Effect of ornithine on the ileal histology, nitric oxide production and lipid peroxidation in LPS-induced endotoxemia.

Effect of ornithine which is known to inhibit L-arginine uptake via cationic amino acid transport system has been tested, and compared to aminoguanidine, an iNOS inhibitor in lypopolysaccharide (LPS)-induced endotoxemia in rats. Serum nitrite/nitrate and malondialdehyde (MDA) level have been measured, and ileal histology has also been examined. Endotoxin increased serum nitrite/nitrate and MDA levels from 15.7+/- 2.4 micromol/ml and 2.1 +/-0.2 nmol/ml to 23.1 +/- 1.0 micromol/ml and 5.2+/- 0.3 nmol/ml (both P&#60;0.05), respectively. In addition, LPS caused ileal degeneration. L-ornithine (500 mg/kg) did not improve septic manifestations, i.e., serum nitrite/nitrate and MDA levels did not differ from those in endotoxemia. Neither does it have an improving action on ileal histology. However, higher dose of L-ornithine (2,500 mg/kg) lowered the increased level of nitrite/nitrate and MDA by LPS. Moreover, it restored ileal histology from grade 3 (median) to 0 (median) (P&#60;0.05). On the other hand, aminoguanidine (100 mg/kg) normalized serum nitrite/nitrate and MDA levels but not ileal histology in endotoxemic rats. In conclusion, high dose of L-ornithine could improve endotoxemic parameters in LPS-treated rats.</p

Evidence that the nitrergic neurotransmitter and endothelium-derived relaxing factor might be S-nitrosothiols in the mouse corpus cavernosum.

The effects of thimerosal, a sulfhydryl oxidizing agent on nitrergic, endothelium-dependent and -independent relaxations were investigated to examine the possibility that the nitrergic neurotransmitter and endothelium-derived relaxing factor (EDRF) could be S-nitrosothiol or free nitric oxide (NO) in the isolated mouse corpus cavernosum. Thimerosal (5 x 10(-6)-2 x 10(-5) M) inhibited or almost abolished electrical field stimulation--(EFS, 30V, 0.5 ms, 15 sec, 1, 2, 4, 8, 16 Hz), acetylcholine--(ACh, 5 x 10(-8)-1.25 x 10(-6) M), glyceryl trinitrate--(GTN, 3 x 10(-7)-3 x 10(-6) M), and S-nitrosoglutathione--(GSNO, 5 x 10(-6)-1.25 x 10(-4) M) induced relaxations. Thiomerosal inhibition seems to be specific to L-arginine NO pathways since it had no effect on acidified sodium nitrite--(10(-4)-5 x 10(-4) M), photoactivated sodium nitrite--(2 x 10(-4) M), isoprenaline--(10(-6) M), or papaverine--(10(-4) M) elicited relaxations. Moreover, the inhibitory effect of thimerosal on the nitrergic, ACh- or GTN-induced relaxations were partly reversed by sulfhydryl-containing compounds, L-cysteine (10(-3) M), dithiothreitol (10(-3) M), or glutathione (10(-3) M). However L-methionine (10(-3) M), which contains a methyl group on the sulphur atom, failed to restore the thimerosal inhibition. Thimerosal did not change the contraction produced by 10(-4) M NG-nitro-L-arginine methyl ester. These findings indicate that the nitrergic neurotransmitter as well as EDRF may not be free NO but NO-transferring molecules, probably S-nitrosothiols, in the mouse corpus cavernosum.</p

Comparative investigation of antitumoral effectiveness of Rho-kinase inhibitor Y-27632, pravastatin and atorvastatin in anaplastic thyroid cancer cell culture

Anaplastic thyroid cancer is an aggressive malignancy with a poor prognosis. In metastatic cases instead of treatment alternatives including surgery, radiotherapy, and chemotherapeutic regimens, targeted treatments should be sought for. Statins are 3-hidroxy-3 methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitors, and inhibit conversion of HMG-CoA into mevalonate. Inhibition of mevalonate pathway Ras prenylation, can also inhibit tumoral growth. Rho/Rho kinase pathway has an important role in tumoral proliferation, and metastasis in which activity of ROCK increases leading to tumoral invasion. Herein we investigated antitumoral effectiveness of two HMG-CoA reductase inhibitor statins namely pravastatin, and atorvastatin, and Rho-kinase inhibitor Y-27632 in anaplastic thyroid cancer cell cultures through suppression of cellular proliferation. Various concentrations of pravastatin (20, and 60 μM), and atorvastatin (10, and 30 μM), Y-27632 (10, and 30 μM), and their combinations including pravastatin -Y-27632 (20 μM + 10 μM; 60 μM + 30 μM), atorvastatin - Y-27632 (10 μM + 10 μM, and 30 μM + 30 μM) solutions were prepared. Anaplastic thyroid cancer cell culture media were treated with these more water-soluble drug solutions of pravastatin which induced lower dose-, and time-dependent decreases in cellular indices relative to more lipid-soluble atorvastatin which also markedly suppressed cellular proliferation. Y-27632 also decreased cell indices in a dose-, and time-dependent manner. Combination of Y-27632 with pravastatin, and atorvastatin did not demonstrate additive, synergic or antagonistic interactions. HMG-CoA reductase, and also Rho-kinase inhibitors are promising treatment alternatives of anaplastic thyroid cancers. Further in vivo, and clinical studies are needed on this issue

Nitric oxide does not downregulate Rho-kinase (ROCK-2) expression in rat coronary endothelial cells

WOS: 000253874000005PubMed ID: 18287881Rho kinase (ROCK) and nitric oxide (NO) are important targets in cardiovascular diseases. Therefore, we investigated the possible influence of NO on Rho kinase (ROCK-2 isoform) expressions in cultured rat coronary microvascular endothelial cells. The cells were isolated from Wistar rats on a Langendorff system, and were incubated overnight (similar to 16 h) with an NO generator, A-23187 (10(-7) to 10(-6) M) NO donors, such as sodium nitroprusside (10(-7) to 10-6 M) glyceryl trinitrate (10(-7) to 10-6 M), 2,2'-(hydroxynitro- sohydrazono)bis-ethanimine (10(-7) to 10(-6) M), and NaNO2 (10(-4) to 10(-3) M) or a nitric oxide synthase (NOS) inhibitor, N-G-nitro-L-arginme methylester (2x10(-4) M), or two ROCK inhibitors, (-+)(R)-trans-4-(1-aminoethyl)- N-(4-pyridyl) cyclohexanecarboxamide dihydrochloride monohydrate (Y-27632, 10(-5) M) and fasudil (10(-5) M) in the absence or presence of thrombin (4 U/mL). ROCK-2 and endothelial NOS (eNOS) expressions were detected by Western blotting. Moreover, nitrite/nitrate levels were detected by Griess method in the presence of the ROCK inhibitors. The NO donors and the NO generator had no significant effects on ROCK-2 expression. Y-27632 and fasudil did not alter eNOS expression and NO production. Nitrite/nitrate levels were 4.4 +/- 0.32 mu M in control and 4.0 +/- 0.93 mu M and in Y-27632 group. These results demonstrate that prolong NO donation could not suppress the expression of ROCK-2 protein, and the ROCK inhibitor did not change e-NOS expression and NO production in the cultured rat coronary microvascular endothelial cells

Role of Rho-kinase in contractions of ureters from rabbits with unilateral ureteric obstruction

WOS: 000249953400037PubMed ID: 17711509To investigate the expression of two isoforms of Rho-kinase (ROCK) and its functional role in the pathophysiological control of smooth muscle contraction in rabbits with unilateral ureteric obstruction (UUO). Left UUO was created in 14 rabbits and eight other rabbits (controls) had sham operations. After 2 weeks all the rabbits were killed. Ureteric strips suspended in an organ bath were used for functional studies and the effects of Y-27632, a specific inhibitor of Rhokinase, on spontaneous contractions and electrical field stimulation (EFS; 50 V, 1 ms, 16 Hz, for 20 s), carbachol- (10(-7) - 10(-4) M), phenylephrine- (10(-7) - 10(-4) M) and KCl- (50 mM) induced contractions were analysed. Western blotting was used to determine expression levels of Rho-kinase protein in the ureters of UUO and control rabbits. In the functional analysis, the contractions induced by EFS, KCl, phenylephrine and carbachol in the ureteric strips from rabbits with UUO were significantly greater than those from the control rabbits. Y-27632 considerably suppressed the ureter contractile responses in both UUO and control rabbits. Western blot analysis showed that both ROCK-1 and ROCK-2 proteins were expressed in the rabbit ureter. In accordance with the functional studies, the expression levels of both ROCK-1 and ROCK-2 were significantly greater in the ureters of UUO rabbits than in the controls. Y-27632 suppressed ureteric contractions in the rabbits with UUO. Western blot analysis also confirmed greater expression levels of ROCK-1 and ROCK-2 in the ureters of UUO rabbits. It is important to elucidate by which mechanisms the Rho-kinase pathway affects ureteric function after obstruction