l-NAME inhibits tumor cell progression and pulmonary metastasis of r/m HM-SFME-1 cells by decreasing NO from tumor cells and TNF-α from macrophages

Highly metastatic ras/myc-transformed serum-free mouse embryo (r/m HM-SFME-1) cells were injected subcutaneously to mice and the effects of Nω-nitro-l-arginine methyl ester (l-NAME) on the tumor progression and pulmonary metastasis were investigated. In addition, production of nitric oxide (NO), matrix metalloproteinases (MMPs) and tumor necrosis factor-alpha (TNF-α) in the tumor cells and in a mouse macrophage-like cell line, J774.1 cells, was analyzed. The increase in footpad thickness was significantly smaller in the mice which were fed the l-NAME containing water (4.24 ± 0.39 mg/day/mouse). The number of the tumor cells metastasized to the lungs was smaller in the l-NAME treated mice, although statistical significance was not found. Co-treatment of r/m HM-SFME-1 cells with interferon-gamma (IFN-γ; 100 U/ml) and lipopolysaccharide (LPS; 0.5 μg/ml) significantly enhanced NO production, and the presence of l-NAME at 1 mM significantly decreased this response. In r/m HM-SFME-1 cells, MMP-2 was undetectable and MMP-9 was also very little in the basal level, and both MMPs were unaffected by the IFN-γ and/or LPS treatments, not to mention by the l-NAME treatment. In J774.1 cells, any treatment including LPS appeared to enhance MMP-9 production, however, this upregulation was not inhibited by the additional presence of l-NAME. Production of TNF-α by J774.1 cells was markedly enhanced with LPS treatment, and this enhancement was significantly reduced in the presence of l-NAME. These results indicate that the inhibitory effects of l-NAME on the tumor cell progression and pulmonary metastasis could be due to suppression of NO from tumor cells and TNF-α from macrophages (Mol Cell Biochem, 2007). Hideaki Yamaguchi and Yumi Kidachi contributed equally to this work.     

The Role of Nitric Oxide in the Neuroprotection of Limb Ischemic Preconditioning in Rats

Brief limb ischemia was reported to protect neurons against injury induced by subsequent cerebral ischemia-reperfusion, and this phenomenon is known as limb ischemic preconditioning (LIP). To explore the role of nitric oxide (NO) in neuroprotection of LIP in rats, we observed changes in the content of nitric oxide (NO) and activity of NO synthase (NOS) in the serum and CA1 hippocampus of rats after transient limb ischemic preconditioning (LIP), and the influence of N^G-nitro-l-arginine methylester (l-NAME), a NOS inhibitor, on the neuroprotection of LIP against cerebral ischemia-reperfusion injury. Results showed that NO content and NOS activity in serum increased significantly after LIP compared with the sham group. The increase showed a double peak pattern, in which the first one appeared at time 0 (immediate time point) and the second one appeared at 48 h after the LIP (P < 0.01). The NO content and NOS activity in the CA1 hippocampus in LIP group showed similar change pattern with the changes in the serum, except for the first peak of up-regulation of NO content and NOS activity appeared at 6 h after LIP. Pretreatment with l-NAME before LIP blocked the neuroprotection of LIP against subsequent cerebral ischemic insult. The blocking effect of l-NAME was abolished with pretreatment of l-Arg. These findings indicated that NO may be associated with the tolerance of pyramidal cells in the CA1 hippocampus to ischemia induced by LIP in rats.     

Role of nitric oxide in UV-B-induced activation of PAL and stimulation of flavonoid biosynthesis in Ginkgo biloba callus

The role of nitric oxide (NO) in UV-B-induced secondary metabolite accumulation in Ginkgo biloba callus was investigated. Overall, UV-B irradiation induced multiple biological responses in callus of G. biloba, including increased both NO production and nitric oxide synthase (NOS) activity, and subsequent activation of phenylalanine ammonium lyase (PAL) and synthesis of flavonoids. Application of NO via the donor sodium nitroprusside (SNP) enhanced UV-B-induced PAL activity and increased accumulation of flavonoids in G. biloba callus. Both, the NOS inhibitor l-NAME (N (G)-nitro-l-arginine methyl ester) and the NO scavenger c-PTIO (2-phenyl-4,4,5,5-tetramethyl-imidazoline-1-oxyl-3-oxide) reduced the production of NO. Moreover, UV-B-induced increase of PAL activity and flavonoid accumulation were suppressed by l-NAME and c-PTIO. These findings suggested a causal relationship between NO release and both PAL activity and flavonoid accumulation under UV-B irradiation. In addition, it also indicated that NO, produced via NOS-like activity in ginkgo callus subjected to UV-B irradiation, might act as an essential signaling molecule for triggering the activation of PAL and synthesis of flavonoids. Additionally, a guanylyl cyclase inhibitor 6-anilino-5,8-quinolinequinone (LY-83583) prevented both UV-B- and SNP-induced enhancement of PAL activation and flavonoid biosynthesis thus suggesting that the NO function was mediated by cyclic guanosine 5’-monophosphate. However, these effects of c-PTIO, l-NAME, and LY-83583 were partial, thus suggesting that there were NO-independent pathways in UV-B signaling networks. Gangping Hao and Xihua Du are contributed equally to this article.     

Effect of nitric oxide synthase inhibitors on the temporal coordination of duodenal contractions and pancreatic exocrine secretion in sheep

The present study evaluated the role of nitric oxide in the regulation of duodenal motility and pancreatic exocrine secretion in conscious sheep. Intravenous infusions of nitric oxide synthase inhibitors, Nω-nitro-l-arginine-methyl ester (l-NAME) and Nω-nitro-l-arginine, induced clusters of duodenal contractions like phase III of migrating motor complexes and simultaneously inhibited flow rate, bicarbonate ion and enzyme outputs of pancreatic juice. The effects of l-NAME were inhibited by simultaneous infusion of l-arginine, but not altered by adrenergic blockade using a combined infusion of phentolamine and propranolol. Inhibition of the pancreatic secretion occurred in coincidence with initiation of the duodenal contractions, while the pancreatic secretion was not inhibited when the premature duodenal contractions were abolished by the l-arginine infusion. The initiation of the cluster of duodenal contractions by l-NAME was not abolished by background infusion of atropine, whereas the amplitude of contractions was significantly inhibited by atropine. These results suggest that intrinsic nitric oxide plays a crucial role in the regulation of duodenal tone and maintenance of continuous secretion by the exocrine pancreas in sheep. These results also implied that inhibition of pancreatic exocrine secretion by the nitric oxide synthase inhibitor is presumably mediated in part through the contractile effect on the duodenum. Accepted: 27 June 2000     

Effect of aluminum on the blood-brain barrier permeability during nitric oxide-blockade-induced chronic hypertension in rats

We examined the effect of aluminum on the permeability of the blood-brain barrier (BBB) during nitric oxide-blockade-induced chronic hypertension in rats. Animals were given the inhibitor of nitric oxide synthase, l-NAME (N ^ω-nitro-l-arginine methyl ester), for 4 wk to induce chronic hypertension. Two groups of rats were given an intraperitoneal injection of aluminum chloride. The integrity of the BBB was assessed by a quantitative measurement for Evans blue (EB) dye. The arterial blood pressure in l-NAME- and l-NAME plus aluminum-treated animals was significantly elevated from 115±2.8 and 110±1.7 mm Hg to 174±5.2 and 175±4.8 mm Hg, respectively (p<0.01). The EB dye content in the brain regions of the rats in the l-NAME group was increased, but there was no statistical significance compared to the saline group. The extravasation of EB dye was significantly increased in the brain regions of the animals treated with aluminum compared to the rats treated with saline (p<0.05). A significantly higher EB dye content in the brain regions was observed in the l-NAME plus aluminium group compared to l-NAME, aluminum, and saline groups (p<0.01). These findings indicate that exposure to a high level of aluminum leads to an additional increase in BBB permeability where nitric oxide-blockade-induced chronic hypertension potentiates the effect of aluminum to enhance BBB permeability to EB dye.     

Inhibition of nitric oxide synthase enhances contractile response of ventricular myocytes from streptozotocin-diabetic rats

The contractile hyporesponsiveness of the streptozotocin diabetic rat heart in vitro to β-adrenergic agonists is eliminated when the heart is perfused with N^G-nitro-l-arginine methyl ester (l-NAME), a non-selective inhibitor of nitric oxide synthase (NOS). The following study evaluated the hypothesis that an increased production of NO/cGMP within the diabetic myocyte inhibits the β-adrenergic-stimulated increase in calcium current and contractile response. Male Sprague-Dawley rats were given an intravenous injection of streptozotocin (60 mg/kg). After 8 weeks, L-type calcium currents were recorded in ventricular myocytes using the whole cell voltage-clamp method. Shortening of isolated myocytes was determined using a video edge detection system. cAMP and cGMP were measured using radioimmunoassay. Nitric oxide production was determined using the Griess assay kit. Basal cGMP levels and nitric oxide production were elevated in diabetic myocytes. Shortening of the diabetic myocytes in response to isoproterenol (1 μM) was markedly diminished. However, there was no detectable difference in the isoproterenol-stimulated L-type calcium current or cAMP levels between control and diabetic myocytes. Acute superfusion of the diabetic myocyte with l-NAME (1 mM) decreased basal cGMP and markedly enhanced the shortening response to isoproterenol but did not alter isoproterenol-stimulated calcium current. These data suggest that increased production of NO/cGMP within the diabetic myocyte suppressed β-adrenergic stimulated shortening of the myocyte. However, NO/cGMP apparently does not suppress shortening of the myocyte by inhibition of the β-stimulated calcium current.     

Propofol-induced vascular permeability change is related to the nitric oxide signaling pathway and occludin phosphorylation

The present study was undertaken to elucidate the mechanism of intra-arterial propofol-induced vascular permeability change resulting in tissue edema. The mechanism of propofol-induced hyperpermeability was examined in a rat femoral artery injection model. Vascular permeability was determined by measuring the Evans blue content of the dorsal skin of the infused limb at 15, 30, 45 and 60 min after propofol injection. The total content of the tight junction proteins occludin, ZO-1 and claudin-5 under experimental conditions was also determined by western blotting. Intra-arterial injection with propofol resulted in a marked dose-dependent increase in vascular permeability of the rat hindpaw. Pretreatment with 10 mg/kg of N-nitro-l-arginine methyl ester (l-NAME) but not aminoguanidine significantly inhibited the change in vascular permeability after challenge with propofol. Pretreatment with l-arginine and nitroprusside increased the propofol-induced permeability change. Intra-arterial injection of propofol significantly increased occludin phosphorylation after 15 min, which was consistent with the time profile of the vascular permeability change. l-NAME partially reversed the change in occludin phosphorylation, whereas aminoguanidine had no effect compared with that in the controls. Our observations indicate that nitric oxide (NO) is an important mediator in the induction of vascular permeability induced by propofol. Occludin phosphorylation is a determining factor in the vascular permeability change induced by propofol. NO synthase (NOS) inhibitors might be useful in the treatment of accidental intra-arterial injection of propofol, in the reduction of any adverse effects.     

The influence of short-term <Emphasis Type="SmallCaps">l</Emphasis>-arginine supplementation on rats’ muscular and hepatic cells in ischemia–reperfusion syndrome

Due to the complex mechanisms of l-arginine activity, it is difficult to determine the clinical significance of supplementation with this amino acid. The objective of this study was to determine the influence of short-term supplementation with l-arginine in stress conditions, induced by ischemia–reperfusion syndrome, by assessing the damage to muscular and hepatic cells on the basis of creatine kinase (CK), alanine aminotransferase (ALAT) and aspartic aminotransferase (AspAT) activity in blood and the level of oxygen free radicals in analyzed tissues of rats. We observed that induced ischemia of hind limb caused an increase in CK, ALAT and AspAT activity and an increase in the level of free radicals in liver, but not in skeletal muscle. Supplementation with l-arginine led to a reduction in serum activity of CK and AspAT and reduction of the level of free radicals in analysed tissues. Simultaneous supplementation with l-arginine AND l-NAME resulted in a reversal of changes induced by l-arginine supplementation in the case of AspAT and free radicals in skeletal muscle. The results indicate that under conditions of ischemia–reperfusion, short-term administration of l-arginine has a protective effect on skeletal muscle manifesting itself by reduction of CK in the serum and reduction of free radicals level in THIS tissue.     

Effects of tensile stress on the α1 nicotinic acetylcholine receptor expression in maxillofacial skeletal myocytes

The present study was to test the hypothesis that 11,12-epoxyeicosatrienoic acid (11,12-EET), a metabolic product of arachidonic acid by cytochrome P450 epoxygenase, regulates nitric oxide (NO) generation of the l-arginine/NO synthase (NOS) pathway in human platelets. Human platelets were incubated in the presence or absence of different concentrations of 11,12-EET for 2 h at 37°C, followed by measurements of activities of the l-arginine/NOS pathway. Incubation with 11,12-EET increased the platelet NOS activity, nitrite production, cGMP content, and the platelet uptake of l-[³H]arginine in a concentration-dependent manner. In addition, 11,12-EET attenuated intracellular free Ca²⁺ accumulation stimulated by collagen, which was at least partly mediated by EET-activated l-arginine/NOS pathway. It is suggested that 11,12-EET regulates platelet function through up-regulating the activity of the l-arginine/NOS/NO pathway.     

Influence of nitric oxide synthase inhibition on vasopressin and corticosterone secretion during water deprivation in rats

Nitric oxide (NO) is a short-lived radical that functions as a neurotransmitter in the central nervous system and plays a physiological role in the regulation of hypothalamic–pituitary–adrenal axis and vasopressinergic axis. In the present study, we aimed to investigate the interaction between the generation of NO and vasopressin (AVP) and corticosterone release after 3 days of water deprivation in rats. Animals were previously treated with intraperitoneal (i.p.) saline or l-nitro-arginine methyl ester (L-NAME) injection. l-NAME is a nonspecific inhibitor of nitric oxide synthases. In control rats given i.p. saline or l-NAME, hypothalamic, pituitary, and plasma AVP levels and plasma corticosterone did not change from baseline levels (p > 0.05). Three days of water deprivation increased significantly the corticosterone levels in plasma (p < 0.01) and AVP levels in hypothalamus and plasma (p < 0.01), but not in pituitary, which showed a significant decrease. These variations were concomitant with the elevation of nitrates/nitrates in plasma. l-NAME injection abolished significantly (p < 0.01) the elevation of plasma corticosterone and hypothalamic AVP levels induced by water deprivation. These findings showed that in water-deprived rats, nitric oxide synthase inhibition by l-NAME inhibits corticosterone and vasopressin release, suggesting a potent stimulatory role of NO.     

Arginine increases genotoxicity induced by methyl methanesulfonate in human lymphocytes

Nitric oxide (NO) is a free radical that is produced in cells from l-arginine. NO is involved in the physiological control of different tissues, but it can act as a toxic mediator in the cells. In this study we investigated the effect of l-arginine on the genotoxicity induced by methyl methanesulfonate (MMS) in human lymphocytes. Blood was treated with N^G-nitro-l-arginine methyl ester (l-NAME) as an inhibitor of nitric oxide synthase for finding out the role of NO in this effect. Human whole blood was treated with l-arginine (50, 100 and 250 μM) and/or l-NAME, then it was treated in vitro with MMS after 24 h of culture. The lymphocytes were stimulated by phytohemagglutinin to find out the micronuclei in cytokinesis-blocked binucleated cells. DNA fragmentation of lymphocytes was detected by using a fluorescence microscope after propidium iodide staining. These data showed that arginine increased the frequency of MMS-induced micronuclei in lymphocytes. However, the genotoxicity was decreased by using l-NAME. Arginine and l-NAME have not shown any DNA damage in cultured human lymphocytes. In conclusion, addition of l-arginine to MMS as an alkylating agent caused an increase of DNA damage in human lymphocytes. This enhancement of genotoxicity was reduced by NAME as NO inhibitor. It is thus cleared that an increase of DNA damage by arginine and MMS is related to NO production.     

l-NAME has Opposite Effects on the Productions of S-adenosylhomocysteine and S-adenosylmethionine in V12-H-Ras and M-CR3B-Ras Pheochromocytoma Cells

Homocysteine is a sulfur-containing, nonproteinogenic, neurotoxic amino acid biosynthesized during methyl cycles after demethylation of S-adenosylmethionine (SAM) to S-adenosylhomocysteine (SAH) and subsequent hydrolysis of SAH into homocysteine and adenosine. Formed homocysteine is either catabolized into cystathionine (transsulfuration pathway) by cystathionine β-synthase, or remethylated into methionine (remethylation pathway) by methionine synthase. To demonstrate the specificity of Ras-elicited effects on the activity of methyl cycles, wild-type pheochromocytoma PC12, mutant oncogenic rasH gene (MVR) expressing PC12 pheochromocytoma and normal c-rasH stably transfected M-CR3B cells were incubated with the N^ω-nitro-l-arginine methyl ester (l-NAME), and manumycin, (inhibitors of nitric oxide synthase and farnesyltransferase, respectively). We have found that l-NAME significantly changes the SAM/SAH ratio in both MCR and MVR cells. Moreover, these alterations have reciprocal character; in the MCR cells, the SAM/SAH ratio was raised, whereas in the MVR cells this ratio was decreased. We conclude that depletion of endogenous NO with l-NAME increased the production of SAH only in cells with mutated oncogenic RasH, possibly through enhancement of production of reactive oxygen species (ROS). Oxidative stress can increase cystathionine β-synthase activity that switches methyl cycles from remethylation into transsulfuration pathway to maintain the intracellular glutathione pool (essential for the redox-regulating capacity of cells) via an adaptive process.     

The Potential Role of Nitric Oxide Synthase in Survival and Regeneration of Magnocellular Neurons of Hypothalamo-Neurohypophyseal System

Previous investigations from this laboratory have demonstrated that hypophysectomy induces up-regulation of neuronal nitric oxide synthase (nNOS) in magnocellular neurons of the mammalian hypothalamo-neurohypophyseal system (HNS). Accompanied by this upregulation of nNOS, both neuronal regeneration and degeneration are also observed in this system following hypophysectomy. The specific aim of this study was to determine the potential role of nNOS upregulation in neuronal survival and regeneration after hypophysectomy in the adult Sprague–Dawley (SD) rat by using a competitive nitric oxide synthase blocker, N(G)-nitrol-l-arginine methyl ester (l-NAME). We found that l-NAME treatment effectively blocked the regeneration of magnocellular neurons of the rodent hypothalamus as observed in the lumen of the third cerebral ventricle following hypophysectomy. However, l-NAME had no effect on the survival of magnocellular neurons in the supraoptic (SON) and paraventricular (PVN) nuclei after hypophysectomy. These results suggest that the induced increase of nNOS expression enhance the regenerative ability of magnocellular neurons of the HNS following hypophysectomy.     

Antioxidative defense organization in retroperitoneal white adipose tissue during acclimation to cold—The involvement of l-arginine/NO pathway

1. Retroperitoneal white adipose tissue (RpWAT) antioxidative defense was investigated in untreated, l-arginine-treated and N^ω-nitro-l-arginine methyl ester (l-NAME)-treated rats kept at 4±1 °C (1, 3, 7, 12, 21 and 45 days) and compared to control rats at 22±1 °C.

2. Cold-acclimation-induced RpWAT weight decrease was accompanied by a decline in glutathione level and increased activity of manganese superoxide dismutase (MnSOD), glutathione S-transferase (GST), catalase, glutathione peroxidase and glutathione reductase at different time-points.

3. l-arginine accelerated RpWAT weight decrease, the increase in MnSOD and GST activities and the prolonged increase of catalase, MnSOD and GST activities. l-NAME delayed cold-induced catalase activity increase and tissue weight decrease. Prolonged l-NAME-treatment had a similar effect on RpWAT as l-arginine.

4. Results suggest the involvement of l-arginine/NO pathway in RpWAT oxidative metabolic augmentation induced by cold-acclimation.

Nitric oxide modulates cadmium influx during cadmium-induced programmed cell death in tobacco BY-2 cells

Nitric oxide (NO) is a bioactive gas and functions as a signaling molecule in plants exposed to diverse biotic and abiotic stresses including cadmium (Cd²⁺). Cd²⁺ is a non-essential and toxic heavy metal, which has been reported to induce programmed cell death (PCD) in plants. Here, we investigated the role of NO in Cd²⁺-induced PCD in tobacco BY-2 cells (Nicotiana tabacum L. cv. Bright Yellow 2). In this work, BY-2 cells exposed to 150 μM CdCl₂ underwent PCD with TUNEL-positive nuclei, significant chromatin condensation and the increasing expression of a PCD-related gene Hsr203J. Accompanied with the occurring of PCD, the production of NO increased significantly. The supplement of NO by sodium nitroprusside (SNP) had accelerated the PCD, whereas the NO synthase inhibitor Nω-nitro-l-arginine methyl ester hydrochloride (l-NAME) and NO-specific scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO) alleviated this toxicity. To investigate the mechanism by which NO exerted its function, Cd²⁺ concentration was measured subsequently. SNP led more Cd²⁺ content than Cd²⁺ treatment alone. By contrast, the prevention of NO by l-NAME decreased Cd²⁺ accumulation. Using the scanning ion-selective electrode technique, we analyzed the pattern and rate of Cd²⁺ fluxes. This analysis revealed the promotion of Cd²⁺ influxes into cells by application of SNP, while l-NAME and cPTIO reduced the rate of Cd²⁺ uptake or even resulted in net Cd²⁺ efflux. Based on these founding, we concluded that NO played a positive role in CdCl₂-induced PCD by modulating Cd²⁺ uptake and thus promoting Cd²⁺ accumulation in BY-2 cells.     

Dietary Zn deficiency does not influence systemic blood pressure and vascular nitric oxide signaling in normotensive rats

Because zinc (Zn) is an important component for cell protection against certain oxygen species, it has been suggested that Zn deficiency impairs the potent oxidant defense capacity, which is constitutively provided in the vascular system. However, the influence of dietary Zn deficiency on systemic blood pressure and vascular system is controversial and unclear. We therefore examine the effect of dietary Zn deficiency on systemic blood pressure, a potent superoxide scavenger, aortic Cu/Zn superoxide dismutase (SOD) activity, a most representative synthase of the endothelium-derived relaxing factor, and aortic endothelial nitric oxide synthase (eNOS) expression. Furthermore, the direct effects of intravenous administration of NOS inhibitor, N ^ω-nitro-l-arginine methyl ester (l-NAME), and a SOD mimetic compound, tempol, in normotensives were tested in Wistar-Kyoto (WKY) rats. A Zn-deficient diet (4 wk) contributed to growth retardation, the decrease in thymus weight, and the lower levels of serum Zn compared with the standard diet group. However, no significant difference in conscious systolic and diastolic blood pressure was found in the Zn-deficiency group. The administration of l-NAME caused an increase in the mean arterial pressure (MAP) levels in the two groups of rats and the involvement of the vasodilator nitric oxide (NO) in the regulation of systemic BP in the normotensive state. On the other hand, administration of the superoxide scavenger, tempol, led to a decrease in MAP levels in the two groups of rats, indicating the participation of the oxygen free radical, superoxide, in the maintenance of the systemic BP in a normotensive state. There were no significant differences between the Zn-deficient diet group and the standard diet group in the normotensive state. eNOS expression and Cu/Zn SOD activity in the aorta were also intact in Zn-deficient normotensive rats. These findings suggest that the 4 wk of Zn deficiency was inadequate to alter systemic blood pressure and focal NO signaling in the normotensive state. Long-term Zn deficiency affects the neuronal, immune, and hematopoietic systems, which contribute to systemic and/or local circulation. However, Zn deficiency alone does not cause hypertension and local vascular dysfunction in the normotensive state.     

Antimicrobial activity of various cationic molecules on foodborne pathogens

Antibacterial effects of various arginine- and lysine-rich polycationic proteins and polymers were evaluated by broth and solid dilution assay on a range of foodborne pathogens, Gram-positive and Gram-negative bacteria. The Minimum Inhibitory Concentration (MIC) and the Minimum Bactericidal Concentration (MBC) of α-poly-l-lysine (poly-lys), α-poly-l-arginine (poly-arg) and protamines from herring sperm (clupeine sulphate) and salmon sperm (salmine sulphate) were determined on Bacillus subtilis, Bacillus cereus, Staphylococcus aureus, Listeria monocytogenes, Salmonella typhimurium, Shigella sonnei, Escherichia coli O157:H7 and Pseudomonas aeruginosa. All these molecules showed antibacterial activity on all strains with different MIC and MBC values. The molecular mechanisms underlying the effect of α-poly-l-arginine might be related to the entrance of the molecule into the cell. In fact α-poly-l-arginine labelled with 7-Diethylamino coumarin-3-carboxylic acid, succinimidyl ester (DEAC,SE) showed ability to permeate the cell membrane of B. cereus and E. coli O157:H7.     

Interferon-γ- and lipopolysaccharide-induced tumor necrosis factor-α is required for nitric oxide production: Tumor necrosis factor-α and nitric oxide are independently involved in the killing ofMycobacterium microti in interferon-γ-and lipopolysaccharide-treated J774A.1 cells

A comparative study was done using J774A.1 and J774A. 1-derived transfected cells (J774A.1 C.1) containing antisense tumor necrosis factor α (TNF-α) plasmid to determine the role of endogenous TNF-α on nitric oxide production as well as on the growth ofMycobacterium microti in interferon γ (IFN-γ)- and lipopolysaccharide (LPS)-treated cells. On stimulation with IFN-γ and LPS a higher level of NO was observed in J774A.1 cells compared to J774A.1 C.1 which indicated that endogenous TNF-α is required for the production of NO. Comparing the effect of IFN-γ and LPS on the intracellular growth ofM. microti, the growth-reducing activity was higher in J774A.1 cells than in J774A.1 C.1 cells and was not completely abrogated in the presence of the nitric oxide inhibitorN ^G-methyl-l-arginine (l-NMA). J774A.1 C.1 cells infected withM. microti produced a significant amount of NO when exogenous TNF-α was added along with IFN-γ and LPS and the concentration of intracellular bacteria decreased almost to that in IFN-γ and LPS treated parental J774A.1 cells. Addition of exogenous TNF-α even in the presence ofl-NMA in J774.1 C.1 cells could also partially restore intracellular growth inhibition ofM. microti caused by IFN-γ and LPS. TNF-α is probably required for the production of NO in J774A.1 cells by IFN-γ and LPS but TNF-α and NO are independently involved in the killing of intracellularM. microti with IFN-γ and LPS.