Sleep spindles are reduced in children with Down syndrome and sleep-disordered breathing

Background Children with Down syndrome (DS) are at increased risk of sleep-disordered breathing (SDB). We investigated sleep spindle activity, as a marker of sleep quality, and its relationship with daytime functioning in children with DS compared to typically developing (TD) children. Methods Children with DS and SDB (n = 44) and TD children matched for age, sex and SDB severity underwent overnight polysomnography. Fast or Slow sleep spindles were identified manually during N2/N3 sleep. Spindle activity was characterized as spindle number, density (number of spindles/h) and intensity (density × average duration) on central (C) and frontal (F) electrodes. Parents completed the Child Behavior Check List and OSA-18 questionnaires. Results In children with DS, spindle activity was lower compared to TD children for F Slow and F Slow&Fast spindles combined (p < 0.001 for all). Furthermore, there were no correlations between spindle activity and CBCL subscales; however, spindle activity for C Fast and C Slow&Fast was negatively correlated with OSA-18 emotional symptoms and caregiver concerns and C Fast activity was also negatively correlated with daytime function and total problems. Conclusions Reduced spindle activity in children with DS may underpin the increased sleep disruption and negative effects of SDB on quality of life and behavior. Impact Children with Down syndrome (DS) are at increased risk of sleep-disordered breathing (SDB), which is associated with sleep disruption affecting daytime functioning. Sleep spindles are a sensitive marker of sleep quality. We identified for the first time that children with DS had reduced sleep spindle activity compared to typically developing children matched for SDB severity. The reduced spindle activity likely underpins the more disrupted sleep and may be associated with reduced daytime functioning and quality of life and may also be an early biomarker for an increased risk of developing dementia later in life in children with DS.

Parents completed a medical history form and demographics questionnaire.All TD children were born at term and were otherwise healthy with no comorbidities such as craniofacial syndromes, developmental disability or genetic syndromes as assessed by the referring pediatrician.Both TD and children with DS were well at the time of the PSG study and none of the TD children were taking medications known to affect sleep or breathing.Of the children with DS, two had undergone surgery for non-cyanotic congenital heart disease as an infant but were considered to have no active cardiac disease at the time of the study, four had undergone an adenotonsillectomy, six were on thyroxine for hypothyroidism and one was taking melatonin for sleep-onset insomnia.

Sleep and respiratory analysis
All PSG studies were scored manually in 30 s epochs for sleep stages (N1, N2, N3 and REM), respiratory events > 2 breaths in duration and arousals by trained pediatric sleep scientists using Compumedics ProFusion software according to American Academy of Sleep Medicine pediatric guidelines 1 .An obstructive apnea was defined as the cessation of airflow in association with ongoing respiratory effort; an obstructive hypopnea was defined as ≥30% decrease in nasal pressure signal amplitude, associated with increased work of breathing and an arousal or ≥3% decrease in oxygen saturation; a central apnea was defined as cessation of airflow without inspiratory effort lasting either ≥20 seconds or at least the duration of two breaths and associated with an arousal or ≥3% oxygen desaturation; and a central hypopnea as ≥30% decrease in nasal pressure signal amplitude with reduced inspiratory effort throughout the entire duration of the event.A mixed apnea was defined if an event was associated with absent respiratory effort during one portion of the event and the presence of obstructed inspiratory efforts in another portion, regardless of which portion came first 1 .The OAHI, defined as the total number of obstructive apneas, mixed apneas, and obstructive hypopneas per hour of total sleep time (TST) was used to define SDB severity.Other respiratory parameters included the respiratory disturbance index (RDI), defined as the total number of respiratory events including obstructive and central apneas, mixed apneas, obstructive and central hypopneas; the REM RDI; the arousal index (ArI), defined as the number of cortical EEG arousals per hour of TST; the central apnea hypopnea index (CnAHI), defined as the number of central apneas and hypopneas per hour of TST.Desaturation measures included the average SpO2 drop, defined as the average SpO2 desaturation with scored respiratory events; SpO2 nadir, the lowest oxygen saturation associated with a respiratory event; ODI 4%, defined as the number of times the SpO2 dropped by greater than or equal to 4% per hour of TST; ODI 90%, defined as the number of times the SpO2 dropped below 90% per hour of TST; and the average transcutaneous pCO2 during TST (Av TcCO2).

Sleep Macro-architecture analysis
Standard measures of sleep quality were calculated for each participant and included the following parameters.The duration of each sleep stage (N1, N2, N3, REM) was expressed as a % of TST.Wake after sleep onset (WASO) was calculated as the percentage of time awake during the sleep period time (SPT), defined as the amount of time in minutes from sleep onset until lights on at the end of the study, including all periods of wake in between.TST was defined as SPT excluding all periods of wake.Time in bed (TIB) was defined as the time between lights off and lights on.Sleep latency was defined as the period from lights off to the first 3 consecutive epochs of N1 sleep or an epoch of any other stage.Sleep efficiency was defined as the ratio of TST to TIB.

Sleep spindles analysis
Sleep spindles were identified manually using Compumedics Profusion 3.0 software (Melbourne, Australia).Spindles were identified from the C4-A1 and F4-A1 EEG channels during N2 and N3 sleep from raw EEG signals recorded using a band-pass filter of 0.3 Hz to 100 Hz and a sampling frequency of 512 Hz.Each 30 s epoch was studied with the EEG gain set to 25 μV per mm, the high pass band filtered at 0.3 Hz and low pass band filtered at 35 Hz 2 .Spindles were categorized according to criteria reported by Chatburn et al, where identified spindles were required to be relatively equal in amplitude and with waves of a consistent frequency and a narrow conical shape with a duration of > 0.5 s 3 .Candidate spindles were not included if the amplitudes of the waves were not consistent and the waves were not conical, as if waves were too broad, the minimum of 5 peaks within 0.5 s would not be met and if too variable then this was most likely to be artefact.Waveforms that could otherwise be defined as a vertex or sawtooth wave or of highly variable amplitudes or frequencies were also excluded 3 .Spindles were then categorized and labelled as either being a F4 or C4 Slow or F4 or C4 Fast spindles, and F4 or C4 Slow&Fast spindles combined.Slow spindles consisted of 5-6 peaks while fast spindles had ≥7 peaks within 0.5 s.Broken spindles (spindles within 1 s of each other) were counted as one spindle 3 .
Total spindle number was the total number of each category of spindle found in N2 and N3 sleep.The duration of every individual spindle was automatically recorded by Profusion and when exported, verified to have a duration >0.5 s before calculating the average duration for each spindle category.Spindle density was calculated as the number of spindles per minute of N2 or N3 sleep; spindle intensity was calculated as the product of spindle density and average spindle duration.The majority of spindles were identified in N2 as expected, but as including spindles from N3 increased the number of children with spindles in the various categories, data are presented with spindles from N2 and N3 combined.

Questionnaires.
The OSA-18 assesses quality of life in children with OSA 4 .It comprises 18 questions categorized into five domains: sleep disturbance, physical symptoms, emotional symptoms, daytime function and caregiver concerns.Scores are rated using a Likert scale about how frequently the child has experienced symptoms in the previous four weeks with scores ranging from "1 -None of the time" to "7 -All of the time."Scores <60 suggest a small impact, scores between 60 and 80 suggest a moderate impact and scores >80 suggest a significant impact on QOL 5 .
The Child Behavior Checklist (CBCL) 6 for children 6 -18 years old contains 118 questions.
Scales include internalizing and externalizing behavior and total problems 7 .Questions are rated on a Likert scale as either 0 (not true), 1 (somewhat or sometimes true) or 2 (very true or often true).The raw scores were calculated by totaling the items under each scale and then converting to normal-referenced T-scores.T-scores ≥ 70, which are equivalent to scores above the 98th percentile, are considered to be of clinical concern 8 .
In addition, the parents of the children with DS also completed the Adaptive Behavior Assessment System, Second Edition (ABAS II) 9 , the Pediatric Sleep Problem Survey Instrument (PSSI) 10 and the Epworth Sleepiness Scale for Children and Adolescents (ESS-CHAD) 11 .
The ABAS-II was used to assess skills necessary for daily functioning with separate versions used for children aged under 5 years of age and those over 5 years.The ABAS-II has three major adaptive domains: conceptual, social and practical tasks, covering eleven skill areas.Each skill area is scored according to a 4-point Likert scale with scores rated as 0 (is not able to), 1 (never when needed), 2 (sometimes when needed) or 3 (always when needed).The general adaptive composite is a composite score of all adaptive skill areas.Scaled scores for the three adaptive domains and general adaptive composite scores are classified as very superior (130 or more), superior (120-129), above average (110-119), average (90-109), below average (80-89), low (71-79) and extremely low (70 or less) 9 .
The PSSI contains 26 questions assessing a range of sleep related disorders in children which are rated on a 4-point Likert scale of "Never," "Rarelyonce a week," Sometimes -2 to 4 times a week," or "Usually -5 to 7 times a week."The relative items are summed and scores are assigned to assess subscales: sleep routine, bedtime anxiety, morning tiredness, night arousals, sleep disordered breathing (SDB) and restless sleep.A T-score of ≥70, which is equivalent to a score above the 95 th percentile, is indicative to be of clinical concern 10 .
The ESS-CHAD is a measure of daytime sleepiness.It contains eight common daily situations, for which the parent scores how likely their child is to doze or fall asleep from a scale of 0 (would never doze or sleep) to 3 (high chance of dozing or sleeping).The total scores are summed and a score ≥ 10 suggests excessive daytime sleepiness 12 .