Jigme M. Sethi

Exhaled nitric oxide in the diagnosis and management of asthma: Clinical implications

Exhaled nitric oxide (eNO) used as an aid to the diagnosis and management of lung disease is receiving attention from pulmonary researchers and clinicians alike because it offers a noninvasive means to directly monitor airway inflammation. Research evidence suggests that eNO levels significantly increase in individuals with asthma before diagnosis, decrease with inhaled corticosteroid administration, and correlate with the number of eosinophils in induced sputum. These observations have been used to support an association between eNO levels and airway inflammation. This review presents an update on current opportunities regarding use of eNO in patient care, and more specifically on its potential usage for asthma diagnosis and monitoring. The review will also discuss factors that may complicate use of eNO as a diagnostic tool, including changes in disease severity, symptom response, and technical measurement issues. Regardless of the rapid, convenient, and noninvasive nature of this test, additional well-designed, long-term longitudinal studies are necessary to fully evaluate the clinical utility of eNO in asthma management.Introduction Although socioeconomic differences in prevalence of obesity are well documented, whether patterns of weight gain during key periods of growth and development differ among youth from different socioeconomic backgrounds is unknown. This study examines socioeconomic disparities in overweight status and 5-year weight gain among adolescents. Methods Project EAT (Eating Among Teens)-II followed a socioeconomically and ethnically diverse sample of 2,516 adolescents from 1999 through 2004. Mixed-model regression analyses examined longitudinal trends in overweight status as a function of socioeconomic status (SES). Results Girls and boys in the low-SES category were more likely to be overweight than were those in the high-SES category. Boys in the high-SES category showed a significant decrease (P = .006) in overweight prevalence between 1999 and 2004, whereas boys in the low- and middle-SES categories showed no significant change. Girls in the low-SES category showed a significant 5-year increase (P = .004) in overweight prevalence compared with a stable prevalence of overweight among girls in the middle- and high-SES categories. Conclusion Our data show continued and, in some cases, increasing socioeconomic disparities in risk for overweight. Youth from low-SES backgrounds are at increased risk for overweight and are more likely to remain overweight (boys) or become overweight (girls). Designing obesity prevention and treatment interventions that reach and address the unique needs of youth and families from less-advantaged socioeconomic backgrounds is a public health priority.

Carbon monoxide-dependent signaling.

It has become accepted that nitric oxide serves important functions in biological systems as a second messenger. Another diatomic gaseous molecule, carbon monoxide (CO), is also rapidly gaining acceptance as a signaling agent. Some of the activities of CO are analogous to those of nitric oxide in the vascular system and the brain, but CO also behaves in novel ways. Like nitric oxide, CO is capable of activating soluble guanylyl cyclase. This mechanism of CO signaling is important in vasodilation and neurotransmission. There is growing evidence, however, that CO also acts independently of soluble guanylyl cyclase. CO has been shown to protect against septic shock and lung injury in animal models, and the mitogen-activated protein kinase system appears to mediate this cytoprotective effect. Although much remains to be elucidated about the mechanisms of cell signaling by CO, the pace of discovery in this field is making the picture clearer with every passing day.

Discovery of the gene signature for acute lung injury in patients with sepsis

The acute respiratory distress syndrome (ARDS)/acute lung injury (ALI) was described 30 yr ago, yet making a definitive diagnosis remains difficult. The identification of biomarkers obtained from peripheral blood could provide additional noninvasive means for diagnosis. To identify gene expression profiles that may be used to classify patients with ALI, 13 patients with ALI + sepsis and 20 patients with sepsis alone were recruited from the Medical Intensive Care Unit of the University of Pittsburgh Medical Center, and microarrays were performed on peripheral blood samples. Several classification algorithms were used to develop a gene signature for ALI from gene expression profiles. This signature was validated in an independently obtained set of patients with ALI + sepsis (n = 8) and sepsis alone (n = 1). An eight-gene expression profile was found to be associated with ALI. Internal validation found that the gene signature was able to distinguish patients with ALI + sepsis from patients with sepsis alone with 100% accuracy, corresponding to a sensitivity of 100%, a specificity of 100%, a positive predictive value of 100%, and a negative predictive value of 100%. In the independently obtained external validation set, the gene signature was able to distinguish patients with ALI + sepsis from patients with sepsis alone with 88.9% accuracy. The use of classification models to develop a gene signature from gene expression profiles provides a novel and accurate approach for classifying patients with ALI.

Differential Modulation by Exogenous Carbon Monoxide of TNF-α Stimulated Mitogen-Activated Protein Kinases in Rat Pulmonary Artery Endothelial Cells

Heme oxygenase-1 (HO-1) is an enzyme that is highly inducible by various cellular stressors, especially oxidant injury. Our laboratory and others have demonstrated that induction of HO-1 exerts an antiinflammatory effect both in vitro and in vivo. We hypothesized that carbon monoxide (CO), a major catalytic byproduct of heme catalysis by HO-1, may mediate this antiinflammatory effect by modulating signal transduction pathways, in particular the mitogen-activated protein (MAP) kinase pathway. Confluent primary cultures of rat pulmonary artery endothelial cells (RPAEC) were treated with tumor necrosis factor-alpha (TNF-alpha; 50 ng/ml), and whole-cell extracts were assayed for phosphorylated ERK1/2, JNK1/2, and p38 MAP kinases. These three major MAP kinase pathways were activated by TNF-alpha in a time-dependent manner. RPAEC treated with TNF-alpha in the presence of a low concentration of CO (1%) exhibited marked attenuation of the phosphorylation of ERK1/2 MAP kinase when compared with cells treated with TNF-alpha alone. A similar effect was seen on the upstream MEK 1/2 kinase. Interestingly, CO (1%) accentuated TNF-alpha-induced phosphorylated p38 MAP kinase. These effects of exogenous CO on the ERK1/2 and p38 systems could be replicated by overexpression of HO-1 in RPAEC, using an adenoviral vector. As these MAP kinases are implicated in the regulation of various inflammatory molecules and adhesion molecules, our data provide a potential mechanism by which HO-1, acting via CO, may modulate the inflammatory response by differential activation of the MAP kinase pathway.

Markers of lung disease in exhaled breath: nitric oxide.

Management of airway inflammation requires proper monitoring and treatment to improve long-term outcomes. However, achieving this goal is difficult, as current methods have limitations. Although nitric oxide (NO) was first identified 200 years ago, its physiological importance was not recognized until the early 1980s. Many studies have established the role of NO as an essential messenger molecule in body systems. In addition, studies have demonstrated a significant relationship between changes in exhaled NO levels and other markers of airway inflammation. The technique used to measure NO in exhaled breath is noninvasive, reproducible, sensitive, and easy to perform. Consequently, there is growing interest in the use of exhaled NO in the management of asthma and other pulmonary conditions. The purpose of this review is to promote a basic understanding of the physiologic actions of NO, measurement techniques, and ways that research findings might translate to future application in clinical practice. Specifically, the article will review the role of exhaled NO in regard to its historical background, mechanisms of action, measurement techniques, and implications for clinical practice and research.

Non-invasive measurements of exhaled NO and CO associated with methacholine responses in mice

BackgroundNitric oxide (NO) and carbon monoxide (CO) in exhaled breath are considered obtainable biomarkers of physiologic mechanisms. Therefore, obtaining their measures simply, non-invasively, and repeatedly, is of interest, and was the purpose of the current study.MethodsExpired NO (ENO) and CO (ECO) were measured non-invasively using a gas micro-analyzer on several strains of mice (C57Bl6, IL-10-/-, A/J, MKK3-/-, JNK1-/-, NOS-2-/- and NOS-3-/-) with and without allergic airway inflammation (AI) induced by ovalbumin systemic sensitization and aerosol challenge, compared using independent-sample t-tests between groups, and repeated measures analysis of variance (ANOVA) within groups over time of inflammation induction. ENO and ECO were also measured in C57Bl6 and IL-10-/- mice, ages 8–58 weeks old, the relationship of which was determined by regression analysis. S-methionyl-L-thiocitrulline (SMTC), and tin protoporphyrin (SnPP) were used to inhibit neuronal/constitutive NOS-1 and heme-oxygenase, respectively, and alter NO and CO production, respectively, as assessed by paired t-tests. Methacholine-associated airway responses (AR) were measured by the enhanced pause method, with comparisons by repeated measures ANOVA and post-hoc testing.ResultsENO was significantly elevated in naïve IL-10-/- (9–14 ppb) and NOS-2-/- (16 ppb) mice as compared to others (average: 5–8 ppb), whereas ECO was significantly higher in naïve A/J, NOS-3-/- (3–4 ppm), and MKK3-/- (4–5 ppm) mice, as compared to others (average: 2.5 ppm). As compared to C57Bl6 mice, AR of IL-10-/-, JNK1-/-, NOS-2-/-, and NOS-3-/- mice were decreased, whereas they were greater for A/J and MKK3-/- mice. SMTC significantly decreased ENO by ~30%, but did not change AR in NOS-2-/- mice. SnPP reduced ECO in C57Bl6 and IL-10-/- mice, and increased AR in NOS-2-/- mice. ENO decreased as a function of age in IL-10-/- mice, remaining unchanged in C57Bl6 mice.ConclusionThese results are consistent with the ideas that: 1) ENO is associated with mouse strain and knockout differences in NO production and AR, 2) alterations of ENO and ECO can be measured non-invasively with induction of allergic AI or inhibition of key gas-producing enzymes, and 3) alterations in AR may be dependent on the relative balance of NO and CO in the airway.

Alterations in Nitric Oxide and Cytokine Production with Airway Inflammation in the Absence of IL-10

IL-10 is an anti-inflammatory cytokine that suppresses NO synthase (NOS) and production of NO; its lack may promote NO production and alterations in cytokines modulated by NO with allergic airway inflammation (AI), such as IL-18 and IL-4. Therefore, we induced AI in IL-10 knockout (−/−) and IL-10-sufficient C57BL/6 (C57) mice with inhaled OVA and measured airway NO production, as exhaled NO (ENO) and bronchoalveolar lavage fluid nitrite levels. ENO and nitrite levels were elevated significantly in naive IL-10−/− mice as compared with C57 mice. With AI, ENO and nitrite levels increased in C57 mice and decreased in IL-10−/− mice. IL-18 production fell with both AI and addition of S-nitroso-N-acetyl-d,l-penicillamine (a NO donor) but was not significantly increased by chemical NOS inhibition by l-N5-(1-iminoethyl)-ornithine. IL-4 AI was increased significantly (up to 10-fold greater) in the absence of IL-10 but was reduced significantly with chemical inhibition of NOS. Airway responsiveness was lower in IL-10−/− mice and was associated with alteration in production of NO and IL-4. Thus, IL-4 production was increased, and likely decreased NO production, in a way not predicted by the absence of IL-10. Inhibition of IL-4 production, with inhibition of NOS in the absence of IL-10, demonstrated the importance of a NO and IL-4 feedback mechanism regulating this interaction.

Exhaled carbon monoxide as a biomarker of inflammatory lung disease

Carbon monoxide (CO) can be detected on the exhaled breath of humans. Exhaled CO (E-CO) originates from the inspiration of ambient CO and from endogenous metabolic sources that include heme metabolism catalyzed by heme oxygenase (HO) enzymes. HO occurs in both constitutive (HO-2) and inducible (HO-1) forms; the latter responds to pro-inflammatory or pro-oxidative stimuli. E-CO may arise in the airways from inducible HO-1 activity in the bronchial epithelium, alveolar macrophages and other lung cell types, as a consequence of local inflammation, and from the alveolae in equilibrium with carboxyhemoglobin (Hb-CO) in the pulmonary circulation. Elevations in Hb-CO in turn may reflect increases in ambient CO, as well as increased HO activity in systemic tissues. E-CO increases dramatically in active smokers and can be used to monitor the smoking habit. Elevations in E-CO have been observed in critically ill or post-surgical patients and those with various pulmonary diseases associated with inflammation, including chronic obstructive pulmonary disease (COPD), asthma, cystic fibrosis and infections. Despite improvements in the standardization and sensitivity of methods to detect E-CO, the predictive value of this measurement as a diagnostic tool remains unclear.

Heme Oxygenase-1 in Acute Lung Injury

Heme oxygenase (HO) was first described, in 1968, as the enzyme responsible for the rate-limiting step in the catalytic breakdown of the heme moiety of hemoglobin, and understandably, initial work focused on the regulation and function of this enzyme in heme metabolism (1). That HO could be important for much more than the color changes of a bruise as heme was broken down through biliverdin to bilirubin, was not suspected until the work of Tyrrell and his colleagues. These researchers showed that HO-1 could be vigorously induced not just by heme, its natural substrate, but by a variety of agents that had as their common feature the ability to generate reactive oxidant species (ROS) (2). Examples of the broad spectrum of agents capable of inducing HO include lipopolysaccharide (LPS), phorbol esters, sodium arsenite, hydrogen peroxide, ultraviolet radiation, hyperthermia, hyperoxia, sulfhydryl reagents, heat shock, and heavy metals. This extraordinary variety of inducers is surprising for an enzyme subserving a seemingly housekeeping function of heme turnover, and speculation arose that the enzyme was also vital to cellular homeostasis.