scholarly journals Fasting-induced intestinal apoptosis is mediated by inducible nitric oxide synthase and interferon-γ in rat

2010 ◽  
Vol 298 (6) ◽  
pp. G916-G926 ◽  
Author(s):  
Junta Ito ◽  
Hiroyuki Uchida ◽  
Takayuki Yokote ◽  
Kazuo Ohtake ◽  
Jun Kobayashi
Keyword(s):  
Nitric Oxide ◽  
Protein Expression ◽  
Interferon Γ ◽  
No Synthase ◽  
Histological Evaluation ◽  
No Production ◽  
Ros Production ◽  
Nitrite Production ◽  
Ifn Γ

Nitric oxide (NO) is associated with intestinal apoptosis in health and disease. This study aimed to investigate the role of intestinal NO in the regulation of apoptosis during fasting in rats. Male Wistar rats were divided into two groups and subcutaneously injected with saline (SA) or aminoguanidine (AG), followed by fasting for 24, 48, 60, and 72 h. At each time point, the jejunum was subjected to histological evaluation for enterocyte apoptosis by histomorphometric assessment and TUNEL analysis. We performed immunohistochemistry for inducible NO synthase (iNOS) expression in the jejunum and measured tissue nitrite levels using HPLC and 8-hydroxydeoxyguanosine adduct using ELISA, indicative of endogenous NO production and reactive oxygen species (ROS) production, respectively. Jejunal transcriptional levels of iNOS, neuronal NO synthase (nNOS), and interferon-γ (IFN-γ) were also determined by RT-PCR. Fasting caused significant jejunal mucosal atrophy due to attenuated cell proliferation and enhanced apoptosis with increase in iNOS transcription, its protein expression in intestinal epithelial cells (IEC), and jejunal nitrite levels. However, AG treatment histologically reduced apoptosis with inhibition of fasting-induced iNOS transcription, protein expression, and nitrite production. We also observed fasting-induced ROS production and subsequent IFN-γ transcription, which were all inhibited by AG treatment. Furthermore, we observed reduced transcriptional levels of nNOS, known to suppress iNOS activation physiologically. These results suggest that fasting-induced iNOS activation in IEC may induce apoptosis mediators such as IFN-γ via a ROS-mediated mechanism and also a possible role of nNOS in the regulation of iNOS activity in fasting-induced apoptosis.

scholarly journals Role of cytokines and nitric oxide in the induction of tuberculostatic macrophage functions

2000 ◽  
Vol 9 (6) ◽  
pp. 261-269 ◽  
Author(s):  
Vera L. Petricevich ◽  
Rosely C. B. Alves
Keyword(s):  
Nitric Oxide ◽  
Culture Media ◽  
Peritoneal Macrophages ◽  
Interferon Γ ◽  
Enzyme Linked Immunosorbent Assay ◽  
No Production ◽  
Phenotypic Differences ◽  
Ifn Γ ◽  
Necrosis Factor

The aim of this study was to determine phenotypic differences when BCG invades macrophages. Bacilli prepared from the same BCG primary seed, but produced in different culture media, were analysed with respect to the ability to stimulate macrophages and the susceptibility to treatment with cytokines and nitric oxide (NO). Tumour necrosis factor (TNF) activity was assayed by measuring its cytotoxic activity on L-929 cells, interleukin-6 (IL-6) and interferon γ (IFN-γ) were assayed by enzyme-linked immunosorbent assay (ELISA), whereas NO levels were detected by Griess colorimetric reactions in the culture supernatant of macrophages incubated with IFN-γ , TNF or NO and subsequently exposed to either BCG-I or BCG-S. We found that BCG-I and BCGS bacilli showed different ability to simulate peritoneal macrophages. Similar levels of IL-6 were detected in stimulated macrophages with lysate from two BCG samples. The highest levels of TNF and IFN-γ were observed in macrophages treated with BCG-S and BCG-I, respectively. The highest levels of NO were observed in cultures stimulated for 48h with BCG-S. We also found a different susceptibility of the bacilli to ex ogenous treatm ent w ith IFN-γ and TNF which were capable of killing 60 and 70% of both bacilli, whereas NO was capable of killing about 98 and 47% of BCG-I and BCG-S, respectively. The amount of bacilli proportionally decreased with IFN-γ and TNF, suggesting a cytokine-related cytotox ic effect. Moreover, NO also decreased the viable number of bacilli. Interestingly, NO levels of peritoneal macrophages were significantly increased after cytokine treatment. This indicates that the treatment of macrophages with cytokines markedly reduced bacilli number and presented effects on NO production. The results obtained here emphasize the importance of adequate stimulation for guaranteeing efficient killing of bacilli. In this particular case, the IFN-γ and TNF were involved in the activation of macrophage bactericidal activity.

scholarly journals Inducible nitric oxide synthase gene deletion exaggerates MAPK-mediated cyclooxygenase-2 induction by inflammatory stimuli

2010 ◽  
Vol 299 (3) ◽  
pp. H613-H623 ◽  
Author(s):  
Brian D. Lamon ◽  
Rita K. Upmacis ◽  
Ruba S. Deeb ◽  
Hilal Koyuncu ◽  
David P. Hajjar
Keyword(s):  
Nitric Oxide ◽  
Atherosclerotic Lesion ◽  
No Synthase ◽  
No Production ◽  
Nitrite Production ◽  
Inflammatory Stimuli ◽  
Ifn Γ ◽  
Cox 2 ◽  
The Impact ◽  
Inducible Nitric Oxide

Cyclooxygenase (COX)-2 and inducible nitric oxide (NO) synthase (iNOS) are responsive to a wide array of inflammatory stimuli, have been localized to vascular smooth muscle cells (SMCs), and are intimately linked to the progression of vascular disease, including atherosclerotic lesion formation. We and others have shown that the production and subsequent impact of COX products appear to be correlative with the status of NO synthesis. This study examined the impact of inflammation-driven NO production on COX-2 expression in SMCs. Concurrent stimulation of quiescent rat aortic SMCs with lipopolysaccharide (LPS) and interferon (IFN)-γ increased COX-2, iNOS, and nitrite production. Pharmacological inhibition of NO synthase ( NG-monomethyl-l-arginine) concentration- and time-dependently magnified LPS + IFN-γ-mediated COX-2 mRNA and protein induction in a cGMP-independent manner. COX-2 induction was associated with activation of the ERK, p38, and JNK mitogen-activated protein kinase (MAPK) pathways. Interestingly, NO synthase inhibition enhanced ERK, p38, and to a lesser extent JNK phosphorylation but suppressed MAPK phosphatase (MKP)-1 induction in response to LPS + IFN-γ. Similarly, the exposure of SMCs from iNOS−/− mice to LPS + IFN-γ produced an enhancement of COX-2 induction, p38, and JNK phosphorylation and an attenuated upregulation of MKP-1 versus their wild-type counterparts. Taken together, our data indicate that NO, in part derived from iNOS, negatively regulates the immediate early induction of COX-2 in response to inflammatory stimuli.

scholarly journals Acquired interferon γ responsiveness during Caco-2 cell differentiation: effects on iNOS gene expression

Gut ◽  
1999 ◽  
Vol 44 (5) ◽  
pp. 659-665 ◽  
Author(s):  
A M Chavez ◽  
M J Morin ◽  
N Unno ◽  
M P Fink ◽  
R A Hodin
Keyword(s):  
Nitric Oxide ◽  
Cell Differentiation ◽  
Interferon Γ ◽  
Intestinal Barrier Function ◽  
Inos Mrna ◽  
Pyrrolidine Dithiocarbamate ◽  
No Production ◽  
Mrna Induction ◽  
Ifn Γ ◽  
Inos Induction

BACKGROUNDImpairment of intestinal barrier function occurs under a variety of inflammatory conditions and is mediated at least in part by interferon γ (IFN-γ) induced nitric oxide (NO) production. Previous in vivo studies have shown that systemic lipopolysaccharide treatment caused an induction of the rat inducible nitric oxide synthase (iNOS) mRNA primarily in villus cells, rather than in undifferentiated crypt cells.AIMSTo examine iNOS induction by IFN-γ in vitro as a function of enterocyte differentiation.METHODSPreconfluent and postconfluent Caco-2 cells were treated with IFN-γ in the presence or absence of various inhibitors. Northern analyses were performed to assess the magnitude of iNOS mRNA induction. IFN-γ receptor mRNA and protein levels were determined.RESULTSiNOS mRNA induction by IFN-γ occurred at two hours and was not blocked by cycloheximide, indicating that it is an immediate early response. iNOS induction and nitrite/nitrate increases were inhibited by dexamethasone and pyrrolidine dithiocarbamate, supporting an important role for the NF-κB transcription factor in this process. The stimulated iNOS induction was seen almost exclusively under conditions of cellular differentiation—that is, in postconfluent Caco-2 cells. This increased IFN-γ responsiveness seen in postconfluent Caco-2 cells correlated with an increased expression of IFN-γ receptor, whereas T84 and HT-29 cells did not show any significant alterations in either iNOS induction or IFN-γ receptor levels as a function of postconfluent growth.CONCLUSIONSWith regard to iNOS mRNA induction, IFN-γ responsiveness is acquired during Caco-2 cell differentiation, perhaps related to an increase in the numbers of IFN-γ receptors.

scholarly journals Nitric oxide production in murine spleen cells: role of interferons and prostaglandin E2in the generation of cytotoxic activity

1996 ◽  
Vol 5 (1) ◽  
pp. 62-68 ◽  
Author(s):  
Dominique Vaillier ◽  
Richard Daculsi ◽  
Norbert Gualdel
Keyword(s):  
Nitric Oxide ◽  
Cytotoxic Activity ◽  
Minor Role ◽  
No Production ◽  
Spleen Cells ◽  
Kinase C ◽  
Murine Spleen ◽  
Ifn Γ ◽  
A Minor

The production of nitric oxide (NO) was measured in cultures of spleen cells stimulated by lipopolysaccharide (LPS), IL-2 or LPS + IL-2. We observed that NO synthesis is increased by IFN-γ but inhibited by IFN-α/β. This is not the case when IL-2 is present in the cultures, since interferons play a minor role in the regulation of the NO production. When IL-2 and LPS were associated in the cultures, the IFN-α/β role seems more important than that of IFN-γ. PGE2inhibits NO production in LPS supplemented cultures but has a slight effect in the presence of IL-2 and no effect with IL-2 + LPS. 3-isoButyl-1-methylxanthine (IBMX), an inhibitor of phosphodiesterases, induces a decrease of IFN production. In the presence of H-7, an inhibitor of protein kinase C (PKC), NO production is reduced when the cultures are supplemented by LPS or IL-2 but not when IL-2 and LPS are both added. H-7 also reduced IFN production. In the presence of NG-monomethyl-L-arginine (N-MMA), an inhibitor of NO synthesis, IFN production was increased, with no change in the cytotoxic activity. Hence, interferons regulate NO production by mouse spleen cells and, in return, NO modulates the generation of IFN.

Free Arginase-I in Human Pulmonary Microvascular Endothelial Cell Culture Leads to Endothelial Dysfunction and Increased Reactive Oxygen Species.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3041-3041
Author(s):  
John S. Corns ◽  
Leif D Nelin
Keyword(s):  
Nitric Oxide ◽  
Fold Change ◽  
No Production ◽  
Ros Production ◽  
Oxygen Species ◽  
Protein Levels ◽  
Nitrite Production ◽  
Urea Production ◽  
The Media ◽  
Arginase I

Abstract Abstract 3041 Poster Board II-1017 Introduction Endothelial dysfunction in chronic hemolytic diseases (e.g., sickle cell disease) is believed to lead to hemolysis-associated pulmonary hypertension. Dysregulated arginine metabolism, via free arginase-I released during hemolysis, is one mechanism proposed in prior studies. However, the effects of free arginase-I on endothelial cell function have not previously been reported. We hypothesized that free arginase-I would decrease hPMVEC nitric oxide (NO) production and therefore limit NO bioavailability. Methods Cultured human pulmonary microvascular endothelial cells (hPMVEC), at passage 6-8 and approximately 80% confluence, with increasing arginase-I concentrations added to the media (CN: 0 ng prot/mL, A30: 30 ng prot/mL, A90: 90 ng prot/mL) were studied. To maximize arginase-I activity, 0.1 mM MnCl2 (an important cofactor for arginase-I) and 3mM of L-arginine were added to each condition. The hPMVEC were incubated for 24h in normoxia (n=8) or hypoxia (1% O2; n=4), then media was harvested for urea/nitrite analyses and cell protein was harvested for Western blot analyses. ANOVA analyses were conducted to determine statistical significance. Results Urea production significantly increased with increasing media arginase-I in both normoxia (CN: 1.47±0.22, A30: 1.46±0.15, A90: 2.09±0.20 umol/mgPr; p<0.05) and hypoxia (CN: 0.913±0.22, A30: 1.01±0.04, A90: 1.99±0.14 umol/mgPr; p<0.005); interestingly, hypoxia means were significantly lower than those in normoxia (p<0.01). However, the percent change from controls in hypoxia (CN: 100±24, A30: 110±4, A90: 218±15) was significantly higher (p<0.05) than in normoxia conditions (CN: 100±15, A30: 99±10, A90: 142±13). Nitrite production showed no significant change with increasing media arginase-I in normoxia (CN: 17.6±2.4, A30: 20.8±4.6, A90: 18.8±2.1 nmol/mgPr; p=0.79); however, in hypoxia nitrites trended down with increasing media arginase-I (CN: 5.26±0.42, A30: 4.35±0.27, A90: 3.69±0.51 nmol/mgPr; p=0.068). In addition, means for nitrite production in hypoxia were significantly less than in normoxia conditions (p<0.0001). In normoxia, Western blot analyses showed significantly decreased 3-nitrotyrosine (3-NT) levels with increasing media arginase-I (CN: 1.00±0.03, A30: 0.88±0.07, A90: 0.77±0.07 [fold change]; p<0.05). 3-NT expression in hypoxia showed no differences with increasing media arginase-I (CN: 1.90±0.16, A30: 1.88±0.19, A90: 1.75±0.27 [fold change, normalized to normoxia control]; p=0.87); however, hypoxic means were significantly higher than in normoxia conditions (p<0.0001). In normoxia, NAD(P)H oxidase isoform 4 (Nox4) show a decreasing trend in protein levels with increasing media arginase-I (CN: 1.00±0.03, A30: 0.93±0.10, A90: 0.86±0.06 [fold change]), with an increase in Nox4 protein levels in hypoxia with increasing media arginase-I (CN: 1.18±0.02, A30: 1.33±0.08, A90: 1.47±0.01 [fold change, normalized to normoxia control]). The hypoxic levels of Nox4 were also higher than in normoxia conditions. Conclusions As expected, increasing free media arginase-I concentrations in hPMVEC culture resulted in significant increases in urea production in both normoxia and hypoxia. Hypoxia resulted in lower nitrite production by endothelial nitric oxide synthase (eNOS). However, increasing amounts of free arginase-I only affected nitrite production in hypoxia, suggesting that increased arginase-I serum concentrations may also become an important regulator of NO production in clinical cases of hypoxia (e.g., acute chest syndrome and vaso-occlusive crises in sickle cell patients). The concurrent increased expression of 3-NT in hypoxia, despite lower nitrites, implies an increase in reactive oxygen species (ROS). Our preliminary data for Nox4 expression, the predominant source of ROS in endothelial cells, indicates that ROS production via this pathway may increase in hypoxia with increasing free arginase-I. Further studies on how free arginase-I in the media affects endothelial function are warranted, particularly in regards to the mechanism by which free arginase-I leads to greater Nox4 expression and ROS production. We speculate that arginase-I inhibitors may represent a potential therapeutic target in patients with chronic hemolysis and acute hypoxic events. Disclosures No relevant conflicts of interest to declare.

Role of airway nitric oxide on the regulation of pulmonary circulation by carbon dioxide

2001 ◽  
Vol 91 (3) ◽  
pp. 1121-1130 ◽  
Author(s):  
Yasushi Yamamoto ◽  
Hitoshi Nakano ◽  
Hiroshi Ide ◽  
Toshiyuki Ogasa ◽  
Toru Takahashi ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Pulmonary Arterial Pressure ◽  
Hypoxic Pulmonary Vasoconstriction ◽  
No Synthase ◽  
No Production ◽  
Pulmonary Vasoconstriction ◽  
Exhaled No ◽  
Progressive Hypoxia ◽  
Pulmonary Arterial

The effects of hypercapnia (CO2) confined to either the alveolar space or the intravascular perfusate on exhaled nitric oxide (NO), perfusate NO metabolites (NOx), and pulmonary arterial pressure (Ppa) were examined during normoxia and progressive 20-min hypoxia in isolated blood- and buffer-perfused rabbit lungs. In blood-perfused lungs, when alveolar CO2concentration was increased from 0 to 12%, exhaled NO decreased, whereas Ppa increased. Increments of intravascular CO2levels increased Ppa without changes in exhaled NO. In buffer-perfused lungs, alveolar CO2 increased Ppa with reductions in both exhaled NO from 93.8 to 61.7 (SE) nl/min ( P < 0.01) and perfusate NOx from 4.8 to 1.8 nmol/min ( P < 0.01). In contrast, intravascular CO2 did not affect either exhaled NO or Ppa despite a tendency for perfusate NOx to decline. Progressive hypoxia elevated Ppa by 28% from baseline with a reduction in exhaled NO during normocapnia. Alveolar hypercapnia enhanced hypoxic Ppa response up to 50% with a further decline in exhaled NO. Hypercapnia did not alter the apparent K m for O2, whereas it significantly decreased the V max from 66.7 to 55.6 nl/min. These results suggest that alveolar CO2 inhibits epithelial NO synthase activity noncompetitively and that the suppressed NO production by hypercapnia augments hypoxic pulmonary vasoconstriction, resulting in improved ventilation-perfusion matching.

Effect of corticosteroids on nitric oxide production in inflammatory bowel disease: are leukocytes the site of action?

2005 ◽  
Vol 288 (2) ◽  
pp. G261-G267 ◽  
Author(s):  
John D. Linehan ◽  
George Kolios ◽  
Vassilis Valatas ◽  
Duncan A. F. Robertson ◽  
John Westwick
Keyword(s):  
Nitric Oxide ◽  
Ulcerative Colitis ◽  
Mononuclear Cells ◽  
Intestinal Inflammation ◽  
Inos Mrna ◽  
No Production ◽  
Nitrite Production ◽  
Ifn Γ ◽  
Colonic Biopsies ◽  
Ht 29

Nitric oxide (NO) production is increased in the human colonic mucosa in intestinal inflammation. We examined the effect of corticosteroids and the role of mononuclear cells in this production. Colonic biopsies from patients with ulcerative colitis and normal controls were cultured with either budesonide or prednisolone in the presence of proinflammatory cytokines. Human mixed mononuclear cells (MMCs) were cocultured with HT-29 cells stimulated with IFN-γ and LPS in the presence or absence of corticosteroids. Nitrite production was measured in supernatants by a modification of the Griess reaction, and inducible NO synthase (iNOS) mRNA expression was studied in colonic tissue by RT-PCR. Both steroids significantly suppressed the nitrite production and iNOS mRNA expression in inflamed colonic biopsies from ulcerative colitis patients and in cytokine-stimulated normal colonic biopsies but not in cytokine-stimulated HT-29 cells. Nitrite production by HT-29 cells was significantly increased ( P < 0.01) in cocultures with MMCs stimulated with IFN-γ and LPS. The presence of either prednisolone or budesonide significantly ( P < 0.01) suppressed nitrite production from cocultures of HT-29 cells and MMCs but not from cultures of HT-29 cells stimulated with conditioned media from activated MMCs. Interestingly, stimulation of HT-29 with conditioned media from MMCs pretreated with steroids before stimulation with LPS and IFN-γ induced a significantly ( P < 0.01) lower nitrite production. These results suggest that the inhibitory effect of corticosteroids on the NO production in the intestinal inflammation might be via the inhibition of MMC-produced mediators responsible for NO production by colonic epithelial cells.

scholarly journals Salt-induced hypertension in Dahl salt-resistant and salt-sensitive rats with NOS II inhibition

1999 ◽  
Vol 277 (2) ◽  
pp. H732-H739 ◽  
Author(s):  
M. Audrey Rudd ◽  
Maria Trolliet ◽  
Susan Hope ◽  
Anne Ward Scribner ◽  
Geraldine Daumerie ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Function ◽  
No Synthase ◽  
No Production ◽  
Induced Hypertension ◽  
Blood Pressures ◽  
Diameter Change ◽  
Inducible Nos ◽  
Tail Cuff

Although recent evidence suggests that reduced nitric oxide (NO) production may be involved in salt-induced hypertension, the specific NO synthase (NOS) responsible for the conveyance of salt sensitivity remains unknown. To determine the role of inducible NOS (NOS II) in salt-induced hypertension, we treated Dahl salt-resistant (DR) rats with the selective NOS II inhibitor 2-amino-5,6-dihydro-6-methyl-4H-1,3-thiazine (AMT) for 12 days. Tail-cuff systolic blood pressures rose 29 ± 6 and 42 ± 8 mmHg in DR rats given 150 and 300 nmol AMT/h, respectively ( P < 0.01, 2-way ANOVA) after 7 days of 8% NaCl diet. We observed similar results with two other potent selective NOS II inhibitors, S-ethylisourea (EIT) and N-[3-(aminomethyl)benzyl]acetamidine hydrochloride (1400W). Additionally, AMT effects were independent of alterations in endothelial function as assessed by diameter change of mesenteric arterioles in response to methacholine using videomicroscopy. We, therefore, conclude from these data that NOS II is important in salt-induced hypertension.

Sex differences in renal injury and nitric oxide production in renal wrap hypertension

2005 ◽  
Vol 288 (1) ◽  
pp. H43-H47 ◽  
Author(s):  
Hong Ji ◽  
Carlo Pesce ◽  
Wei Zheng ◽  
James Kim ◽  
Yinghua Zhang ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Sex Differences ◽  
Protein Expression ◽  
Renal Injury ◽  
Renal Cortex ◽  
No Synthase ◽  
Female Rats ◽  
No Production ◽  
Nitric Oxide Production ◽  
Renal Disease Progression

To investigate the faster rate of renal disease progression in men compared with women, we addressed the following questions in the renal wrap (RW) model of hypertension: 1) Do sex differences exist in RW-induced renal injury, which are independent of sex differences in blood pressure? 2) Do sex differences in nitric oxide (NO) production exist in RW hypertension? Male (M) and female (F) rats underwent sham-operated (M-Sham, n = 7; F-Sham, n = 10) or RW (M-RW, n = 13; F-RW, n = 14) surgery for 9 wk. Markers of renal injury, including the glomerulosclerosis index (F-RW, 0.70 ± 0.1 vs. M-RW, 2.2 ± 0.6; P < 0.05), mean glomerular volume (F-RW, 1.05 ± 0.050 × 106 vs. M-RW, 1.78 ± 0.15 × 106 μm3; P < 0.001), and proteinuria (F-RW, 68.7 ± 15 vs. M-RW, 124 ± 7.7 mg/day; P < 0.001) were greater in RW males compared with RW females. Endothelial NO synthase protein expression was elevated in the renal cortex (3.2-fold) and medulla (2.2-fold) 9 wk after RW in males, whereas no differences were observed in females. Neuronal NO synthase protein expression was unchanged in the renal cortex in males and in both the renal cortex and medulla in females, whereas in the male medulla, neuronal NOS was decreased by 57%. These data suggest the degree of renal injury is greater in male compared with female rats in RW hypertension despite similar degrees of hypertension and renal function and may involve sex differences in renal NO metabolism.

scholarly journals Gastrointestinal nitric oxide generation in germ-free and conventional rats

2004 ◽  
Vol 287 (5) ◽  
pp. G993-G997 ◽  
Author(s):  
Tanja Sobko ◽  
Claudia Reinders ◽  
Elisabeth Norin ◽  
Tore Midtvedt ◽  
Lars E. Gustafsson ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Gastrointestinal Tract ◽  
Intestinal Microflora ◽  
No Synthase ◽  
No Production ◽  
Germ Free ◽  
Qualitative And Quantitative ◽  
Nitrate Load ◽  
Nos Inhibitor

Nitric oxide (NO) is a central mediator of various physiological events in the gastrointestinal tract. The influence of the intestinal microflora for NO production in the gut is unknown. Bacteria could contribute to this production either by stimulating the mucosa to produce NO, or they could generate NO themselves. Using germ-free and conventional rats, we measured gaseous NO directly in the gastrointestinal tract and from the luminal contents using a chemiluminescence technique. Mucosal NO production was studied by using an NO synthase (NOS) inhibitor, and to evaluate microbial contribution to the NO generation, nitrate was given to the animals. In conventional rats, luminal NO differed profoundly along the gastrointestinal tract with the greatest concentrations in the stomach [>4,000 parts per billion (ppb)] and cecum (≈200 ppb) and lower concentrations in the small intestine and colon (≤20 ppb). Cecal NO correlated with the levels in incubated luminal contents. NOS inhibition lowered NO levels in the colon, without affecting NO in the stomach and in the cecum. Gastric NO increased greatly after a nitrate load, proving it to be a substrate for NO generation. In germ-free rats, NO was low (≤30 ppb) throughout the gastrointestinal tract and absent in the incubated luminal contents. NO also remained low after a nitrate load. Our results demonstrate a pivotal role of the intestinal microflora in gastrointestinal NO generation. Distinctly compartmentalized qualitative and quantitative NO levels in conventional and germ-free rats reflect complex host microbial cross talks, possibly making NO a regulator of the intestinal eco system.

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