The mechanism of vasodilation effect of GS-389, 1-(4'-methoxybenzyl)-6,7 -dimethoxy-1,2,3,4-tetrhydroisoquinoline hydrocholoride, a possible cyclic GMP specific phosphodiesterase inhibitor, on rat and mouse thoracic aorta ring has been investigated. GS-389 relaxed rat and mouse thoracic aorta precontracted with phenylephrine and high K+(60mM) in concentration dependent manner. Presence or absence of endothelium did not alter the relaxing effects of it. GS-389 inhibited Ca2+-induced contraction of the high K+ or 1 uM phenylephrine. Initial phasic contraction induced by phenylephrine and caffeine in Ca2+ free solution was inhibited by GS-389. Methylene blue pretreatment suppressed relaxation effect of GS-389. Relaxation by isoproterenol or sodium nitroprusside and by acetylcholine in endotheilium preserved aorta was potentiated by concurrentadministration of GS-389. GS-389 inhibited phenylephrine-induced phosphatidylinositide hydrolysis. It is suggested that inhibition of phosphoinositide turnover associated with elevated cyclic nucleotide by GS-389 may be the possible vascular relaxation mechanism of it.
Acetylcholine ; Animals ; Aorta ; Aorta, Thoracic ; Caffeine ; Cyclic GMP ; Endothelium ; Hydrolysis ; Isoproterenol ; Methylene Blue ; Mice ; Nitroprusside ; Phenylephrine ; Rats ; Relaxation* ; Vasodilation

BACKGROUND/AIM: Abnormality in GB motility is related with many gallbladder diseases including GB stone. Gallbladder motility is controlled by both hormonal and neural mechanisms. CCK, gastrin, motilin play a role on gallbladder contraction and VIP, somatostatin are inhibitory agents. Nitric oxide(NO) is known to account for the biologic properties of endothelium dependant relaxing factor. It also plays an important role in mediation of relaxation in various types of non-vascular smooth muscle of GI tract. The objective of this study was to determine the effect of nitric oxide in human gallbladder muscle. METHOD: In this study, nitric oxide was generated by photolysis using long wave-length UV lamp(366 nm) on NO carrying molecule, streptozotocin. GB muscle strips were obtained from 10 cholecystectomized patients and contracted by potassium or CCK-8. We also investigated the effect of methylene blue, which is a inhibitor of guanylate cyclase, after addition of methylene blue to the organ bath containing streptozotocin. Gallbladder movements were recorded using Polygraph(Grass model 79E, USA). And we identified the production of nitric oxide using nitrite assay in our No generating system. RESULTS: 1. Streptozotocin, No containing compound, released NO when UV irradiated. The longer UVR and the higher concentration of STZ, the larger is the amount of produced NO. 2. The human GB muscle was relaxed immediately by photo-induced NO and rapidly disappeared. The maximal relaxation under STZ, 60 sec UVR was 23.1 % comparing potassium or CCK induced contraction. 3. The relaxation was significantly inhibited by methylene blue, the inhibitor of gualylate cyclase. CONCLUSION: According to above results, we confirmed that nitric oxide relaxed human gallbladder muscle. And we think that the further study should be done to examine whether the L-arginine/NO pathway exist in human GB.
Baths ; Endothelium ; Gallbladder Diseases ; Gallbladder* ; Gastrins ; Gastrointestinal Tract ; Guanylate Cyclase ; Humans ; Methylene Blue ; Motilin ; Muscle, Smooth ; Negotiating ; Nitric Oxide* ; Photolysis ; Potassium ; Relaxation ; Sincalide ; Somatostatin ; Streptozocin

The mechanism underlying the bladder relaxation during filling is not fully understood. Nitric oxide played an important role in mediation of relaxation in between vascular and various types of non-vascular smooth muscle. Theoretically, an increased activity of nitric oxide-releasing inhibitory nerves to the detrusor could be a factor keeping the bladder relaxed during the filling phase. The role of nitric oxide in detrusor muscle is still not fully characterized. The objective of this study was to determine the effect and action mechanism of nitric oxide in the rat detrusor. In this experiment, nitric oxide was generated by photolysis using a long wavelength UV lamp ( 366nm) under nitric oxide, NO2-carrying molecules( streptozotocin, NG-nitro-L-arginine). This study were consisted of in vitro examination using Polygraph(Grass Model 79E, Quincy, MA, U.S.A.), radioimmunoassay for guanosine 3', 5'-cyclic monophosphate and nitrite assay to identify the production of nitric oxide. Nitric oxide was generated by photolysis from nitric oxide and NO-containing compounds. The longer UV irradiated and the stronger photo energy of light source, the more produced the amount of nitric oxide. The relaxation induced by nitric oxide-containing compound ( streptozotocin) is much more than that by NO2-containing compound(NG-nitro-L-arginine). Nitric oxide was a potent but labile relaxing substance to the rat detrusor strip. The maximal relaxation under streptozotocin treatment and 60 second UV irradiation was approximately 70.0 % comparing to carbachol induced contraction. That relaxation was significantly inhibited by pyrogallol, methylene blue treatment. The photo-induced nitric oxide directly activated soluble form of guanylate cyclase resulting in increased concentration of guanosine 3' 5'-cyclic monophosphate in detrusor strips. From the above results. it was confirmed that nitric oxide relaxed the detrusor muscle, which means that L-arginine/nitric oxide pathway may be present in the mechanism of relaxation. Therefore, new therapeutic approach using exogenous nitric oxide will be an attractive modality in treating functional voiding disturbances such as detrusor hyperreflexia and bladder instability.
Animals ; Carbachol ; Guanosine ; Guanylate Cyclase ; Methylene Blue ; Muscle, Smooth ; Negotiating ; Nitric Oxide* ; Photolysis* ; Pyrogallol ; Radioimmunoassay ; Rats* ; Reflex, Abnormal ; Relaxation ; Streptozocin ; Urinary Bladder

A clinical analysis of vasospasm as seen angiographically after the onset of subarachnoid hemorrhage was carried out in 22 out of 4 cases of ruptured intracranial aneurysms experienced in our department from January, 1978 to May, 1979. The following results were obtained. The incidence of vasospasm was 50%. 2) Angiographic filling abnormalities(vasospasm) were morphologically classified into 4 types:tapering, diffuse, segmental and nodular types. 3) No significant difference was found in the location of the arteries with vasospasm in relation to the site of the ruptured aneurysm. In this study, the most frequent occurance of spasm was noted in cases of internal carotid aneurysm(69.2%). 4) Discrepancy in the incidence of angiographic spasm between the group with only one subarachnoid hemorrhage(41.9% out of 31 cases) and the other group with more than one hemorrhage(50% out of 44 cases) was demonstrated. While vasospasm occurred frequently within 9 days after the last hemorrhage in cases with more than two hemorrhage, the appearance of vasospasm was frequently seen angiographically between 10 to 17 days after hemorrhage in cases with only one hemorrhage. 5) There was no relationship between the age and the incidence of spasm, and most cases with spasm showed bloody and xanthochromic C.S.F. 6) Neurological findings were as follows:mental disturbance 12 cases, cranial nerve palsy 3 cases, paresis 3 cases, seizure 2 cases and visual disturbance 2 cases. These neurologic deficits may not be related to the spasm. 7) Follow up carotid angiography was performed in 9 cases of those treated by direct approach. Among 5 cases with vasospasm in preoperative angiogram, the spasm persited in 2 cases and was not no longer visualized in 3 cases. On the other hand, in 2 cases without preoperative spasm, postoperative spasm was detected. 8) Of 23 cases of ruptured aneurysms treated surgically(including 3 cases of carotid ligation), 2 out of 15 cases with spasm expired and all 8 cases without spasm were cured. 9) The diffuse type(using above classification) had the worst postoperative prognosis:i.e., among 9 cases with that type of spasm, 2 cases expired and one case resulted in a mild neurologic deficit. Other types of spasm had no special relationship with the postoperative prognosis.
Aneurysm, Ruptured ; Angiography ; Arteries ; Cranial Nerve Diseases ; Follow-Up Studies ; Hand ; Hemorrhage ; Incidence ; Intracranial Aneurysm* ; Neurologic Manifestations ; Paresis ; Prognosis* ; Seizures ; Spasm ; Subarachnoid Hemorrhage

Nitric oxide (NO)-mediated relaxation in vascular smooth muscle involves not only activation of guanylate cyclase but also hyperpolarization of the membrane. It has been shown that depolarization decreases the (Ca2+) sensitivity of myosin light chain kinase in arterial smooth muscle, and nitric oxide (NO)-mediated relaxation was attenuated in this situation. However, why potassium inhibits or attenuates the action of EDRF/NO is not clear. Therefore, we investigated the magnitude of relaxation and cGMP contents using measures known to release NO, such as photorelaxation, photo activated NO-mediated relaxation, and NO-donor (SNP)-mediated relaxation in porcine coronary arterial rings in which contractile conditions were made by different degree of depolarization, i.e., contraction in response to U46619 or U46619 plus KCl. In all cases, the magnitude of relaxation was significantly greater (P<0.05) in U46619-contracted rings than in U46619+KCl-contracted ones. Although accumulation of cGMP was evident with three measures employed in the present study, no difference was found in cGMP contents between U46619 and U46619+KCl conditions, indicating that the diminished relaxation in KCl containing solution is cGMP-independent mechanism(s). To understand this further, cytosolic Ca2+ changes due to NO were compared in rat thoracic aorta by exploiting photoactivated NO using streptozotocin (STZ) that was contracted with either NE or KCl. Fura-3 (Ca)cyt signal caused by NO was small and transient in high K+-, but large and sustained in NE-contracted aorta. The inhibitory potency of STZ expressed in terms Of IC50 was 5.14 and 3.88 gM in NE and in high K+, respectively. These results suggest that modification of the cellular mobilization of Ca2+ rather than cGMP levels may be an important mechanism for the NO-mediated relaxation when vascular membrane is depolarized, such as atherosclerosis and hypertension.
15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid ; Animals ; Aorta ; Aorta, Thoracic ; Atherosclerosis ; Cytosol* ; Guanylate Cyclase ; Hypertension ; Inhibitory Concentration 50 ; Membranes ; Muscle, Smooth ; Muscle, Smooth, Vascular* ; Myosin-Light-Chain Kinase ; Nitric Oxide ; Potassium ; Rats ; Relaxation* ; Streptozocin

Oxidative stress and methylglyoxal (MG), a reactive dicarbonyl metabolites produced by enzymatic and non-enzymatic reaction of normal metabolism, induced aldose reductase (AR) expression in rat aortic smooth muscle cells (SMC). AR expression was induced in a time-dependent manner and reached at a maximum of 4.5-fold in 12 h of MG treatment. This effect of MG was completely abolished by cyclohemide and actinomycin D treatment suggesting AR was synthesized by de novo pathway. Pretreatment of the SMC with N-acetyl-L-cysteine significantly down-regulated the MG-induced AR mRNA. Furthermore, DL-Buthionine-(S,R)-sulfoximine, a reagent which depletes intracellular glutathione levels, increased the levels of MG-induced AR mRNA. These results indicated that MG induces AR mRNA by increasing the intracellular peroxide levels. Aminoguanidine, a scanvenger of dicarbonyl, significantly down-regulated the MG-induced AR mRNA. In addition, the inhibition of AR activities with statil, an AR inhibitor, enhanced the cytotoxic effect of MG on SMC under normal glucose, suggesting a protective role of AR against MG-induced cell damages. These results imply that the induction of AR by MG may contribute to an important cellular detoxification of reactive aldehyde compounds generated under oxidative stress in extrahepatic tissues.
Acetylcysteine ; Aldehyde Reductase* ; Animals ; Dactinomycin ; Glucose ; Glutathione ; Metabolism ; Muscle, Smooth, Vascular* ; Myocytes, Smooth Muscle ; Oxidative Stress* ; Pyruvaldehyde ; Rats ; RNA, Messenger

Oxygen-derived free radicals have been implicated in many important functions in the biological system. Electrical field stimulation (EFS) causes arterial relaxation in animal models. We found that EFS applied to neither muscle nor nerve but to Krebs solution caused a relaxation of rat aorta that had been contracted with phenylephrine. In the present study, therefore, we investigated the characteristics of this EIRF (electrolysis-induced relaxing factor) using rat isolated aorta. Results indicated that EIRF acts irrespective of the presence of endothelium. EIRF shows positive Griess reaction and is diffusible and quite stable. EIRF-induced relaxation was stronger on PE-contracted aorta than on KCl-contracted one, and inhibited by the pretreatment with methylene blue. Zaprinast, a cGMP-specific phosphodiesterase inhibitor, potentiated the EIRF-induced relaxation. NG-nitro-L-arginine, NO synthase inhibitor, did not inhibit the EIRF-induced relaxation. Deferroxamine, but not ascorbic acid, DMSO potentiated the EIRF-induced relaxation. These results indicate that electrolysis of Krebs solution produces a factor that relaxes vascular smooth muscle via cGMP-mediated mechanism.
Animals ; Aorta ; Ascorbic Acid ; Dimethyl Sulfoxide ; Electrolysis* ; Endothelium ; Free Radicals ; Methylene Blue ; Models, Animal ; Muscle, Smooth, Vascular* ; Nitric Oxide Synthase ; Nitroarginine ; Phenylephrine ; Rats ; Reactive Oxygen Species ; Relaxation

BACKGROUND: Recent clinical observation reported that there was a significant correlation between change in circulating vascular endothelial growth factor (VEGF) levels and the occurrence of severe acute graft-versus-host disease (GVHD) following allogeneic hematopoietic stem cell transplantation (allo-HSCT), but the action mechanisms of VEGF in GVHD have not been demonstrated. METHODS: This study investigated whether or not blockade of VEGF has an effect on acute GVHD in a lethally irradiated murine allo-HSCT model of B6 (H-2b)-->B6D2F1 (H-2b/d). Syngeneic or allogeneic recipient mice were injected subcutaneously with anti-VEGF peptides, dRK6 (50 microg/dose) or control diluent every other day for 2 weeks (total 7 doses). RESULTS: Administration of the dRK6 peptide after allo-HSCT significantly reduced survival with greaterclinical GVHD scores and body weight loss. Allogeneic recipients injected with the dRK6 peptide exhibited significantly increased circulating levels of VEGF and expansion of donor CD3+ T cells on day +7 compared to control treated animals. The donor CD4+ and CD8+ T-cell subsets have differential expansion caused by the dRK6 injection. The circulating VEGF levels were reduced on day +14 regardless of blockade of VEGF. CONCLUSION: Together these findings demonstrate that the allo-reactive responses after allo-HSCT are exaggerated by the blockade of VEGF. VEGF seems to be consumed during the progression of acute GVHD in this murine allo-HSCT model.
Animals ; Body Weight ; Graft vs Host Disease ; Hematopoietic Stem Cell Transplantation ; Hematopoietic Stem Cells ; Humans ; Mice ; Oligopeptides ; Peptides ; T-Lymphocyte Subsets ; T-Lymphocytes ; Tissue Donors ; Vascular Endothelial Growth Factor A

BACKGROUND: Clinically apparent thyroid nodule is a very common disease in adults. Patients are often treated with thyroxine in order to reduce the size of the nodule, but the efficacy of thyroxine-suppressive therapy with thyroxine remains uncertain. We investigated the efficacy of thyroxine-suppressive therapy and the hormonal characteristics during thyroxine therapy to find out whether such measurement could be used to determine the effectiveness of this therapy in patients with benign solitary thyroid nodule proved by aspiration biopsy. METHODS: In this study, 54 patients were randomly assigned to receive L-thyroxine (Group I, n=24) or placebo (GroupII, n=30) for 1 year. High resolution ultrasonography (10MHz) was used to measure the size of the nodules at six month intervals. Thyroid hormones and thyroglobulin assay and FNA (fine needle aspiration) was done at the same time. The dose of thyroxine was 200ug/day. Patients were followed at 6 month intervals. RESULTS: The results were as follows: 1) 13 (54.1%) out of 24 Group I patients after adequate TSH suppression had a statistically significant reduction of nodule size and before-to-after nodule volume ratios were significantly different between the Group I and Group II patients. 2) In the responders among Group I patients, the before-to-after therapy ratio of the nodule volume was not related to the pretreatment nodule size, thyroid hormones and thyroglobulin levels. CONCLUSION: Thus we concluded that an adequate suppressive dose of L-thyroxine significantly altered the volume of the benign solitary thyroid nodules 12 months later.
Adult ; Biopsy, Needle ; Humans ; Needles ; Thyroglobulin ; Thyroid Gland* ; Thyroid Hormones ; Thyroid Nodule* ; Thyroxine* ; Ultrasonography

Growing evidence indicates that enhanced generation or actions of nitric oxide (NO) are implicated in the pathogenesis of hypertension in spontaneously hypertensive rats and diabetic nephropathy in streptozotocin (STZ)-induced diabetic rats. We investigated whether inducible nitric oxide synthase (iNOS) expression in STZ-induced diabetic rats is responsible for the alterations of vascular reactivity. Diabetic state was confirmed 28 days after injection of STZ (i.p) in rats by measuring blood glucose. In order to evaluate whether short term (4 weeks) diabetic state is related with altered vascular reactivity caused by iNOS expression, isometric tension experiments were performed. In addition, plasma nitrite/nitrate (NOx) levels and expression of iNOS in the lung and aorta of control and STZ-treated rats were compared by using Griess reagent and Western analysis, respectively. Results indicated that STZ-treated rats increased the maximal contractile response of the aorta to phenylephrine (PE), and high K+, while the sensitivity remained unaltered. Endothelium-dependent relaxation, but not SNP-mediated relaxation, was reduced in STZ-treated rats. Plasma nitrite/nitrates are significantly increased in STZ-treated rats compared to controls. The malondialdehyde (MDA) contents of liver, serum, and aorta of diabetic rats were also significantly increased. Furthermore, nitrotyrosine, a specific foot print of peroxynitrite, was significantly increased in endothelial cells and smooth muscle layers in STZ-induced diabetic aorta. Taken together, the present findings indicate that enhanced release of NO by iNOS along with increased lipid peroxidation in diabetic conditions may be responsible, at least in part, for the augmented contractility, possibly through the modification of endothelial integrity or ecNOS activity of endothelium in STZ-diabetic rat aorta.
Animals ; Aorta ; Blood Glucose ; Diabetic Nephropathies ; Endothelial Cells ; Endothelium ; Foot ; Hypertension ; Lipid Peroxidation ; Liver ; Lung ; Malondialdehyde ; Muscle, Smooth ; Muscle, Smooth, Vascular ; Nitric Oxide ; Nitric Oxide Synthase Type II* ; Peroxynitrous Acid ; Phenylephrine ; Plasma ; Rats* ; Rats, Inbred SHR ; Relaxation ; Streptozocin