- Email: [email protected]
Urine concentrations changes of cysteinyl leukotrienes in non-obese children with obstructive sleep apnea undergoing adenotonsillectomy
Accepted Manuscript Urine concentrations changes of cysteinyl leukotrienes in non-obese children with obstructive sleep apnea undergoing adenotonsillectomy George Κ. Mousailidis, MD, MSc, Vasileios A. Lachanas, MD, PhD, Afroditi Vasdeki, MD, Emmanuel I. Alexopoulos, MD, PhD, Athanasios G. Kaditis, MD, PhD, Petinaki Efthymia, MD, PhD, Nick Balatsos, PhD, John G. Bizakis, MD, PhD, Charalampos E. Skoulakis, MD, PhD PII:
S0165-5876(18)30505-6
DOI:
10.1016/j.ijporl.2018.10.002
Reference:
PEDOT 9204
To appear in:
International Journal of Pediatric Otorhinolaryngology
Received Date: 20 June 2018 Revised Date:
1 October 2018
Accepted Date: 1 October 2018
Please cite this article as: G.Κ. Mousailidis, V.A. Lachanas, A. Vasdeki, E.I. Alexopoulos, A.G. Kaditis, P. Efthymia, N. Balatsos, J.G. Bizakis, C.E. Skoulakis, Urine concentrations changes of cysteinyl leukotrienes in non-obese children with obstructive sleep apnea undergoing adenotonsillectomy, International Journal of Pediatric Otorhinolaryngology (2018), doi: https://doi.org/10.1016/ j.ijporl.2018.10.002. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT 1 Title: Urine concentrations changes of cysteinyl leukotrienes in non-obese children with obstructive sleep apnea undergoing adenotonsillectomy.
RI PT
Authors:
George Κ. Mousailidis, MD, MSca; Vasileios A. Lachanas, MD, PhDa; Afroditi Vasdeki, MDb; Emmanuel I. Alexopoulos, MD, PhDc; Athanasios G. Kaditis, MD, PhDd; Petinaki
SC
Efthymia, MD, PhDb; Nick Balatsos, PhDe; John G. Bizakis, MD, PhDa; Charalampos E.
M AN U
Skoulakis, MD, PhDa
Institutions:
a. Department of Otorhinolaryngology, University Hospital of Larissa – Greece b. Department of Microbiology, University of Thessaly, University Hospital of Larissa
TE D
c. Sleep Disorders Laboratory, University Hospital of Larissa, Greece d. First Department of Pediatrics, National and Kapodistrian University of Athens School of Medicine and Aghia Sophia Children's Hospital, Athens, Greece
EP
e. Department of Biochemistry and Biotechnology, University of Thessaly
AC C
Corresponding Author:
Vasileios A. Lachanas, MD, PhD, Consultant University Hospital of Larissa, Viopolis, PC 41110, PO Box 1425, Larissa, Greece Tel.: +30 2413 502740, Fax: +30 2413 501569, Email: [email protected]
ACCEPTED MANUSCRIPT 2 Funding source: This research was financially supported by the University of Thessaly Research Committee
RI PT
Financial disclosure: None of the authors has any financial relationships relevant to this article to disclose
SC
Conflicts of interest: None of the authors has any potential conflicts of interest to
AC C
EP
TE D
M AN U
disclose
ACCEPTED MANUSCRIPT 1 Abstract Objective: The main objective of the study was to compare preoperative to postoperative levels
of
urine-Cysteinyl
leukotrienes
(uCysLT)
in
children
undergoing
RI PT
adenotonsillectomy (AT) for obstructive sleep apnea (OSA) in order to investigate whether exaggerated leukotriene activity is the cause or consequence of OSA.
Methods and materials: A prospective study was conducted on non-obese children (4-
SC
10 years old) referred for overnight PSG. Children with moderate/severe OSA treated with AT were included. A second PSG study performed 2 months postoperatively to
M AN U
confirm OSA resolution, and those with residual OSA were excluded. Morning urine specimens after both PSG studies were obtained and pre-operative uCysLT levels were compared to postoperative levels.
Results: 27 children fulfilled the criteria and underwent a post-operative PSG study with
TE D
three exclusions for residual OSA (postop-AHI>2), so the study group consisted of 24 children (mean age: 5.7±2.1 years). Mean preoperative and postoperative AHI was 10.96±5.93 and 1.44±0.56 respectively. Mean preop-uCysLT were 21.14±4.65, while
EP
after AT they significantly reduced to 12.62±2.67 (P<0.01). Conclusion: uCysLT levels are significantly reduced after AT in non-obese children with
AC C
moderate/severe OSA, suggesting that exaggerated leukotriene activity is mainly a consequence of OSA.
ACCEPTED MANUSCRIPT 2 Keywords: Urine cysteinyl leukotriene, polysomnography, pediatric obstructive sleep
AC C
EP
TE D
M AN U
SC
RI PT
apnea, OSA, adenotonsillectomy
ACCEPTED MANUSCRIPT 3 1. Introduction Pediatric obstructive sleep apnea (OSA) is a disorder affecting 1-5% of children. It is characterized by recurrent episodes of complete and/or partial upper airway obstruction
RI PT
during sleep, resulting in intermittent hypoxemia and hypercapnia, frequent arousals, and sleep fragmentation. It has unique characteristics not seen in adults, such as hyperactivity, poor school performance and growth retardation, while it differs from adult OSA in
SC
pathophysiology, clinical presentation, polysomnographic features, and outcomes. Moreover, children with OSA are more likely to have hypertension and metabolic
M AN U
syndrome as adults, while both neuropsychological behavior and quality of life tend to normalize after airway obstruction has resolved [1-5].
Currently, overnight polysomnography (PSG) is the gold standard for diagnosis of pediatric OSA. It confirms OSA and quantifies its severity objectively that can be used to
TE D
decide proper treatment. On the other hand, it has several limitations, since it is expensive, time-consuming, extensive collaboration is required, imposes discomfort on children and their families, is relatively inaccessible to children, while it does not predict
EP
complications associated with SDB such as adverse effects on quality of life [1-6]. Therefore, investigators are trying to identify and develop novel and reliable diagnostic
AC C
tools to evaluate and diagnose OSA. These tools should be cheap and easy to use permitting expanded screening, able for accurate identification of the presence or absence of OSA with adequate sensitivity and specificity, and well correlated with OSA severity [6].
There is evidence in the literature supporting that local and systemic inflammation play a role to pediatric OSA pathogenesis, while increased inflammatory markers in the
ACCEPTED MANUSCRIPT 4 upper airway tissue were found [5-11]. Cysteinyl leukotrienes (CysLT) are key inflammatory mediators and modulators in the respiratory system, have been associated with respiratory disorders such as asthma and allergic rhinitis [12-19], and recent data
RI PT
suggest that they may play an important role in pediatric OSA pathophysiology by promoting adenotonsillar hypertrophy [19-22]. Furthermore, many studies reported increased levels of CysLT in adenotonsillar tissue, exhaled breath, blood and urine of
SC
children with OSA [14-26] and CysLT correlation with OSA severity [16-19], suggesting CysLT as a potential biomarker for pediatric OSA. It should be noted that yet, it is
M AN U
unknown whether inflammation is the cause or result of OSA, while little is known about what happens to inflammatory biomarker levels after treatment of pediatric OSA [14, 19]. Enlarged adenotonsillar tissues have emerged as the major pathophysiological contributor in children OSA, followed by obesity as the second most important [6].
TE D
Accordingly, unless contraindicated, the guidelines of the American Academy of Pediatrics recommend adenotonsillectomy (AT) as the first line of treatment if a child is determined to have OSAS and has a clinical examination consistent with adenotonsillar
EP
hypertrophy [27]. AT has a high success rate, even though sometimes its indications remain controversial and, in some cases, residual OSA remain postoperatively [14].
AC C
To our knowledge, no study until now has evaluated uCysLT levels before and
after AT on OSA children. In the present study, we hypothesized that urine CysLT (uCysLT) is a biomarker for pediatric OSA and exaggerated leukotriene activity is a result of pediatric OSA, therefore after OSA resolution with AT uCysLT levels will be significantly reduced. So, the main objective of the present study was to investigate whether the exaggerated leukotriene activity is the cause or the consequence of OSA by
ACCEPTED MANUSCRIPT 5 comparing preoperative to postoperative levels of uCysLT in children undergoing AT for OSA.
RI PT
2. Material and Methods 2.1 Study participants and setting
A prospective study was conducted on children 4-10 years old, referred to the Sleep
SC
Disorders Laboratory for overnight PSG in University Hospital of Larissa - Greece due to symptoms of obstructive sleep disordered breathing (SDB). A detailed history was taken
M AN U
and physical examination was performed, including a complete ENT examination and endoscopy. Weight and standing height were measured, and body mass index (BMI) zscore was calculated.
Children underwent a preoperative PSG study, and those non-obese (BMI z-score <
TE D
1.65) with moderate/severe OSA and adenotonsillar hypertrophy who consented to surgical therapy with AT were included. Children were operated within 4 weeks afterwards, and in order to assess OSA resolution a second PSG study performed 2
healing).
EP
months after surgery (to minimize confounding effects of post-operative swelling and
AC C
In order to have an homogeneous study group in terms of SDB pathophysiology the
following exclusion criteria were established: (1) children with a BMI z-score > 1.65 (2) prior tonsil, adenoid, cleft palate or previous pharyngeal surgery, (3) partial tonsillectomy, (4) residual OSA according to postoperative PSG study (5) Down or other syndrome or genetic disorders involving the head and neck, (6) cardiovascular disorders, (7) neuromuscular disorders, (8) symptoms or signs of asthma or respiratory tract
ACCEPTED MANUSCRIPT 6 infection, (9) symptoms or signs of other acute or chronic inflammatory disorders, (10) eczema, and (11) use of antihistamines, nasal corticosteroids, or CysLT receptor
postoperative PSG.
RI PT
inhibitors for one month prior to preoperative PSG and within the study period until
In order to compare CysLT levels all children provided a morning urine specimen immediately after preoperative and postoperative PSG studies.
SC
The study was approved by the University of Thessaly - School of Medicine Review Board (approval No. 5654/06-12-2012), and informed consent was obtained from
published study from our department. 2.2 Polysomnography
M AN U
parents or legal caretaker. No subjects of the present study were participants in a related
Children underwent overnight PSG at the University Hospital of Larisa Sleep
TE D
Disorders Laboratory. The following channels were recorded during polysomnography: EEG (F4/M1, C4/M1, O2/M1), right and left oculogram, submental and tibial electromyogram, body position, electrocardiogram, thoracic and abdominal wall motion
EP
(piezoelectric transducers), oronasal airflow (three-pronged thermistor and nasal pressure transducer), and oxygen saturation of hemoglobin. Sleep stages, arousals, and respiratory
AC C
events were scored and polysomnography indices were defined using the American Academy of Sleep Medicine manual [28, 29]. Apnea-hypopnea index (AHI) was calculated as the mean number of obstructive and mixed apneas and hypopneas per hour of total sleep time. Children with an AHI > 1 episode/h were diagnosed with OSA [30] and the degree of OSA was further classified as normal/mild (AHI: < 5), moderate (AHI: 5-10), or severe (AHI: >10). To date, no consensus exists on the threshold value of AHI
ACCEPTED MANUSCRIPT 7 to determine residual OSA in children. AHI > 1, AHI > 2 and AHI > 5 has been proposed to define residual OSA [31]. In our study we have defined a postoperative AHI > 2 as residual OSA.
RI PT
2.3. Measurement of Urine cysteinyl leukotriene
All children provided a urine specimen in the morning immediately after preoperative and postoperative PSG. Urine samples were centrifuged and the supernatant
SC
was transferred to clean test tubes, while part of each sample was used for measurement of urine concentration of creatinine. In the remaining urine, indomethacin was added at a
M AN U
concentration up to 10 µg/dL, and then the sample was frozen at -70°C until assayed. To determine CysLT concentrations, the Cysteinyl leukotriene ELISA kit (Enzo© Life Sciences, Inc.) was used. This is a competitive enzyme-linked immunosorbent assay for the quantitative determination of CysLT in urine. The kit measures three of the major
TE D
metabolites of LTA4; LTC4, LTD4 and LTE4 and uses a monoclonal antibody to the CysLT (LTC4, LTD4 and LTE4) to bind, in a competitive manner, the CysLT in the sample or an alkaline phosphatase molecule which has LTC4 covalently attached to it.
EP
The measured optical density of LTC4 is used to calculate the concentration of CysLT in the sample. The assay detection limit is 26.6 pg/mL. For the calculation of the
AC C
concentration of CysLT in the samples, our data were handled by an immunoassay software
package
utilizing
a
four-parameter
logistic
curve
fitting
program
(AssayBlaster™).
The results were normalized to the creatinine concentration determined in the same
sample and the LTC4/creatinine ratios were used for subsequent analysis. 2.4. Data analysis
ACCEPTED MANUSCRIPT 8 Data analysis was performed with the SPSS 20 statistical software (IBM, Chicago, IL, USA) and values of p< 0.05 were considered as significant results. Data were presented as mean ± SD and non-parametric Wilcoxon Signed Rank Test
RI PT
was used to compare pre- to post-operative uCysLT levels. Furthermore, potential correlations of preoperative AHI and uCysLT levels were explored by using non-
SC
parametric Spearman’s correlation coefficient.
3. Results
M AN U
Twenty-seven (27) children fulfilled the criteria and obtained a postoperative PSG study. Three children with residual OSA (postop-AHI > 2) were excluded, thus 24 children were finally included in our study group (table 1). Nine (9) were female and 15 male, mean age was 5.7 ± 2.1 years and mean zBMI was 0.28 ± 1.08.
TE D
Mean preop-AHI was 10.96 ± 5.93 and mean postop-AHI was 1.44 ± 0.56. Preoperative and postoperative mean uCysLT levels were 21.14 ± 4.65 and 12.62 ± 2.67 pg/mL per mg/dL of urine creatinine respectively (Table 2).
EP
Wilcoxon signed rank test showed significant reduction of urine CysLT levels after adenotonsillectomy (Z: -4.286; p < 0.01). There was a significant positive correlation
AC C
between preoperative AHI and uCysLT (Spearman's rho: 0.534; p < 0.01)
4. Discussion
Lately, a lot of studies are trying to identify potential biomarkers as diagnostic tools
and indicators for pediatric OSA. As biomarker is defined a ‘biological molecule found in blood, other body fluids, or tissues that is a sign of a normal or abnormal process, or of a
ACCEPTED MANUSCRIPT 9 condition or disease’ [6]. Local and systemic inflammation play an important role to pediatric OSA pathogenesis, while increased markers of inflammation in the upper airway tissue have been found [5-11]. C-reactive protein, interleukins, tumor necrosis
RI PT
factor alpha, and CysLT have been associated with inflammation in serum, urine, and exhaled breath condensate of OSA children and have been identified as potential biomarkers [10, 11, 14-26].
SC
Leukotrienes are key inflammatory mediators in respiratory system propagating upper airway inflammation. Especially CysLT (LTC4, LTD4, and LTE4) are playing an
M AN U
important role in regulation of the immune system and upper airway inflammation of SDB children [12-19]. CysLT are derived by leukocytes from arachidonic acid through the 5-lipoxygenase pathway, while increased systemic levels and local CysLT’ receptor expression in adenotonsillar tissues of OSA children has been reported [18, 20-25].
TE D
Dayyat et al [23], in an in vitro study of tonsillar tissue, reported that LTD4 stimulates adenotonsillar cell proliferation with a dose-dependent pattern. Furthermore, there are invivo studies reporting increased CysLT levels in exhaled breath [26], blood and urine of
EP
children with OSA [14, 16-18] and supporting the in-vitro findings. As a result, CysLT were suggested as an important biomarker candidate for pediatric OSA.
AC C
Kaditis et al [16] and Shen et al [17] have found increased urine CysLT levels in
OSA children compared to controls as well as positive correlation of uCysLT levels with OSA severity [16-18]. Our results also showed a significant correlation of uCysLT with AHI in non-obese children with moderate/severe OSA. On the other hand, Biyani et al [15] did not identify correlation between AHI and urinary LTE4 levels in children with
ACCEPTED MANUSCRIPT 10 SDB, but this may be due to inclusion of children with primary snoring and inflammatory conditions, such as asthma and allergic rhinitis, who were taking inhaled corticosteroids. Inconsistent results have been also reported regarding association between uCysLT
RI PT
levels and obesity. Obesity is the second most important major pathophysiological contributor in children with OSA [6] and obese OSA children also improve with AT even though they have a higher prevalence of residual OSA [31]. Kaditis et al [16] found that
SC
uCysLT levels were not correlated with the degree of adiposity, whilst Shen et al [17] reported that urinary LTE4 concentrations were significantly higher in the
M AN U
overweight/obese SDB group than the normal weight or underweight patients. Satdhabudha et al [14] also reported a potential association between obesity and urinary LTE4 levels, while obese subjects were less likely to have improvements in LTE4 levels after surgery. In our study since our main goal was to investigate if the inflammatory
TE D
mechanism is the cause or consequence of OSA in childhood, due to these inconsistent results and in order to have a homogeneous study group in terms of SDB pathophysiology we have included only non-obese children.
EP
Little is known about what happen to CysLT levels after treatment of OSA. Satdhabudha et al studied 33 children with SDB and reported that urinary LTE4 levels
AC C
significantly decrease after adenotonsillectomy, so they suggested that increased leukotriene activity is a consequence of pediatric SDB. However, the main limitation of this study was that PSG was not performed, therefore participants were not objectively diagnosed with OSA, could not be classified for OSA severity and OSA resolution could not be confirmed [14]. To our knowledge, no study so far has evaluated uCysLT levels before and after AT on children who diagnosed with OSA by means of PSG. In our
ACCEPTED MANUSCRIPT 11 study PSG was used for OSA diagnosis, staging and resolution after treatment. Moreover, we established as strict inclusion and excision criteria as possible, in order to provide a homogenous study group in terms of OSA pathophysiology. Our results showed that
RI PT
uCysLT levels of children with OSA, significantly reduce after OSA resolution with adenotonsillectomy. These results suggest that our hypothesis is true, thus that exaggerated leukotriene activity noticed in children with OSA is a consequence of OSA
SC
rather than a cause of it. Moreover, our findings further support the potential use of uCysLT as a pediatric OSA biomarker, since it seems that uCysLT levels respond to
M AN U
therapy with AT. However, more research is needed since there are fluctuations of uCysLT levels depending upon several factors, while there are no clinical standards for normal ranges and thresholds for OSA severity groups [14, 15].
Prospective design, the use of PSG to diagnose OSA and confirm its resolution, and
TE D
strict inclusion and excision criteria in order to provide a homogenous study group are the strengths of this study. There are also some limitations. The relative small sample size is the main limitation. However, there were difficulties in recruiting children especially for
EP
the postoperative PSG study due to PSG discomfort and cost. Another limitation is that we have not evaluated children with mild OSA. Mild OSA children were excluded in our
AC C
study because in this group pathophysiology and natural history are still not fully understood and surgical treatment is still controversial [15, 32].
5. Conclusion
In conclusion our study showed that uCysLT levels of non-obese children with moderate/severe OSA significantly correlate with AHI and significantly decrease after
ACCEPTED MANUSCRIPT 12 adenotonsillectomy. These findings support the concept that exaggerated leukotriene activity is a consequence of OSA. Additionally, further support uCysLT as biomarker
AC C
EP
TE D
M AN U
SC
be established as biomarkers in OSA clinical practice.
RI PT
candidates for pediatric OSA, but more research is needed in the future until uCysLT can
ACCEPTED MANUSCRIPT 13 References [1] V. Certal, E. Catumbela, J.C. Winck, I. Azevedo, A. Teixeira-Pinto, A. Costa-Pereira,
meta-analysis, Laryngoscope. 122 (2012) 2105-2114.
RI PT
Clinical assessment of pediatric obstructive sleep apnea: a systematic review and
[2] Z. Ehsan, S.L. Ishman, Pediatric Obstructive Sleep Apnea, Otolaryngol Clin North Am. 49 (2016) 1449-1464.
SC
[3] G.K. Mousailidis, V.A. Lachanas, C.E. Skoulakis, A. Sakellariou, S.T. Exarchos, A.G. Kaditis, J.G. Bizakis, Cross-cultural adaptation and validation of the Greek
M AN U
OSA-18 questionnaire in children undergoing polysomnography, Int J Pediatr Otorhinolaryngol. 78 (2014) 2097-2102.
[4] V.A. Lachanas, G.K. Mousailidis, C.E. Skoulakis, N. Papandreou, S. Exarchos, E.I. Alexopoulos, J.G. Bizakis, Validation of the Greek OSD-6 quality of life in
children
undergoing
polysomnography,
Int
J
Pediatr
TE D
questionnaire
Otorhinolaryngol. 78 (2014) 1342-1347.
[5] A.D. Goldbart, A. Tal, Inflammation and sleep disordered breathing in children: a
EP
state-of-the-art review, Pediatr. Pulmonol. 43 (12) (2008) 1151-1160. [6] D. Gozal, Serum, urine, and breath related biomarkers in the diagnosis of obstructive
AC C
sleep apnea in children: is it for real?, Curr Opin Pulm Med. 18 (2012) 561-567.
[7] D. Gozal, Sleep, sleep disorders and inflammation in children, Sleep. Med. 10 (Suppl 1) (2009) S12-S16.
[8] I. Rubinstein, Nasal inflammation in patients with obstructive sleep apnea, Laryngoscope 105 (1995) 175-177.
ACCEPTED MANUSCRIPT 14 [9] M. Sekosan, M. Zakkar, B.L. Wenig, C.P. Olopade, I. Rubinstein, Inflammation in the uvula mucosa of patients with obstructive sleep apnea, Laryngoscope 106 (1996) 1018-1020. R. Tauman, A. Ivanenko, L.M. O'Brien, D. Gozal, Plasma C-reactive protein
RI PT
[10]
levels among children with sleep-disordered breathing, Pediatrics 113 (2004) 564569.
D. Gozal, V.M. Crabtree, O. Sans Capdevila, L.A. Witcher, L. Kheirandish-
SC
[11]
Gozal, C-reactive protein, obstructive sleep apnea, and cognitive dysfunction in
[12]
M AN U
school-aged children, Am. J. Respir. Crit. Care. Med. 176 (2007) 188-193. A. Marmarinos, P. Saxoni-Papageorgiou, D. Cassimos, E. Manoussakis, C.
Tsentidis, A. Doxara, I. Paraskakis, D. Gourgiotis, Urinary leukotriene E4 levels in atopic and non atopic preschool children with recurrent episodic (viral) wheezing: a
[13]
TE D
potential marker?, J Asthma. 52 (2015) 554-559.
R. Divekar, J. Butterfield, Diagnostic utility of urinary LTE4 in asthma, allergic
rhinitis, chronic rhinosinusitis, nasal polyps, and aspirin sensitivity, J Allergy Clin
[14]
EP
Immunol Pract. 4 (2016) 665-670.
A. Satdhabudha, P. Sritipsukho, S. Manochantr, W. Chanvimalueng, U.
AC C
Chaumrattanakul, P. Chaumphol, Urine cysteinyl leukotriene levels in children with sleep disordered breathing before and after adenotonsillectomy, Int J Pediatr Otorhinolaryngol. 94 (2017) 112-116.
[15]
S. Biyani, M.J. Benson, S.C. DeShields, T.D. Cunningham, C.M. Baldassari,
Urinary Leukotriene E4 Levels in Children with Sleep-Disordered Breathing, Otolaryngol Head Neck Surg. 158 (2018) 947-951.
ACCEPTED MANUSCRIPT 15 [16]
A.G. Kaditis, E. Alexopoulos, K. Chaidas, G. Ntamagka, A. Karathanasi, I.
Tsilioni, et al., Urine concentrations of cysteinyl leukotrienes in children with obstructive sleep-disordered breathing, Chest 135 (2009) 1496-1501. Y. Shen, Z. Xu, K. Shen, Urinary leukotriene E4, obesity, and adenotonsillar
RI PT
[17]
hypertrophy in Chinese children with sleep disordered breathing, Sleep 34 (2011) 1135-1141.
Y. Shen, Z. Xu, Z. Huang, J. Xu, Q. Qin, K. Shen, Increased cysteinyl leukotriene
SC
[18]
concentration and receptor expression in tonsillar tissues of Chinese children with
[19]
M AN U
sleep-disordered breathing, Int. Immunopharmacol. 13 (2012) 371-376. K. Sunkonkit, S. Sritippayawan, M. Veeravikrom, J. Deerojanawong, N.
Prapphal, Urinary cysteinyl leukotriene E4 level and therapeutic response to montelukast in children with mild obstructive sleep apnea Asian Pac J Allergy
[20]
TE D
Immunol. 35 (2017) 233-238.
A.G. Kaditis, M.G. Ioannou, K. Chaidas, E.I. Alexopoulos, M. Apostolidou, T.
Apostolidis, et al., Cysteinyl leukotriene receptors are expressed by tonsillar T cells
[21]
EP
of children with obstructive sleep apnea, Chest 134 (2008) 324-331. M. Tsaoussoglou, L. Lianou, P. Maragozidis, S. Hatzinidolaou, M. Mavromati, N.
AC C
Orologas, et al., Cysteinyl leukotriene receptors in tonsilar B- and T-lymphocytes from children with obstructive sleep apnea, Sleep. Med. 13 (2012) 879-885.
[22]
J. Kim, R. Bhattacharjee, E. Dayyat, A.B. Snow, L. Kheirandish-Gozal, J.L.
Goldman, R.C. Li, L.D. Serpero, H.B. Clair, D. Gozal, Increased cellular proliferation and inflammatory cytokines in tonsils derived from children with obstructive sleep Apnea, Pediatr Res. 66 (2009) 423-428.
ACCEPTED MANUSCRIPT 16 [23]
E. Dayyat, L.D. Serpero, L. Kheirandish-Gozal, J.L. Goldman, A. Snow, R.
Bhattacharjee, et al., Leukotriene pathways and in vitro adenotonsillar cell proliferation in children with obstructive sleep apnea, Chest 135 (2009) 1142-1149. M. Tsaoussoglou, S. Hatzinikolaou, G.E. Baltatzis, L. Lianou, P. Maragozidis,
RI PT
[24]
N.A. Balatsos, et al., Expression of leukotriene biosynthetic enzymes in tonsillar tissue of children with obstructive sleep apnea A prospective nonrandomized study,
[25]
SC
JAMA. Otolaryngol. Head. Neck. Surg. 140 (2014) 944-950.
A.D. Goldbart, J.L. Goldman, R.C. Li, K.R. Brittian, R. Tauman, D. Gozal,
M AN U
Differential expression of cysteinyl leukotriene receptors 1 and 2 in tonsils of children with obstructive sleep apnea syndrome or recurrent infection, Chest 126 (2004) 1318. [26]
A.D. Goldbart, J. Krishna, R.C. Li, L.D. Serpero, D. Gozal, Inflammatory
TE D
mediators in exhaled breath condensate of children with obstructive sleep apnea syndrome, Chest 130 (2006) 143-148. [27]
C.L. Marcus, L.J. Brooks, K.A. Draper, D. Gozal, A.C. Halbower, J. Jones, M.S.
EP
Schechter, S.H. Sheldon, K. Spruyt, S.D. Ward, C. Lehmann, R.N. Shiffman, American Academy of Pediatrics, Diagnosis and management of childhood
AC C
obstructive sleep apnea syndrome, Pediatrics. 130 (2012) 576-584
[28]
R.B. Berry, R. Budhiraja, D.J. Gottlieb, D. Gozal, C. Iber, V.K. Kapur, C.L.
Marcus, R. Mehra, S. Parthasarathy, S.F. Quan, S. Redline, K.P. Strohl, S.L. Davidson Ward, M.M Tangredi, American Academy of Sleep M, Rules for scoring respiratory events in sleep: update of the 2007 AASM Manual for the Scoring of Sleep and Associated Events. Deliberations of the Sleep Apnea Definitions Task
ACCEPTED MANUSCRIPT 17 Force of the American Academy of Sleep Medicine, J Clin Sleep Med. 8 (2012) 597619. [29]
C. Iber, S. Ancoli-Israel, A. Chesson, S.F. Quan, for the American Academy of
RI PT
Sleep Medicine. The AASM Manual for the Scoring of Sleep and Associated Events: Rules, Terminology and Technical Specifications, 1st edition. City: American Academy of Sleep Medicine, 2007.
M.H. Wagner, D.M. Torrez. Interpretation of the polysomnogram in children.
Otolaryngol Clin North Am. 40 (2007) 745-759.
M. Imanguli, S.O Ulualp, Risk factors for residual obstructive sleep apnea after
M AN U
[31]
SC
[30]
adenotonsillectomy in children, Laryngoscope. 126 (2016) 2624-2629. [32]
C.M. Baldassari, S. Choi, Mild Obstructive Sleep Apnea in Children: What is the
AC C
EP
TE D
Best Management Option?, Laryngoscope. (2018) doi: 10.1002/lary.27157
ACCEPTED MANUSCRIPT 1
preoperative AHI
preoperative uCysLT
postoperative AHI
postoperative uCysLT
1
5,90
17,33
0,50
9,25
2
7,30
18,80
1,30
3
11,40
20,27
2,00
4
17,30
26,05
2,00
5
15,50
19,68
0,40
6
16,00
22,12
7
8,60
23,95
8
6,60
16,17
2,00
9,45
9
6,00
20,67
1,80
10,15
10
7,30
19,15
1,50
10,67
11
6,20
20,22
1,80
10,51
12
22,10
28,05
1,30
18,07
13
7,80
19,39
1,60
13,85
14
7,20
20,99
1,50
12,46
15
5,70
17,39
2,00
8,78
16
8,20
2,00
13,17
17
EP
16,79
16,00
24,57
2,00
16,41
18
5,60
14,26
0,90
6,76
19
21,80
34,29
2,00
15,39
20
7,50
19,15
1,20
12,60
21
12,20
20,71
0,70
15,31
22
7,00
29,89
0,60
13,00
23
25,40
17,55
2,00
11,05
24
8,50
19,82
0,60
15,49
SC
RI PT
patient
AC C
Table 1. Polysomnography data and urinary findings each Study Group patient.
13,53 13,91 13,50
1,30
14,15
1,50
13,58
M AN U
TE D
11,92
ACCEPTED MANUSCRIPT 2
Abbreviations apnea-hypopnea index
uCysLT
urine Cysteinyl leukotrienes (pg/mL per mg/dL of urine creatinine)
AC C
EP
TE D
M AN U
SC
RI PT
AHI
ACCEPTED MANUSCRIPT 1 Table 2. Characteristics of Study Group (means ± SD) Study group: n=24 (9 female /15 male)
age
5.7 ± 2.1 years
BMI z-score
0.28 ± 1.08
preoperative AHI
10.96 ± 5.93 episodes/h
postoperative AHI
1.44 ± 0.56 episodes/h
preoperative uCysLT
21.14 ± 4.65 pg/mL per mg/dL of urine creatinine
SC
RI PT
Variable
Abbreviations
M AN U
postoperative uCysLT 12.62 ± 2.67 pg/mL per mg/dL of urine creatinine
apnea-hypopnea index
BMI
body mass index
uCysLT
urine Cysteinyl leukotrienes
AC C
EP
TE D
AHI
S0165-5876(18)30505-6
DOI:
10.1016/j.ijporl.2018.10.002
Reference:
PEDOT 9204
To appear in:
International Journal of Pediatric Otorhinolaryngology
Received Date: 20 June 2018 Revised Date:
1 October 2018
Accepted Date: 1 October 2018
Please cite this article as: G.Κ. Mousailidis, V.A. Lachanas, A. Vasdeki, E.I. Alexopoulos, A.G. Kaditis, P. Efthymia, N. Balatsos, J.G. Bizakis, C.E. Skoulakis, Urine concentrations changes of cysteinyl leukotrienes in non-obese children with obstructive sleep apnea undergoing adenotonsillectomy, International Journal of Pediatric Otorhinolaryngology (2018), doi: https://doi.org/10.1016/ j.ijporl.2018.10.002. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT 1 Title: Urine concentrations changes of cysteinyl leukotrienes in non-obese children with obstructive sleep apnea undergoing adenotonsillectomy.
RI PT
Authors:
George Κ. Mousailidis, MD, MSca; Vasileios A. Lachanas, MD, PhDa; Afroditi Vasdeki, MDb; Emmanuel I. Alexopoulos, MD, PhDc; Athanasios G. Kaditis, MD, PhDd; Petinaki
SC
Efthymia, MD, PhDb; Nick Balatsos, PhDe; John G. Bizakis, MD, PhDa; Charalampos E.
M AN U
Skoulakis, MD, PhDa
Institutions:
a. Department of Otorhinolaryngology, University Hospital of Larissa – Greece b. Department of Microbiology, University of Thessaly, University Hospital of Larissa
TE D
c. Sleep Disorders Laboratory, University Hospital of Larissa, Greece d. First Department of Pediatrics, National and Kapodistrian University of Athens School of Medicine and Aghia Sophia Children's Hospital, Athens, Greece
EP
e. Department of Biochemistry and Biotechnology, University of Thessaly
AC C
Corresponding Author:
Vasileios A. Lachanas, MD, PhD, Consultant University Hospital of Larissa, Viopolis, PC 41110, PO Box 1425, Larissa, Greece Tel.: +30 2413 502740, Fax: +30 2413 501569, Email: [email protected]
ACCEPTED MANUSCRIPT 2 Funding source: This research was financially supported by the University of Thessaly Research Committee
RI PT
Financial disclosure: None of the authors has any financial relationships relevant to this article to disclose
SC
Conflicts of interest: None of the authors has any potential conflicts of interest to
AC C
EP
TE D
M AN U
disclose
ACCEPTED MANUSCRIPT 1 Abstract Objective: The main objective of the study was to compare preoperative to postoperative levels
of
urine-Cysteinyl
leukotrienes
(uCysLT)
in
children
undergoing
RI PT
adenotonsillectomy (AT) for obstructive sleep apnea (OSA) in order to investigate whether exaggerated leukotriene activity is the cause or consequence of OSA.
Methods and materials: A prospective study was conducted on non-obese children (4-
SC
10 years old) referred for overnight PSG. Children with moderate/severe OSA treated with AT were included. A second PSG study performed 2 months postoperatively to
M AN U
confirm OSA resolution, and those with residual OSA were excluded. Morning urine specimens after both PSG studies were obtained and pre-operative uCysLT levels were compared to postoperative levels.
Results: 27 children fulfilled the criteria and underwent a post-operative PSG study with
TE D
three exclusions for residual OSA (postop-AHI>2), so the study group consisted of 24 children (mean age: 5.7±2.1 years). Mean preoperative and postoperative AHI was 10.96±5.93 and 1.44±0.56 respectively. Mean preop-uCysLT were 21.14±4.65, while
EP
after AT they significantly reduced to 12.62±2.67 (P<0.01). Conclusion: uCysLT levels are significantly reduced after AT in non-obese children with
AC C
moderate/severe OSA, suggesting that exaggerated leukotriene activity is mainly a consequence of OSA.
ACCEPTED MANUSCRIPT 2 Keywords: Urine cysteinyl leukotriene, polysomnography, pediatric obstructive sleep
AC C
EP
TE D
M AN U
SC
RI PT
apnea, OSA, adenotonsillectomy
ACCEPTED MANUSCRIPT 3 1. Introduction Pediatric obstructive sleep apnea (OSA) is a disorder affecting 1-5% of children. It is characterized by recurrent episodes of complete and/or partial upper airway obstruction
RI PT
during sleep, resulting in intermittent hypoxemia and hypercapnia, frequent arousals, and sleep fragmentation. It has unique characteristics not seen in adults, such as hyperactivity, poor school performance and growth retardation, while it differs from adult OSA in
SC
pathophysiology, clinical presentation, polysomnographic features, and outcomes. Moreover, children with OSA are more likely to have hypertension and metabolic
M AN U
syndrome as adults, while both neuropsychological behavior and quality of life tend to normalize after airway obstruction has resolved [1-5].
Currently, overnight polysomnography (PSG) is the gold standard for diagnosis of pediatric OSA. It confirms OSA and quantifies its severity objectively that can be used to
TE D
decide proper treatment. On the other hand, it has several limitations, since it is expensive, time-consuming, extensive collaboration is required, imposes discomfort on children and their families, is relatively inaccessible to children, while it does not predict
EP
complications associated with SDB such as adverse effects on quality of life [1-6]. Therefore, investigators are trying to identify and develop novel and reliable diagnostic
AC C
tools to evaluate and diagnose OSA. These tools should be cheap and easy to use permitting expanded screening, able for accurate identification of the presence or absence of OSA with adequate sensitivity and specificity, and well correlated with OSA severity [6].
There is evidence in the literature supporting that local and systemic inflammation play a role to pediatric OSA pathogenesis, while increased inflammatory markers in the
ACCEPTED MANUSCRIPT 4 upper airway tissue were found [5-11]. Cysteinyl leukotrienes (CysLT) are key inflammatory mediators and modulators in the respiratory system, have been associated with respiratory disorders such as asthma and allergic rhinitis [12-19], and recent data
RI PT
suggest that they may play an important role in pediatric OSA pathophysiology by promoting adenotonsillar hypertrophy [19-22]. Furthermore, many studies reported increased levels of CysLT in adenotonsillar tissue, exhaled breath, blood and urine of
SC
children with OSA [14-26] and CysLT correlation with OSA severity [16-19], suggesting CysLT as a potential biomarker for pediatric OSA. It should be noted that yet, it is
M AN U
unknown whether inflammation is the cause or result of OSA, while little is known about what happens to inflammatory biomarker levels after treatment of pediatric OSA [14, 19]. Enlarged adenotonsillar tissues have emerged as the major pathophysiological contributor in children OSA, followed by obesity as the second most important [6].
TE D
Accordingly, unless contraindicated, the guidelines of the American Academy of Pediatrics recommend adenotonsillectomy (AT) as the first line of treatment if a child is determined to have OSAS and has a clinical examination consistent with adenotonsillar
EP
hypertrophy [27]. AT has a high success rate, even though sometimes its indications remain controversial and, in some cases, residual OSA remain postoperatively [14].
AC C
To our knowledge, no study until now has evaluated uCysLT levels before and
after AT on OSA children. In the present study, we hypothesized that urine CysLT (uCysLT) is a biomarker for pediatric OSA and exaggerated leukotriene activity is a result of pediatric OSA, therefore after OSA resolution with AT uCysLT levels will be significantly reduced. So, the main objective of the present study was to investigate whether the exaggerated leukotriene activity is the cause or the consequence of OSA by
ACCEPTED MANUSCRIPT 5 comparing preoperative to postoperative levels of uCysLT in children undergoing AT for OSA.
RI PT
2. Material and Methods 2.1 Study participants and setting
A prospective study was conducted on children 4-10 years old, referred to the Sleep
SC
Disorders Laboratory for overnight PSG in University Hospital of Larissa - Greece due to symptoms of obstructive sleep disordered breathing (SDB). A detailed history was taken
M AN U
and physical examination was performed, including a complete ENT examination and endoscopy. Weight and standing height were measured, and body mass index (BMI) zscore was calculated.
Children underwent a preoperative PSG study, and those non-obese (BMI z-score <
TE D
1.65) with moderate/severe OSA and adenotonsillar hypertrophy who consented to surgical therapy with AT were included. Children were operated within 4 weeks afterwards, and in order to assess OSA resolution a second PSG study performed 2
healing).
EP
months after surgery (to minimize confounding effects of post-operative swelling and
AC C
In order to have an homogeneous study group in terms of SDB pathophysiology the
following exclusion criteria were established: (1) children with a BMI z-score > 1.65 (2) prior tonsil, adenoid, cleft palate or previous pharyngeal surgery, (3) partial tonsillectomy, (4) residual OSA according to postoperative PSG study (5) Down or other syndrome or genetic disorders involving the head and neck, (6) cardiovascular disorders, (7) neuromuscular disorders, (8) symptoms or signs of asthma or respiratory tract
ACCEPTED MANUSCRIPT 6 infection, (9) symptoms or signs of other acute or chronic inflammatory disorders, (10) eczema, and (11) use of antihistamines, nasal corticosteroids, or CysLT receptor
postoperative PSG.
RI PT
inhibitors for one month prior to preoperative PSG and within the study period until
In order to compare CysLT levels all children provided a morning urine specimen immediately after preoperative and postoperative PSG studies.
SC
The study was approved by the University of Thessaly - School of Medicine Review Board (approval No. 5654/06-12-2012), and informed consent was obtained from
published study from our department. 2.2 Polysomnography
M AN U
parents or legal caretaker. No subjects of the present study were participants in a related
Children underwent overnight PSG at the University Hospital of Larisa Sleep
TE D
Disorders Laboratory. The following channels were recorded during polysomnography: EEG (F4/M1, C4/M1, O2/M1), right and left oculogram, submental and tibial electromyogram, body position, electrocardiogram, thoracic and abdominal wall motion
EP
(piezoelectric transducers), oronasal airflow (three-pronged thermistor and nasal pressure transducer), and oxygen saturation of hemoglobin. Sleep stages, arousals, and respiratory
AC C
events were scored and polysomnography indices were defined using the American Academy of Sleep Medicine manual [28, 29]. Apnea-hypopnea index (AHI) was calculated as the mean number of obstructive and mixed apneas and hypopneas per hour of total sleep time. Children with an AHI > 1 episode/h were diagnosed with OSA [30] and the degree of OSA was further classified as normal/mild (AHI: < 5), moderate (AHI: 5-10), or severe (AHI: >10). To date, no consensus exists on the threshold value of AHI
ACCEPTED MANUSCRIPT 7 to determine residual OSA in children. AHI > 1, AHI > 2 and AHI > 5 has been proposed to define residual OSA [31]. In our study we have defined a postoperative AHI > 2 as residual OSA.
RI PT
2.3. Measurement of Urine cysteinyl leukotriene
All children provided a urine specimen in the morning immediately after preoperative and postoperative PSG. Urine samples were centrifuged and the supernatant
SC
was transferred to clean test tubes, while part of each sample was used for measurement of urine concentration of creatinine. In the remaining urine, indomethacin was added at a
M AN U
concentration up to 10 µg/dL, and then the sample was frozen at -70°C until assayed. To determine CysLT concentrations, the Cysteinyl leukotriene ELISA kit (Enzo© Life Sciences, Inc.) was used. This is a competitive enzyme-linked immunosorbent assay for the quantitative determination of CysLT in urine. The kit measures three of the major
TE D
metabolites of LTA4; LTC4, LTD4 and LTE4 and uses a monoclonal antibody to the CysLT (LTC4, LTD4 and LTE4) to bind, in a competitive manner, the CysLT in the sample or an alkaline phosphatase molecule which has LTC4 covalently attached to it.
EP
The measured optical density of LTC4 is used to calculate the concentration of CysLT in the sample. The assay detection limit is 26.6 pg/mL. For the calculation of the
AC C
concentration of CysLT in the samples, our data were handled by an immunoassay software
package
utilizing
a
four-parameter
logistic
curve
fitting
program
(AssayBlaster™).
The results were normalized to the creatinine concentration determined in the same
sample and the LTC4/creatinine ratios were used for subsequent analysis. 2.4. Data analysis
ACCEPTED MANUSCRIPT 8 Data analysis was performed with the SPSS 20 statistical software (IBM, Chicago, IL, USA) and values of p< 0.05 were considered as significant results. Data were presented as mean ± SD and non-parametric Wilcoxon Signed Rank Test
RI PT
was used to compare pre- to post-operative uCysLT levels. Furthermore, potential correlations of preoperative AHI and uCysLT levels were explored by using non-
SC
parametric Spearman’s correlation coefficient.
3. Results
M AN U
Twenty-seven (27) children fulfilled the criteria and obtained a postoperative PSG study. Three children with residual OSA (postop-AHI > 2) were excluded, thus 24 children were finally included in our study group (table 1). Nine (9) were female and 15 male, mean age was 5.7 ± 2.1 years and mean zBMI was 0.28 ± 1.08.
TE D
Mean preop-AHI was 10.96 ± 5.93 and mean postop-AHI was 1.44 ± 0.56. Preoperative and postoperative mean uCysLT levels were 21.14 ± 4.65 and 12.62 ± 2.67 pg/mL per mg/dL of urine creatinine respectively (Table 2).
EP
Wilcoxon signed rank test showed significant reduction of urine CysLT levels after adenotonsillectomy (Z: -4.286; p < 0.01). There was a significant positive correlation
AC C
between preoperative AHI and uCysLT (Spearman's rho: 0.534; p < 0.01)
4. Discussion
Lately, a lot of studies are trying to identify potential biomarkers as diagnostic tools
and indicators for pediatric OSA. As biomarker is defined a ‘biological molecule found in blood, other body fluids, or tissues that is a sign of a normal or abnormal process, or of a
ACCEPTED MANUSCRIPT 9 condition or disease’ [6]. Local and systemic inflammation play an important role to pediatric OSA pathogenesis, while increased markers of inflammation in the upper airway tissue have been found [5-11]. C-reactive protein, interleukins, tumor necrosis
RI PT
factor alpha, and CysLT have been associated with inflammation in serum, urine, and exhaled breath condensate of OSA children and have been identified as potential biomarkers [10, 11, 14-26].
SC
Leukotrienes are key inflammatory mediators in respiratory system propagating upper airway inflammation. Especially CysLT (LTC4, LTD4, and LTE4) are playing an
M AN U
important role in regulation of the immune system and upper airway inflammation of SDB children [12-19]. CysLT are derived by leukocytes from arachidonic acid through the 5-lipoxygenase pathway, while increased systemic levels and local CysLT’ receptor expression in adenotonsillar tissues of OSA children has been reported [18, 20-25].
TE D
Dayyat et al [23], in an in vitro study of tonsillar tissue, reported that LTD4 stimulates adenotonsillar cell proliferation with a dose-dependent pattern. Furthermore, there are invivo studies reporting increased CysLT levels in exhaled breath [26], blood and urine of
EP
children with OSA [14, 16-18] and supporting the in-vitro findings. As a result, CysLT were suggested as an important biomarker candidate for pediatric OSA.
AC C
Kaditis et al [16] and Shen et al [17] have found increased urine CysLT levels in
OSA children compared to controls as well as positive correlation of uCysLT levels with OSA severity [16-18]. Our results also showed a significant correlation of uCysLT with AHI in non-obese children with moderate/severe OSA. On the other hand, Biyani et al [15] did not identify correlation between AHI and urinary LTE4 levels in children with
ACCEPTED MANUSCRIPT 10 SDB, but this may be due to inclusion of children with primary snoring and inflammatory conditions, such as asthma and allergic rhinitis, who were taking inhaled corticosteroids. Inconsistent results have been also reported regarding association between uCysLT
RI PT
levels and obesity. Obesity is the second most important major pathophysiological contributor in children with OSA [6] and obese OSA children also improve with AT even though they have a higher prevalence of residual OSA [31]. Kaditis et al [16] found that
SC
uCysLT levels were not correlated with the degree of adiposity, whilst Shen et al [17] reported that urinary LTE4 concentrations were significantly higher in the
M AN U
overweight/obese SDB group than the normal weight or underweight patients. Satdhabudha et al [14] also reported a potential association between obesity and urinary LTE4 levels, while obese subjects were less likely to have improvements in LTE4 levels after surgery. In our study since our main goal was to investigate if the inflammatory
TE D
mechanism is the cause or consequence of OSA in childhood, due to these inconsistent results and in order to have a homogeneous study group in terms of SDB pathophysiology we have included only non-obese children.
EP
Little is known about what happen to CysLT levels after treatment of OSA. Satdhabudha et al studied 33 children with SDB and reported that urinary LTE4 levels
AC C
significantly decrease after adenotonsillectomy, so they suggested that increased leukotriene activity is a consequence of pediatric SDB. However, the main limitation of this study was that PSG was not performed, therefore participants were not objectively diagnosed with OSA, could not be classified for OSA severity and OSA resolution could not be confirmed [14]. To our knowledge, no study so far has evaluated uCysLT levels before and after AT on children who diagnosed with OSA by means of PSG. In our
ACCEPTED MANUSCRIPT 11 study PSG was used for OSA diagnosis, staging and resolution after treatment. Moreover, we established as strict inclusion and excision criteria as possible, in order to provide a homogenous study group in terms of OSA pathophysiology. Our results showed that
RI PT
uCysLT levels of children with OSA, significantly reduce after OSA resolution with adenotonsillectomy. These results suggest that our hypothesis is true, thus that exaggerated leukotriene activity noticed in children with OSA is a consequence of OSA
SC
rather than a cause of it. Moreover, our findings further support the potential use of uCysLT as a pediatric OSA biomarker, since it seems that uCysLT levels respond to
M AN U
therapy with AT. However, more research is needed since there are fluctuations of uCysLT levels depending upon several factors, while there are no clinical standards for normal ranges and thresholds for OSA severity groups [14, 15].
Prospective design, the use of PSG to diagnose OSA and confirm its resolution, and
TE D
strict inclusion and excision criteria in order to provide a homogenous study group are the strengths of this study. There are also some limitations. The relative small sample size is the main limitation. However, there were difficulties in recruiting children especially for
EP
the postoperative PSG study due to PSG discomfort and cost. Another limitation is that we have not evaluated children with mild OSA. Mild OSA children were excluded in our
AC C
study because in this group pathophysiology and natural history are still not fully understood and surgical treatment is still controversial [15, 32].
5. Conclusion
In conclusion our study showed that uCysLT levels of non-obese children with moderate/severe OSA significantly correlate with AHI and significantly decrease after
ACCEPTED MANUSCRIPT 12 adenotonsillectomy. These findings support the concept that exaggerated leukotriene activity is a consequence of OSA. Additionally, further support uCysLT as biomarker
AC C
EP
TE D
M AN U
SC
be established as biomarkers in OSA clinical practice.
RI PT
candidates for pediatric OSA, but more research is needed in the future until uCysLT can
ACCEPTED MANUSCRIPT 13 References [1] V. Certal, E. Catumbela, J.C. Winck, I. Azevedo, A. Teixeira-Pinto, A. Costa-Pereira,
meta-analysis, Laryngoscope. 122 (2012) 2105-2114.
RI PT
Clinical assessment of pediatric obstructive sleep apnea: a systematic review and
[2] Z. Ehsan, S.L. Ishman, Pediatric Obstructive Sleep Apnea, Otolaryngol Clin North Am. 49 (2016) 1449-1464.
SC
[3] G.K. Mousailidis, V.A. Lachanas, C.E. Skoulakis, A. Sakellariou, S.T. Exarchos, A.G. Kaditis, J.G. Bizakis, Cross-cultural adaptation and validation of the Greek
M AN U
OSA-18 questionnaire in children undergoing polysomnography, Int J Pediatr Otorhinolaryngol. 78 (2014) 2097-2102.
[4] V.A. Lachanas, G.K. Mousailidis, C.E. Skoulakis, N. Papandreou, S. Exarchos, E.I. Alexopoulos, J.G. Bizakis, Validation of the Greek OSD-6 quality of life in
children
undergoing
polysomnography,
Int
J
Pediatr
TE D
questionnaire
Otorhinolaryngol. 78 (2014) 1342-1347.
[5] A.D. Goldbart, A. Tal, Inflammation and sleep disordered breathing in children: a
EP
state-of-the-art review, Pediatr. Pulmonol. 43 (12) (2008) 1151-1160. [6] D. Gozal, Serum, urine, and breath related biomarkers in the diagnosis of obstructive
AC C
sleep apnea in children: is it for real?, Curr Opin Pulm Med. 18 (2012) 561-567.
[7] D. Gozal, Sleep, sleep disorders and inflammation in children, Sleep. Med. 10 (Suppl 1) (2009) S12-S16.
[8] I. Rubinstein, Nasal inflammation in patients with obstructive sleep apnea, Laryngoscope 105 (1995) 175-177.
ACCEPTED MANUSCRIPT 14 [9] M. Sekosan, M. Zakkar, B.L. Wenig, C.P. Olopade, I. Rubinstein, Inflammation in the uvula mucosa of patients with obstructive sleep apnea, Laryngoscope 106 (1996) 1018-1020. R. Tauman, A. Ivanenko, L.M. O'Brien, D. Gozal, Plasma C-reactive protein
RI PT
[10]
levels among children with sleep-disordered breathing, Pediatrics 113 (2004) 564569.
D. Gozal, V.M. Crabtree, O. Sans Capdevila, L.A. Witcher, L. Kheirandish-
SC
[11]
Gozal, C-reactive protein, obstructive sleep apnea, and cognitive dysfunction in
[12]
M AN U
school-aged children, Am. J. Respir. Crit. Care. Med. 176 (2007) 188-193. A. Marmarinos, P. Saxoni-Papageorgiou, D. Cassimos, E. Manoussakis, C.
Tsentidis, A. Doxara, I. Paraskakis, D. Gourgiotis, Urinary leukotriene E4 levels in atopic and non atopic preschool children with recurrent episodic (viral) wheezing: a
[13]
TE D
potential marker?, J Asthma. 52 (2015) 554-559.
R. Divekar, J. Butterfield, Diagnostic utility of urinary LTE4 in asthma, allergic
rhinitis, chronic rhinosinusitis, nasal polyps, and aspirin sensitivity, J Allergy Clin
[14]
EP
Immunol Pract. 4 (2016) 665-670.
A. Satdhabudha, P. Sritipsukho, S. Manochantr, W. Chanvimalueng, U.
AC C
Chaumrattanakul, P. Chaumphol, Urine cysteinyl leukotriene levels in children with sleep disordered breathing before and after adenotonsillectomy, Int J Pediatr Otorhinolaryngol. 94 (2017) 112-116.
[15]
S. Biyani, M.J. Benson, S.C. DeShields, T.D. Cunningham, C.M. Baldassari,
Urinary Leukotriene E4 Levels in Children with Sleep-Disordered Breathing, Otolaryngol Head Neck Surg. 158 (2018) 947-951.
ACCEPTED MANUSCRIPT 15 [16]
A.G. Kaditis, E. Alexopoulos, K. Chaidas, G. Ntamagka, A. Karathanasi, I.
Tsilioni, et al., Urine concentrations of cysteinyl leukotrienes in children with obstructive sleep-disordered breathing, Chest 135 (2009) 1496-1501. Y. Shen, Z. Xu, K. Shen, Urinary leukotriene E4, obesity, and adenotonsillar
RI PT
[17]
hypertrophy in Chinese children with sleep disordered breathing, Sleep 34 (2011) 1135-1141.
Y. Shen, Z. Xu, Z. Huang, J. Xu, Q. Qin, K. Shen, Increased cysteinyl leukotriene
SC
[18]
concentration and receptor expression in tonsillar tissues of Chinese children with
[19]
M AN U
sleep-disordered breathing, Int. Immunopharmacol. 13 (2012) 371-376. K. Sunkonkit, S. Sritippayawan, M. Veeravikrom, J. Deerojanawong, N.
Prapphal, Urinary cysteinyl leukotriene E4 level and therapeutic response to montelukast in children with mild obstructive sleep apnea Asian Pac J Allergy
[20]
TE D
Immunol. 35 (2017) 233-238.
A.G. Kaditis, M.G. Ioannou, K. Chaidas, E.I. Alexopoulos, M. Apostolidou, T.
Apostolidis, et al., Cysteinyl leukotriene receptors are expressed by tonsillar T cells
[21]
EP
of children with obstructive sleep apnea, Chest 134 (2008) 324-331. M. Tsaoussoglou, L. Lianou, P. Maragozidis, S. Hatzinidolaou, M. Mavromati, N.
AC C
Orologas, et al., Cysteinyl leukotriene receptors in tonsilar B- and T-lymphocytes from children with obstructive sleep apnea, Sleep. Med. 13 (2012) 879-885.
[22]
J. Kim, R. Bhattacharjee, E. Dayyat, A.B. Snow, L. Kheirandish-Gozal, J.L.
Goldman, R.C. Li, L.D. Serpero, H.B. Clair, D. Gozal, Increased cellular proliferation and inflammatory cytokines in tonsils derived from children with obstructive sleep Apnea, Pediatr Res. 66 (2009) 423-428.
ACCEPTED MANUSCRIPT 16 [23]
E. Dayyat, L.D. Serpero, L. Kheirandish-Gozal, J.L. Goldman, A. Snow, R.
Bhattacharjee, et al., Leukotriene pathways and in vitro adenotonsillar cell proliferation in children with obstructive sleep apnea, Chest 135 (2009) 1142-1149. M. Tsaoussoglou, S. Hatzinikolaou, G.E. Baltatzis, L. Lianou, P. Maragozidis,
RI PT
[24]
N.A. Balatsos, et al., Expression of leukotriene biosynthetic enzymes in tonsillar tissue of children with obstructive sleep apnea A prospective nonrandomized study,
[25]
SC
JAMA. Otolaryngol. Head. Neck. Surg. 140 (2014) 944-950.
A.D. Goldbart, J.L. Goldman, R.C. Li, K.R. Brittian, R. Tauman, D. Gozal,
M AN U
Differential expression of cysteinyl leukotriene receptors 1 and 2 in tonsils of children with obstructive sleep apnea syndrome or recurrent infection, Chest 126 (2004) 1318. [26]
A.D. Goldbart, J. Krishna, R.C. Li, L.D. Serpero, D. Gozal, Inflammatory
TE D
mediators in exhaled breath condensate of children with obstructive sleep apnea syndrome, Chest 130 (2006) 143-148. [27]
C.L. Marcus, L.J. Brooks, K.A. Draper, D. Gozal, A.C. Halbower, J. Jones, M.S.
EP
Schechter, S.H. Sheldon, K. Spruyt, S.D. Ward, C. Lehmann, R.N. Shiffman, American Academy of Pediatrics, Diagnosis and management of childhood
AC C
obstructive sleep apnea syndrome, Pediatrics. 130 (2012) 576-584
[28]
R.B. Berry, R. Budhiraja, D.J. Gottlieb, D. Gozal, C. Iber, V.K. Kapur, C.L.
Marcus, R. Mehra, S. Parthasarathy, S.F. Quan, S. Redline, K.P. Strohl, S.L. Davidson Ward, M.M Tangredi, American Academy of Sleep M, Rules for scoring respiratory events in sleep: update of the 2007 AASM Manual for the Scoring of Sleep and Associated Events. Deliberations of the Sleep Apnea Definitions Task
ACCEPTED MANUSCRIPT 17 Force of the American Academy of Sleep Medicine, J Clin Sleep Med. 8 (2012) 597619. [29]
C. Iber, S. Ancoli-Israel, A. Chesson, S.F. Quan, for the American Academy of
RI PT
Sleep Medicine. The AASM Manual for the Scoring of Sleep and Associated Events: Rules, Terminology and Technical Specifications, 1st edition. City: American Academy of Sleep Medicine, 2007.
M.H. Wagner, D.M. Torrez. Interpretation of the polysomnogram in children.
Otolaryngol Clin North Am. 40 (2007) 745-759.
M. Imanguli, S.O Ulualp, Risk factors for residual obstructive sleep apnea after
M AN U
[31]
SC
[30]
adenotonsillectomy in children, Laryngoscope. 126 (2016) 2624-2629. [32]
C.M. Baldassari, S. Choi, Mild Obstructive Sleep Apnea in Children: What is the
AC C
EP
TE D
Best Management Option?, Laryngoscope. (2018) doi: 10.1002/lary.27157
ACCEPTED MANUSCRIPT 1
preoperative AHI
preoperative uCysLT
postoperative AHI
postoperative uCysLT
1
5,90
17,33
0,50
9,25
2
7,30
18,80
1,30
3
11,40
20,27
2,00
4
17,30
26,05
2,00
5
15,50
19,68
0,40
6
16,00
22,12
7
8,60
23,95
8
6,60
16,17
2,00
9,45
9
6,00
20,67
1,80
10,15
10
7,30
19,15
1,50
10,67
11
6,20
20,22
1,80
10,51
12
22,10
28,05
1,30
18,07
13
7,80
19,39
1,60
13,85
14
7,20
20,99
1,50
12,46
15
5,70
17,39
2,00
8,78
16
8,20
2,00
13,17
17
EP
16,79
16,00
24,57
2,00
16,41
18
5,60
14,26
0,90
6,76
19
21,80
34,29
2,00
15,39
20
7,50
19,15
1,20
12,60
21
12,20
20,71
0,70
15,31
22
7,00
29,89
0,60
13,00
23
25,40
17,55
2,00
11,05
24
8,50
19,82
0,60
15,49
SC
RI PT
patient
AC C
Table 1. Polysomnography data and urinary findings each Study Group patient.
13,53 13,91 13,50
1,30
14,15
1,50
13,58
M AN U
TE D
11,92
ACCEPTED MANUSCRIPT 2
Abbreviations apnea-hypopnea index
uCysLT
urine Cysteinyl leukotrienes (pg/mL per mg/dL of urine creatinine)
AC C
EP
TE D
M AN U
SC
RI PT
AHI
ACCEPTED MANUSCRIPT 1 Table 2. Characteristics of Study Group (means ± SD) Study group: n=24 (9 female /15 male)
age
5.7 ± 2.1 years
BMI z-score
0.28 ± 1.08
preoperative AHI
10.96 ± 5.93 episodes/h
postoperative AHI
1.44 ± 0.56 episodes/h
preoperative uCysLT
21.14 ± 4.65 pg/mL per mg/dL of urine creatinine
SC
RI PT
Variable
Abbreviations
M AN U
postoperative uCysLT 12.62 ± 2.67 pg/mL per mg/dL of urine creatinine
apnea-hypopnea index
BMI
body mass index
uCysLT
urine Cysteinyl leukotrienes
AC C
EP
TE D
AHI