Determinants of nitric oxide in exhaled gas in the isolated rabbit lung

Nitric oxide concentrations in the exhaled gas (NOe) increases during various inflammatory conditions in humans and animals. Little is known about the sources and factors that influence NOe. NOe at end expiration was measured by chemiluminescence in an isolated, blood-perfused rabbit lung. The average end-expiratory concentration over 10 breaths was used. The effect of positive end-expiratory pressure (PEEP), flow rate, pH, hypoxia, venous pressure, and flow pulsatility on NOe were determined. At constant blood flow, increasing PEEP from 1 to 5 cm H2O elicited a reproducible increase in NOe from 49 +/- 7 to 53 +/- 8 parts per billion (ppb) (p < 0.05). When blood pH was increased from 7.40 to 7.74 by breathing low CO2 gas, NOe rose from 45 +/- 7 to 55 +/- 7 ppb (p < 0.001). Hypoxia caused a dose-dependent decrease in NOe from 37 +/- 3 during baseline to 23 +/- 2 during ventilation with 0% O2 (p < 0.01). Venous pressure elevation from 0 to 5 and 10 mm Hg decreased NOe from 32 +/- 5, to 26 +/- 5 and 24 +/- 5 ppb, respectively (p < 0.05). Switching from steady to pulsatile flow (same man flow) resulted in a small, albeit significant reduction in NOe; 30 +/- 4 to 28 +/- 4 ppb (p < 0.05). Changes in flow rate between 200 and 20 ml/min were associated with small changes in NOe; however, when flow was stopped, NOe rose substantially to 56 +/- 6 ppb (p < 0.05). The changes in NOe were rapid (1 to 2 min) and reversible. The results suggest that NOe is influenced by ventilatory and hemodynamic variables, pH, and hypoxia. We suggest that caution must be taken when interpreting changes in exhaled NO in humans or experimental animals. Changes in total and regional blood flow, capillary blood volume, ventilation, hypoxia, and pH should not be overlooked.     

Proteasome inhibition attenuates nitric oxide synthase expression, VCAM-1 transcription and the development of chronic colitis

The objectives of this study were to (1) assess the role of the 26S proteasome complex in regulating the expression of the inducible isoform of nitric oxide synthase (iNOS) and vascular cell adhesion molecule-1 (VCAM-1) in a model of chronic granulomatous colitis in vivo and (2) determine the role of the proteasome in regulating the inflammatory response observed in this model of chronic gut inflammation. The selective proteasome inhibitor MG-341 (0.3 mg/kg) was administered by gavage beginning immediately before the induction of colitis and continuing daily thereafter for the entire 14-day experimental period. We found that chronic proteasome inhibition using MG-341 significantly attenuated the peptidoglycan/polysaccharide (PG/PS)-induced up-regulation of iNOS in the colon and spleen and the consequent increase in plasma levels of nitrate and nitrite. Furthermore, we found that the proteasome inhibitor suppressed the up-regulation of the adhesion molecule VCAM-1 in the colon. We also found that MG-341 attenuated PG/PS-induced increases in macroscopic colonic inflammation, bowel wall thickness, colonic dry weight and colonic MPO activity. Treatment with MG-341 also significantly reduced PG/PS-induced increases in macroscopic spleen inflammation, spleen weight and spleen MPO activity. We conclude that the 26S proteasome complex plays an important role in regulating the PG/PS-induced up-regulation of iNOS and VCAM-1 in vivo and appears to be important in regulating colonic and splenic inflammation.     

Thrombolysis plus aortic counterpulsation: improved survival in patients who present to community hospitals with cardiogenic shock

OBJECTIVES: We sought to explore the potential benefit of combining intraaortic balloon counterpulsation (IABP) with thrombolysis for acute myocardial infarction (MI) complicated by cardiogenic shock. BACKGROUND: In community hospitals, this condition is usually managed with thrombolysis alone. METHODS: We reviewed the charts of 335 patients from two community hospitals who presented with acute MI and had cardiogenic shock between 1985 and 1995. RESULTS: Of 46 patients who underwent thrombolysis within 12 h of acute infarction with confirmed cardiogenic shock, 27 underwent IABP and 19 did not. Age, systolic blood pressure with shock, pulmonary artery catheter use, pulmonary capillary wedge pressure and the incidence of diabetes mellitus and anterior MI did not differ between groups. Patients treated with IABP were somewhat more likely to have prior MI and had a significantly greater cardiac index (2.0 vs. 1.5 liters/min per m2, p = 0.04). Although no deaths occurred within 2 h of presentation, patients not treated with IABP tended to die earlier (6.8 +/- 5 vs. 23.8 +/- 19 h, p = 0.13). Patients treated with IABP had a significantly higher rate of community hospital survival (93% vs. 37%, p = 0.0002), and more of them were transferred for revascularization (85% vs. 37%). Of 30 patients transferred for revascularization, 27 underwent angioplasty or bypass surgery; hospital survival was 74%. Patients treated with IABP also had a significantly higher overall hospital and 1-year survival rate (67% vs. 32%, p = 0.019). CONCLUSIONS: Survival may be enhanced and transfer for revascularization facilitated when community hospitals use both thrombolysis and IABP to treat patients with acute MI complicated by cardiogenic shock.     

The critical role of tissue angiotensin-converting enzyme as revealed by gene targeting in mice

Angiotensin-converting enzyme (ACE) generates the vasoconstrictor angiotensin II, which plays a critical role in maintenance of blood pressure in mammals. Although significant ACE activity is found in plasma, the majority of the enzyme is bound to tissues such as the vascular endothelium. We used targeted homologous recombination to create mice expressing a form of ACE that lacks the COOH-terminal half of the molecule. This modified ACE protein is catalytically active but entirely secreted from cells. Mice that express only this modified ACE have significant plasma ACE activity but no tissue-bound enzyme. These animals have low blood pressure, renal vascular thickening, and a urine concentrating defect. The phenotype is very similar to that of completely ACE-deficient mice previously reported, except that the renal pathology is less severe. These studies strongly support the concept that the tissue-bound ACE is essential to the control of blood pressure and the structure and function of the kidney.     

Increasing the immunogenicity of a trivalent influenza virus vaccine with adjuvant-active nonionic block copolymers for potential use in the elderly

High molecular weight nonionic block copolymers consisting of a large hydrophobic core made from repeat oxypropylene units and smaller hydrophilic blocks of oxyethylene repeat units were evaluated as adjuvants in experimental influenza virus vaccine formulations. The goal was to identify a copolymer that would increase the immunogenicity of the commercial Fluogen trivalent influenza virus vaccine. Vaccine experiments done using BALB/c mice provided data that allowed us to identify a copolymer that increased both antibody titers specific for total virus proteins as well as antibodies with hemagglutination inhibition (HAI) activity. This copolymer, termed CRL1005, increased the production of IgG1, IgG2a and IgG2b which suggested it increased the activity of both Type-1 and Type-2 T-helper lymphocytes. The CRL1005 copolymer was tested further in rhesus monkeys with similar results. Levels of antibodies specific for total virus protein preparations were increased as were HAI antibody titers following vaccination with the copolymer-supplemented Fluogen vaccine. Thus, the CRL1005 copolymer adjuvant appears to be compatible for use with the current generation of inactivated viron-based influenza vaccines and useful for increasing the immunogenicity. A more potent influenza virus vaccine could well be more efficacious in the aged segment of our population than current vaccines.     

Identification of darlin, a Dictyostelium protein with Armadillo-like repeats that binds to small GTPases and is important for the proper aggregation of developing cells

We purified from Dictyostelium lysates an 88-kDa protein that bound to a subset of small GTPases, including racE, racC, cdc42Hs, and TC4ran, but did not bind to R-ras or rabB. Cloning of the gene encoding this 88-kDa protein revealed that it contained multiple armadillo-like repeats most closely related to the mammalian GTP exchange factor smgGDS. We named this protein darlin (Dictyostelium armadillo-like protein). Disruption of the gene encoding darlin demonstrated that this protein is not essential for cytokinesis, pinocytosis, phagocytosis, or development. However, the ability of darlin null cells to aggregate in response to starvation is severely affected. When starved under liquid medium, the mutant cells were unable to form aggregation centers and streams, possibly because of a defect in cAMP relay signaling. This defect was not due to an inability of the darlin mutants to activate adenylate cyclase in response to G protein stimulation. These results suggest that the darlin protein is involved in a signaling pathway that may modulate the chemotactic response during early development.     

Comparison of three different experimental methods for the assessment of peripheral compartment pharmacokinetics in humans

In many cases the concentration reached in a peripheral effect compartment rather than in plasma determines the clinical outcome of therapy. Therefore, several experimental approaches have been developed for direct assessment of drug kinetics in peripheral compartments. Particularly saliva sampling, skin blister fluid sampling, and in vivo microdialysis are frequently employed for measuring peripheral drug concentrations. However, data derived from these techniques have never been directly compared. In the present study, the tissue kinetics of theophylline were measured following single dose administration simultaneously in cantharides induced skin blisters, saliva and microdialysates of subcutaneous- and skeletal muscle- tissue and compared to plasma concentrations. Theophylline was administered to 9 healthy volunteers as an i.v. infusion of 240 mg. Mean ratio (AUCsaliva/AUCplasma) was 0.63 +/- 0.05, mean ratio (AUCblister/AUCplasma) was 0.69 +/- 0.12, mean ratio (AUCmuscle/AUCplasma) was 0.41 +/- 0.10, mean ratio (AUCsubcutaneous/AUCplasma) was 0.34 +/- 0.07. The time course of the concentration(peripheral)/concentration(plasma)-ratios showed that tissue concentrations obtained by microdialysis were closely correlated to free plasma levels, whereas saliva- and cantharides blister data overestimated the corresponding free plasma concentrations. It is concluded that microdialysis represents a reliable technique for the measurement of unbound peripheral compartment concentrations and is superior to saliva- and skin blister concentration measurements.     

Monoaminergic effects of high-dose corticotropin in corticotropin-responsive pediatric opsoclonus-myoclonus

Children with the opsoclonus-myoclonus syndrome (OMS) usually respond to corticotropin (adrenocorticotrophic hormone, ACTH) treatment but the mechanism of benefit is unknown. We previously showed that both cerebrospinal fluid (CSF) homovanillic acid (HVA) and 5-hydroxyindole-acetic acid (5-HIAA) concentrations are low in pediatric OMS. In this study, we measured levels of CSF Dopa, catecholamines, deaminated metabolites of catecholamines, as well as HVA and 5-HIAA in eight patients before and during treatment with ACTH. All the children were ACTH-responsive with 50-70% improvement in multiple clinical features of OMS. ACTH treatment reduced the HVA concentration in every child by a mean of 21% (p < 0.001). Treatment with ACTH was associated with significant correlations between dopaminergic markers such as HVA, dihydroxyphenylacetic acid (DOPAC), and Dopa. There were no significant changes in the CSF concentrations of the noradrenergic markers norepinephrine (NE) and dihydroxyphenylglycol (DHPG), or the serotonergic marker 5-HIAA. The only child with a marked inflammatory pattern in CSF, which was reversed by ACTH, was atypical for a large increase in NE and decrease in 5-HIAA during ACTH treatment. Beneficial effects of ACTH in OMS are not associated with normalization of HVA or 5-HIAA levels. The pattern of decreased HVA and unchanged DOPAC levels could reflect decreased extraneuronal uptake of catecholamines (which steroids inhibit) or decreased 0-methylation of catecholamines in nonneuronal cells.