Low alveolar and bronchial nitric oxide in severe uncomplicated obesity

Obesity Research & Clinical Practice (2015) 9, 603—608

ORIGINAL ARTICLE

Low alveolar and bronchial nitric oxide in severe uncomplicated obesity Mauro Maniscalco a,∗, Anna Zedda a, Stanislao Faraone a, Stefano Cristiano b, Matteo Sofia c, Andrea Motta d a

Section of Respiratory Medicine, S. Maria della Pietà Hospital, Casoria, Naples, Italy Department of Surgery, S. Maria della Pietà Hospital, Casoria, Naples, Italy c Department of Respiratory Medicine, Monaldi Hospital, University Federico II of Naples, Italy d Institute of Biomolecular Chemistry, National Research Council, Pozzuoli, Naples, Italy b

Received 24 September 2014 ; received in revised form 18 February 2015; accepted 19 March 2015

KEYWORDS Obesity; Exhaled nitric oxide; Airway; BMI; Lung

∗

Summary Background: Fractional concentration of exhaled nitric oxide (FeNO) is a recognized biomarker of the lower respiratory tract, where it is produced by the proximal conducting airways and the expansible peripheral bronchoalveolar compartment. We have previously shown that large increase in body mass decreases FeNO. Here we evaluated bronchial and alveolar components of the NO output of the lower respiratory tract in subjects with severe uncomplicated obesity (OB). Methods: Fifteen OB subjects (BMI 45.3 ± 5.6 kg/m2 ), 15 healthy controls (HC) (BMI 22.4 ± 2.4 kg/m2 ) and 10 obese subjects who experienced weight loss after bariatric surgery (OBS) (BMI 31.2 ± 3.4 kg/m2 ), were examined. Anthropometry and respiratory lung tests were performed. Exhaled NO was assessed using multiple single-breath NO analysis at different constant expiratory flow rates. From the fractional NO concentration measured at each flow-rate, the total NO flux between tissue and gas phase in the bronchial lumen (J’awNO), and the alveolar NO concentration (CANO) were extrapolated. Results: Measured FeNO levels at 50 mL/s were lower in OB compared with HC and OBS (11.6 ± 2.8 ppb, 18.0 ± 4.1 ppb and 17.6 ± 2.9 ppb, respectively, p < 0.05). In OB, both J’awNO and CANO resulted significantly lower than OBS and HC values. Conclusions: Respiratory NO output is decreased in severe uncomplicated obesity for the reduction of both large/central airway maximal NO flux and alveolar NO

Corresponding author at: Largo delle Mimose 1, 80131 Naples, Italy. Tel.: +39 0817411457; fax: +39 0817411457. E-mail address: [email protected] (M. Maniscalco).

http://dx.doi.org/10.1016/j.orcp.2015.03.004 1871-403X/© 2015 Asian Oceanian Association for the Study of Obesity. Published by Elsevier Ltd. All rights reserved.

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M. Maniscalco et al. concentration. The pathophysiological relevance of airway NO abnormalities in severe obese phenotype remains to be investigated. © 2015 Asian Oceanian Association for the Study of Obesity. Published by Elsevier Ltd. All rights reserved.

Introduction

Materials and methods

Fractional exhaled nitric oxide (FeNO) concentration is recognized as a useful biomarker of airway inflammation for many pulmonary diseases [1]. FeNO can be partitioned in its bronchial and alveolar components, which may represent the contribution to the NO respiratory output from both the proximal non expansible conducting airways compartment and the expansible alveolar small airways compartment [2]. Mathematical models and measurement of NO at different exhalation flow rates [2,3] partitioned peripheral (peripheral/small airway/alveolar NO concentration [CANO])and central (large/central airway maximal NO flux [J’awNO]) airways NO production. This approach requires a minimum of three exhalation flows, and both J’awNO and CANO are calculated by a linear regression analysis. The results provided information on abnormalities of the respiratory NO output in several diseases including asthma, allergic alveolitis, cystic fibrosis, scleroderma, allergic rhinitis, chronic obstructive pulmonary disease, and Sjögren’s syndrome [3]. Some of the above diseases are often associated with obesity [4], which may influence single breath FeNO by interacting with changes in conducting airway and lung volumes [5,6]. Two main reasons are put forward: impaired systemic NO production and sub-clinical low-grade inflammation, and change in both airway and alveolar compartments with consequent changes in respiratory NO output. However, studies on exhaled NO in obesity have shown controversial results [7—9]. These results might be due to the reduction of NO concentration in lung periphery due to the large ‘‘sink effect’’ of the pulmonary circulation, and the high affinity of NO for haemoglobin [10], which may cancel out any change in NO production by the lung parenchyma. To assess the origin of that abnormality within the respiratory tract, we performed extended FeNO analysis by multiple flow in severe obese subjects.

Fifteen consecutive severe obese patients (OB), evaluated for bariatric surgery (Table 1) were studied. Exclusion criteria included: current smoking, respiratory infection within the past 3 weeks; atopy and/or chronic respiratory diseases; FEV1 /FVC ratio < 0.7; heart disease; sleep apnea syndrome; regular medications. Ten obese patients fulfilling the same exclusion criteria, who had undergone gastric banding for weight reducing at least 1 year before (OBS), and 15 healthy controls (HC) were also studied (Table 1). The study was approved by the local Ethic Committee and informed written consent was obtained from each patient. Patients were evaluated the first day with anthropometry, spirometry and pulse oximetry, and the second day with measurement of exhaled NO. All anthropometric measurements were determined in the morning with the subjects wearing very light clothing. The body mass index (BMI) was calculated as the ratio of weight to height squared (kg/m2 ). Lung volumes and flow rates were determined using automated equipment (V Max 22 System SensorMedics, Milan, Italy) as previously described [8]. Recommendations for standardized procedures for various lung function tests were followed [11].

Exhaled NO measurement Exhaled NO concentrations were measured by chemiluminescence (280 NOA Sievers Instruments, Boulder, Sensor Medics, Milan, Italy). Daily 2-point calibration was performed with an external zero filter (Zero Air Filter, Sievers) and a certified NO gas mixture at 1.01 ppm (SIAD Osio, Italy). NO was exhaled using a ‘restricted breath technique’ following the method recommended for online measurement of the fractional exhaled NO concentration in adult [12]. All subjects performed a vital capacity manoeuvre and then a slow (20 s) exhalation against three separate resistances in turn while maintaining the same expiratory

Exhaled nitric oxide and severe obesity

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Table 1 Demographic, anthropometrics and spirometric data in obese subjects (OB), healthy controls (HC) and obese patients after weight loss for bariatric surgery (OBS).

Age (yr) Sex (M/F) Height (m) BMI (kg/m2 ) BMI pre-surgery FVC (% pred) FEV1 (% pred) TLC (% pred) FRC (% pred) RV (% pred)

OB n = 15

HC n = 15

OBS n = 10

p Value

33.8 (10.2) 13/2 1.62 (0.07) 45.3 (5.6)

31.8 (9.6) 13/2 1.67 (0.08) 22.4 (2.5)

0.2 <0.001

100.5 (11.1) 99.4 (11.5) 92.1 (14.5) 69.0 (19.9) 78.2 (24.1)

103.7 (7.1)

41.7 (9.2) 7/3 1.71 (0.07) 31.2 (3.4) 41.1 (1.1) 104.5 (7.1)

102.8 (10.3) 100.1 (8.1) 95.4 (8.6) 102.2 (10.5)

102.3 (7.9) 99.2 (10.1) 87.4 (17.3) 82.2 (26.5)

0.6 0.4 <0.001 <0.001

0.4

0.5

Values given as mean (SD). BMI, body mass index; FVC, forced vital capacity; FEV1 , forced volume at 1 s; TLC, total lung capacity; FRC, functional residual capacity; RV, residual volume.

pressure of 20 mbar mouth resistance, thus yielding three different flow rates of 50, 100 and 150 mL/s. We used a custom-made system including three resistors and a computer programme to achieve a visual feedback for the subjects to expire at a pressure of 20 mbar, which is effective in closing the soft palate [12]. For all resistors, non-linear flow-pressure curves were assessed using a calibrated bell spirometer as flow integrator, so that deviations from the target pressure/flow could be taken into account. All NO measurements were performed when patients were fasting. Ambient NO levels were always low than 10 ppb. The bronchial production of NO (J’awNO) and its alveolar concentration (CANO) were calculated by linear regression according to the equation of Tsoukias and George [2], where the slope and the intercept of the regression line between NO output and exhalation flow indicates CANO and J’awNO, respectively. In addition, J’awNO and CANO were also calculated using the Condorelli adjustment for the axial diffusion of NO as follows [13]: J’awNO = 1.7 × (I) and CANO = [(S) − (I)]/(740 mL/s), where S is the slope and I is the y-intercept by simple linear regression.

Statistics Data are presented as mean ± SD. As the data presented a normal distribution, parametric tests one-way analysis of variance and unpaired student t test were used to analyze the data. Correlations were tested by Pearson test. The validity of the slope—intercept model was tested by mean of Pearson correlation test. A p-value < 0.05 was considered to be significant.

Statistical analyses were performed using GraphPAD Prism, version 4 (GraphPAD Inc., San Diego, CA, USA).

Results Clinical and lung function tests Table 1 shows the main anthropometrics and spirometric data of OB and HS. Obese patients had a higher weight and BMI as compared to HC and OBS groups, while the height, and FVC and FEV1 were not significantly different among the three groups. Lung volumes measurements revealed that FRC and RV were lower in OB as compared to OBS. Total lung capacity did not differ significantly between OB and OBS groups.

Exhaled NO measurements FeNO50 values were significantly lower in OB compared with HC and OBS (11.6 ± 2.8 ppb, 18.0 ± 4.11 ppb and 7.6 ± 2.9 ppb respectively; p = 0.01). J’awNO was significantly reduced in OB with respect to HC and OBC (820 ± 250 pL/s, 1334 ± 362 pL/s and 1205 ± 271 pL/s respectively; p = 0.002), indicating decreased conducting airway flux of NO into the airway lumen (Fig. 1). Similarly, the slopes of the OB patients were significantly reduced compared with the control groups (1.4 ± 1.2 ppb, 2.0 ± 1.3 ppb and 3.1 ± 1.7 ppb, respectively; p = 0.0035), indicating a reduced CANO (Fig. 1). No differences in J’awNO and CANO between HC and OBS were found (p always >0.1).

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M. Maniscalco et al. 7.5

2500

* CANO (ppb)

J'awNO (pl/s)

2000 1500 1000

5.0

*

2.5

500 0

OB

HC

OBS

0.0

OB

HC

OBS

Figure 1 Exhaled nitric oxide with calculated maximal bronchial NO diffusion (J’awNO), and alveolar NO concentration (CANO) in obese subjects (OB), healthy controls (HC) and obese patients after bariatric surgery (OBS). *p < 0.005. Bars correspond to mean values.

Table 2 Correlation (r value) among exhaled nitric oxide parameters and spirometric parameters in obese patients (always p > 0.1).

FVC (% pred) FEV1 (% pred) TLC (% pred) FRC (% pred) RV (% pred) FEV1 /VC RV/TLC

FeNO50 (ppb)

CANO (ppb)

J’awNO (pL/s)

−0.18 −0.14 −0.28 −0.16 −0.08 −0.15 −0.15

−0.03 0.03 −0.19 −0.09 −0.07 −0.07 −0.12

−0.25 −0.21 −0.08 −0.30 −0.06 −0.11 −0.05

FVC, forced vital capacity; FEV1 , forced volume at 1 s; TLC, total lung capacity; FRC, functional residual capacity; RV, residual volume.

No correlation was found among FeNO50 , CANO and J’awNO and BMI. Similarly FeNO50 , CANO and J’awNO did not correlate with either maximal static or dynamic lung volumes (Table 2).

Discussion In this study we have shown that the reduction in FeNO found in severely obese subjects as compared to normo-weighted HC and obese subjects after bariatric surgery is due to both a reduced large/central airway maximal NO flux and alveolar NO concentration. We used multiple single-breath FeNO measurements at different expiratory flow-rates, ranging from 50 to 150 mL/s to obtain additional flowindependent parameters of NO respiratory output like the large/central airway maximal NO flux and the alveolar NO concentration [2]. In agreement with theoretical [2] and experimental [14] data, the relationship between NO output and the flowrate was linear in the range of the expiratory flows used, allowing J’awNO and CANO calculation as the

intercept and the slope, respectively. J’awNO calculated with linear method shows high correlation with the value obtained with the nonlinear method [15], which, on the other hand, allows for separate calculation of diffusing NO and airway wall NO concentration. Considering axial back-diffusion of NO in the gas phase, J’awNO and CANO were calculated using the Condorelli adjustment for the axial diffusion of NO [13]. To rule out possible over-adjustment correction as that reported for lean healthy subjects [16], we obtained unadjusted values for J’awNO and CANO, which were very similar to those obtained with correction (data not shown). We are aware that our study lacks the analysis of the relationship between exhaled NO and airflow rates < 50 mL, which could have added relevant information on the NO maximal flux (rate of radial transport) in the airway compartment. Indeed, NO measurement at lower flow rate could characterize the significant decrease found at 50 mL/s. However, our aim was to analyze the effect of obesity on NO changes in the alveolar compartment, which should better estimate respiratory consequences if

Exhaled nitric oxide and severe obesity any low-grade systemic inflammation and endothelial dysfunction are present in obesity [17,18]. Our results are in contrast with data from Chow and colleagues [19], who found increased exhaled NO in obese non-asthmatic children in comparison with non-obese non-asthmatic children [19]. A possible explanation could be related to the age difference and BMI of the studied groups (children with BMI ranging between 24 and 33 kg/m2 , and adults with BMI > 40 kg/m2 ). Confirming our previous study [8] and more recent findings [9,20], we did not find any correlation between FeNO and BMI, supporting the idea that the large variation in body size or body weight has no significant effect on eosinophilic airway inflammation, and that hormones and systemic inflammation derived from adipose tissue do not affect eosinophilic airway inflammation. On the contrary, obesity is considered a chronic inflammatory state that could favour a neutrophilic airway inflammation, which, in turn, may hide/replace a normal airway eosinophilic pattern in healthy individuals, thus influencing FeNO measurements [21]. As suggested [22], the FeNO decrease in obese subjects might originate from differences in airway calibre or lung volumes between the examined groups. In fact, a significant correlation between FEV1 and FeNO in patients with low lung volumes in comparison to healthy subjects was found [22]. An animal model of continuous low-volume mechanical ventilation with physiological tidal volume lends support to the mechanical hypothesis. FeNO was found to decrease in the absence of significant change in the levels of pro-inflammatory cytokines, suggesting an abnormal mechanical shear stress related to the cycling closure and opening of peripheral airways [23]. Accordingly, in intubated healthy subjects, positive end-expiratory pressure (PEEP) increased FeNO, probably by a pulmonary recruitment effect that reduces atelectasis or altered resistance and compliance. This was also shown in rabbits, where stretch-dependent mechanisms and reduced cardiac output was postulated to cause these effects [24]. We cannot exclude that reported results might be consequence of a simple mechanical obstructive effect on airways that favours a ventilation perfusion (VA/Q) imbalance [25]. Probably obesity can cause some areas to have low/no ventilation (with airways obstruction) leading to pulmonary gas exchange abnormalities and therefore, in this case, to low FeNO values. After bariatric surgery, an improvement of VA/Q inequality has been observed [25], and this may determine a normalization of FeNO with important clinical consequences.

607 The clinical implication of a uniformly low exhaled NO in obese subjects is not known. Nitric oxide is regarded as physiological modulator of bronchial tone in airways [26]. Accordingly, obesity is associated with reduced lung volume (also linked to airway narrowing), which, at least in male, is greater than that due to reduced lung volume alone [27]. Furthermore, during sedation and paralysis, obese subjects are characterized by hypoxaemia and marked alterations of respiratory system mechanical properties, largely explained by a reduction in lung volume due to the excessive unopposed intra-abdominal pressure. Remarkably, during anaesthesia and paralysis, PEEP improves respiratory function in morbidly obese patients but not in normal subjects [28]. There is also much debate if obesity, a recognized important comorbid factor in severe asthma, may act as proinflammatory trigger or an aggravating mechanical factor limiting airway flow and lung volumes. Our findings on impaired NO large/central airway maximal NO flux in uncomplicated severe obesity, extend our previous data on limited effect of inhaled steroid on FeNO in severe obese asthmatic women with the beneficial effect on asthma control exerted by a consistent weight loss [29]. In conclusion, respiratory NO output is decreased in severe uncomplicated obesity for the reduction of both large/central airway maximal NO flux and alveolar NO concentration. The pathophysiological relevance of airway NO abnormalities in severe obese phenotype remains to be investigated. An NMR-based metabolomic study on possible discrimination of clinical obese phenotype (‘‘metabotype’’) is in progress in our lab.

Conflict of interest All the authors disclose any commercial interest that they may have in the subject of study and the source of any financial or material support.

References [1] Barnes PJ, Dweik RA, Gelb AF, Gibson PG, George SC, Grasemann H, et al. Exhaled nitric oxide in pulmonary diseases: a comprehensive review. Chest 2010;138(3):682—92. [2] Tsoukias NM, George SC. A two-compartment model of pulmonary nitric oxide exchange dynamics. J Appl Physiol (1985) 1998;85(2):653—66. [3] George SC, Hogman M, Permutt S, Silkoff PE. Modeling pulmonary nitric oxide exchange. J Appl Physiol (1985) 2004;96(3):831—9. [4] De Winter-de Groot KM, Van der Ent CK, Prins I, Tersmette JM, Uiterwaal CS. Exhaled nitric oxide: the missing link

608

[5]

[6]

[7]

[8]

[9]

[10]

[11]

[12]

[13]

[14]

[15]

[16]

M. Maniscalco et al. between asthma and obesity? J Allergy Clin Immunol 2005;115(2):419—20. Tsang KW, Ip SK, Leung R, Tipoe GL, Chan SL, Shum IH, et al. Exhaled nitric oxide: the effects of age, gender and body size. Lung 2001;179(2):83—91. Jilma B, Kastner J, Mensik C, Vondrovec B, Hildebrandt J, Krejcy K, et al. Sex differences in concentrations of exhaled nitric oxide and plasma nitrate. Life Sci 1996;58(6): 469—76. Depalo A, Carpagnano GE, Spanevello A, Sabato R, Cagnazzo MG, Gramiccioni C, et al. Exhaled NO and iNOS expression in sputum cells of healthy, obese and OSA subjects. J Intern Med 2008;263(1):70—8. Maniscalco M, de Laurentiis G, Zedda A, Faraone S, Giardiello C, Cristiano S, et al. Exhaled nitric oxide in severe obesity: effect of weight loss. Respir Physiol Neurobiol 2007;156(3):370—3. Kim SH, Kim TH, Lee JS, Koo TY, Lee CB, Yoon HJ, et al. Adiposity, adipokines, and exhaled nitric oxide in healthy adults without asthma. J Asthma 2011;48(2): 177—82. Toledo Jr JC, Augusto O. Connecting the chemical and biological properties of nitric oxide. Chem Res Toxicol 2012;25(5):975—89. Standardization of spirometry, 1994 update. American Thoracic Society. Am J Respir Crit Care Med 1995;152(3):1107—36. 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(8):912—30. 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 (1985) 2007;102(1):417—25. Maniscalco M, Vatrella A, Cremona G, Carratu L, Sofia M. Exhaled nitric oxide after inhalation of isotonic and hypotonic solutions in healthy subjects. Clin Sci (Lond) 2001;101(6):645—50. Mahut B, Delacourt C, Zerah-Lancner F, De Blic J, Harf A, Delclaux C. Increase in alveolar nitric oxide in the presence of symptoms in childhood asthma. Chest 2004;125(3):1012—8. Heijkenskjold-Rentzhog C, Nordvall L, Janson C, Borres MP, Alving K, Malinovschi A. Alveolar and exhaled NO in relation to asthma characteristics — effects of correction for axial diffusion. Allergy 2014;69(8):1102—11.

[17] Roytblat L, Rachinsky M, Fisher A, Greemberg L, Shapira Y, Douvdevani A, et al. Raised interleukin-6 levels in obese patients. Obes Res 2000;8(9):673—5. [18] Nawrocki AR, Scherer PE. The delicate balance between fat and muscle: adipokines in metabolic disease and musculoskeletal inflammation. Curr Opin Pharmacol 2004;4(3):281—9. [19] Chow JS, Leung AS, Li WW, Tse TP, Sy HY, Leung TF. Airway inflammatory and spirometric measurements in obese children. Hong Kong Med J 2009;15(5):346—52. [20] Singleton MD, Sanderson WT, Mannino DM. Body mass index, asthma and exhaled nitric oxide in U.S. adults, 2007—2010. J Asthma 2014;51(7):756—61. [21] Schneider A, Schwarzbach J, Faderl B, Welker L, KarschVolk M, Jorres RA. FENO measurement and sputum analysis for diagnosing asthma in clinical practice. Respir Med 2013;107(2):209—16. [22] Ho LP, Wood FT, Robson A, Innes JA, Greening AP. The current single exhalation method of measuring exhales nitric oxide is affected by airway calibre. Eur Respir J 2000;15(6):1009—13. [23] D’Angelo E, Pecchiari M, Della Valle P, Koutsoukou A, Milic-Emili J. Effects of mechanical ventilation at low lung volume on respiratory mechanics and nitric oxide exhalation in normal rabbits. J Appl Physiol (1985) 2005;99(2):433—44. [24] Persson MG, Lonnqvist PA, Gustafsson LE. Positive endexpiratory pressure ventilation elicits increases in endogenously formed nitric oxide as detected in air exhaled by rabbits. Anesthesiology 1995;82(4):969—74. [25] Rivas E, Arismendi E, Agusti A, Sanchez M, Delgado S, Gistau C, et al. Ventilation-perfusion distribution abnormalities in morbidly obese subjects before and after bariatric surgery. Chest 2014, http://dx.doi.org/10.1378/CHEST 14-1749. [26] Ricciardolo FL, Sterk PJ, Gaston B, Folkerts G. Nitric oxide in health and disease of the respiratory system. Physiol Rev 2004;84(3):731—65. [27] King GG, Brown NJ, Diba C, Thorpe CW, Munoz P, Marks GB, et al. The effects of body weight on airway calibre. Eur Respir J 2005;25(5):896—901. [28] Pelosi P, Luecke T, Rocco PR. Chest wall mechanics and abdominal pressure during general anaesthesia in normal and obese individuals and in acute lung injury. Curr Opin Crit Care 2011;17(1):72—9. [29] Maniscalco M, Zedda A, Faraone S, Cerbone MR, Cristiano S, Giardiello C, et al. Weight loss and asthma control in severely obese asthmatic females. Respir Med 2008;102(1):102—8.

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