s-nitro-n-acetylpenicillamine and pimagedine

We tested whether endotoxin (lipopolysaccharide, LPS) inhibits IGF-I gene expression in hepatocytes and the possible role of Kupffer cells and nitric oxide (NO) in this effect. LPS decreased IGF-I mRNA in hepatocyte cultures and increased the nitrite + nitrate levels in the culture medium. Furthermore, there was a negative correlation between the IGF-I mRNA and the nitrite+nitrate levels. When hepatocytes were cocultured with Kupffer cells, the inhibitory effect of LPS on IGF-I mRNA was higher than in hepatocyte cultures, but the stimulatory effect on nitrite+nitrate was similar in both conditions. The exogenous NO donated by S-nitroso-n-acetyl-d,l-penicillamide also decreased the IGF-I gene expression in hepatocyte cultures. In addition, two specific inducible NO synthase (iNOS) inhibitors, l-N6-(1-iminoethyl)lysine (l-NIL) and aminoguanidine, prevented the effect of LPS on nitrite+nitrate levels and on IGF-I gene expression in hepatocyte cultures. These data indicate that iNOS-derived NO may cause downregulation of IGF-I gene expression in hepatocytes. However, in cocultures, the iNOS inhibitor l-NIL prevented the effect of LPS on nitrite+nitrate levels, but only attenuated the LPS-induced decrease in IGF-I gene expression. We conclude that in hepatocytes, LPS-induced decrease in IGF-I is mainly due to induction of iNOS, whereas in the presence of Kupffer cells LPS inhibits IGF-I through NO release and through other inhibitory pathways.

Topics: Animals; Blotting, Northern; Cells, Cultured; Coculture Techniques; Dose-Response Relationship, Drug; Endotoxins; Gene Expression; Guanidines; Hepatocytes; Insulin-Like Growth Factor I; Kupffer Cells; Lipopolysaccharides; Male; Nitrates; Nitric Oxide; Nitric Oxide Donors; Nitric Oxide Synthase Type II; Nitrites; Penicillamine; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

Effects of endotoxemia-induced NO production on rat liver and hepatocytes in culture were investigated. Rats were treated intraperitoneally with saline, lipopolysaccharide (LPS, 10 mg/kg), L-nitroarginine methyl ester (L-NAME)+LPS, aminoguanidine (AG)+LPS, FK 506+LPS, S-nitroso-N-acetyl penicillamine (SNAP)+L-NAME+LPS and SNAP+FK 506+LPS. Mortality, hepatocyte viability and liver function test were estimated. Liver morphology was observed by light and electron microscopy. Hepatocyte cultures were treated with LPS, cytokine mixture (CM) with or without FK 506, L-NAME or AG. Hepatocyte function and inducible form of NOS (iNOS) expression were evaluated. Twenty-four hours after treatments with saline, LPS, L-NAME+LPS, AG+LPS, FK 506+LPS, SNAP+L-NAME+LPS and SNAP+FK 506+LPS, rat mortalities were 0%, 10%, 48%, 8%, 20%, 38% and 0%, and hepatocyte viabilities were 93+/-3%, 80+/-3%, 52+/-8%, 88+/-1%, 70+/-3%, 80+/-4% and 82+/-3%, respectively. AG+LPS or L-NAME+LPS administration was followed by excessive vacuolization of hepatocytes with lesions in the intermediary lobule zone characterized by features of secondary necrosis as a continuation of apoptotic processes. SNAP+L-NAME+LPS resulted in a well-preserved structure of central vein lobules with sparse signs of apoptosis. Treatment with LPS or CM increased iNOS expression in hepatocyte culture, which was inhibited by L-NAME, FK 506 or AG. AG reduced LPS-induced rise in alanine aminotransferase leakage. LPS-induced NO exerts cytoprotective effects in vivo, while LPS-induced NO in vitro appears to be toxic. Based on the data of this report, one cannot use in vitro results to predict in vivo responses to LPS-induced NO production. The pharmacological modulation of iNOS expression or NO production in vivo or in vitro, therefore, by the development of specific NO donors or inhibitors is promising for improvement of hepatocyte functions under the two experimental conditions, respectively.

Topics: Alanine Transaminase; Animals; Cell Survival; Cells, Cultured; Enzyme Inhibitors; Gene Expression Regulation, Enzymologic; Guanidines; Hepatocytes; Interferon-gamma; Interleukin-1; Lipopolysaccharides; Liver; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitrites; Penicillamine; Rats; Rats, Wistar; RNA, Messenger; Tacrolimus; Tumor Necrosis Factor-alpha

Lipopolysaccharide (LPS)-regulated contractility in pericytes may play an important role in mediating pulmonary microvascular fluid hemodynamics during inflammation and sepsis. LPS has been shown to regulate inducible nitric oxide (NO) synthase (iNOS) in various cell types, leading to NO generation, which is associated with vasodilatation. The purpose of this study was to test the hypothesis that LPS can regulate relaxation in lung pericytes and to determine whether this relaxation is mediated through the iNOS pathway. As predicted, LPS stimulated NO synthesis and reduced basal tension by 49% (P < 0.001). However, the NO synthase inhibitors N (omega)-nitro-L-arginine methyl ester, aminoguanidine, and N (omega)-monomethyl-L-arginine did not block the relaxation produced by LPS. In fact, aminoguanidine and N (omega)-monomethyl-L-arginine potentiated the LPS response. The possibility that NO might mediate either contraction or relaxation of the pericyte was further investigated through the use of NO donor compounds; however, neither sodium nitroprusside nor S-nitroso-N-acetylpenicillamine had any significant effect on pericyte contraction. The inhibitory effect of aminoguanidine on LPS-stimulated NO production was confirmed. This ability of LPS to inhibit contractility independent of iNOS was also demonstrated in lung pericytes derived from iNOS-deficient mice. This suggests the presence of an iNOS-independent but as yet undetermined pathway by which lung pericyte contractility is regulated.

Topics: Animals; Capillaries; Cell Size; Cells, Cultured; Dose-Response Relationship, Drug; Enzyme Activation; Enzyme Inhibitors; Gene Expression Regulation, Enzymologic; Guanidines; Lipopolysaccharides; Lung; Male; Mice; Mice, Inbred C57BL; NG-Nitroarginine Methyl Ester; Nitric Oxide Donors; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitrogen Oxides; Nitroprusside; omega-N-Methylarginine; Penicillamine; Pulmonary Circulation; Rats; Rats, Sprague-Dawley; Spermine; Vasodilator Agents

The synaptic basal lamina protein, agrin, is required for the formation of the neuromuscular junction. Agrin signals through a muscle-specific receptor tyrosine kinase (MuSK) initiating a cascade of events that lead to the aggregation of acetylcholine receptors (AChR) at the postsynaptic site. Another important synaptic signalling molecule is nitric oxide (NO), which is produced by the enzyme, nitric oxide synthase (NOS). We investigated the interaction between the agrin signalling cascade and the NO signalling cascade by treating cultured myotubes with agrin, NOS inhibitors, and NO donors. NOS inhibitors prevented agrin induced AChR aggregation and phosphorylation of the AChR beta subunit. Furthermore, NO donors induced AChR aggregation in the absence of agrin, as well as phosphorylation of the AChR beta subunit. These results demonstrate a role for NO as a downstream mediator of agrin induced AChR aggregation and AChR beta subunit phosphorylation at the neuromuscular junction.

Topics: Agrin; Animals; Cells, Cultured; Chick Embryo; Enzyme Inhibitors; Guanidines; Muscle Fibers, Skeletal; Neuromuscular Junction; Nitric Oxide; Nitric Oxide Donors; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitroprusside; Penicillamine; Phosphorylation; Receptors, Cholinergic; Signal Transduction; Vasodilator Agents

This paper aimed to study the mechanism of vascular hyporeactivity during severe hemorrhagic shock. Rats were divided into control and shock group. Membrane potential of arteriolar strips was measured with intracellular recording method and membrane potential changes in arteriolar smooth muscle cells (ASMC) were recorded with membrane potential sensitive fluorescent dye (DiBAC4) and confocal microscopy. Hyperpolarization of ASMC membrane appeared at the late stage of shock, which correlated to low vasoreactivity. Glybenclamide, an inhibitor of K(ATP) channel reversed the hyperpolarizing effect. S-nitroso-N-acetylpenicillamine (SNAP), a donor of NO, in a higher concentration (400 mol/l) caused membrane hyperpolarization in control and shock group, which was completely reversed by application of Tiron, a scavenger of O2-. The hyperpolarizing effect of SNAP was decreased by ODQ, glybenclamide and (or) charybdotoxin. It is concluded that hyperpolarization of ASMC leads to vascular hyporeactivity. Peroxynitrite (OONO-) involves in the development of hyperpolarization in severe shock. The production of cGMP and activation of K(ATP) and K(Ca) channel contribute to the hyperpolarizing effect of OONO-*.

Topics: 1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt; Animals; Arginine; Arterioles; Charybdotoxin; Cyclic GMP; Drug Interactions; Female; Free Radical Scavengers; Glyburide; Guanidines; Ion Transport; Male; Membrane Potentials; Mesenteric Arteries; Microscopy, Confocal; Microscopy, Fluorescence; Muscle, Smooth, Vascular; Nitrates; Nitric Oxide Donors; Norepinephrine; Oxadiazoles; Penicillamine; Potassium; Potassium Channels; Quinoxalines; Rats; Rats, Sprague-Dawley; Shock, Hemorrhagic; Vasodilator Agents

To clarify the role of nitric oxide (NO) in the development and progression of acute pancreatitis, we investigated the effect of different NO synthase inhibitors and NO donors on experimental pancreatitis in rats. Closed duodenal loop (CDL)-induced pancreatitis was produced in male Wistar rats, and the animals were treated with normal saline, the NO-synthase substrate L-arginine, the NO donor S-nitroso-N-acetylpenicillamine, aminoguanidine, which is a more powerful inhibitor of inducible NO synthase (iNOS) than is endothelial NO synthase (eNOS), and N-nitro-L-arginine methyl ester (L-NAME), a more powerful inhibitor of eNOS than of iNOS. All drugs were infused intravenously during a period of 6 or 12 h in each group. Pancreatic tissue was removed at 6 and 12 h after creating the CDL. L-Arginine, S-nitroso-N-acetyl-penicillamine, and aminoguanidine treatment had no effect on the elevation of serum pancreatic enzymes, whereas L-NAME administration significantly exacerbated their elevation. Pathologically, L-NAME treatment resulted in a significantly worse histologic score at 6 and 12 h, especially in terms of the degree of hemorrhage, acinar cell necrosis, and microvascular thrombosis. Addition of L-arginine clearly reversed the effect of L-NAME. Neither the NO substrate nor NO donor could inhibit the progression of hemorrhagic pancreatitis in CDL-induced pancreatitis. Aminoguanidine had no effect on the severity of the pancreatitis. We therefore concluded that NO production by eNOS may play a significant role in preventing the development and progression of acute pancreatitis.

Topics: Acute Disease; Animals; Arginine; Ascitic Fluid; Duodenum; Edema; Enzyme Inhibitors; Guanidines; Ligation; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide Donors; Nitric Oxide Synthase; Nitric Oxide Synthase Type III; Pancreatitis; Penicillamine; Rats; Rats, Sprague-Dawley