thiourea and aminoethyl-isothiourea

The existence of cancer stem cells, stem-like cancer cells (SLCCs), or tumor-initiating cells is considered as the cause of tumor formation and recurrence, indicating the importance of studying novel therapy that targets SLCCs. The origin of SLCCs is controversial because of two competing hypotheses: SLCCs are either transformed from tissue adult stem cells or dedifferentiated from transformed progenitor cells. Our previous research demonstrates that SLCCs are inducible by increasing genomic instability in cancer cells. In this study, to block the emergence of SLCCs, aminoethyl isothiourea (AET), a compound that clears free radicals and is used to protect patients from radioactive exposure, was used as an agent that maintains genomic stability in combination with mitomycin C (MMC), a commonly used chemotherapeutic drug that damages DNA. Using a rabbit tumor model with VX2 hepatic carcinoma, we found that MMC alone increased lung metastases and disadvantaged survival outcome, but the combination of MMC and AET reversed this effect and even prolonged overall survival. Moreover, in a VX2 xenograft model by immunocompromised mice, MMC alone enriched tumor-initiating cells, but the administration of MMC in combination with AET eliminated tumor cells effectively. Furthermore, MMC alone enhanced genomic instability, but MMC combined with AET attenuated the extent of genomic instability in primary VX2 tumor tissue. Taken together, our data suggest that the genomic protector AET can inhibit the induction of SLCCs, and this combination treatment by AET and cytotoxic agents should be considered as a promising strategy for future clinical evaluation.

Topics: Animals; Carcinoma, Hepatocellular; Disease Models, Animal; Genomic Instability; Humans; Mice; Mitomycin; Neoplastic Processes; Neoplastic Stem Cells; Nitric Oxide Synthase; Nucleic Acid Synthesis Inhibitors; Rabbits; Radiation-Protective Agents; Reactive Oxygen Species; Thiourea; Treatment Outcome

Some evidence indicates that nitric oxide (NO) contributes to inflammation, while other evidence supports the opposite conclusion. To clarify the role of NO in inflammation, we studied carrageenin-induced pleurisy in rats treated with an NO donor (NOC-18), a substrate for NO formation (L-arginine), and/or an NO synthase inhibitor (S-(2-aminoethyl) isothiourea or N(G)-nitro-L-arginine). We assessed inflammatory cell migration, nitrite/nitrate values, lipid peroxidation and pro-inflammatory mediators. NOC-18 and L-arginine reduced the migration of inflammatory cells and edema, lowered oxidative stress, and normalized antioxidant enzyme activities. NO synthase inhibitors increased the exudate formation and inflammatory cell number, contributed to oxidative stress, induced an oxidant/antioxidant imbalance by maintaining high O(2) (-), and enhanced the production of pro-inflammatory mediators. L-arginine and NOC-18 reversed the proinflammatory effects of NO synthase inhibitors, perhaps by reducing the expression of adhesion molecules on endothelial cells. Thus, our results indicate that NO is involved in blunting-not enhancing-the inflammatory response.

Topics: Animals; Antioxidants; Arginine; Carrageenan; Disease Models, Animal; Male; Malondialdehyde; Nitrates; Nitric Oxide; Nitric Oxide Donors; Nitric Oxide Synthase; Nitrites; Pleurisy; Rats; Rats, Wistar; Thiourea

To investigate the expression and the role of iNOS expression in hepatic ischemia-reperfusion (I/R) injury.. Male Wistar rats were subjected to 30-minute hepatic ischemia, then iNOS protein and iNOS mRNA expression in liver tissue was assessed by Western blot and RT-PCR analysis respectively at different time points after reperfusion. The effects of L-NAME (Nomega-nitro-L-arginine methyl ester, a nonselective NOS inhibitor) or AE-ITU (aminoethytl-isothiourea, a relative selective inhibitor of iNOS) treatment were also evaluated.. High levels of iNOS protein and mRNA expression were detected in the liver tissue subjected to I/R, but not in the sham-operated rats. iNOS protein and iNOS mRNA expression reached a maximum on the first day after reperfusion and decreased later. The levels of iNOS protein and iNOS mRNA disappeared on 7th, 3rd day after reperfusion respectively. The high iNOS expression was correlated with hepatic dysfunction. L-NAME administration worsened hepatic dysfunction induced by hepatic I/R. In contrast, AE-ITU administration showed mild protective effects against hepatic dysfunction induced by hepatic I/R.. Ischemia-reperfusion may induce or up-regulate the expression of iNOS protein and iNOS mRNA, which is detrimental to hepatic I/R injury

Topics: Alanine Transaminase; Animals; Aspartate Aminotransferases; Enzyme Inhibitors; Gene Expression Regulation, Enzymologic; Liver Diseases; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Rats; Rats, Wistar; Reperfusion Injury; RNA, Messenger; Thiourea

Nitric oxide (NO), produced via inducible nitric oxide synthase (iNOS), is implicated in the pathophysiology of renal ischemia/reperfusion (I/R) injury. The aim of this study was to investigate the effects of the iNOS inhibitors L-N6-(1-iminoethyl)lysine (L-NIL) and aminoethyl-isothiourea (AE-ITU) on (a) renal dysfunction and injury mediated by bilateral I/R of rat kidneys in vivo and (b) cytokine-stimulated NO production by primary cultures of rat proximal tubule (PT) cells.. Male Wistar rats subjected to bilateral renal ischemia (45 min) followed by reperfusion (6 h). Rats were administered either L-NIL (3 mg/kg IV bolus 15 min prior to I/R followed by 1 mg/kg/h throughout I/R) or AE-ITU (1 mg/kg IV bolus 15 min prior to I/R followed by 1 mg/kg/h throughout I/R). Serum and urinary biochemical indicators of renal dysfunction and injury were measured; serum creatinine (SCr, glomerular dysfunction), fractional excretion of Na+ (FENa, tubular dysfunction), serum aspartate aminotransferase (sAST, I/R injury) and urinary N-acetyl-beta-d-glucosaminidase (uNAG, tubular injury). Additionally, renal sections were used for histological grading of renal injury and for immunological evidence of nitrotyrosine formation. Nitrate/nitrate levels in plasma were measured using the Griess assay and used as an indicator of NO production. Primary cultures of rat PT cells were incubated with interferon-gamma(IFN-gamma, 100 IU/mL) and lipopolysaccharide (LPS, 10 microg/mL) for 24 h, either in the absence or presence of increasing concentrations of L-NIL or AE-ITU (0.001 to 1 mmol/L) after which nitrite/nitrate levels were measured using the Griess assay.. L-NIL and AE-ITU significantly reduced the I/R-mediated increases in SCr, FENa, sAST and uNAG, indicating attenuation of I/R-mediated renal dysfunction and injury. Specifically, L-NIL and AE-ITU reduced the I/R-mediated glomerular and tubular dysfunction and biochemical and histological evidence of tubular injury. Both L-NIL and AE-ITU attenuated the plasma levels of nitrate (indicating reduced NO production) and the immunohistochemical evidence of the formation of nitrotyrosine. In vitro, L-NIL and AE-ITU both significantly reduced cytokine-stimulated NO production by primary cultures of rat PT cells in a dose-dependent manner.. These results suggest that L-NIL and AE-ITU reduce the renal dysfunction and injury associated with I/R of the kidney, via inhibition of iNOS activity and subsequent reduction of NO (and peroxynitrite) generation. We propose that selective and specific inhibitors of iNOS activity may be useful against the NO-mediated renal dysfunction and injury associated with I/R of the kidney.

Topics: Animals; Cells, Cultured; Cytokines; Enzyme Inhibitors; Ischemia; Kidney Glomerulus; Kidney Tubules; Kidney Tubules, Proximal; Lysine; Male; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Rats; Rats, Wistar; Renal Circulation; Reperfusion Injury; Thiourea; Tyrosine

Aminoethylisothiourea (AET) is a potent inhibitor of inducible nitric oxide synthase (NOS). The present study was performed to investigate whether AET and its rearrangement products might modulate vascular contraction independently of its effects as a NOS inhibitor in rat small femoral arteries. AET caused an endothelium-independent increase in contraction induced by norepinephrine (NE). This effect was not affected by either N(omega)-nitro-L-arginine methyl ester, nitro-L-arginine, indomethacin or propanolol, but it was suppressed in Ca(2+)-free medium. AET enhanced extracellular Ca(2+) component of NE-induced contraction, and this effect was prevented by the receptor-mediated Ca(2+) entry blocker, 1-{beta-[3-(p-methoxyphenyl)-propyloxyl]-p-methoxyphenetyl}- 1H-imidaz ole hydrochloride (SK&F 96365), but not by the voltage-dependent Ca(2+) channel blocker, nitrendipine. AET did not alter the response to CaCl(2) in vessels exposed to KCl depolarization. The protein kinase C (PKC) inhibitor, 2-[1-(3-dimethylaminopropyl)indol-3-yl]-3-(indol-3-yl) (GF 109203X), prevented the potentiating effect of AET on the NE response. AET failed to produce an increase in tone in the presence of NE and GTP in permeabilized arteries. Among the AET rearrangement products, mercaptoethylguanidine produced an endothelium-independent increase in the NE response. 2-aminothiazoline had no effect, and guanidinoethyldisulphide produced relaxation. The effect of mercaptoethylguanidine was dependent on extracellular Ca(+) and was prevented by GF 109203X. These results indicate that AET is able to potentiate the contraction to NE in rat femoral resistance arteries independently of its inhibitory effect on either NOS or cyclo-oxygenase. Its effect occurs via an enhancement of SK&F 96365-sensitive Ca(2+) entry. A PKC inhibitor-sensitive mechanism also appears to be involved in the AET effect. Mercaptoethylguanidine potentiates NE response through a mechanism similar to AET.

Topics: Animals; Calcium; Drug Synergism; Endothelium, Vascular; Enzyme Inhibitors; Femoral Artery; Guanidines; In Vitro Techniques; Male; Nitric Oxide Synthase; Norepinephrine; Prostaglandin-Endoperoxide Synthases; Protein Kinase C; Radiation-Protective Agents; Rats; Rats, Wistar; Thiazoles; Thiourea; Vasoconstriction

The bulk of published data has shown that NO is proinflammatory. However, there also exists the conflicting notion that NO may be protective during an inflammatory insult. In an attempt to resolve this issue, we have compared the effects on inflammation of a range of NO synthase (NOS) inhibitors given either directly to the site of the inflammatory lesion or systemically. It was found that in the carrageenin-induced pleurisy, a single intrapleural injection of the selective inducible NO inhibitors S-(2-aminoethyl) isothiourea (AE-ITU; 3 and 10 mg/kg) and N-(3-(aminomethyl)-benzyl) acetamidine (1400W; 10 mg/kg) or the selective endothelial cell NOS inhibitor L-N(5)(1-iminoethyl)-ornithine (10 mg/kg) not only exacerbated inflammation at the very early stages of the lesion (1-6 h), but also prevented inflammatory resolution. By contrast, administering NOS inhibitors systemically ameliorated the severity of inflammation throughout the reaction. To elucidate the mechanisms by which inhibition of NO synthesis locally worsened inflammation, we found an increase in histamine, cytokine-induced neutrophil chemoattractant, superoxide, and leukotriene B(4) levels at the inflammatory site. In conclusion, this work shows that the local production of NO is protective by virtue of its ability to regulate the release of typical proinflammatory mediators and, importantly, that NOS inhibitors have differential anti-inflammatory effects depending on their route of administration.

Topics: Acute Disease; Amidines; Animals; Antioxidants; Benzylamines; Carrageenan; Disease Models, Animal; Drug Administration Schedule; Edema; Enzyme Inhibitors; Free Radical Scavengers; Inflammation; Inflammation Mediators; Injections; Injections, Intraperitoneal; Male; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; omega-N-Methylarginine; Pleura; Pleurisy; Rats; Rats, Wistar; Superoxides; Thiourea

Topics: Animals; Enzyme Inhibitors; Gram-Positive Bacterial Infections; Humans; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Shock, Septic; Thiourea

To elucidate the possible roles of nitric oxide (NO), endothelin-1 (ET-1), and reactive oxygen species (ROS) in the pathophysiology of serogroup A streptococcal (GAS) peritoneal sepsis, we investigated the effects of aminoethylisothiourea (AE-ITU), an inducible NO synthase (iNOS) inhibitor, and a ROS scavenger, and the ET-1 receptor antagonist bosentan. In rats, live GAS inocula, 3 x 10(8) and 1 x 10(9) cfu/kg, entailed a 24-h mortality of 10% and 90%, respectively. GAS caused increases in tissue iNOS activity (9 h), in serum nitrite/nitrate (9-24 h), and in intracellular leukocyte ROS levels (3-6 h). These changes were all prevented by the pre-treatment with AE-ITU. A novel finding was that AE-ITU also prevented the GAS-induced marked increase in plasma ET-1 at 6 h. Short-term (7-h) survival was improved by both AE-ITU and by bosentan. The mechanism(s) for the beneficial effects of AE-ITU may possibly be a combined mode of action; iNOS inhibition, ROS scavenging, and inhibition of the increase in plasma ET-1 caused by GAS.

Topics: Animals; Antihypertensive Agents; Blood Pressure; Bosentan; Endothelin Receptor Antagonists; Endothelin-1; Enzyme Inhibitors; Male; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Rats; Rats, Sprague-Dawley; Receptor, Endothelin A; Shock, Septic; Streptococcal Infections; Streptococcus pyogenes; Sulfonamides; Survival Rate; Thiourea; Time Factors

The incidence of severe invasive disease caused by serogroup A streptococci (GAS) is increasing, and to elucidate the role of streptococcal cell wall components in the inflammatory response, human whole blood was stimulated with lipoteichoic acid (LTA, 0.005-50 microg/mL) and peptidoglycan (10 and 100 microg/ml) from Streptococcus pyogenes. Both stimulants increased dose dependently the leukocyte release of cytokines many thousand fold: tumor necrosis factor alpha (0 to 158,000+/-4,900 pg/mL), interleukin (IL)-1beta (85+/-56 to 31,000+/-4,600 pg/mL), IL-6 (30+/-11 to 34,800+/-15,000 pg/mL), and IL-8 (300+/-150 to 29,000+/-14,000 pg/mL). Intracellular leukocyte levels of reactive oxygen species (ROS) as measured by flow cytometry increased 15-20 fold, from 25 to 400-500 mean fluorescence intensity. Aminoethyl-isothiourea (AE-ITU), a relatively selective inhibitor of the inducible nitric oxide synthase (iNOS) and a ROS scavenger, reduced the cytokine production by 70-100%, and intracellular leukocyte ROS levels by 50-70% (all P < 0.05). The non-selective NOS inhibitor N-nitro-L-arginine methyl ester (L-NAME) did not affect intracellular ROS levels, but it caused a moderate selective inhibition of IL-8 production. Leukocyte NO production (measured up to 36 h) was not enhanced by LTA, peptidoglycan, inactivated streptococci, or cytokine combinations. The mechanisms for the anti-inflammatory effects of AE-ITU may be through a reduction of intracellular ROS levels, or through a direct effect on signal transduction, whereas NO modulation is an unlikely mechanism.

Topics: Cell Survival; Cell Wall; Cytokines; Dose-Response Relationship, Drug; Enzyme Inhibitors; Humans; In Vitro Techniques; Inflammation; Interleukin-1; Interleukin-6; Interleukin-8; Leukocytes; Lipopolysaccharides; NG-Nitroarginine Methyl Ester; Nitrates; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitrites; Peptidoglycan; Reactive Oxygen Species; Streptococcus pyogenes; Teichoic Acids; Thiourea; Tumor Necrosis Factor-alpha

To test the effect of a continuous infusion of the nitric oxide (NO) synthase (S) inhibitor aminoethyl-isothiourea (AE-ITU) on survival time, hemodynamics, and oxygen transport in a porcine model of live group A streptococcal (GAS) sepsis. Furthermore, to examine the role of endothelin-1, histamine, and reactive oxygen species (ROS) in streptococcal shock.. Prospective, randomized trial.. Laboratory at a university hospital.. Twenty-eight pigs with an average weight of 25 kg.. Sixteen animals received a continuous infusion of live Streptococcus pyogenes 1.3 x 10(10) colony forming units/hr: eight received fluids only, and the other eight received an intravenous infusion of AE-ITU 10 mg/kg/hr starting 30 mins before the GAS challenge. Six control pigs received AE-ITU 10 mg/kg/hr iv for 5 hrs. Another six animals received half the dose of GAS over 5 hrs.. GAS infusion caused a rapid increase in pulmonary, hepatic, and systemic vascular resistance, followed by hypotension with a 90% lethality at 4 hrs. Treatment with AE-ITU increased 4-hr survival in septic animals from 1/8 to 8/8 and 5-hr survival from 0/8 to 5/8, prevented hypotension, and increased urine output. AE-ITU attenuated the decrease in cardiac output, liver blood flow, and oxygen delivery, and hepatic arterial blood flow as a fraction of cardiac output increased (all p < .05). Plasma nitrate/nitrite levels decreased in all animals. Inducible NOS and endothelial constitutive NOS activities in liver, gut, and lung were not increased during sepsis, nor were they decreased after AE-ITU. Plasma levels of endothelin-1 and methylhistamine increased in all septic animals and were not modified by AE-ITU. AE-ITU prevented the increase in monocyte ROS production caused by GAS. In control animals, AE-ITU caused an increase in mean arterial pressure, liver blood flow, and oxygen delivery.. In this model of porcine GAS-induced septic shock, which was not associated with enhanced NO production, infusion of the NOS inhibitor AE-ITU prolonged survival, prevented hypotension, and improved cardiac contractility, organ perfusion, and tissue oxygenation. These beneficial effects of AE-ITU might be a result of the combined effect of ROS scavenging and modulation of local NO production, thus improving the balance of vasodilator and vasoconstrictor forces and reducing oxidative stress.

Topics: Animals; Disease Models, Animal; Female; Hemodynamics; Isothiuronium; Male; Nitric Oxide; Random Allocation; Shock, Septic; Streptococcal Infections; Streptococcus pyogenes; Survival Rate; Swine; Thiourea