Predicting reamputation risk in patients undergoing lower extremity amputation due to the complications of peripheral artery disease and/or diabetes.

BACKGROUND: Patients undergoing amputation of the lower extremity for the complications of peripheral artery disease and/or diabetes are at risk of treatment failure and the need for reamputation at a higher level. The aim of this study was to develop a patient-specific reamputation risk prediction model. METHODS: Patients with incident unilateral transmetatarsal, transtibial or transfemoral amputation between 2004 and 2014 secondary to diabetes and/or peripheral artery disease, and who survived 12 months after amputation, were identified using Veterans Health Administration databases. Procedure codes and natural language processing were used to define subsequent ipsilateral reamputation at the same or higher level. Stepdown logistic regression was used to develop the prediction model. It was then evaluated for calibration and discrimination by evaluating the goodness of fit, area under the receiver operating characteristic curve (AUC) and discrimination slope. RESULTS: Some 5260 patients were identified, of whom 1283 (24·4 per cent) underwent ipsilateral reamputation in the 12 months after initial amputation. Crude reamputation risks were 40·3, 25·9 and 9·7 per cent in the transmetatarsal, transtibial and transfemoral groups respectively. The final prediction model included 11 predictors (amputation level, sex, smoking, alcohol, rest pain, use of outpatient anticoagulants, diabetes, chronic obstructive pulmonary disease, white blood cell count, kidney failure and previous revascularization), along with four interaction terms. Evaluation of the prediction characteristics indicated good model calibration with goodness-of-fit testing, good discrimination (AUC 0·72) and a discrimination slope of 11·2 per cent. CONCLUSION: A prediction model was developed to calculate individual risk of primary healing failure and the need for reamputation surgery at each amputation level. This model may assist clinical decision-making regarding amputation-level selection.

Mortality prediction following non-traumatic amputation of the lower extremity.

BACKGROUND: Patients who undergo lower extremity amputation secondary to the complications of diabetes or peripheral artery disease have poor long-term survival. Providing patients and surgeons with individual-patient, rather than population, survival estimates provides them with important information to make individualized treatment decisions. METHODS: Patients with peripheral artery disease and/or diabetes undergoing their first unilateral transmetatarsal, transtibial or transfemoral amputation were identified in the Veterans Affairs Surgical Quality Improvement Program (VASQIP) database. Stepdown logistic regression was used to develop a 1-year mortality risk prediction model from a list of 33 candidate predictors using data from three of five Department of Veterans Affairs national geographical regions. External geographical validation was performed using data from the remaining two regions. Calibration and discrimination were assessed in the development and validation samples. RESULTS: The development sample included 5028 patients and the validation sample 2140. The final mortality prediction model (AMPREDICT-Mortality) included amputation level, age, BMI, race, functional status, congestive heart failure, dialysis, blood urea nitrogen level, and white blood cell and platelet counts. The model fit in the validation sample was good. The area under the receiver operating characteristic (ROC) curve for the validation sample was 0·76 and Cox calibration regression indicated excellent calibration (slope 0·96, 95 per cent c.i. 0·85 to 1·06; intercept 0·02, 95 per cent c.i. -0·12 to 0·17). Given the external validation characteristics, the development and validation samples were combined, giving a total sample of 7168. CONCLUSION: The AMPREDICT-Mortality prediction model is a validated parsimonious model that can be used to inform the 1-year mortality risk following non-traumatic lower extremity amputation of patients with peripheral artery disease or diabetes.

Recent trends in testosterone testing, low testosterone levels, and testosterone treatment among Veterans.

Low serum testosterone (T) is common and increasingly prevalent with increased age. Recent studies report an 'epidemic' of T prescribing and concern about unnecessary T treatment. We investigated the number of men tested for T, the prevalence of low serum T levels, and initiation of T treatment among those with low T levels in men treated at Veterans Affairs (VA) facilities in the Northwest US (VISN 20). We identified male Veterans aged 40-89 years and examined yearly proportions of men tested for T, found to have low T levels (total T < 280 ng/dL, free T < 34 pg/mL, or bioavailable T < 84 ng/dL), and subsequently treated with T from 2002 to 2011. We excluded men who had T treatment in the year prior and men with diagnoses of prostate or breast cancer. Treatment initiation was defined as the first prescription for T within a year following a low T test. From 2002 to 2011, the yearly population of eligible men in VISN 20 increased from 129 247 to 163 572. The proportion of men who had serum T tests increased from 3.2% in 2002 to 5.8% in 2011. Among the tested men, the percentage of men with low T levels increased from 35.0 to 47.3%. However, the proportion of men with low T levels who were given T treatment within a year decreased from 31.0 to 28.0%. Despite large increases in T testing, and detection of men with low T levels, there was a slight decrease in the proportion of men with low T levels who were treated with T. The decrease in T treatment during this time period contrasts with other studies and may be related to higher comorbidity in Veterans and/or VA formulary restrictions on the use of transdermal T formulations.

Role of the nitric oxide pathway and the endocannabinoid system in neurogenic relaxation of corpus cavernosum from biliary cirrhotic rats.

BACKGROUND AND PURPOSE: Relaxation of corpus cavernosum, which is mediated by nitric oxide (NO) released from non-adrenergic non-cholinergic (NANC) neurotransmission, is critical for inducing penile erection and can be affected by many pathophysiological conditions. However, the peripheral effect of liver cirrhosis on erectile function is as yet unknown. The aim of the present study was to investigate the effect of biliary cirrhosis on NANC-mediated relaxation of rat corpus cavernosum and the possible roles of endocannabinoid and nitric oxide systems in this model. EXPERIMENTAL APPROACH: Cirrhosis was induced by bile duct ligation. Controls underwent sham operation. Four weeks later, strips of corpus cavernosum were mounted in a standard organ bath and NANC-mediated relaxations were obtained by applying electrical field stimulation. KEY RESULTS: The NANC-mediated relaxation was enhanced in corporal strips from cirrhotic animals. Anandamide potentiated the relaxations in both groups. Either AM251 (CB(1) antagonist) or capsazepine (vanilloid VR(1) antagonist), but not AM630 (CB(2) antagonist), prevented the enhanced relaxations of cirrhotic strips. Either the non-selective NOS inhibitor L-NAME or the selective neuronal NOS inhibitor L-NPA inhibited relaxations in both groups, but cirrhotic groups were more resistant to the inhibitory effects of these agents. Relaxations to sodium nitroprusside (NO donor) were similar in tissues from the two groups. CONCLUSIONS AND IMPLICATIONS: Cirrhosis potentiates the neurogenic relaxation of rat corpus cavernosum probably via the NO pathway and involving cannabinoid CB(1) and vanilloid VR(1) receptors.

Naltrexone, an opioid receptor antagonist, attenuates liver fibrosis in bile duct ligated rats.

AIM: The aim of this study was to investigate the hypothesis that the opioid system is involved in the development of hepatic fibrosis. METHODS: The effect of naltrexone (an opioid receptor antagonist) on hepatic fibrosis in bile duct ligated (BDL) or sham rats was assessed by histology and hepatic hydroxyproline levels. Liver matrix metalloproteinase 2 (MMP-2) was measured by zymography, and alpha smooth muscle actin (alpha-SMA) and CD45 (leucocyte common antigen) by immunohistochemistry. The redox state of the liver was assessed by hepatic glutathione (GSH)/oxidised glutathione (GSSG) and S-nitrosothiol levels. Subtypes of opioid receptors in cultured hepatic stellate cells (HSCs) were characterised by reverse transcriptase-polymerase chain reaction, and the effects of selective delta opioid receptor agonists on cellular proliferation, tissue inhibitor of metalloproteinase 1 (TIMP-1), and procollagen I expression in HSCs determined. RESULTS: Naltrexone markedly attenuated the development of hepatic fibrosis as well as MMP-2 activity (p<0.01), and decreased the number of activated HSCs in BDL rats (p<0.05). The development of biliary cirrhosis altered the redox state with a decreased hepatic GSH/GSSG ratio and increased concentrations of hepatic S-nitrosothiols, which were partially or completely normalised by treatment with naltrexone, respectively. Activated rat HSCs exhibited expression of delta1 receptors, with increased procollagen I expression, and increased TIMP-1 expression in response to delta(1) and delta(2) agonists, respectively. CONCLUSIONS: This is the first study to demonstrate that administration of an opioid antagonist prevents the development of hepatic fibrosis in cirrhosis. Opioids can influence liver fibrogenesis directly via the effect on HSCs and regulation of the redox sensitive mechanisms in the liver.

Low plasma protein nitrotyrosine levels distinguish primary Raynaud's phenomenon from scleroderma.

OBJECTIVE: To investigate the hypothesis that increased formation of reactive nitrogen species may contribute to the vascular pathology that develops in patients with connective tissue disease such as scleroderma. PATIENTS AND METHODS: The level of protein-bound nitrotyrosine in plasma was measured by stable isotope dilution gas chromatography/negative ion chemical ionisation mass spectrometry in 11 patients with primary Raynaud's phenomenon, 37 with scleroderma, 13 with chronic renal impairment, and in 23 healthy controls. RESULTS: Plasma protein-bound nitrotyrosine was markedly decreased in patients with primary Raynaud's phenomenon (mean (SEM) 0.60 (0.06) ng/mg dry protein) compared with patients with scleroderma (1.78 (0.21) ng/mg protein), chronic renal impairment (1.42 (0.17) ng/mg protein) or healthy controls (1.63+/-0.15 ng/mg protein, ANOVA p<0.001). CONCLUSION: These data suggest that there is decreased nitration of plasma proteins, or increased degradation of nitrated proteins from the circulation of patients with primary but not secondary Raynaud's phenomenon.

Inactivation of xanthine oxidoreductase is associated with increased joint swelling and nitrotyrosine formation in acute antigen-induced arthritis.

OBJECTIVE: Arthritis is associated with increased articular formation of nitrotyrosine, which may contribute to injury. Nitrotyrosine is formed by nitration of tyrosine by reactive nitrogen species such as peroxynitrite, the formation of which may be enhanced by xanthine oxidoreductase (XOR), since it can generate nitric oxide from nitrite/nitrate, and superoxide during xanthine metabolism. We hypothesized that inactivation of XOR would protect against antigen-induced arthritis (AIA) and decrease nitrotyrosine formation. METHODS: AIA was induced with methylated bovine serum albumin (mBSA) in three groups of Wistar rats: animals fed on (1) tungsten-enriched chow (0.7 g/kg) (TG), which inactivates XOR, (2) standard chow (SG), and (3) rats treated with allopurinol (50 mg/kg/day; p.o.) (AG). Nitrotyrosine in patella-synovium was quantified by mass spectrometry three weeks after intra-articular (i.a.) antigen injection. RESULTS: Treatment with tungsten, but not allopurinol, suppressed plasma and articular XOR activity at < or = 0.9% of normal levels. XOR inactivation was associated with increased knee swelling 24-48 hrs post i.a. mBSA, compared with controls (mean increase +/- SEM of knee diameter from baseline of 3.3 +/- 0.5, 2.0 +/- 0.3 and 1.9 +/- 0.2 mm in TG, SG and AG (n = 14 each group), respectively; p < 0.05, TG vs SG, ANOVA). Mean ratio of articular nitrotyrosine-tyrosine (+/- SEM) was increased in the XOR-inactivated group, compared with controls: 12.3 +/- 0.7, 9.6 +/- 0.8 and 10.4 +/- 0.5 pg/microg in TG, SG and AG, respectively; p < 0.05, TG vs SG. CONCLUSION: Contrary to expectation, XOR inactivation was associated with increased joint swelling and articular tyrosine nitration in acute AIA, suggesting a novel, protective role for XOR in inflammatory arthritis.

Endothelial transcytosis of myeloperoxidase confers specificity to vascular ECM proteins as targets of tyrosine nitration.

Nitrotyrosine formation is a hallmark of vascular inflammation, with polymorphonuclear neutrophil-derived (PMN-derived) and monocyte-derived myeloperoxidase (MPO) being shown to catalyze this posttranslational protein modification via oxidation of nitrite (NO(2)(-)) to nitrogen dioxide (NO(2)(*)). Herein, we show that MPO concentrates in the subendothelial matrix of vascular tissues by a transcytotic mechanism and serves as a catalyst of ECM protein tyrosine nitration. Purified MPO and MPO released by intraluminal degranulation of activated human PMNs avidly bound to aortic endothelial cell glycosaminoglycans in both cell monolayer and isolated vessel models. Cell-bound MPO rapidly transcytosed intact endothelium and colocalized abluminally with the ECM protein fibronectin. In the presence of the substrates hydrogen peroxide (H(2)O(2)) and NO(2)(-), cell and vessel wall-associated MPO catalyzed nitration of ECM protein tyrosine residues, with fibronectin identified as a major target protein. Both heparin and the low-molecular weight heparin enoxaparin significantly inhibited MPO binding and protein nitrotyrosine (NO(2)Tyr) formation in both cultured endothelial cells and rat aortic tissues. MPO(-/-) mice treated with intraperitoneal zymosan had lower hepatic NO(2)Tyr/tyrosine ratios than did zymosan-treated wild-type mice. These data indicate that MPO significantly contributes to NO(2)Tyr formation in vivo. Moreover, transcytosis of MPO, occurring independently of leukocyte emigration, confers specificity to nitration of vascular matrix proteins.

Pathogenesis and treatment of renal dysfunction in rhabdomyolysis.

Rhabdomyolysis is a major cause of acute renal failure, and recent experimental data have provided a better understanding of the pathophysiology of the renal dysfunction. Renal failure is due to renal vasoconstriction, tubular damage caused by oxidant injury, and possibly tubular obstruction. Recent studies have provided greater insight into the rationale behind current therapy and potential treatment strategies. This review thus aims to summarise current understanding of the causes, pathogenesis and treatment of renal failure caused by rhabdomyolysis.

Analysis of free and protein-bound nitrotyrosine in human plasma by a gas chromatography/mass spectrometry method that avoids nitration artifacts.

Measurement of nitrotyrosine in biological fluids and tissues is increasingly being used to monitor the production of reactive nitrogen species in vivo. The detection of nitrotyrosine in vivo has been reported with the use of a variety of methods including immunoassay, HPLC and GLC/MS. The validity of HPLC and immunoassays have been questioned with regard to their selectivity and sensitivity limits. In principle, the measurement of nitrotyrosine by GLC/MS permits a highly specific, highly sensitive and fully quantitative assay. The nitration of tyrosine under acidic conditions in the presence of nitrite is well documented. Derivatization for the full quantification of nitrotyrosine by using GLC/MS can lead to the artifactual nitration of tyrosine if performed under acidic conditions in the presence of nitrite. We describe a novel alkaline method for the hydrolysis and derivatization of nitrotyrosine and tyrosine, and demonstrate its applicability to the measurement of plasma concentrations of both free and protein-bound nitrotyrosine and tyrosine. A detection limit of 1 pg for nitrotyrosine and 100 pg for tyrosine has been achieved. Our method allows, for the first time, the analysis of free and protein-bound nitrotyrosine and tyrosine in biological samples. The plasma concentrations (means+/-S.E.M.) of free tyrosine and nitrotyrosine in eight normal subjects were 12+/-0.6 microg/ml and 14+/-0.7 ng/ml respectively. Plasma proteins contained tyrosine and nitrotyrosine at 60.7+/-1.7 microg/mg and 2.7+/-0.4 ng/mg respectively.

A causative role for redox cycling of myoglobin and its inhibition by alkalinization in the pathogenesis and treatment of rhabdomyolysis-induced renal failure.

Muscle injury (rhabdomyolysis) and subsequent deposition of myoglobin in the kidney causes renal vasoconstriction and renal failure. We tested the hypothesis that myoglobin induces oxidant injury to the kidney and the formation of F2-isoprostanes, potent renal vasoconstrictors formed during lipid peroxidation. In low density lipoprotein (LDL), myoglobin induced a 30-fold increase in the formation of F2-isoprostanes by a mechanism involving redox cycling between ferric and ferryl forms of myoglobin. In an animal model of rhabdomyolysis, urinary excretion of F2-isoprostanes increased by 7.3-fold compared with controls. Administration of alkali, a treatment for rhabdomyolysis, improved renal function and significantly reduced the urinary excretion of F2-isoprostanes by approximately 80%. EPR and UV spectroscopy demonstrated that myoglobin was deposited in the kidneys as the redox competent ferric myoglobin and that it's concentration was not decreased by alkalinization. Kinetic studies demonstrated that the reactivity of ferryl myoglobin, which is responsible for inducing lipid peroxidation, is markedly attenuated at alkaline pH. This was further supported by demonstrating that myoglobin-induced oxidation of LDL was inhibited at alkaline pH. These data strongly support a causative role for oxidative injury in the renal failure of rhabdomyolysis and suggest that the protective effect of alkalinization may be attributed to inhibition of myoglobin-induced lipid peroxidation.

Early closure of a randomized trial: surgery and postoperative radiotherapy versus radiotherapy in the management of intra-oral tumours.

Tumours of the oral cavity/oropharynx occur relatively infrequently in the UK. The management of such lesions, especially the squamous cell carcinomas, is still a little controversial. Some centres advocate radiotherapy while others adopt surgery and radiotherapy. In an attempt to resolve the question of which approach gives the better results, a multicentre randomized trial was established to compare surgery plus postoperative radiotherapy with radical radiotherapy alone. It was anticipated that 350 patients would be required to give a statistically significant result, but, after 35 patients had been entered, the trial was closed prematurely with a marked difference in overall survival in favour of the combination arm (P = 0.0006). At this analysis, carried out 23 months after trial closure, the survival difference between the two arms remains statistically significant for all causes of mortality (P = 0.001; relative death rate = 0.24; 95% CI 0.10-0.59).

Effect of the isoprostanes, 8-iso prostaglandin E2 and 8-iso prostaglandin F2 alpha on the rabbit lung in vivo.

8-Iso-prostaglandin (PG)E2 and 8-iso-PGF2 alpha are members of the isoprostane class of prostanoids which are formed by free radical mediated oxidation of arachidonic acid. Both E2- and F2-isoprostanes are potent vasoconstrictors and are believed to act through the prostanoid TP-receptors or a closely related receptor. In lightly anaesthetised, spontaneously breathing rabbits, aerosolised administration of histamine (1.25-40 mg ml-1, n = 8) caused a modest dose-dependent increase in total lung resistance (RL) and a concomitant fall in dynamic lung compliance (CL dyn). Aerosolised methacholine (0.625-20 mg ml-1, n = 6) caused considerable bronchoconstriction, with a dose-dependent increase in RL, and a corresponding fall in CL dyn. In contrast, intratracheal administration of either 8-iso PGE2 or 8-iso-PGF2 alpha (1 ng ml(-1)-100 micrograms ml-1, n = 8) had no significant effect on lung function. The TP-receptor agonist, U-46619, was similarly inactive in this model when given by aerosol. Intravenous administration of histamine or 8-iso PGF2 omega had no significant effect on the lung indices, RL and CL dyn, or on the pulmonary and systemic vasculature (n = 4 per drug group). 8-Iso-PGE2 caused a concentration-dependent decrease in the right ventricular systolic pressure from 3 nmol kg-1 to 100 nmol kg-1 (n = 43, p < 0.05), but showed no other activity. In contrast, U-46619 given intravenously caused an increase in transpulmonary pressure (n = 4, p < 0.05), but had no effect on airflow. At higher doses, it did cause a significant drop in both systemic and right ventricular systolic pressures (n = 4, p < 0.05), which were probably due to an interaction with platelets. The isoprostanes had no effect on the rabbit airway up to a concentration of 3 microM. In contrast, 3 microM U-46619 caused a modest contraction of tracheal smooth muscle, whilst 3 microM methacholine was at least five-fold more potent in contracting the same tissues. We conclude that the aerosolised isoprostanes are not broncho-constricting agents in the rabbit in vivo.

Identification of the major urinary metabolite of the F2-isoprostane 8-iso-prostaglandin F2alpha in humans.

F2-isoprostanes are prostaglandin-like products of nonenzymatic lipid peroxidation. Measurement of levels of endogenous unmetabolized F2-isoprostanes has proven to be a valuable approach to assess oxidative stress in vivo. However, measurement of levels of urinary metabolites of F2-isoprostanes in timed urine collections offers an advantage over measuring unmetabolized F2-isoprostanes, e. g. in a plasma sample, in that it can provide an integrated index of isoprostane production over time. Therefore, we sought to identify the major urinary metabolite in humans of one of the more abundant F2-isoprostanes produced, 8-iso-prostaglandin F2alpha (8-iso-PGF2alpha). 20 microCi of tritiated 8-iso-PGF2alpha was infused over 1 h into a male volunteer. 75% of the infused radioactivity was excreted into the urine during the following 4.5 h and was combined with urine collected for 4 h from a rhesus monkey following infusion of 500 microg of unlabeled 8-iso-PGF2alpha. Urinary metabolites were isolated and purified by adsorption chromatography and high pressure liquid chromatography. The major urinary metabolite, representing 29% of the total extractable recovered radioactivity in the urine, was structurally identified by gas chromatography and mass spectrometry as 2,3-dinor-5, 6-dihydro-8-iso-prostaglandin F2alpha. The identification of 2, 3-dinor-5,6-dihydro-prostaglandin F2alpha as the major urinary metabolite of 8-iso-prostaglandin F2alpha provides the basis for the development of methods of assay for its quantification as a means to obtain an integrated assessment of oxidative stress status in humans.

Vasoconstrictor responsiveness of the rat mesenteric arterial bed in cirrhosis.

1. The effects of cirrhosis on mesenteric vascular reactivity were assessed in constantly perfused mesenteric arterial beds isolated from cirrhotic rats (carbon tetrachloride with phenobarbitone, n = 6), and from phenobarbitone-treated and untreated age-matched controls (n = 4,5). 2. At a constant flow rate of 5 ml min-1 there was no difference in basal perfusion pressure between the groups. Electrical field stimulation (EFS; 4-32 Hz, 90V, 1 ms, 30 s) of perivascular nerves caused frequency-dependent increases in perfusion pressure which were not different between the groups. Dose-dependent vasoconstrictor responses to exogenous noradrenaline (NA), methoxamine (an alpha 1-adrenoceptor agonist), adenosine 5'-triphosphate (ATP) and vasopressin were also similar between the groups. 3. The nitric oxide (NO) synthesis inhibitor NG-nitro-L-arginine methyl ester (L-NAME; 30 microM) augmented constrictor responses to NA, EFS, methoxamine and vasopressin in all groups, and as shown for EFS and NA, this was reversed by L-arginine (300 microM). However, the maximum constrictor responses of cirrhotic preparations in the presence of L-NAME were significantly lower than those of both groups of control animals at the highest frequency of EFS (32 Hz) and highest doses of NA (0.15 and 0.5 mumol) and, compared to phenobarbitone-treated controls, methoxamine (5 mumol). Responses to ATP were significantly augmented by L-NAME only in the cirrhotic group. 4. A step-wise increase in perfusate flow to 10, 15 and 20 ml min-1 produced a broadly similar increase in perfusion pressure within each group. At increased flow rates, cirrhotic preparations were hyporesponsive to NA (15 nmol) compared to the phenobarbitone-treated animals but not the untreated controls. Glibenclamide (5 microM) or L-NAME (30 microM) had no significant effect on the relationship between flow and perfusion pressure or on responses to NA at the different flow rates. 5. We conclude that sympathetic neurotransmission is unchanged in cirrhosis. Endogenous NO is important in modulation of constriction in both normal and cirrhotic states. Changes in NO may occur in cirrhosis, although the role of this in hyporesponsiveness of cirrhotic preparations to NA at higher flow rates and to the greater potentiation of ATP-mediated constriction in the presence of L-NAME, together with the impact of factors such as changes in calcium and potassium channels, is not entirely clear.

Pain relief with low-dose intravenous clonidine in a child with severe burns.

The case of an 11-year-old boy who suffered second and third degree burns to 78% of his body is reported. The large doses of morphine used as analgesia resulted in severe side effects: ventilatory dependence, impairment of gastrointestinal function and psychological disturbance. Intravenous lignocaine was added without benefit. The addition of low-dose intravenous clonidine, however, precipitated a dramatic reduction in morphine consumption with an attendant improvement in ventilatory, gastrointestinal and psychological functions.

Mesenteric vasodilator responses in cirrhotic rats: a role for nitric oxide?

The contribution of nitric oxide to mesenteric arterial vasodilator responses was investigated in the isolated perfused mesenteric arterial bed of cirrhotic rats (carbon tetrachloride/phenobarbitone; n = 6). Age-matched (n = 9) and phenobarbitone-treated rats (n = 9) served as controls. Responses to the endothelium-dependent dilators acetylcholine and adenosine 5'-triphosphate (ATP) and the smooth muscle dilator (NO donor) sodium nitroprusside were investigated after tone was raised by continuous infusion of methoxamine, before and during infusion of the NO synthesis inhibitor NG-nitro-L-arginine methyl ester (L-NAME; 30 mumol/L) +/- L-arginine (1 mmol/L). A significant hyporesponsiveness to methoxamine infusion in cirrhotic preparations (P < .05) was not fully corrected by L-NAME. There was no difference in the percentage vasodilator response to acetylcholine in the cirrhotic group compared with controls; L-NAME significantly and reversibly inhibited the dilator response in all groups. ATP elicited dose-dependent vasodilation that, in the absence of L-NAME, did not differ between the groups. By contrast, in the presence of L-NAME, ATP (5 x 10(-8) mol) produced pronounced, reversible vasoconstriction only in cirrhotic animals (P < .02). Vasodilatation attributable to sodium nitroprusside (5 x 10(-8) mol) was significantly attenuated in cirrhotic rats. The methoxamine data support the concept of mesenteric hyposensitivity to vasoconstrictor agents in cirrhosis that may be at least partly NO mediated. Increased NO activity in smooth muscle leading to decreased guanylate cyclase availability may account for the diminished vasodilator responses to sodium nitroprusside in cirrhotic preparations. The unchanged responsiveness to vasodilatation by acetylcholine (ACh) and the vasoconstriction to ATP observed during NO blockade in cirrhotic animals indicate that mesenteric endothelial NO is unchanged or possibly diminished.