BACKGROUND: According to target cells, apoptosis-inducing agents, and NO concentration, NO concentration, NO shows both pro- and antiapoptotic effects. OBJECTS: Our study was perfermed to verify the role of NO in UVB-induced apoptosis in HaCaT cells. METHODS: After UVB irradiation, FACS using propidium iodide, LDH cytotoxicity assay, and nitrite assay based on Griess reaction were done in HaCaT cells. These procedures were repeated after UVB irradiation and NG-monomethyl-arginine (L-NMMA), a NO synthase (NOS) inhibitor, addition. RESULTS: 1) UVB irradiation (5-80mJ/cm2) induced apoptosis in HaCaT cells dose-dependently. 2) UVB irradiation (80mJ/cm2) stimulated NO production 30-50% more over baseline level, and this was inhibited by 500 micrometer L-NMMA. 3) 500 micrometer L-NMMA did not inhibit UVB-induced apoptosis. CONCLUSION: UVB irradiation evokes apoptosis in HaCaT keratinocytes through NO-independent mechanism.
Apoptosis* ; Keratinocytes* ; Nitric Oxide Synthase ; Nitric Oxide* ; omega-N-Methylarginine ; Propidium

PURPOSE: We investigate the effects of intravesical BCG therapy on the occurance of superficial bladder cancer induced by N-butyl-N-(4-hydroxybutyl) nitrosamine(BBN) in Fisher 344 rats in vivo. We also examine whether NO mediated the antitumor activity of BCG against superficial bladder cancer in Fisher 344 rats. MATERIALS AND METHODS: BBN(0.1%) is orally administered for 20 weeks and it is combined with BCG(0.27mg) or NG-monomethyl-L-arginine (NG MMA, 10mg). once every week from 8th week to 19th week. The rats are sacrified at 20th week. NO secretion in urine for 24 ours is significantly increased in the BCG treated rats compared to the animals treated with saline or NGMMA. RESULTS: Pathologic findings demonstrate that the incidence of carcinoma is not statistically different in saline, BCG, NGMMA(p>0.05). However the size and number of tumor is decreased in the BCG treated rats compared with saline or NGMMA treated rats bearing bladder cancer induced by BBN(p<0.05). Inducible NO synthase(iNOS) is strongly induced in bladder tissue of rats treated with BCG and NGMMA but not in saline. CONCLUSIONS: Intravesical instillation of BCG on bladder cancer induced by BBN does not decrease the cancer occurrence but reduces the number and size of bladder cancer. Our results suggest that nitric oxide induced by intravesical instillation of BCG may mediate antitumor activity against the occupance of superficial bladder cancer.
Administration, Intravesical ; Animals ; Incidence ; Mycobacterium bovis* ; Nitric Oxide ; omega-N-Methylarginine ; Rats ; Urinary Bladder Neoplasms* ; Urinary Bladder*

BACKGROUND: Aging is associated with increased cardiovascular risk and firmly established as a risk factor for the development of atherosclerosis. However the exact mechanism of age-related damage to the arterial wall and its relation to the atherosclerotic process are not well known.The endothelium plays an important role for the regulation of vascular tone and the endothelial function is impaired in the presence of risk factors early in the process of atherosclerosis. Assessment of endothelial function appears to be a valuable tool for the diagnosing and therapeutic monitoring of coronary artery disease. Anti-oxidants are known to improve endothelial dysfunction in atherosclerosis patients. The aim of this study was, (1) to evaluate the endothelial function in elderly, (2) to investigate whether vitamin C administration has benefit on the endothelial function in elderly. METHODS: The endothelial function was estimated using venous occlusion plethysmography(VOP) in 7 elderly and 7 young healthy volunteers. The strain guage was connected to plethysmograph to record the forearm volume change. A rapid cuff inflator was used to inflate the arm cuff to 40 mmHg instantaneuosly thus occluding venous return from the forearm. The measurement of forearm volume change was repeated for 7 times each stage. The change of the forearm blood flow(FBF) was measured with the acetylcholine infusion through brachial artery and also with intra-arterial vitamin C. RESULTS: Endothelium-dependent vasodilatation was significantly impaired in the elderly group compared to the young group(321 +/-17% in elderly group vs 509 +/-81%, mean+/-SEM) Forearm blood flow response to acetylcholine was significantly enhanced with inraarterial infusion of vitamin C in elderly group(321+/-17% in elderly group vs 78% in vitamin C) Coinfusion of L-NMMA, an inhibitor of nitric oxide synthase, blunted forearm blood flow response to acetylcholine. CONCLUSIONS: Even though the mechanisms leading to drpressed endothelial function in elderly remains to be elucidated, our study shows that vitamin C result in demonstrable improvement by a mechanism that is probably related to antioxidant activity.
Acetylcholine ; Aged* ; Aging ; Arm ; Ascorbic Acid* ; Atherosclerosis ; Brachial Artery ; Coronary Artery Disease ; Endothelium ; Forearm ; Healthy Volunteers ; Humans ; Nitric Oxide Synthase ; omega-N-Methylarginine ; Risk Factors ; Vasodilation* ; Vitamins*

OBJECTIVETo investigate the effects of repeated injections of L-NMMA on a goat model with osteoarthrotic temporomandibular joint (TMJ) disease.

METHODSEight goats were selected in this study. Bilateral TMJ osteoarthrosis (OA) was induced by injecting 0.5% collagenase. L-NMMA was injected into one side of TMJs at 4 weeks after collagenase injection (one time every three days). Another joint as control was simultaneously injected using 0.9% saline solution. All goats were killed at 12 weeks after collagenase injection. The TMJ specimens were harvested and processed for histological examination. Modified Mankin's grading score system was used for evaluating changes in the TMJ.

RESULTSThe control side of TMJs showed severe osteoarhrotic changes in the condyle whereas the L-NMMA-treated TMJs showed less degenerative alterations. The histologic score was 3.83 in the L-NMMA treated side, and 6.33 in the control. There was a significant difference in osteoarthrotic changes between the L-NMMA-treated and control TMJs (P < 0.01).

CONCLUSIONSRepeated intra-articular injection of L-NMMA into TMJ may play a role in inhibiting TMJOA progression.

Animals ; Enzyme Inhibitors ; administration & dosage ; therapeutic use ; Female ; Goats ; Injections, Intra-Articular ; Male ; Osteoarthritis ; drug therapy ; Temporomandibular Joint Disorders ; drug therapy ; omega-N-Methylarginine ; administration & dosage ; therapeutic use

To examine the role of nitric oxide in the beta-adrenergic vasodilation of epicardial coronary arteries in dogs, 12 dogs were instrumented for measurement of left anterior descending coronary artery diameter by transthoracic echocardiography before and after dobutamine (5 microg/kg/min IV) with and without intracoronary infusion of NG-monomethyl-L-arginine (L-NMMA) (1 mg/kg). In all 12 dogs, the diameter of left anterior descending coronary artery increased significantly from 2.35 +/- 0.25 mm to 2.59 +/- 0.24 mm (P<0.001) after dobutamine administration. In 6 of the 12 dogs, the percent change in left anterior descending coronary artery diameter induced by dobutamine decreased significantly from 12.5% +/- 8.6% to -1.5% +/- 5.4% (P<0.05) after the administration of intracoronary L-NMMA (1 mg/kg for 5 min) to block nitric oxide synthesis from L-arginine. The study demonstrated that nitric oxide formation contributes to the beta-adrenergic dilatory response of epicardial coronary arteries to dobutamine in dogs.
Adrenergic beta-Agonists ; pharmacology ; Animals ; Coronary Vessels ; physiology ; Dobutamine ; pharmacology ; Dogs ; Echocardiography ; Female ; Male ; Nitric Oxide ; physiology ; Receptors, Adrenergic, beta ; physiology ; Vasodilation ; physiology ; omega-N-Methylarginine ; pharmacology

Vascular endothelial growth factor (VEGF) is an important regulator of neovascularization. Hypoxia inducible nitric oxide (NO) enhanced the expression of VEGF and thymosin beta-4 (Tbeta4), actin sequestering protein. Here, we investigated whether NO-mediated VEGF expression could be regulated by Tbeta4 expression in HeLa cervical cancer cells. Hypoxia inducible NO production and VEGF expression were reduced by small interference (si) RNA of Tbeta4. Hypoxia response element (HRE)-luciferase activity and VEGF expression were increased by the treatment with N-(beta-D-Glucopyranosyl)-N2-acetyl-S-nitroso-D, L-penicillaminamide (SNAP-1), to generate NO, which was inhibited by the inhibition of Tbeta4 expression with Tbeta4-siRNA. In hypoxic condition, HRE-luciferase activity and VEGF expression were inhibited by the treatment with N(G)-monomethyl-L-arginine (L-NMMA), an inhibitor to nitric oxide synthase (NOS), which is accompanied with a decrease in Tbeta4 expression. VEGF expression inhibited by L-NMMA treatment was restored by the transfection with pCMV-Tbeta4 plasmids for Tbeta4 overexpression. Taken together, these results suggest that Tbeta4 could be a regulator for the expression of VEGF via the maintenance of NOS activity.
Actins ; Anoxia ; Nitric Oxide Synthase ; Nitric Oxide* ; omega-N-Methylarginine ; Plasmids ; Response Elements ; RNA ; Thymosin* ; Transfection ; Uterine Cervical Neoplasms* ; Vascular Endothelial Growth Factor A*

BACKGROUND: When used to reverse the anticoagulant effect of heparin, protamine administration after cardiovascular bypass often can lead to systemic hypotension. During the reversal of heparin-induced anticoagulation, the effects of protamine on both a heparin-protamine complex and free protamine on the cardiovascular system should be considered. METHOD: To determine whether the hypotensive effect of heparin-protamine and/or protamine could be caused by endothelium-dependent and-independent component, we studied rings of the arotic arteries in rats suspended in organ chambers containing Tris Tyrode solution at 37oC and 100% O2. Arterial rings with or without endothelium were contracted with 40 mM KCl or 3 +/- 10-6M phenylephrine and then exposed to increasing concentrations of protamine (final organ bath concentration, 40~400 g/ml) both in the absence and presence of heparin (200 U/ml). RESULTS: Protamine induced concentration-dependent relaxation in arterial rings with endothelium, which were significantly greater than in rings without endothelium. The endothelium-dependent relaxation induced by protamine was inhibited by NG-monomethyl-L-arginine (L-NMMA) (10-5M) pretreatment, but was not inhibited by indomethacin (3x10-6M) pretreatment on rings with endothelium. Furthermore, the contractile inhibition was enhanced by superoxide dismutase (100 U/ml). Also, such vasodilating actions were not influenced in the presence of heparin (200 U/ml). In endothelium-denuded strips, protamine (400ug/ml) inhibited Ca++ induced contraction, which was evoked in Ca++-free solution containing 40 mM K+, and also inhibited the norepinephrine (NE)-induced contraction. Protamine inhibited on the NE-induced contraction, but not the caffein-induced contration in Ca++ free, 2 mM EGTA solution. Also, such inhibition of contracions were not inluenced in the presence of heparin (40 U/ml). CONCLUSION: This study demonstrates that protamine (in the presence or absence of heparin) acts on endothelial cell receptors to stimulate the production of nitric oxide and inhibits both Ca++-influx and the NE-induced Ca++ release from intracellular stores.
Animals ; Arteries* ; Baths ; Cardiovascular System ; Egtazic Acid ; Endothelial Cells ; Endothelium ; Heparin* ; Hypotension ; Indomethacin ; Nitric Oxide ; Norepinephrine ; omega-N-Methylarginine ; Phenylephrine ; Rats* ; Relaxation ; Superoxide Dismutase ; Vasodilation*

Externally applied acetylcholine(Ach) in corpus cavernosum has been shown to cause endothelium dependent smooth muscle relaxation. ATP is accepted as a relaxant of smooth muscle by both a direct action and more powerful indirect action via the endothelial cells. Endothelium-derived relaxing factor(EDRF) is released with stimulation of acetylcholine or other endothelium dependent substances raise cGMP level within the smooth muscle cell and cause relaxation of smooth muscle. EDRF is known as nitric oxide(NO) and its actions are abolished by specific inhibitor of nitric oxide synthesis, such as L-n-monomethyl arginine(L-NMMA) or inhibitors of cyclic GMP synthesis, such as methylene blue(MB). In this study, we evaluated the action of ATP related with NO and compared effect of ATP with acetylcholine and bethanechol chloride in rabbit corpus cavernosal smooth muscle under organ bath. Changes in isometric tension of corporal strips were monitored. With pretreatment L-NMMA or MB, relaxing effects of acetylcholine or bethanechol chloride in corporal strips were completely inhibited, but relaxing effects of ATP were not altered. These data suggested that nitric oxide plays a crucial role in cholinergically induced cavernosal smooth muscle relaxation. ATP mediated rabbit corporal smooth muscle relaxation was not affected by inhibitors of nitric oxide synthesis and independent of cyclic GMP accumulation.
Acetylcholine ; Adenosine Triphosphate* ; Adenosine* ; Baths ; Bethanechol ; Cyclic GMP ; Endothelial Cells ; Endothelium ; Muscle, Smooth* ; Myocytes, Smooth Muscle ; Nitric Oxide ; omega-N-Methylarginine ; Relaxation*

One of the pathophysiologic change of priapism is known as hypoxic condition of corpus cavernosum. In vitro study of corpus cavernosum under hypoxia showed suppressed endothelium- dependent relaxation caused by cholinergic and nonadrenergic noncholinergic neuroeffector system, but in vivo study it is not fully evaluated yet. So this study aimed to identify the changes of corpus cavernosum related to cholinergic neuroeffector system and endothelium derived relaxation factor (EDRF) under hypoxia in animal study and to understand the Physiologic change of priapism. Under the general anesthesia with tracheostomy, adult male cats were conditioned at normoxia and hypoxia with ventilation. Acetylcholine, Nc-monomethyl-L- arginine (L-NMMA) and L-arginine was infused via internal pudendal artery. The change of intracavernosal pressure in response to drugs were monitored with physiograph (Gilson, IC-MP) in both normoxic and hypoxic state. The relaxation effect of acetylcholine under hypoxia was weaker than under normoxia (n=5, p suppressed by L-NMMA under normoxia but not under hypoxia. L-arginine showed the relaxation effect under normoxia but, no relaxation under hypoxia. These result suggest that acetylcholine induced relaxation was influenced in a some by hypoxic condition but not suppressed completely. EDRF pathway via nitric oxide synthesis does not play a role in relaxation of cat corpus cavernosum under hypoxia.
Acetylcholine ; Adult ; Anesthesia, General ; Animals ; Anoxia* ; Arginine ; Arteries ; Cats* ; Endothelium ; Humans ; Male ; Nitric Oxide* ; omega-N-Methylarginine ; Priapism ; Relaxation* ; Tracheostomy ; Ventilation

Recently oxygen free radicals and nitric oxide (NO) are known to play an important role in neuronal reperfusion injury. This study was aimed to investigate the role of oxygen f ree radicals and NO during cerebral ischemia/reperfusion, using dimethylthiourea (DMTU) and NG-monomethyl-L-arginine (NMMA), an oxygen f ree radical scavenger and a competitive NOS inhibitor respectively. In the in vivo experiment, the ischemia/reperfusion-induced changes of cerebral biogenic amines were examined in Mongolian gerbil (Meriones unguiculatus) pre-treated with NMMA and/or DMTU. To induce cerebral ischemia/reperfusion, bilateral common carotid arteries were clamped for 10 minutes and then released for 15 minutes. The biogenic amines were measured by using HPLC-ECD(High Performance Liquid Chromatography-Electrochemical detection). To confirm the results from the in vivo experiments, the effect of NMMA and/or DMTU on [3H]dopamine release from striatal slices exposed to hypoxia was investigated. The results are as follows; 1) Ischemia/reperfusion increased the ratio of DOPAC/dopamine and HVA/dopamine as well as the concentrations of DOPAC and HVA, which were evident only in corpus striatum. 2) NMMA attenuated the ischemia/reperfusion-induced increase in the ratio of DOPAC/dopamine in corpus striatum. However, the change of DOPAC or HVA was minimal. 3) DMTU attenuated the ischemia/reperfusion-induced increase of DOPAC and HVA, and the ratio ofDOPAC / dopa- mine and HVA/dopamine in corpus striatum. 4) Simultaneous pre-treatment with NMMA and DMTU attenuated the ischemia/reperfusion-induced increase of DOPAC and HVA, and the ratio Of DOPAC/dopamine and HVA/dopamine in corpus striatum. The extent of attenuation was greater than the single treatment group with NMMA or DMTU. 5) Exposure to hypoxia markedly increased the release of [3H]dopamine in the striatal slices. 6) The administration of either NMMA or DMTU attenuated the increase of [3H]dopamine release induced by hypoxia in the striatal slices. 7) The administration of both NMMA or DMTU markedly attenuated the increase of [3H]dopamine release induced by hypoxia to the extent of the control in the striatal slices. These results suggest that oxygen free radicals play an important role in cerebral ischemia/reperfusion injury, for which NO seems to be responsible.
3,4-Dihydroxyphenylacetic Acid ; Anoxia ; Biogenic Amines* ; Carotid Artery, Common ; Corpus Striatum ; Dopamine ; Free Radicals* ; Gerbillinae ; Ischemia ; Neurons ; Nitric Oxide* ; omega-N-Methylarginine ; Oxygen* ; Reperfusion ; Reperfusion Injury