-arginine plus LPS (l-NNA, group 3), and normal saline as the control group. Hemodynamic and oxygenation data as well as extravascular lung water (EVLW) were measured or calculated. The results showed thatl-arginine increases cardiac output index (CI) and decreased the peripheral vascular resistance index (PVRI) without a significant influence on oxygen extraction ratio (O₂ER), oxygen delivery (DO₂), or oxygen consumption (VO₂). All of the untoward hemodynamic effects of LPS were exacerbated by the addition ofl-NNA. Therefore, as DO₂ was significantlys decreased byl-NNA, and although O₂ER was increased (insufficiently), VO₂ was still decreased significantly. EVLW was markedly increased byl-NNA. These results support the hypothesis that inhibition of nitric oxide synthesis may exacerbate hemodynamic and oxygenation consequences in septic shock.     

The Effect of <Emphasis Type="Italic">N</Emphasis>ω-Nitro-<Emphasis Type="SmallCaps">l</Emphasis>-Arginine Methyl Ester and <Emphasis Type="SmallCaps">l</Emphasis>-Arginine on Lung Injury Induced by Abdominal Aortic Occlusion–Reperfusion

Purpose

The aim of this study was to examine the effects of Nω-nitro-l-arginine methyl ester (l-NAME) and l-arginine on lung injury after aortic ischemia–reperfusion (IR).

Methods

Twenty-four Wistar-Albino rats were randomized into four groups (n = 6) as follows: Control (sham laparotomy), Aortic IR (30?min ischemia and 120?min reperfusion), l-Arginine (intraperitoneal 100?mg?kg<συπ>?1 live weight)+aortic IR, and l-NAME (intraperitoneal 10?mg?kg<συπ>?1 live weight)+aortic IR. In the lung specimens, the tissue levels of malondialdehyde (MDA), vascular endothelial growth factor (VEGF), and nitric oxide (NO) were measured and a histological examination was done.

Results

Aortic IR increased MDA, VEGF, and NO. l-Arginine further significantly increased MDA and NO, and decreased VEGF (P < 0.05 vs aortic IR). l-NAME significantly decreased MDA and NO (P < 0.05 vs l-arginine+aortic IR) and increased VEGF (P < 0.05 vs other groups). A histological examination showed the aortic IR to significantly increase (P < 0.05 vs control) while l-arginine also further increased (P > 0.05 vs aortic IR), whereas l-NAME caused a significant decrease in pulmonary leukocyte infiltration (P < 0.05 vs aortic IR).

Conclusions

Our results indicate that l-arginine aggravates the lung injury induced by aortic IR, while l-NAME attenuates it.

     

7-Nitroindazole, a selective inhibitor of neuronal nitric oxide synthase: effect on sevoflurane MAC and cerebellar cyclic GMP in mice

Purpose Considerable evidence suggests that nitric oxide (NO) plays a role in synaptic transmission in the central and peripheral nervous systems. However, whether inhibition of NO synthesis decreases minimum alveolar concentration (MAC) of inhalational anesthetics is controversial. We examined the effects of 7-nitroindazole (7-NI), a selective inhibitor of neuronal NOS (nNOS), on the MAC of sevoflurane and cerebellar cyclic guanosine monophosphate (cGMP) levels in mice. Methods Sevoflurane MAC and cerebellar cGMP levels were determined in mice after acute intraperitoneal or weeklong gavage feeding of 7-NI. Sevoflurane MAC and cerebellar cGMP levels after chronic treatment were measured on days 1, 4, and 7 and were repeated after an acute intraperitoneal dose of nitro ^g -l-arginine methylester (l-NAME). Results Acute and chronic treatment with 7-NI decreased the sevoflurane MAC by 20%–30%. Reduction of cerebellar cGMP levels was greater after intraperitoneal administration of NOS inhibitors than after gavage feeding of 7-NI. Conclusion Acute or chronic selective inhibition of neuronal NOS decreases the sevoflurane MAC and cerebellar cGMP levels in mice. 7-NI permitted probing of the role of NO in perception of noxious stimuli.     

-Arginine given after ischaemic preconditioning can enhance cardioprotection in isolated rat hearts

Objective: Ischaemic or pharmacological preconditioning with -arginine has been reported to be insufficient for optimal cardioprotection. The ability of nitric oxide (NO) to enhance ischaemic preconditioning was assessed, and the role of -arginine-induced ischaemic preconditioning in myocardial protection was determined. Methods: Isolated rat hearts were prepared and divided into six groups: control hearts (control, n=6) were perfused without global ischaemia at 37°C for 160 min; global ischaemia hearts (GI, n=6) were subjected to ischaemia for 20 min and reperfusion for 120 min; ischaemic preconditioned hearts (IP, n=6) received 2 min of zero-flow global ischaemia followed by 5 min reperfusion, before 20 min of global ischaemia; -arginine hearts (ARG, n=6) received 1 mmol/l -arginine for 5 min, before 20 min of global ischaemia; ischaemic preconditioning plus nitro- -arginine methyl ester hearts (IP+ -NAME, n=6) received 2 min of ischaemic preconditioning and 5 min reperfusion with 3 mmol/l -NAME in Krebs–Henseleit buffer, before 20 min of global ischaemia; and ischaemic preconditioning plus -arginine hearts (IP+ARG, n=6) received 2 min of ischaemic preconditioning and 5 min reperfusion with 1 mmol/l -arginine in Krebs–Henseleit buffer. Haemodynamic parameters and coronary flow were recorded continuously. Nitrites and nitrates (NOx) were measured 5 and 60 min after reperfusion, and infarct size was also determined. Results: In the IP+ARG group, significant amelioration and preservation of left ventricular peak developed pressure and coronary flow was observed compared with the GI, IP, ARG and IP+ -NAME groups. Infarct size in the IP+ARG group was reduced significantly compared with that in the GI, IP, ARG and IP+ -NAME groups. Significant preservation of NOx was observed during reperfusion in the IP+ARG group compared with the GI group. Conclusions: Inhibition of NO synthase with -NAME had little impact on ischaemic preconditioning, suggesting that endogenous NO is not a major mediator of ischaemic preconditioning. Nevertheless, enhancement of the effects of ischaemic preconditioning can be achieved with -arginine, a precursor of NO, improving post-ischaemic functional recovery and infarct size in the isolated rat heart.     

An in vitro study of nonadrenergic-noncholinergic activity on the cavernous tissue of mouse

The relaxant effects of electrical field stimulation (EFS) and exogenously applied acetylcholine (ACh) or acidified NaNO₂ (a-NaNO₂) were investigated in the isolated mouse corpus cavernosum precontracted with phenylephrine hydrochloride (PE). Tetrodotoxin (TTX) blocked the relaxant effects of EFS completely, whereas it had no effect on the responses to ACh or a-NaNO₂. Guanethidine and indomethacin failed to affect the electrically or ACh-induced relaxations. Atropine completely blocked the effect of ACh; however, it caused a slight reduction in the relaxation evoked by EFS.N ^G- Nitro-l-arginine (l-NOARG) reduced the effects of EFS and ACh significantly, but it was ineffective on the relaxations induced by a-NaNO₂. The inhibitory action ofl-NOARG was partly restored byl-arginine, but not byd-arginine. Methylene blue (MB) and hydroxocobalamin (HC) exhibited significant inhibition on the relaxations evoked by EFS, ACh and a-NaNO₂. Hydroquinone (HQ) reduced relaxation due to a-NaNO₂, but did not affect that of EFS and ACh. Our findings suggest that EFS-induced relaxations of mouse cavernosal tissue are mediated by a transmitter which probably resembles an organic nitrate.     

Effects of nitric oxide synthase inhibitors on bone formation in rats

NO is synthesized from -arginine by at least three isoforms of nitric oxide synthase (NOS) and is known to function as a vasodilator and neurotransmitter. NO is produced by bone cells but its function in bone biology is, as yet, unclear. We hypothesized that NOS mediates bone formation in remodeling regions of the skeleton. We studied the effects of two NOS inhibitors: N^G-nitro- -arginine methyl ester ( -NAME), which is a general inhibitor of NOS activity and is known to inhibit the vasodilatory effects of the endothelial NOS (eNOS) isoform; and aminoguanidine, which is a selective inhibitor of the inducible NOS (iNOS) isoform. Our hypothesis was tested by treating rats with NOS inhibitors and measuring bone formation rates in the tibial epiphysis and diaphysis. Bone formation indices were measured using standard bone histomorphometry. -NAME treatment significantly raised mean arterial blood pressure (MAP). This effect was partially reversed by addition of -arginine. Aminoguanidine had no significant effect on MAP, indicating that it did not block eNOS. The treatments also had substantial effects on bone formation in remodeling trabecular bone. -NAME did not significantly change trabecular bone formation rate, whereas aminoguanidine reduced bone formation rate in the tibial epiphysis by 79% compared with control. This reduction was completely reversed by -arginine, suggesting that bone formation during remodeling is, in part, mediated through -arginine metabolism. No effect of aminoguanidine on bone formation was seen in the tibial diaphysis, a site that undergoes minimal bone remodeling. This finding suggests that the -arginine-NO pathway is important in bone remodeling.     

Physiologic Studies on Nitric Oxide in Rat Small Bowel Isografts

The enteric nervous system (ENS), especially the nonadrenergic noncholinergic (NANC) inhibitory nerves, is an important factor in intestinal peristalsis. Recently, it was established that nitric oxide (NO) is released after stimulation of NANC inhibitory nerves. Inhibitory nerves such as NANC inhibitory nerves in the ENS are more easily damaged than excitatory nerves by reperfusion or ischemic injuries during small bowel transplantation (SBT). To evaluate the effects of reperfusion and ischemic injuries to the ENS in the transplanted small bowel, we examined the ENS responses, including the effects of NO in the isografted rat jejunum, using the nontransplanted jejunum as a control. To avoid potentially confounding immune phenomena, we used syngeneic Lewis (LEW) rats. Orthotopic entire SBT with portocaval drainage was performed from LEW rats to LEW rats. Isografted muscle strips were obtained from 8 LEW rats 130 days after SBT (n = 24). As controls, normal muscle strips of the jejunum were obtained from 20 nontransplanted LEW rats (n = 60). A mechanograph was used to evaluate in vitro jejunal responses to electrical field stimulation of the adrenergic and cholinergic nerves before and after treatment with various autonomic nerve blockers, N ^g -nitro-l-arginine (L-NNA), and l-arginine. The results indicated that excitatory nerves, especially NANC excitatory nerves, were more dominant in the isografted jejunum than in the normal jejunum (p < 0.01). NANC inhibitory nerves were found to act on the normal jejunum and to a lesser extent on the isografted jejunum (p < 0.05). NO mediates the relaxation reaction of NANC inhibitory nerves in the normal jejunum and to a lesser extent in the isografted jejunum. These results indicated that the intrinsic intestinal innervation contains excitatory and inhibitory nerves and that the former, especially NANC excitatory nerves, are more dominant in the isografted jejunum than in the normal jejunum. In addition, reduction of the action of NANC inhibitory nerves such as that by NO may be largely related to impaired motility in the isografted jejunum. Thus over a long period of time (more than 130 days after SBT) transplanted small bowel dysmotility may be influenced by reperfusion or ischemic injury to the ENS (especially NANC inhibitory nerves) via NO in the transplanted jejunum after syngeneic SBT.     

Role of endogenous endothelin in renal haemodynamics of newborn rabbits

Endothelin is a potent vasoconstrictor peptide produced by vascular endothelial cells which could play a role in the physiological regulation of the renal microcirculation. To test this hypothesis, experiments were performed in 24 anaesthetized and mechanically-ventilated newborn rabbits. In 8 newborn rabbits (group 1), a bolus injection of 5 nmol/kg endothelin caused a marked increase in mean blood pressure (MBP) and renal vascular resistance (RVR), leading to a significant fall in glomerular filtration rate (GFR) (by 12%±4%), and renal blood flow (RBF) (by 16%±3%). A second group of animals (n=8) was used for testing the in vivo neutralizing activity of an endothelin-1 antiserum. The antiserum was thereafter infused into 8 additional newborn rabbits (group 3) in order to define the role of endogenous endothelin in modulating the function of the immature kidney. The antiserum induced a surprising increase in RVR (by 34%±9%,P<0.05) associated with a fall in GFR (by 21%±4%,P<0.05) and RBF (by 25%±4%,P<0.05), while the filtration fraction and MBP remained unchanged. The occurrence of a vasoconstrictive response to both high-dose endothelin and to its antiserum could be explained by the recent demonstration that high levels of endothelin lead to renal vasoconstriction, while lower levels induce renal vasodilation. The present results suggest that endogenous endothelin is active at low levels under normal conditions and that this peptide plays a role in the physiological control of renal function, but not MBP.     

Protective effect of trapidil and l-arginine against renal and hepatic toxicity induced by cyclosporine in rats

Rationale: Cyclosporine A (CsA) leads to renal and liver injury, production of free radicals and nitric oxide (NO) deficiency. This study investigates the possible protective effects of trapidil and l-arginine against CsA-induced tissue injury. Objectives: Forty adult male Wistar rats (180 ± 20 g) were divided into five groups, eight animals in each. The first group served as control, second group served as CsA group, third group served as CsA + trapidil group, fourth group served as CsA + l-arginine group, and fifth group served as CsA + trapidil + l-arginine group. Kidney and liver functions, inflammatory mediators, cytokines, oxidant and antioxidant parameters as well as histopathological studies of renal and liver tissue were assessed in all groups. Main findings: CsA induced renal and hepatic dysfunction, which was confirmed by laboratory and histopathological examination. Administration of trapidil diminished the renal and liver injury and significantly attenuated the levels of serum creatinine, urea, aspartate aminotransferase (AST), alanine aminotransferase (ALT), interleukin-1β (IL-1β), tumor necrosis factor alpha (TNF-α), monocyte chemoattractant protein-1 (MCP-1), and oxidative stress, while it significantly elevated the level of serum nitric oxide and the activity of antioxidative stress. l-Arginine gave the same trend as trapidil, but trapidil effect was more pronounced. Coadministration of trapidil + l-arginine significantly ameliorated the toxic effect of CsA, but did not differ significantly from the effect of trapidil alone. Conclusions: Treatment with trapidil or l-arginine diminished the renal and hepatic CsA-induced toxicity. However, the effect of trapidil was more pronounced. Therefore, treatment with trapidil alone may be the most economic and effective as a potential therapeutic agent in CsA injury.     

The Effect of <Emphasis Type="SmallCaps">l</Emphasis>-arginine and Aprotinin on Intestinal Ischemia–reperfusion Injury

Background Intestinal ischemia/reperfusion (I/R) results in local mucosal injury, systemic injuries, and organ dysfunction. These injuries are characterized by altered microvascular and epithelial permeability and villous damage. Activation of neutrophils, platelets, and endothelial factors are known to be involved in this process. Cytokines such as TNF-α, IL-1, IL-6, and oxygen-derived free radicals are believed to be important pathogenic mediators. Capillary no-reflow is also known to play a role in I/R. The aim of our study was to examine the role of l-arginine, a known nitric oxide (NO) donor, and aprotinin, a protease inhibitor with multiple effects, on intestinal I/R. Methods Pigs weighing 20–25 kg were used. Ischemia was established by clamping the superior mesenteric artery (SMA) at its origin and was sustained for 2 hours. Duration of reperfusion was 2 hours. The animals were divided into four groups: group A, the control group, which was submitted to I/R injury only; group B, in which l-arginine was administered at a rate of 5 mg/kg/min during ischemia and continuing throughout reperfusion; group C, in which aprotinin was administered with an initial bolus dose of 20,000 U/kg during ischemia followed by a continuous dose at 50 U/hour throughout reperfusion; and group D in which both substances were administered. In all groups TNF-α, IL-1, and IL-6 levels were measured using ELISA at baseline, 2 hours of ischemia, and 1 hour and 2 hours of reperfusion. SMA blood flow was measured with a Doppler probe at baseline, 10 min, 1 hour, and 2 hours of reperfusion. Histological changes of the intestinal mucosa were examined and graded on a five-point scale in all groups. Results In the control group, levels of TNF-α, IL-1, and IL-6 were significantly increased during reperfusion (p < 0.05) compared to baseline. Administration of l-arginine and aprotinin led to suppression of the release of TNF-α, IL-1, and IL-6 during reperfusion in a statistically significant manner (all p < 0.05). A synergistic or additive effect of l-arginine and aprotinin was not observed. SMA blood flow in the control group was decreased (p > 0.05) during reperfusion compared to baseline. In animals treated with l-arginine and aprotinin, SMA blood flow during reperfusion was significantly increased (p < 0.05) compared to the control group. Histologic examination of the intestinal mucosa was characterized by flattening of the villi and necrosis in the control group. In the treated animals, less severe histological changes were noted. Conclusions Administration of l-arginine and aprotinin may lead to amelioration of intestinal I/R injury. We did not note a synergistic or additive effect of these two substances. These findings warrant further studies in clinical settings for future treatment efforts. This paper was presented as a poster at the 47th Annual Meeting of the Society for Surgery of the Alimentary Tract, Los Angeles, California, May 20–24, 2006.     

The effect of intraoperative colonic lavage withN G -nitro-l-arginine methyl ester (l-NAME) on anastomotic healing in the presence of left-sided colonic obstruction in the rat

-arginine methyl ester (L-NAME) on the healing of colonic anastomosis in the presence of a left-sided obstruction in the rat was investigated. Left-sided colonic obstruction was created in 144 Wistar rats. The obstruction site was excised 24 h later and anastomosis was performed after either no irrigation or colonic lavage with either saline, povidone iodine (PI), short-chain fatty acids (SCFA), L-NAME, or glutamine, in 24 animals each. Animals were killed on days 3 and 6, and a 4-cm colonic segment with the anastomosis at the center was excised. Bursting pressure (BP) and hydroxyproline (HP) content were measured. In the saline, PI, and SCFA groups, BP was higher (P < 0.05, P < 0.05, and P < 0.001, respectively) and HP concentration was similar compared with controls. Both the BP and HP concentrations were higher in the glutamine group compared with controls (P < 0.001). BP was lower (P < 0.05) and HP concentration was similar in the L-NAME group compared with the control group. Colonocyte nutrition and tissue perfusion are the mainstays of anastomotic healing. Intraoperative colonic lavage with L-NAME suppresses colonic anastomotic healing in the presence of a left-sided obstruction. (Received for publication on Mar. 29, 1999; accepted on Nov. 11, 1999)     

The role of intrapulmonary nitric oxide generation in the development of adult respiratory distress syndrome

N ^G-monomethyl-l-arginine. These observations indicate that the intrapulmonary generation of NO may play an important role in the development of ARDS. (Received for publication on Aug. 12, 1998; accepted on May 28, 1999)     

A possible role for nitric oxide in the regulation of human ureteral smooth muscle tone in vitro

There is ample evidence that nitric oxide (NO) is an important neurotransmitter in many tissues of the urogenital tract. The aim of the present study was to examine the possible role of NO in ureteral relaxation. Human ureteral rings were mounted in organ bath chambers and precontracted with KCl. Increasing doses of the NO donor linsidomine (SIN-1) were added with and without prior blockade of the NO/cGMP pathway by methylene blue and protein kinase (PK) inhibitors Rp-8-pCPT-cGMPS and Rp-8-CPT-cAMPS. Electrical field stimulation (EFS) was done before and after incubation with L-NOARD (N ^G-nitro-L-arginine) and TTX (tetratodoxin). For detection of neuronal NO synthase (NOS), ureters were stained immunohistochemically. Ureteral strips were dose dependently relaxed by SIN-1; preincubation with methylene blue and protein kinase G inhibitor significantly reduced the SIN-1-induced relaxations. No effects of L-NOARG and TTX on EFS-induced tone alterations were found. NOS-positive neuronal axons and nerve-ending-like structures were found in the muscular layers. Our in vitro findings suggest that ureteral relaxation may involve the NO pathway.