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Added Value with Extended NO Analysis
PART
Nitric Oxide, NO, and Carbon Monoxide, CO
D
CHAPTER
Added Value with Extended NO Analysis
11
Marieann Högman*,† and Pekka Meriläinen* * Department
of Medical Sciences, Respiratory Medicine and Allergology, Uppsala University, SE 75105 Uppsala, Sweden † Centre for Research and Development, Uppsala University/County Council of Gävleborg, SE 801 88 Gävle, Sweden
11.1 BACKGROUND NO is involved in many processes in the respiratory system in health1,2 and can be altered during disease.3 Individuals with atopic asthma have higher than normal NO concentrations4 and this is due to increased expression of the inducible NO synthase (iNOS) and availability of the substrate L-arginine in airway epithelial cells of asthmatics.5 There is an abundant expression in the airways of iNOS6 due to continuous transcriptional activation.5 Asthma is an obstructive inflammatory disease that effects the airways and NO has been said to be an inflammatory marker in asthma. Lately it has been put forward that asthma can also be a systemic disease.7 Since the disease is characterized by recurrent attacks and varies in frequency and severity from person to person and is due to inflammation, the need for monitoring is obvious. However, it was recently concluded in a Cochrane review that, at present, defining the dose of inhaled corticosteroids based on exhaled nitric oxide cannot be routinely advocated.8 In severe asthma the peripheral airways are also affected and treatment given by inhalation may not reach these airways. Therefore, it is desirable to have insight into how widespread the inflammation is. One way of doing this is to model NO in the respiratory system and thereby get NO values from the alveolar region as well as from the airways. This has been done by Lehtimäki et al. and they have found in more severe asthma and in nocturnal asthma that NO from the alveolar region is increased.9,10 The discovery of the flow dependency of exhaled NO was made with the new generation of fast responding NO analyzers and led to the opportunity to model NO production. This discovery shed light on the different results published in healthy controls as in disease and was done by two research groups independently.11,12 A European task force was formed in 1997 and the latest published guidelines is an ERS and ATS joint statement which was to set the flow rate at 50 mL/s (FE NO0.05 ), but other flow rates could be used but must be stated.13 As can be seen in Figure 11.1, middle panel; higher flow rates results in lower NO values. FE NO0.05 was chosen because it gives a good reading of the analyzer and it is easy for the patient Volatile Biomarkers. http://dx.doi.org/10.1016/B978-0-44-462613-4.00011-8 © 2013 Elsevier B.V. All rights reserved.
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0
2
4
6
8
10 12 14 16
Time seconds
-1
40 35 30 25 20 15 10 5 0
NO output nL·min
FE NO ppb
90
FE NO ppb
196
0
0.1
0.2
0.3
0.4 -1
Expiratory flow rate L.s
80 70 60 50 40
0
0.05
0.1
0.15
0.2
Expiratory flow rate L.s
0.25
0.3
-1
FIGURE 11.1 The characteristics of an exhaled breath at a flow rate of 0.050 L/s (FE NO0.05 , left panel). The peak represents the inhaled NO not taken up and with a steady exhalation flow the NO levels out and with the inhalation of low concentration of NO the curve rapidly goes toward environment concentrations. The middle panel shows the flow dependency and each plateau value and plateau values at the different exhalation flow rates are plotted. The right panel shows the output of NO for each flow rate and this is the plot that is used for both the linear and the non-linear models.
to perform. The studies included in the previously mentioned Cochrane review are all done with one exhalation flow rate. Exhalation at different flow rates gives different NO concentrations in the exhaled gas. Contrary to the CO2 measurement, the concentration of which peaks at the end exhalation, NO concentrations peaked at the beginning of the exhalation; see Figure 11.1 left panel. Therefore, less NO must be present in the alveoli so the gradient in NO number density allows the application of Fick’s first law of diffusion.14 For simplicity, low concentration enters a tube and travels along the tube while molecules diffuse into the tube depending on the transfer rate across the tube and the concentration surrounding the tube. Hence, during an inhalation air is drawn down to the lung periphery by pressure difference (convection), the alveoli expand, and gas exchange takes place driven by a concentration gradient (diffusion). With the movement of air and the addition of NO from conducting airways into the alveolar region, equilibrium for the different gases is soon reached and the exhalation begins. In health, the concentration of NO in the alveolar gas phase (CA NO) is most likely in the same order as for the alveolar tissue (Calv NO), i.e. close to zero, since blood is a sink for NO. A bolus of near-zero NO gas starts the transport upwards to be exhaled. During the passage in the airway the bolus picks up NO from the small and large airways and the increase of NO in the bolus depends on the time of contact. From a system theory perspective, the conducting airways can be modeled as a black box adding NO to the gas flow from the lungs. The experimentally obtained mutual relationships between NO output and NO input for a set of exhalation flows can be used to construct a transfer function able to predict the system NO output for any other flow. However, instead of a purely mathematical approach it is important to be able to link all the information defining the system to physical/physiological properties of the human airways
Högman and Meriläinen
parameters. In Figure 11.1, right panel, the NO output from the lung is illustrated and is used for both linear and non-linear NO models of the lung.
11.2 A TWO COMPARTMENT MODEL A simplification of the lung can be made based on a two compartment model (2CM), consisting of one rigid compartment representing the airways (the tube), and one expansile compartment representing the gas exchange areas,15 see Figure 11.2. Fick’s first law of diffusion can then be applied to the mechanism controlling the NO transfer to the airway lumen. Measurements of exhaled NO at three or more flow rates can be exploited to determine three different parameters defining the NO production. The abbreviations have been agreed upon16 and are given in Table 11.1. The expansile compartment of the lung holds the fraction of NO in the gas phase of the alveolar region, CA NO in ppb. The rigid conducting airway system lumped into a single tube is characterized by the airway tissue concentration of NO (wall concentration, Caw NO in ppb) and the total airway compartment diffusing capacity, transfer factor, or conductance for radial mass transfer of NO from the airway wall to the gas stream (Daw NO in mL/s). The flux of NO from the wall to the lumen depends on the distance z along the airway and it has a total value Jaw NO = Daw NO · (Caw NO − CA NO) at flows high enough to keep the local lumen NO value close to CA NO all the way along the airway. The highest potential total
Exhaled NO = FE NO
Airway flux = J’aw NO
(((
Alveolarfraction = CA NO
Transfer factor = Daw NO
Airway wall fraction = Caw NO
)))
FIGURE 11.2 A simplified model of the NO production in the lung. It consists of an expansile compartment representing the gas exchange areas, and a rigid compartment representing the airways.
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Table 11.1 Abbreviations for the NO parameters of the lung. FE NO
Fractional concentration of exhaled NO in the gas phase (ppb) The exhalation flow rate is determined in L/s and is given as a subscript; for example, at the recommended flow rate of 50 mL/s the abbreviation becomes FE NO0.05 (ppb)
CA NO
Calculated fractional concentration of NO in the gas phase of the alveolar region (ppb)
Calv NO
Calculated alveolar tissue concentration of NO (ppb)
Caw NO
Calculated tissue concentration of NO of the airway wall (ppb)
Daw NO
Calculated airway compartment diffusing capacity, transfer factor, or conductance for radial mass transfer of NO from the airway wall to the gas stream (mL/s)
Jaw NO
Calculated total maximum flux (rate of radial transport) of NO in the airway compartment (nL/s), which occurs when the gas phase concentration in the alveolar compartment is close to zero, and is equal to the product Caw NO · Daw NO
Jaw NO
Calculated flux of NO in the airway compartment (nL/s). This flux is dependent on the value of CA NO. If CA NO is close to zero the flux is equal to the total flux
V˙ NO
Elimination rate of NO from the breath during exhalation (nL/min)
V˙ E
Exhalation flow rate (mL/s)
NO = D NO · C NO, and is obtained when C NO is zero. flux is then Jaw aw aw A NO, The definitions and physical meanings of the two NO fluxes, Jaw NO and Jaw have been inadequately described and understood in several publications on exhaled NO. This is related to the attempts to solve the three independent parameters from measurements at only two flows, which is possible only by assuming CA NO to be NO. zero, making Jaw NO equal to Jaw As seen in Figure 11.1, right panel, the curve shows two phases where the steeper curve represents the airways portion and the gentle slope the alveolar portion. These two approximately linear curves meet around 50–60 mL/s, just at the recommended flow rate, irrespective of a high FE NO0.05 value or low (unpublished observations). CA NO shows no correlations with FE NO0.05 but has a good correlation with Jaw NO, which also strengthen the two compartment model. Other findings supporting the two compartment model are from Verbanck et al. who have shown that a position change from sitting to prone or supine position only changes the CA NO value and not the NO produced in the airway.17 It is well known that the alveolar volume changes with body position and that the airways are not affected by this change.
Högman and Meriläinen
11.3 DIFFERENT NO MODELS Different approaches have been used to calculate the NO parameters and most of the models were presented during 1998–2000.15,18–21 Comparisons of the different models have been published and the non-linear model has been found to provide a better fit to experimental22 data and is, therefore, suggested to be the best method.23 The advantage of the multiple flow analysis relies on the simplest calculations and may therefore be the most robust.24
11.3.1 The Tsoukias and George two compartment model (2CM) The Tsoukias and George two compartment model (2CM) is a two parameter model giving Jaw NO and CA NO,15 where NO concentrations at multiple constant exhalation flow rates between 100 and 500 mL/s are obtained25 and V˙NO is plotted versus the flow rates; see Figure 11.3. The slope and the intercept of the resulting linear regression line provide estimates of CA NO and Jaw NO according to Eq. (11.1) and the r -value of the line gives the quality of the fit. V˙NO = CA NO · V˙E + Jaw NO
(11.1)
11.3.2 The Silkoff technique (ST) The Silkoff technique (ST) utilizes a non-linear regression technique using Eq. (11.2) which is the basic equation of the 2CM for presenting the dependence of exhaled
NO output nL/min
90 80 70
FENO0.05
60 50 40
0
0.05
0.1
0.15
0.2
0.25
Exhaled flow rate L/s
FIGURE 11.3 2CM model by Tsoukias and George. The slope of the regression line will give the CA NO and the intercept on the Y -axis gives the NO-flux, Jaw NO. FE NO0.05 is not one of these flow rates.
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NO values as a function of flow.21 Silkoff et al. used nine flow rates between 4 and 1550 mL/s to estimate the three NO parameters. FE NO = Caw NO + (CA NO − Caw NO) · exp (−Daw NO/V˙E )
(11.2)
Consequently, the NO output is obtained from Eq. (11.2) by multiplying both sides by V˙E : V˙NO = V˙E · FE NO = V˙E · (Caw NO + (CA NO − Caw NO) · exp (−Daw NO/V˙E ))
(11.3)
For all the flows V˙E Daw NO, the exponential in Eq. (11.3) can be linearized and replaced by (1 − Daw NO/V˙E ); then Eq. (11.3) becomes: V˙NO = V˙E · FE NO = CA NO · V˙E + (Caw NO − CA NO) · Daw NO = CA NO · V˙E + Jaw NO · Daw NO (11.4) NO can be estimated from two low flow Silkoff et al. also suggests that Daw NO and Jaw NO/Daw NO of rates (<50 mL/s) by using the slope (−Daw NO) and the intercept Jaw a plot of V˙NO versus FE NO. However, this is only a semi-quantitative approximation which can be shown by obtaining V˙E as a function of FE NO using Eq. (11.2) and substituting the result in Eq. (11.3).
11.3.3 The Pietropaoli technique (PT) The Pietropaoli technique (PT) includes six constant-exhalation vital capacity maneu NO are estimated vers with flow rates between 6 and 1355 mL/s.20 CA NO and Jaw together with the estimation of Daw NO. The theory of this technique was first published by Hyde et al.26 A plot of FE NO versus 1/V˙E for V˙E > 200 mL/s renders the intercept as an estimation of CA NO according to Eq. (11.5). FE NO = CA NO + Jaw NO · 1/V˙E
(11.5)
NO is When comparing Eq. (11.5) to Eq. (11.4), they become identical if Jaw replaced by Jaw NO, which is the accurate expression for non-zero CA NO values. CA NO and Jaw NO can be solved from two data points if flow values high enough to allow linear approximation of the exponential are used; see Figure 11.4. With more data points, any non-linear curve-fitting algorithm can be used allowing Caw NO and Daw NO to be separated.
11.3.4 The Högman and Meriläinen algorithm (HMA) The Högman and Meriläinen algorithm (HMA) was first described in 1999 and is a non-linear model18,19 with a set of three exhalation flow rates of typical values of 10 mL/s (low), 100 mL/s (medium), and 300 mL/s (high), see Figure 11.5. The low limit is presently under investigation. The high flow rate previously was 500 mL/s, at
Högman and Meriläinen
150
FENO ppb
125 100 75 50 25 0
0
0.05
0.1
0.15
. 1/VE mL s-1
0.2
0.25
FIGURE 11.4 CA NO can be estimated by the Pietropaoli-technique if flow rates above 200 mL/s are used. The intercept on the Y -axis is the CA NO. Jaw NO is the slope of the line.
NO output nL/min
90 80 70
FENO0.05 60 50 40
0
0.05
0.1
0.15
0.2
0.25
0.3
Exhaled flow rate L/s
FIGURE 11.5 The Högman-Meriläinen Algorithm (HMA). To create the non-linear flow-volume curve at least tree flow rates are needed and these flow rates should be at both ends and one in the middle of the curve. FE NO0.05 is not one of these flow rates.
which it was difficult to find a plateau; this can be lowered to 300 mL/s but no lower.27 Therefore, it can be stated that the so-called flow independent NO parameters are actually flow dependent. These high flow rates might still be too high for children to perform. The V˙NO versus flow rate is plotted and a line set through the medium and
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Table 11.2 NO parameters obtained from the different NO models generated by multiple flow rates. Note that the Tsoukias and George model needs an additional flow rate to measure FE NO0.05 while this value can be calculated with the HMA model. CA NO
NO Jaw
Jaw NO
Caw NO
Daw NO
Tsoukias and Two VC at George 2CM15 100–500
×
×
–
–
–
Silkoff technique21
Nine breaths at 4–1550
×
×
×
×
×
Pietropaoli technique20
Six breaths at 6–1355
×
×
×
×
×
HögmanMeriläinen algorithm19
Three breaths at 10, 100, 300
×
×
×
×
×
Model
Range of Flow Rates (mL/s)
high flow rates gives estimates of CA NO and Jaw NO. Daw NO and Caw NO, are then separated from Jaw NO by using all three flow rates employing an iterative algorithm with a third-order correction term added to the first-order linear approximation of the exponential in Eqs. (11.2) and (11.3). A specially added feature of this approach is to use an algorithm to test if the measured set of data points is mathematically consistent with the model. This is to avoid the potential problem of standard curve fitting algorithms when applied to three data points only, when a good fit can be found to Eq. (11.2), even if some of the data points are corrupted by measurement errors of technical origin. The condition required for data allowing reliable separation of Daw NO and Caw NO, can be written in a general form as in Eq. (11.6) (see Table 11.2): V˙E,high V˙E,medium − V˙E,low FE NOlow − FE NOmedium < · FE NOmedium − FE NOhigh V˙E,low V˙E,high − V˙E,medium
(11.6)
If this is not the case, the data set suffers from measurement errors either in the flow or NO concentration, or both. Additionally, it should naturally be required that CA NO be positive, since a concentration of a gas cannot be negative!
11.4 CORRECTIONS FOR AXIAL BACK DIFFUSION During the past few years the role of axial back diffusion of NO at low flows from airway generation close to the alveolar compartment has been under investigation.28–31
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A trumpet-shaped geometry for the airways is utilized instead of a constant diameter tube, as used in earlier models. Even a multi-compartment model has been suggested.32 It still remains to be confirmed if the single or double trumpet model is enough to assess the magnitude of axial diffusion, because the relationships between the local airway wall area and the airway cross section vary a lot in the real fractal airway tree from a single tube even if its cross section is matched with the sum of all the branches. Axial back diffusion models presently in use are those by Condorelli et al.28 and by Kerckx et al.29 The first one is a model combining 2CM with a consideration of axial diffusion and the trumpet shape of the airway tree. Four different flow rates between 100 and 250 mL/s are used to estimate Jaw NO and CA NO. The model predicts that a plot of V˙NO versus V˙E produces a linear relationship in which the slope is equal to NO · 0.00078 and the intercept is equal to J NO/1.7, according to Eqs. CA NO + Jaw aw (11.7) and (11.8): Jaw NOcorrected = 1.7 · Jaw NO CA NOcorrected = CA NO − Jaw NO/740
(11.7) (11.8)
Kerckx et al. also use the 2CM but generate the regression line with flow rates of 50, 175, and 300 mL/s. They conclude that the CA NO should be corrected according to Eq. (11.9): (11.9) CA NOcorrected = (CA NO − 0.08 · FE NO0.05 )/0.92 Endogenous NO from the upper and lower airways is transported into the alveolar region. When the alveolus extends with inhaled gas the process of diffusion of O2 and CO2 takes place. This endogenous inhalation of NO in the ppb range is known to increase oxygenation.33 Exogenous inhalation of NO in the ppm range has also been shown to improve oxygenation,34,35 but most effectively during pulsed delivery in the beginning of the breath.36 Hence, NO is transported together with the highest O2 concentration to reach the pulmonary vessels, dilate them, and thereby facilitate the gas exchange. NO is metabolized and an on-off dilatation is achieved. Continuous NO inhalation will cause endothelin-1 (constriction agent) to increase,37 while pulsed NO delivery does not affect its level.38 Seen from a physiological viewpoint, the movement of NO by axial diffusion is therefore not an optimal solution. During exhalation when efficient gas exchange has taken place, dilatation of vessels by NO axial diffusion will only cause inhomogeneity of the lung.36 Since NO values at multiple flow rates are obtained without breath hold for the extended NO analysis, axial diffusion is not taken into considerations. Another reason for not using the corrections is that they frequently give negative CA NO, especially with increased FE NO0.05 , as can be seen in Table 11.3. This can be caused by the high flow rates at which CA NO was calculated and where axial diffusion might not be significant. Another reason is that back diffusion must (physiologically) be different from individual to individual and especially altered during disease, and so a fixed correction might be inappropriate.
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Table 11.3 Data obtained with three flow rates 10, 100, and 350 mL/s with the HMA method in three subjects with low, normal, and high FE NO0.05 . Corrections for axial back diffusion made by the formulae described by Condorelli et al. correction 128 and by Kerckx et al. correction 2.29 With increasing FE NO0.05 and Jaw NO the corrections become negative. Subjects A B C
FE NO0.05 (ppb) 8 19 47
Jaw NO (nL/s)
CA NO (ppb)
CA NO (ppb) Correction 1
CA NO (ppb) Correction 2
0.4 1.3 2.3
1.0 1.4 1.5
0.5 0.0 −1.6
0.4 −0.1 −2.4
11.5 LIMITATIONS Understanding the limitations of the different theoretical models is important, but it is also very important to know the limitations of the equipment. Some analyzers are built for environmental research (ppm not ppb) and are too slow to be used for modeling, while other analyzers are built for clinical use for high NO concentrations but with low accuracy at low NO concentrations. Extended NO analyses demand an analyzer with an accuracy of 0.1 ppb, which adequately measures low NO concentration at high flow rates. An accuracy of 0.1 mL/s for low flow rates and 1 mL/s for high flow rates is necessary for the flow measurements. The instruments should be easy to calibrate and to verify both the NO signal and flow rate and to have a view of the performance. When using linear regression, R 2 close to 1 will assure the investigator that the data are sound. It is not acceptable to use the wrong flow rates for the chosen model. Most often the model is used with flow rates lower than are recommended; including 50 mL/s, a value that gives a steeper slope and thereby a falsely high CA NO and a lower intercept with a falsely low Jaw NO.22 Vital capacity (VC) maneuvers should be avoided since high pressures in the lung can cause a release of NO in animals.39 It has been shown that performing a VC maneuver in humans results in a FE NO0.1 of 12 ppb with the time to reach a plateau of 16 s. The corresponding values after a deep breath were 12 ppb and 7 s.18 There was no statistical difference in FE NO between the maneuvers, but the time to reach a plateau was significantly longer, due to a down sloping curve before the plateau was reached. This has to be considered when automated programs are designed that often limit the time for exhalation. Avoidance of VC maneuvers is in keeping with the recommendations that NO measurements should be performed before other lung function measurements.13 As with all NO measurements the lips must be closed around the mouthpiece in order to avoid leaks. The system has to be tight, including tubing and fittings. For the measurements at low flow a proper exhaled pressure resistance is required in order to avoid contamination with nasal NO.
Högman and Meriläinen
11.6 VALUES FROM NON-SMOKING HEALTHY SUBJECTS The extended NO analysis (HMA) has been applied to a random sample of the Swedish population.40 Based on these results, it was suggested that the values for healthy subjects should be considered to fall between the following ranges: FE NO0.05 10– 30 ppb; Caw NO 50–250 ppb; Daw NO 5–15 mL/s; Jaw NO 0.8–1.6 nL/s; and CA NO 0–4 ppb. Interestingly, this study showed a correlation between age and CA NO in women (r = 0.51, p = 0.001) but not in men. Explanations for the higher CA NO could possibly then be that the uptake of the inhaled NO is lower, i.e. that there is a ventilation/perfusion mismatch followed by inhomogeneity of the lung with age. Further capillary blood volume in women older than 40 years of age decreases in comparison to younger women.41 The same research group has also shown that the diffusing capacity declines in a linear fashion with age.42 In the study by Gelb et al. using the Tsoukias and George NO model and the Condorelli correction, CA NO was significantly increased in a group of subject above 60 years of age while Jaw NO was similar for all ages.43 The values were in line with the study by Högman et al.40 except for the Jaw NO values which were about 70 % increased in the study by Gelb et al. This is explained by the corrected values suggested by Condorelli et al.28
11.7 THE USEFULNESS OF EXTENDED NO ANALYSIS While extended NO analysis is unlikely to become a diagnostic test for respiratory diseases of itself the analysis will definitely add more information in the diagnostic process and in evaluating treatment, especially corticosteroid treatment in asthma. The recent Cochrane report stated that defining the dose of inhaled corticosteroids (ICS) based on the FE NO0.05 value cannot be routinely supported.8 It has also been said that reference values will not have a role, since clinically stable patients have generally higher values.44 That FE NO0.05 is high in eosinophilic asthma is due to an increased Jaw NO, which is the product of Caw NO and Daw NO. The treatment with ICS will NO and decrease Caw NO without affecting Daw NO,21 resulting in a still elevated Jaw a FE NO0.05 higher than reference values. A key feature in eosinophilic rhinitis and asthma is an elevated Daw NO.45 Hence, if the Daw NO remains high despite treatment with ICS this will hinder the FE NO0.05 value to reach reference values. Instead of using FE NO0.05 as a guide for ISC treatment, the Caw NO value could possibly be beneficial. Due to the simplicity and the instant results from the extended analysis, NO measurements are attractive from a patient’s viewpoint. For example the CA NO values seem to add to the understanding of the severity of asthma9,10 and diseases such as COPD,19,46 alveolitis47 , and scleroderma.48 In Obstructive Sleep Apnea Syndrome the CA NO but not the Jaw NO is reduced, and this low CA NO is associated with an increased risk of hypertension.49 An increase in CA NO could be interpreted as systemic engagement with circulating mediators causing iNOS expression in the alveolar region. In support of this theory is the effect of oral prednisone on the decrease
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of CA NO.50 Another explanation, most likely in COPD but also in severe asthma, is a reduced uptake of inhaled NO due to ventilation/perfusion mismatch. CA NO has been found to be increased in systemic sclerosis and non-invasive NO analysis could possibly be used to assess the extent of interstitial lung disease48 and be a tool for early detection of this disease.51 The modeling of NO production both in asthma and COPD has shown many similarities. With the results from the extended NO analysis two groups of COPD patients could clearly be seen,19 one with increased Daw NO, the characteristic feature of atopic asthma,45 and one group with decreased Daw NO. Notably, non-atopic rhinitis, nonatopic asthma, and healthy individuals have similar values of the NO parameters from the extended NO analysis.45 Modeling of NO production in the lung is one way to see such differences. In the light of the discussion in the European Respiratory Society about focusing on pheno- and endo-types in respiratory diseases, instead of the diagnosis of asthma and COPD, extended NO analysis could be such a tool, either the linear model or the non-linear model. That monitoring of NO is here to stay is clear, but we do need to know how and when to use this tool. NO can be important in indicating the risk of new-onset asthma,52 as well as lung function decline in asthma.53 Atopic rhinitis patients have increased exhaled NO54 due to an increased Daw NO and not an up-regulation of NO production in the airway wall.19 Perhaps the development of atopic asthma could be prevented if there was a way to normalize Daw NO in rhinitis.
11.8 CONCLUSIONS In clinical practice we need simple models/methods that will give us repeatable results. The extended NO analysis will give additional information about the production of NO in the lung, compared to just one NO concentration at one flow rate. NO measurement in exhaled breath is a non-invasive method that gives instant results, gives added value in the diagnostic procedure and is a valuable tool for following the effects of antiinflammatory treatments. To date, there has not been a task force on the modeling of NO, like that for the recommendations on the measurement of exhaled NO.13 The abbreviations of the NO parameters have been agreed on16 and until we get recommendations the researchers and clinicians must follow the flow rates used for the different models and which are also applicable for the models used for correction of different NO parameters. These NO parameters are indeed flow dependent.
References 1. Gaston B, Drazen JM, Loscalzo J, Stamler JS. The biology of nitrogen oxides in the airways. Am J Respir Crit Care Med 1994;149:538–51. 2. Ricciardolo FL. Multiple roles of nitric oxide in the airways. Thorax 2003;58:175–82. 3. Ricciardolo FL, Sterk PJ, Gaston B. Folkerts G. Nitric oxide in health and disease of the respiratory system. Physiol Rev 2004;84:731–65.
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4. Ludviksdottir D, Janson C, Högman M, Hedenström H, Björnsson E, Boman G. Exhaled nitric oxide and its relationship to airway responsiveness and atopy in asthma. BHR-Study Group. Respir Med 1999;93:552–6. 5. Guo FH, Comhair SA, Zheng S, Dweik RA, Eissa NT, Thomassen MJ, et al. Molecular mechanisms of increased nitric oxide (NO) in asthma: evidence for transcriptional and post-translational regulation of NO synthesis. J Immunol 2000;164:5970–80. 6. Kobzik L, Bredt DS, Lowenstein CJ, Drazen J, Gaston B, Sugarbaker D, et al. Nitric oxide synthase in human and rat lung: immunocytochemical and histochemical localization. Am J Respir Cell Mol Biol 1993;9:371–7. 7. Pucci S, Incorvaia C. Allergy as an organ and a systemic disease. Clin Exp Immunol 2008;153(Suppl. 1):1–2. 8. Petsky HL, Cates CJ, Li A, Kynaston JA, Turner C, Chang AB. Tailored interventions based on exhaled nitric oxide versus clinical symptoms for asthma in children and adults. Cochrane Database Syst Rev 2009; CD006340. 9. Lehtimäki L, Kankaanranta H, Saarelainen S, Turjanmaa V, Moilanen E. Increased alveolar nitric oxide concentration in asthmatic patients with nocturnal symptoms. Eur Respir J 2002;20:841–5. 10. Lehtimäki L, Kankaanranta H, Saarelainen S, Turjanmaa V, Moilanen E. Peripheral inflammation in patients with asthmatic symptoms but normal lung function. J Asthma 2005;42:605–9. 11. Högman M, Strömberg S, Schedin U, Frostell C, Hedenstierna G, Gustafsson LE. Nitric oxide from the human respiratory tract efficiently quantified by standardized single breath measurements. Acta Physiol Scand 1997;159:345–6. 12. Silkoff PE, McClean PA, Slutsky AS, Furlott HG, Hoffstein E, Wakita S, et al. Marked flow-dependence of exhaled nitric oxide using a new technique to exclude nasal nitric oxide. Am J Respir Crit Care Med 1997;155:260–7. 13. ATS/ERS recommendations for standardized procedures for the online and offline measurement of exhaled lower respiratory nitric oxide and nasal nitric oxide, 2005. Am J Respir Crit Care Med 2005;171:912–30. 14. Fick A. Braunschweig: druck und Verlag von Friedrich Vieweg und Sohn Die Medizinische. Physik 1856. 15. Tsoukias NM, George SC. A two-compartment model of pulmonary nitric oxide exchange dynamics. J Appl Physiol 1998;85:653–66. 16. George SC, Högman M, Permutt S, Silkoff PE. Modeling pulmonary nitric oxide exchange. J Appl Physiol 2004;96:831–9. 17. Verbanck S, Kerckx Y, Schuermans D, de Bisschop C, Guenard H, Naeije R, et al. The effect of posture-induced changes in peripheral nitric oxide uptake on exhaled nitric oxide. J Appl Physiol 2009;106:1494–8. 18. Högman M, Drca N, Ehrstedt C, Meriläinen P. Exhaled nitric oxide partitioned into alveolar, lower airways and nasal contributions. Respir Med 2000;94:985–91. 19. Högman M, Holmkvist T, Wegener T, Emtner M, Andersson M, Hedenström H, et al. Extended NO analysis applied to patients with COPD, allergic asthma and allergic rhinitis. Respir Med 2002;96:24–30. 20. Pietropaoli AP, Perillo IB, Torres A, Perkins PT, Frasier LM, Utell MJ, et al. Simultaneous measurement of nitric oxide production by conducting and alveolar airways of humans. J Appl Physiol 1999;87:1532–42.
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21. Silkoff PE, Sylvester JT, Zamel N, Permutt S. Airway nitric oxide diffusion in asthma: Role in pulmonary function and bronchial responsiveness. Am J Respir Crit Care Med 2000;161:1218–28. 22. Högman M, Meriläinen P. Extended NO analysis in asthma. J Breath Res 2007;1:024001. 23. Roy K, Borrill ZL, Starkey C, Hazel AL, Morris J, Vestbo J, et al. Use of different exhaled nitric oxide multiple flow rate models in COPD. Eur Respir J 2007;29:651–9. 24. Mahut B, Louis B, Zerah-Lancner F, Delclaux C. Validity criteria and comparison of analytical methods of flow-independent exhaled NO parameters. Respir Physiol Neurobiol 2006;153:148–56. 25. Tsoukias NM, Tannous Z, Wilson AF, George SC. Single-exhalation profiles of NO and CO2 in humans: effect of dynamically changing flow rate. J Appl Physiol 1998;85:642–52. 26. Hyde RW, Geigel EJ, Olszowka AJ, Krasney JA, Forster Jr RE, Utell MJ, et al. Determination of production of nitric oxide by lower airways of humans-theory. J Appl Physiol 1997;82:1290–6. 27. Holmkvist T, Meriläinen P, Högman M. The maximum flow required for the extended NO analysis. Am J Respir Crit Care Med 2003;167:A53. 28. Condorelli P, Shin HW, Aledia AS, Silkoff PE, George SC. A simple technique to characterize proximal and peripheral nitric oxide exchange using constant flow exhalations and an axial diffusion model. J Appl Physiol 2007;102:417–25. 29. Kerckx Y, Michils A, Van Muylem A. Airway contribution to alveolar nitric oxide in healthy subjects and stable asthma patients. J Appl Physiol 2008;104:918–24. 30. Kerckx Y, Van Muylem A. Axial distribution heterogeneity of nitric oxide airway production in healthy adults. J Appl Physiol 2009;106:1832–9. 31. Shin HW, George SC. Impact of axial diffusion on nitric oxide exchange in the lungs. J Appl Physiol 2002;93:2070–80. 32. Shelley DA, Puckett JL, George SC. Quantifying proximal and distal sources of NO in asthma using a multicompartment model. J Appl Physiol 2010;108:821–9. 33. Lundberg JO, Settergren G, Gelinder S, Lundberg JM, Alving K, Weitzberg E. Inhalation of nasally derived nitric oxide modulates pulmonary function in humans. Acta Physiol Scand 1996;158:343–7. 34. Pepke-Zaba J, Higenbottam TW, Dinh-Xuan AT, Stone D, Wallwork J. Inhaled nitric oxide as a cause of selective pulmonary vasodilatation in pulmonary hypertension. Lancet 1991;338:1173–4. 35. Rossaint R, Falke KJ, Lopez F, Slama K, Pison U, Zapol WM. Inhaled nitric oxide for the adult respiratory distress syndrome. N Engl J Med 1993;328:399–405. 36. Heinonen E, Nyman G, Meriläinen P, Högman M. Effect of different pulses of nitric oxide on venous admixture in the anaesthetized horse. Br J Anaesth 2002;88:394–8. 37. Pearl JM, Nelson DP, Raake JL, Manning PB, Schwartz SM, Koons L, et al. Inhaled nitric oxide increases endothelin-1 levels: a potential cause of rebound pulmonary hypertension. Crit Care Med 2002;30:89–93. 38. Grubb TL, Hogman M, Edner A, Frendin JH, Heinonen E, Malavasi LM, et al. Physiologic responses and plasma endothelin-1 concentrations associated with abrupt cessation of nitric oxide inhalation in isoflurane-anesthetized horses. Am J Vet Res 2008;69:423–30. 39. Strömberg S, Lönnqvist PA, Persson MG, Gustafsson LE. Lung distension and carbon dioxide affect pulmonary nitric oxide formation in the anaesthetized rabbit. Acta Physiol Scand 1997;159:59–67.
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40. Högman M, Lafih J, Meriläinen P, Bröms K, Malinovschi A, Janson C. Extended NO analysis in a healthy subgroup of a random sample from a Swedish population. Clin Physiol Funct Imaging 2009;29:18–23. 41. Stam H, Versprille A, Bogaard JM. The components of the carbon monoxide diffusing capacity in man dependent on alveolar volume. Bull Eur Physiopathol Respir 1983;19:17–22. 42. Stam H, Hrachovina V, Stijnen T, Versprille A. Diffusing capacity dependent on lung volume and age in normal subjects. J Appl Physiol 1994;76:2356–63. 43. Gelb AF, George SC, Camacho F, Fraser C, Flynn Taylor C, Shakkottai S. Increased nitric oxide concentrations in the small airway of older normal subjects. Chest 2011;139:368–75. 44. Reddel HK, Taylor DR, Bateman ED, Boulet LP, Boushey HA, Busse WW, et al. An official American Thoracic Society/European Respiratory Society statement: asthma control and exacerbations: standardizing endpoints for clinical asthma trials and clinical practice. Am J Respir Crit Care Med 2009;180:59–99. 45. Högman M, Malinovschi A, Norbäck D, Janson C. Added value with extended NO analysis in atopy and asthma. Clin Physiol Funct Imaging 2011;31:294–9. 46. Brindicci C, Ito K, Resta O, Pride NB, Barnes PJ, Kharitonov SA. Exhaled nitric oxide from lung periphery is increased in COPD. Eur Respir J 2005;26:52–9. 47. Lehtimäki L, Kankaanranta H, Saarelainen S, Hahtola P, Järvenpää R, Koivula T, et al. Extended exhaled NO measurement differentiates between alveolar and bronchial inflammation. Am J Respir Crit Care Med 2001;163:1557–61. 48. Tiev KP, Cabane J, Aubourg F, Kettaneh A, Ziani M, Mouthon L, et al. Severity of scleroderma lung disease is related to alveolar concentration of nitric oxide. Eur Respir J 2007;30:26–30. 49. Foresi A, Leone C, Olivieri D, Cremona G. Alveolar-derived exhaled nitric oxide is reduced in obstructive sleep apnea syndrome. Chest 2007;132:860–7. 50. Gelb AF, Taylor CF, Nussbaum E, Gutierrez C, Schein A, Shinar CM, et al. Alveolar and airway sites of nitric oxide inflammation in treated asthma. Am J Respir Crit Care Med 2004;170:737–41. 51. Wuttge DM, Bozovic G, Hesselstrand R, Aronsson D, Bjermer L, Scheja A, et al. Increased alveolar nitric oxide in early systemic sclerosis. Clin Exp Rheumatol 2010;28:S5-9 . 52. Bastain TM, Islam T, Berhane KT, McConnell RS, Rappaport EB, Salam MT, et al. Exhaled nitric oxide, susceptibility and new-onset asthma in the Children’s Health Study. Eur Respir J 2011;37:523–31. 53. van Veen IH, Ten Brinke A, Sterk PJ, Sont JK, Gauw SA, Rabe KF, et al. Exhaled nitric oxide predicts lung function decline in difficult-to-treat asthma. Eur Respir J 2008;32:344–9. 54. Henriksen AH, Sue-Chu M, Holmen TL, Langhammer A, Bjermer L. Exhaled and nasal NO levels in allergic rhinitis: relation to sensitization, pollen season and bronchial hyperresponsiveness. Eur Respir J 1999;13:301–6.
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Nitric Oxide, NO, and Carbon Monoxide, CO
D
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11
Marieann Högman*,† and Pekka Meriläinen* * Department
of Medical Sciences, Respiratory Medicine and Allergology, Uppsala University, SE 75105 Uppsala, Sweden † Centre for Research and Development, Uppsala University/County Council of Gävleborg, SE 801 88 Gävle, Sweden
11.1 BACKGROUND NO is involved in many processes in the respiratory system in health1,2 and can be altered during disease.3 Individuals with atopic asthma have higher than normal NO concentrations4 and this is due to increased expression of the inducible NO synthase (iNOS) and availability of the substrate L-arginine in airway epithelial cells of asthmatics.5 There is an abundant expression in the airways of iNOS6 due to continuous transcriptional activation.5 Asthma is an obstructive inflammatory disease that effects the airways and NO has been said to be an inflammatory marker in asthma. Lately it has been put forward that asthma can also be a systemic disease.7 Since the disease is characterized by recurrent attacks and varies in frequency and severity from person to person and is due to inflammation, the need for monitoring is obvious. However, it was recently concluded in a Cochrane review that, at present, defining the dose of inhaled corticosteroids based on exhaled nitric oxide cannot be routinely advocated.8 In severe asthma the peripheral airways are also affected and treatment given by inhalation may not reach these airways. Therefore, it is desirable to have insight into how widespread the inflammation is. One way of doing this is to model NO in the respiratory system and thereby get NO values from the alveolar region as well as from the airways. This has been done by Lehtimäki et al. and they have found in more severe asthma and in nocturnal asthma that NO from the alveolar region is increased.9,10 The discovery of the flow dependency of exhaled NO was made with the new generation of fast responding NO analyzers and led to the opportunity to model NO production. This discovery shed light on the different results published in healthy controls as in disease and was done by two research groups independently.11,12 A European task force was formed in 1997 and the latest published guidelines is an ERS and ATS joint statement which was to set the flow rate at 50 mL/s (FE NO0.05 ), but other flow rates could be used but must be stated.13 As can be seen in Figure 11.1, middle panel; higher flow rates results in lower NO values. FE NO0.05 was chosen because it gives a good reading of the analyzer and it is easy for the patient Volatile Biomarkers. http://dx.doi.org/10.1016/B978-0-44-462613-4.00011-8 © 2013 Elsevier B.V. All rights reserved.
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0
2
4
6
8
10 12 14 16
Time seconds
-1
40 35 30 25 20 15 10 5 0
NO output nL·min
FE NO ppb
90
FE NO ppb
196
0
0.1
0.2
0.3
0.4 -1
Expiratory flow rate L.s
80 70 60 50 40
0
0.05
0.1
0.15
0.2
Expiratory flow rate L.s
0.25
0.3
-1
FIGURE 11.1 The characteristics of an exhaled breath at a flow rate of 0.050 L/s (FE NO0.05 , left panel). The peak represents the inhaled NO not taken up and with a steady exhalation flow the NO levels out and with the inhalation of low concentration of NO the curve rapidly goes toward environment concentrations. The middle panel shows the flow dependency and each plateau value and plateau values at the different exhalation flow rates are plotted. The right panel shows the output of NO for each flow rate and this is the plot that is used for both the linear and the non-linear models.
to perform. The studies included in the previously mentioned Cochrane review are all done with one exhalation flow rate. Exhalation at different flow rates gives different NO concentrations in the exhaled gas. Contrary to the CO2 measurement, the concentration of which peaks at the end exhalation, NO concentrations peaked at the beginning of the exhalation; see Figure 11.1 left panel. Therefore, less NO must be present in the alveoli so the gradient in NO number density allows the application of Fick’s first law of diffusion.14 For simplicity, low concentration enters a tube and travels along the tube while molecules diffuse into the tube depending on the transfer rate across the tube and the concentration surrounding the tube. Hence, during an inhalation air is drawn down to the lung periphery by pressure difference (convection), the alveoli expand, and gas exchange takes place driven by a concentration gradient (diffusion). With the movement of air and the addition of NO from conducting airways into the alveolar region, equilibrium for the different gases is soon reached and the exhalation begins. In health, the concentration of NO in the alveolar gas phase (CA NO) is most likely in the same order as for the alveolar tissue (Calv NO), i.e. close to zero, since blood is a sink for NO. A bolus of near-zero NO gas starts the transport upwards to be exhaled. During the passage in the airway the bolus picks up NO from the small and large airways and the increase of NO in the bolus depends on the time of contact. From a system theory perspective, the conducting airways can be modeled as a black box adding NO to the gas flow from the lungs. The experimentally obtained mutual relationships between NO output and NO input for a set of exhalation flows can be used to construct a transfer function able to predict the system NO output for any other flow. However, instead of a purely mathematical approach it is important to be able to link all the information defining the system to physical/physiological properties of the human airways
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parameters. In Figure 11.1, right panel, the NO output from the lung is illustrated and is used for both linear and non-linear NO models of the lung.
11.2 A TWO COMPARTMENT MODEL A simplification of the lung can be made based on a two compartment model (2CM), consisting of one rigid compartment representing the airways (the tube), and one expansile compartment representing the gas exchange areas,15 see Figure 11.2. Fick’s first law of diffusion can then be applied to the mechanism controlling the NO transfer to the airway lumen. Measurements of exhaled NO at three or more flow rates can be exploited to determine three different parameters defining the NO production. The abbreviations have been agreed upon16 and are given in Table 11.1. The expansile compartment of the lung holds the fraction of NO in the gas phase of the alveolar region, CA NO in ppb. The rigid conducting airway system lumped into a single tube is characterized by the airway tissue concentration of NO (wall concentration, Caw NO in ppb) and the total airway compartment diffusing capacity, transfer factor, or conductance for radial mass transfer of NO from the airway wall to the gas stream (Daw NO in mL/s). The flux of NO from the wall to the lumen depends on the distance z along the airway and it has a total value Jaw NO = Daw NO · (Caw NO − CA NO) at flows high enough to keep the local lumen NO value close to CA NO all the way along the airway. The highest potential total
Exhaled NO = FE NO
Airway flux = J’aw NO
(((
Alveolarfraction = CA NO
Transfer factor = Daw NO
Airway wall fraction = Caw NO
)))
FIGURE 11.2 A simplified model of the NO production in the lung. It consists of an expansile compartment representing the gas exchange areas, and a rigid compartment representing the airways.
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Table 11.1 Abbreviations for the NO parameters of the lung. FE NO
Fractional concentration of exhaled NO in the gas phase (ppb) The exhalation flow rate is determined in L/s and is given as a subscript; for example, at the recommended flow rate of 50 mL/s the abbreviation becomes FE NO0.05 (ppb)
CA NO
Calculated fractional concentration of NO in the gas phase of the alveolar region (ppb)
Calv NO
Calculated alveolar tissue concentration of NO (ppb)
Caw NO
Calculated tissue concentration of NO of the airway wall (ppb)
Daw NO
Calculated airway compartment diffusing capacity, transfer factor, or conductance for radial mass transfer of NO from the airway wall to the gas stream (mL/s)
Jaw NO
Calculated total maximum flux (rate of radial transport) of NO in the airway compartment (nL/s), which occurs when the gas phase concentration in the alveolar compartment is close to zero, and is equal to the product Caw NO · Daw NO
Jaw NO
Calculated flux of NO in the airway compartment (nL/s). This flux is dependent on the value of CA NO. If CA NO is close to zero the flux is equal to the total flux
V˙ NO
Elimination rate of NO from the breath during exhalation (nL/min)
V˙ E
Exhalation flow rate (mL/s)
NO = D NO · C NO, and is obtained when C NO is zero. flux is then Jaw aw aw A NO, The definitions and physical meanings of the two NO fluxes, Jaw NO and Jaw have been inadequately described and understood in several publications on exhaled NO. This is related to the attempts to solve the three independent parameters from measurements at only two flows, which is possible only by assuming CA NO to be NO. zero, making Jaw NO equal to Jaw As seen in Figure 11.1, right panel, the curve shows two phases where the steeper curve represents the airways portion and the gentle slope the alveolar portion. These two approximately linear curves meet around 50–60 mL/s, just at the recommended flow rate, irrespective of a high FE NO0.05 value or low (unpublished observations). CA NO shows no correlations with FE NO0.05 but has a good correlation with Jaw NO, which also strengthen the two compartment model. Other findings supporting the two compartment model are from Verbanck et al. who have shown that a position change from sitting to prone or supine position only changes the CA NO value and not the NO produced in the airway.17 It is well known that the alveolar volume changes with body position and that the airways are not affected by this change.
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11.3 DIFFERENT NO MODELS Different approaches have been used to calculate the NO parameters and most of the models were presented during 1998–2000.15,18–21 Comparisons of the different models have been published and the non-linear model has been found to provide a better fit to experimental22 data and is, therefore, suggested to be the best method.23 The advantage of the multiple flow analysis relies on the simplest calculations and may therefore be the most robust.24
11.3.1 The Tsoukias and George two compartment model (2CM) The Tsoukias and George two compartment model (2CM) is a two parameter model giving Jaw NO and CA NO,15 where NO concentrations at multiple constant exhalation flow rates between 100 and 500 mL/s are obtained25 and V˙NO is plotted versus the flow rates; see Figure 11.3. The slope and the intercept of the resulting linear regression line provide estimates of CA NO and Jaw NO according to Eq. (11.1) and the r -value of the line gives the quality of the fit. V˙NO = CA NO · V˙E + Jaw NO
(11.1)
11.3.2 The Silkoff technique (ST) The Silkoff technique (ST) utilizes a non-linear regression technique using Eq. (11.2) which is the basic equation of the 2CM for presenting the dependence of exhaled
NO output nL/min
90 80 70
FENO0.05
60 50 40
0
0.05
0.1
0.15
0.2
0.25
Exhaled flow rate L/s
FIGURE 11.3 2CM model by Tsoukias and George. The slope of the regression line will give the CA NO and the intercept on the Y -axis gives the NO-flux, Jaw NO. FE NO0.05 is not one of these flow rates.
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NO values as a function of flow.21 Silkoff et al. used nine flow rates between 4 and 1550 mL/s to estimate the three NO parameters. FE NO = Caw NO + (CA NO − Caw NO) · exp (−Daw NO/V˙E )
(11.2)
Consequently, the NO output is obtained from Eq. (11.2) by multiplying both sides by V˙E : V˙NO = V˙E · FE NO = V˙E · (Caw NO + (CA NO − Caw NO) · exp (−Daw NO/V˙E ))
(11.3)
For all the flows V˙E Daw NO, the exponential in Eq. (11.3) can be linearized and replaced by (1 − Daw NO/V˙E ); then Eq. (11.3) becomes: V˙NO = V˙E · FE NO = CA NO · V˙E + (Caw NO − CA NO) · Daw NO = CA NO · V˙E + Jaw NO · Daw NO (11.4) NO can be estimated from two low flow Silkoff et al. also suggests that Daw NO and Jaw NO/Daw NO of rates (<50 mL/s) by using the slope (−Daw NO) and the intercept Jaw a plot of V˙NO versus FE NO. However, this is only a semi-quantitative approximation which can be shown by obtaining V˙E as a function of FE NO using Eq. (11.2) and substituting the result in Eq. (11.3).
11.3.3 The Pietropaoli technique (PT) The Pietropaoli technique (PT) includes six constant-exhalation vital capacity maneu NO are estimated vers with flow rates between 6 and 1355 mL/s.20 CA NO and Jaw together with the estimation of Daw NO. The theory of this technique was first published by Hyde et al.26 A plot of FE NO versus 1/V˙E for V˙E > 200 mL/s renders the intercept as an estimation of CA NO according to Eq. (11.5). FE NO = CA NO + Jaw NO · 1/V˙E
(11.5)
NO is When comparing Eq. (11.5) to Eq. (11.4), they become identical if Jaw replaced by Jaw NO, which is the accurate expression for non-zero CA NO values. CA NO and Jaw NO can be solved from two data points if flow values high enough to allow linear approximation of the exponential are used; see Figure 11.4. With more data points, any non-linear curve-fitting algorithm can be used allowing Caw NO and Daw NO to be separated.
11.3.4 The Högman and Meriläinen algorithm (HMA) The Högman and Meriläinen algorithm (HMA) was first described in 1999 and is a non-linear model18,19 with a set of three exhalation flow rates of typical values of 10 mL/s (low), 100 mL/s (medium), and 300 mL/s (high), see Figure 11.5. The low limit is presently under investigation. The high flow rate previously was 500 mL/s, at
Högman and Meriläinen
150
FENO ppb
125 100 75 50 25 0
0
0.05
0.1
0.15
. 1/VE mL s-1
0.2
0.25
FIGURE 11.4 CA NO can be estimated by the Pietropaoli-technique if flow rates above 200 mL/s are used. The intercept on the Y -axis is the CA NO. Jaw NO is the slope of the line.
NO output nL/min
90 80 70
FENO0.05 60 50 40
0
0.05
0.1
0.15
0.2
0.25
0.3
Exhaled flow rate L/s
FIGURE 11.5 The Högman-Meriläinen Algorithm (HMA). To create the non-linear flow-volume curve at least tree flow rates are needed and these flow rates should be at both ends and one in the middle of the curve. FE NO0.05 is not one of these flow rates.
which it was difficult to find a plateau; this can be lowered to 300 mL/s but no lower.27 Therefore, it can be stated that the so-called flow independent NO parameters are actually flow dependent. These high flow rates might still be too high for children to perform. The V˙NO versus flow rate is plotted and a line set through the medium and
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Table 11.2 NO parameters obtained from the different NO models generated by multiple flow rates. Note that the Tsoukias and George model needs an additional flow rate to measure FE NO0.05 while this value can be calculated with the HMA model. CA NO
NO Jaw
Jaw NO
Caw NO
Daw NO
Tsoukias and Two VC at George 2CM15 100–500
×
×
–
–
–
Silkoff technique21
Nine breaths at 4–1550
×
×
×
×
×
Pietropaoli technique20
Six breaths at 6–1355
×
×
×
×
×
HögmanMeriläinen algorithm19
Three breaths at 10, 100, 300
×
×
×
×
×
Model
Range of Flow Rates (mL/s)
high flow rates gives estimates of CA NO and Jaw NO. Daw NO and Caw NO, are then separated from Jaw NO by using all three flow rates employing an iterative algorithm with a third-order correction term added to the first-order linear approximation of the exponential in Eqs. (11.2) and (11.3). A specially added feature of this approach is to use an algorithm to test if the measured set of data points is mathematically consistent with the model. This is to avoid the potential problem of standard curve fitting algorithms when applied to three data points only, when a good fit can be found to Eq. (11.2), even if some of the data points are corrupted by measurement errors of technical origin. The condition required for data allowing reliable separation of Daw NO and Caw NO, can be written in a general form as in Eq. (11.6) (see Table 11.2): V˙E,high V˙E,medium − V˙E,low FE NOlow − FE NOmedium < · FE NOmedium − FE NOhigh V˙E,low V˙E,high − V˙E,medium
(11.6)
If this is not the case, the data set suffers from measurement errors either in the flow or NO concentration, or both. Additionally, it should naturally be required that CA NO be positive, since a concentration of a gas cannot be negative!
11.4 CORRECTIONS FOR AXIAL BACK DIFFUSION During the past few years the role of axial back diffusion of NO at low flows from airway generation close to the alveolar compartment has been under investigation.28–31
Högman and Meriläinen
A trumpet-shaped geometry for the airways is utilized instead of a constant diameter tube, as used in earlier models. Even a multi-compartment model has been suggested.32 It still remains to be confirmed if the single or double trumpet model is enough to assess the magnitude of axial diffusion, because the relationships between the local airway wall area and the airway cross section vary a lot in the real fractal airway tree from a single tube even if its cross section is matched with the sum of all the branches. Axial back diffusion models presently in use are those by Condorelli et al.28 and by Kerckx et al.29 The first one is a model combining 2CM with a consideration of axial diffusion and the trumpet shape of the airway tree. Four different flow rates between 100 and 250 mL/s are used to estimate Jaw NO and CA NO. The model predicts that a plot of V˙NO versus V˙E produces a linear relationship in which the slope is equal to NO · 0.00078 and the intercept is equal to J NO/1.7, according to Eqs. CA NO + Jaw aw (11.7) and (11.8): Jaw NOcorrected = 1.7 · Jaw NO CA NOcorrected = CA NO − Jaw NO/740
(11.7) (11.8)
Kerckx et al. also use the 2CM but generate the regression line with flow rates of 50, 175, and 300 mL/s. They conclude that the CA NO should be corrected according to Eq. (11.9): (11.9) CA NOcorrected = (CA NO − 0.08 · FE NO0.05 )/0.92 Endogenous NO from the upper and lower airways is transported into the alveolar region. When the alveolus extends with inhaled gas the process of diffusion of O2 and CO2 takes place. This endogenous inhalation of NO in the ppb range is known to increase oxygenation.33 Exogenous inhalation of NO in the ppm range has also been shown to improve oxygenation,34,35 but most effectively during pulsed delivery in the beginning of the breath.36 Hence, NO is transported together with the highest O2 concentration to reach the pulmonary vessels, dilate them, and thereby facilitate the gas exchange. NO is metabolized and an on-off dilatation is achieved. Continuous NO inhalation will cause endothelin-1 (constriction agent) to increase,37 while pulsed NO delivery does not affect its level.38 Seen from a physiological viewpoint, the movement of NO by axial diffusion is therefore not an optimal solution. During exhalation when efficient gas exchange has taken place, dilatation of vessels by NO axial diffusion will only cause inhomogeneity of the lung.36 Since NO values at multiple flow rates are obtained without breath hold for the extended NO analysis, axial diffusion is not taken into considerations. Another reason for not using the corrections is that they frequently give negative CA NO, especially with increased FE NO0.05 , as can be seen in Table 11.3. This can be caused by the high flow rates at which CA NO was calculated and where axial diffusion might not be significant. Another reason is that back diffusion must (physiologically) be different from individual to individual and especially altered during disease, and so a fixed correction might be inappropriate.
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Table 11.3 Data obtained with three flow rates 10, 100, and 350 mL/s with the HMA method in three subjects with low, normal, and high FE NO0.05 . Corrections for axial back diffusion made by the formulae described by Condorelli et al. correction 128 and by Kerckx et al. correction 2.29 With increasing FE NO0.05 and Jaw NO the corrections become negative. Subjects A B C
FE NO0.05 (ppb) 8 19 47
Jaw NO (nL/s)
CA NO (ppb)
CA NO (ppb) Correction 1
CA NO (ppb) Correction 2
0.4 1.3 2.3
1.0 1.4 1.5
0.5 0.0 −1.6
0.4 −0.1 −2.4
11.5 LIMITATIONS Understanding the limitations of the different theoretical models is important, but it is also very important to know the limitations of the equipment. Some analyzers are built for environmental research (ppm not ppb) and are too slow to be used for modeling, while other analyzers are built for clinical use for high NO concentrations but with low accuracy at low NO concentrations. Extended NO analyses demand an analyzer with an accuracy of 0.1 ppb, which adequately measures low NO concentration at high flow rates. An accuracy of 0.1 mL/s for low flow rates and 1 mL/s for high flow rates is necessary for the flow measurements. The instruments should be easy to calibrate and to verify both the NO signal and flow rate and to have a view of the performance. When using linear regression, R 2 close to 1 will assure the investigator that the data are sound. It is not acceptable to use the wrong flow rates for the chosen model. Most often the model is used with flow rates lower than are recommended; including 50 mL/s, a value that gives a steeper slope and thereby a falsely high CA NO and a lower intercept with a falsely low Jaw NO.22 Vital capacity (VC) maneuvers should be avoided since high pressures in the lung can cause a release of NO in animals.39 It has been shown that performing a VC maneuver in humans results in a FE NO0.1 of 12 ppb with the time to reach a plateau of 16 s. The corresponding values after a deep breath were 12 ppb and 7 s.18 There was no statistical difference in FE NO between the maneuvers, but the time to reach a plateau was significantly longer, due to a down sloping curve before the plateau was reached. This has to be considered when automated programs are designed that often limit the time for exhalation. Avoidance of VC maneuvers is in keeping with the recommendations that NO measurements should be performed before other lung function measurements.13 As with all NO measurements the lips must be closed around the mouthpiece in order to avoid leaks. The system has to be tight, including tubing and fittings. For the measurements at low flow a proper exhaled pressure resistance is required in order to avoid contamination with nasal NO.
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11.6 VALUES FROM NON-SMOKING HEALTHY SUBJECTS The extended NO analysis (HMA) has been applied to a random sample of the Swedish population.40 Based on these results, it was suggested that the values for healthy subjects should be considered to fall between the following ranges: FE NO0.05 10– 30 ppb; Caw NO 50–250 ppb; Daw NO 5–15 mL/s; Jaw NO 0.8–1.6 nL/s; and CA NO 0–4 ppb. Interestingly, this study showed a correlation between age and CA NO in women (r = 0.51, p = 0.001) but not in men. Explanations for the higher CA NO could possibly then be that the uptake of the inhaled NO is lower, i.e. that there is a ventilation/perfusion mismatch followed by inhomogeneity of the lung with age. Further capillary blood volume in women older than 40 years of age decreases in comparison to younger women.41 The same research group has also shown that the diffusing capacity declines in a linear fashion with age.42 In the study by Gelb et al. using the Tsoukias and George NO model and the Condorelli correction, CA NO was significantly increased in a group of subject above 60 years of age while Jaw NO was similar for all ages.43 The values were in line with the study by Högman et al.40 except for the Jaw NO values which were about 70 % increased in the study by Gelb et al. This is explained by the corrected values suggested by Condorelli et al.28
11.7 THE USEFULNESS OF EXTENDED NO ANALYSIS While extended NO analysis is unlikely to become a diagnostic test for respiratory diseases of itself the analysis will definitely add more information in the diagnostic process and in evaluating treatment, especially corticosteroid treatment in asthma. The recent Cochrane report stated that defining the dose of inhaled corticosteroids (ICS) based on the FE NO0.05 value cannot be routinely supported.8 It has also been said that reference values will not have a role, since clinically stable patients have generally higher values.44 That FE NO0.05 is high in eosinophilic asthma is due to an increased Jaw NO, which is the product of Caw NO and Daw NO. The treatment with ICS will NO and decrease Caw NO without affecting Daw NO,21 resulting in a still elevated Jaw a FE NO0.05 higher than reference values. A key feature in eosinophilic rhinitis and asthma is an elevated Daw NO.45 Hence, if the Daw NO remains high despite treatment with ICS this will hinder the FE NO0.05 value to reach reference values. Instead of using FE NO0.05 as a guide for ISC treatment, the Caw NO value could possibly be beneficial. Due to the simplicity and the instant results from the extended analysis, NO measurements are attractive from a patient’s viewpoint. For example the CA NO values seem to add to the understanding of the severity of asthma9,10 and diseases such as COPD,19,46 alveolitis47 , and scleroderma.48 In Obstructive Sleep Apnea Syndrome the CA NO but not the Jaw NO is reduced, and this low CA NO is associated with an increased risk of hypertension.49 An increase in CA NO could be interpreted as systemic engagement with circulating mediators causing iNOS expression in the alveolar region. In support of this theory is the effect of oral prednisone on the decrease
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of CA NO.50 Another explanation, most likely in COPD but also in severe asthma, is a reduced uptake of inhaled NO due to ventilation/perfusion mismatch. CA NO has been found to be increased in systemic sclerosis and non-invasive NO analysis could possibly be used to assess the extent of interstitial lung disease48 and be a tool for early detection of this disease.51 The modeling of NO production both in asthma and COPD has shown many similarities. With the results from the extended NO analysis two groups of COPD patients could clearly be seen,19 one with increased Daw NO, the characteristic feature of atopic asthma,45 and one group with decreased Daw NO. Notably, non-atopic rhinitis, nonatopic asthma, and healthy individuals have similar values of the NO parameters from the extended NO analysis.45 Modeling of NO production in the lung is one way to see such differences. In the light of the discussion in the European Respiratory Society about focusing on pheno- and endo-types in respiratory diseases, instead of the diagnosis of asthma and COPD, extended NO analysis could be such a tool, either the linear model or the non-linear model. That monitoring of NO is here to stay is clear, but we do need to know how and when to use this tool. NO can be important in indicating the risk of new-onset asthma,52 as well as lung function decline in asthma.53 Atopic rhinitis patients have increased exhaled NO54 due to an increased Daw NO and not an up-regulation of NO production in the airway wall.19 Perhaps the development of atopic asthma could be prevented if there was a way to normalize Daw NO in rhinitis.
11.8 CONCLUSIONS In clinical practice we need simple models/methods that will give us repeatable results. The extended NO analysis will give additional information about the production of NO in the lung, compared to just one NO concentration at one flow rate. NO measurement in exhaled breath is a non-invasive method that gives instant results, gives added value in the diagnostic procedure and is a valuable tool for following the effects of antiinflammatory treatments. To date, there has not been a task force on the modeling of NO, like that for the recommendations on the measurement of exhaled NO.13 The abbreviations of the NO parameters have been agreed on16 and until we get recommendations the researchers and clinicians must follow the flow rates used for the different models and which are also applicable for the models used for correction of different NO parameters. These NO parameters are indeed flow dependent.
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