Challenging the Inflammatory Response System: Are Individuals With Insomnia More Reactive?
Conditions: Insomnia
Sponsor: Beth Israel Deaconess Medical Center

(CCI) Committee on Clinical Investigations 
Page 1 of 13 
 
Study Description – Part B 
CCI Form:  9-2015 
 
 
PI Revision 
Date:02/26/2016 
 
 
 
 
 
 
 
 
 
 
 
 
 
Challenging the inflammatory response system: Are individuals 
with insomnia more reactive? 
 
 
 
 
 
 
NCT02261597 
 
 
Date: 02/26/2016 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
PART B 

(CCI) Committee on Clinical Investigations 
Page 2 of 13 
 
Study Description – Part B 
CCI Form:  9-2015 
 
 
PI Revision 
Date:02/26/2016 
 
STUDY DESCRIPTION 
 
TITLE OF PROTOCOL 
Challenging the inflammatory response system: Are 
individuals with insomnia more reactive? 
Principal Investigator 
Monika Haack PhD 
 
 
 
 
B1. 
PURPOSE OF PROTOCOL 
Primary objective: To investigate the inflammatory response to a physiological stress challenge in 
insomnia disorder. We expect that individuals with insomnia are more reactive, as manifested in a 
stronger response of inflammatory systems markers (interleukin [IL]-6, prostaglandin [PG]E2) to a 
physiological stress challenge (cold pressor test). In accordance, we expect a stronger autonomic 
(blood pressure, heart rate) and hypothalamus-pituitary-adrenal (HPA, cortisol) response to challenge 
in insomnia disorder compared to healthy controls. 
 
Secondary objective 1: To investigate the ability to habituate to a repeated physiological stress 
challenge. We expect that the process of habituation is deteriorated in insomnia compared to healthy 
controls, as indicated by the lack of the inflammatory, autonomic, and HPA responses to decrease 
upon repeated exposure to the cold pressor test.   
 
Secondary objective 2: To investigate whether the inflammatory response to challenge can be 
predicted by the sensitivity of immune cells to glucocorticoids. We expect that in insomnia, monocytes 
are less sensitive to the counter-inflammatory signal of cortisol, resulting in an exaggerated 
inflammatory response to a physiological stress challenge. 
 
      
 
 
B2. 
SIGNIFICANCE AND BACKGROUND FOR THE STUDY 
Sleep is critical for the regulation and maintenance of biological systems, and sleep deficiency, such 
as insomnia, has been shown to be associated with elevated risks for cardiovascular, metabolic, and 
mood disorders11. 
Despite the high prevalence of insomnia in the population23, our understanding of the biological 
consequences of the disorder with respect to inflammatory, autonomic, and HPA system markers is 
limited, and often not consistent. For example, changes in the inflammatory system have been 
indicated by findings of increased levels of interleukin-6 (IL-6)5;33, but not C-reactive protein (CRP)20. 
Changes in the sympatho-adrenal system, as indicated by elevated blood pressure levels or 
increased incidence of hypertension10;19, and increased circulating norepinephrine levels17 have been 
reported in several studies. HPA stress system marker, as measured by cortisol levels in serum or 
plasma, have been found increased in insomnia in some studies26;31, but not others25;30. 
 
All these investigations on the impact of insomnia on biological system markers have in common that 
they measure basal levels of various system markers. However, insomnia may not only alter the 
basal activity of these systems, but may change their reactivity to other stressors and 
challenges. In support of this assumption are findings showing that poor sleep quality in healthy 
individuals is associated with a stronger IL-616, cortisol13, and heart rate32 response to a stressful 
challenge, such as the cold pressor test (CPT). The CPT involves the immersion of the hand in ice-
cold water kept at about 3C for an interval between 30sec to 3min. It is one of the most commonly 
used laboratory physiological challenge tests, provoking not only unpleasantness, but also increases 
in cortisol, norepinephrine, and inflammatory cytokines within 15-50min post-testing18;27;35.  
 

(CCI) Committee on Clinical Investigations 
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Study Description – Part B 
CCI Form:  9-2015 
 
 
PI Revision 
Date:02/26/2016 
 
Investigations of such system’s reactivity to challenge may elucidate systems abnormalities that we 
do not capture by only assessing basal system’s levels. For example, in patients with rheumatoid 
arthritis, basal IL-6 levels are normal, but the response to a physiological stress challenge (cold 
pressor test) is amplified9. Similarly, individuals with poor sleep quality cannot be distinguished from 
good sleepers by their basal IL-levels, only by their IL-6 reactivity to challenge16.  
 
However, to our knowledge, no studies have measured how insomnia may affect the reactivity of 
biological systems to a stressful challenge, which may serve as an important indicator of system’s 
dysregulation and associated disease risk 21 
In this light, the primary objective of this proposal will investigate whether the inflammatory, 
autonomic, and HPA systems are more reactive to a physiological stressful challenge in 
insomnia disorder compared to healthy controls.  
 
Beside the hypothesized changes in the system’s response reactivity in insomnia, the secondary 
objective 1 of this proposal will investigate the ability of system’s responses to habituate to 
repeated stressful challenges. Habituation, or a decreased response following repeated exposures 
to a stressful challenge, is a key feature of the adaptive nature of many biological systems, and has 
been observed in response to a variety of psychological and physiological stressors and challenges in 
both humans and animals15. Considering that most individuals have to deal with stressful challenges 
on a daily basis, failure of systems to habituate to such daily challenges may elevate disease risk in 
the long term15. To test the degree of habituation, we will repeatedly administer the CPT and measure 
the inflammatory, autonomic, and HPA response magnitude after each challenge. 
 
In summary, this proposal focuses on system’s reactivity to challenge (primary objective) and the 
ability of systems to adapt to repeated challenges (secondary objective 1), rather than assessing 
basal systems activity only.  Dysregulation of systems reactivity and habituation are likely to 
constitute processes that may serve as valuable predictors of disease risk in insomnia. 
 
Secondary objectives 2 and 3 investigate potential determinants of the inflammatory response to 
challenge. Is the proposed stronger inflammatory response to challenge in insomnia due to 
less sensitivity of immune cells to the counter-inflammatory actions of cortisol (secondary 
objective 2)? One potential mechanism contributing to increased inflammatory reactivity is a change 
in the sensitivity of monocytes to the glucocorticoid (GC) cortisol. Indeed, impaired GC sensitivity has 
been reported in response under various medical and/or stressful conditions24;28, facilitating synthesis 
and release of inflammatory cytokines by monocytes. Though insomnia disorder has been 
frequently associated with elevated cortisol levels2, the sensitivity of immune cells to the 
cortisol signal surprisingly has never been assessed. Thus, the secondary objective aim 2 will 
investigate whether GC sensitivity in insomnia is a potential predictor of the proposed inflammatory 
response increase to challenge. 
To conclude, this proposal two important concepts that have not been addressed in insomnia: 
Response reactivity to a stressful challenge and the ability to habituate to repeated exposure of such 
challenge. These two concepts may serve as critical indicators of disease risk in the long term15;21. In 
addition, we will investigate a potential determinant of inflammatory reactivity: GC sensitivity that may 
serve as target for future drug development and evaluation. 
      
 

(CCI) Committee on Clinical Investigations 
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Study Description – Part B 
CCI Form:  9-2015 
 
 
PI Revision 
Date:02/26/2016 
 
 
B3. 
DESCRIPTION OF RESEARCH PROTOCOL 
A. 
Study Design – Overview, Methods, Procedures 
 
As seen in the Figure 1, the research design consists of an initial screening visit, followed by an in-
hospital overnight medical/sleep screening. If the participant meets the study recruitment requirements 
they will immediately begin the 1 day in-hospital study visit. 
 
Visit 1. Initial screening. All 
potential participants will have 
the study explained to them in 
detail by the PI or another senior 
member of the research team. If 
interested, potential participants 
will sign the informed consent 
and nursing staff in the Clinical 
Research Center (CRC) will 
take vital signs and collect a 
blood sample.  A short battery of 
screening 
tests 
and 
questionnaires 
will 
also 
be 
collected in order to assess 
health status and ability to 
tolerate ice-cold temperatures (a 
water bath) without adverse 
circulatory 
responses. 
 
In 
addition, participants will be 
asked to record their sleep in 
the home environment, using 
sleep diary and actigraphy. 
  
 
Visit 
2. 
Medical 
history, 
physical, and sleep screening. 
During this visit, urine toxicology 
and pregnancy (for women) 
tests will be performed, and the study physician will meet with the study participant for a physical 
exam and review of blood test results. Participants who remain eligible following medical screen will 
be set up with PSG equipment for a sleep screening night in the CRC, in order to rule out any 
possible sleep disorders.  
If  a participant has done a screening overnight sleep study within the last 6 months, or a sleep diary 
recording, screening blood test (including lipid panel, insulin, glucose, TSH, CBC, PLT, CRP, sodium, 
potassium, billirubin, creatinine, ALT, absolute T Cells, sediment rate), or had a physical  examination 
within the last 3 months, these results will be used for this research. 
 
1-Day CRC Visit: This visit will be scheduled directly following the medical/PSG screen. At 10am, an 
intravenous (iv) line will be placed for blood drawing. Pain threshold measures (heat and pressure) 
will be assessed about an hour later. Baseline assessment will begin at 1am lasting until 12pm, 
during which blood (inflammatory/HPAmarkers), urine (prostaglandin/norepinephinre), and autonomic 
variables (beat-to-beat blood pressure, heart rate) will be collected. . The pain threshold testing will 
start at 1100am.  Following the procedures the heat pain threshold will be assessed by a computer 
controlled thermode (Somatosensory Analyzer, Medoc, Minneapolis, MS) will be placed on their 

(CCI) Committee on Clinical Investigations 
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Study Description – Part B 
CCI Form:  9-2015 
 
 
PI Revision 
Date:02/26/2016 
 
forearm. The participant will click a mouse when they feel the thermode temperature change from 
neutral to warm and then hot to painful. The pressure pain threshold will be measured by electronic 
pressure algometer (Somedic Sales AB, Hörby, Sweden.  Participants will press a control button 
when they experience the first sensation of pain.  The reactivity and habituation assessment will take 
place at 1pm. The cold pressor test (CPT) will be administered 3 times. As detailed under 
procedures, participants will be instructed to immerse their hand in ice-cold water kept at 3 degree C, 
for a duration of 1min. the interval between the three cold pressor tests will be 1.5 hours. Blood will be 
sampled after the 1st, 2nd, and 3rd cold pressor test twice, 20min and 50min after hand removal from 
the cold water bath (these timepoints have been shown sensitive to capture increases of cortisol and 
IL-6). Urine will also be collected during and after the administration of the cold pressor test series, 
starting at 1pm and closing out at 6pm.. Prior to, during, and after the cold pressor series, participants 
will rate their stress levels as well as the unpleasantness of the cold sensation induced by the 
CPTThe final pain modulation tests, temporal summation and conditioned pain modulation will occur 
at 500pm.  Temporal summation of pain is frequently used as an index of central sensitization of 
dorsal horn neurons56, and will be assessed using the Somatosensory Analyzer (Medoc, Minneapolis, 
MS). The participant wil have the thermode placed on forearm volar skin of the arm not used for blood 
draws.   Four sequences each consisting of ten brief consecutive heat pulses will be applied via a 
thermode with a pulse-to-pulse interval of 2.5 sec. Conditioned pain modulation assesses the 
principle of ‘pain inhibits pain’, and is a measure of the pain-inhibitory capacity of the central nervous 
system. The participant will have the thermode placed on the forearm volar skin not used for blood 
draws..For the test, the test stimulus will be the TS sequence at highest tolerable temperature, 
applied to the outer volar surface of the non-dominant forearm, along C8-T1 innervations. Immersion 
of the contralateral foot into a painfully hot water bath (47 °C, Techne® water baths, Bibby Scientific 
US, Burlington, NJ)  Prior to discharge, participants will receive a dinner shortly after 6pm.   
 
Procedures: 
 
Polysomnographic recording (PSG). Sleep will be recorded using the Embla system N7000 (Medcare 
US, Buffalo) on the screening night visit to ensure that participants are free from sleep disorders 
(clinical interview will be used to diagnose insomnia disorder and also to support the diagnoses of 
other sleep disorders, such as nightmare disorder, circadian rhythm disorder). 
 
Blood Sampling. Blood samples will be drawn at intermittent intervals during the experimental visit 
using an indwelling 18-gauge forearm catheter from 1pm to 7pm. The total amount of blood taken will 
not exceed 200ml. 
IL-6 will be measured in plasma in our laboratory, using a high sensitivity enzyme immunosorbent 
assay (ELISA, Quantikine ® HS, R&D Systems, Minneapolis, MN). IL-6 will be measured at baseline, 
after each cold pressor test, and hourly after the last cold pressor test until 7pm. 
PGE2 metabolite will be measured in urine collected during the cold pressor test series (from 3-7pm). 
Urinary PGE2 will be analyzed in our lab using an EIA (Cayman Chemical, Ann Arbor, MI), and levels 
will be adjusted for urinary creatinine levels. 
Cortisol will be measured in plasma and assayed in the Harvard Catalyst Central Lab, using the 
Access Chemiluminescent Immunoassaay (Beckman Coulter Fullerton, CA). Cortisol will be 
measured at baseline, after each cold pressor test, and hourly after the last cold pressor test until 
7pm. 
Glucocorticoid (GC) sensitivity will be assessed in vitro by examining the capacity of monocytes to 
produce IL-6 in response to the synthetic glucocorticoid dexamethasone (DEX). In brief, whole blood 
will be incubated with LPS (100 pg/ml) in the presence of different concentrations of DEX (0-400nM). 
After incubation with fluorescence-conjugated antibodies (CD14, CD45, IL-6), monocyte expression 
of IL-6 will be measured in the Flow Cytometry Core at the BIDMC (Gallios™, Beckman-Coulter Inc). 
This protocol was developed by the PI for our currently ongoing study on pattern of repeated sleep 
restriction and recovery (R01 HL 105544). Lower IL-6 expression in response to DEX would indicate 

(CCI) Committee on Clinical Investigations 
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Study Description – Part B 
CCI Form:  9-2015 
 
 
PI Revision 
Date:02/26/2016 
 
a higher GC sensitivity and this mechanism may contribute to the proposed stronger inflammatory 
reactivity in insomnia disorder. Measures will be performed once at baseline, using the 2pm blood. 
Autonomic markers: Blood pressure (BP) measurements will be collected during baseline, throughout 
the testing sessions and recovery period of the experimental visit, using the dinamap system.  
Physiological stress challenge – cold pressor test (CPT): A trial CPT will be administered during the 
initial screening to identify any previously-unrecognized incidence of Raynaud’s phenomenon or 
inability to tolerate the ice bath. The CPT will be administered three times during the experimental 
visit, starting at 1pm. Throughout the testing and recovery period (until 6pm), the participant will 
remain in a seated position in a comfortable chair. The CPT will be performed using the hand/arm 
that is not being used for blood sampling in order to prevent interference. A series of three CPT tests 
will be applied with an inter-test interval of 60min. Repeated administrations of the cold pressor test 
have been performed in previous studies (with intervals as short as 2min8;13), but without investigating 
system’s responses. For the current study purposes, an inter-test interval of 60 min has been chosen 
to capture response increases of IL-6 and cortisol8;9;16. For each CPT test,  participants will be asked 
to insert their hand in a temperature-controlled water bath (Techne® water baths, Bibby Scientific US, 
Burlington, NJ), kept at 3 degree C, and instructed to leave their hand in for a duration of 1min 
(participants are told that they can take their hand out earlier if the sensation is getting unbearable). 
Blood will be drawn 20 and 50min after hand removal from the cold water bath. These time points 
have been shown sensitive to capture peak cortisol levels as well as significant IL-6 increases8;9;16. 
Participants will rate the unpleasantness of the sensation prior to hand immersion into the cold water 
bath, and continue with ratings at 10sec intervals during and after the testing period (up to 3min post-
testing). 
Pain threshold testing details: 
(a) Heat pain thresholds (HPT) will be assessed using a precise, computer-controlled thermode 
(Somatosensory Analyzer, Medoc, Minneapolis, MS) for generating and recording responses 
to thermal stimuli. The thermode will be attached to the forearm volar skin of the arm not used 
for blood draws, via a Velcro strap, to the palm. From a baseline temperature of 32°C, the 
thermode is heated at a rate of 0.5 °C/sec. The participant is instructed to click a computer 
mouse with their free hand, as soon as heat pain is perceived. To improve accuracy, each 
threshold will be measured 4 times. The inter-stimulus interval is 15 sec, and the average will 
be used for further analysis. 
(b) Pressure pain thresholds (PPT) will be obtained with use of an electronic pressure algometer 
(Somedic Sales AB, Hörby, Sweden), and assessed at the posterior neck and trapezius muscle, 
and pressure will be increased at a rate of 30 kPa/s. Participants will be instructed to press a 
control button when they experienced the first sensation of pain. A series of four pressure-pain 
stimuli will be applied with 15 sec intervals to each side, and the average of these stimuli will 
be calculated. 
Modulation testing details: 
(a) Temporal summation (TS) of pain is frequently used as an index of central sensitization of 
dorsal horn neurons56, and will be assessed using the Somatosensory Analyzer (Medoc, 
Minneapolis, MS). Four sequences each consisting of ten brief consecutive heat pulses will be 
applied via a thermode to the forearm volar skin of the arm not used for blood draws, with a 
pulse-to-pulse interval of 2.5 sec. The temperature used to assess TS will be tailored to each 
person’s tolerance level. The first test sequence of this individually-tailored procedure will have 
a target temperature of 48 °C and an inter-pulse baseline temperature of 32 °C. Depending on 
whether the participant can tolerate the initial 10-pulse sequence, the target temperature of the 
second sequence will be increased or decreased, respectively, by 1.5 °C (i.e., increased to 
49.5 °C or decreased to 46.5 °C). In the third sequence, the target temperature will be again 
increased or decreased by 1.5 °C, depending on whether the participant could tolerate the 
second sequence or not. The inter-sequence interval will be 2 min at a temperature of 32 °C. 
The thermode will be moved systematically between sequences, starting at the thenar 
eminence for a practice trial, and sequentially moving cephalad on the volar aspect of the 

(CCI) Committee on Clinical Investigations 
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Study Description – Part B 
CCI Form:  9-2015 
 
 
PI Revision 
Date:02/26/2016 
 
forearm along the innervation of C5–6 for the remaining three trials. This sequence was 
designed to prevent testing on previously stimulated skin areas (see also25). During each test 
sequence, participants will be prompted to rate the intensity of the 1st, 4th, 7th, and 10th 
thermal pulse using a 0–100 mm visual analogue scale (VAS), presented on separate data 
sheets for each rating. The participant will be instructed to say ‘STOP’ as soon as the 
sensation is no longer tolerable at any point during the testing. 
(b) Conditioned pain modulation (CPM), assesses the principle of ‘pain inhibits pain’, and is a 
measure of the pain-inhibitory capacity of the central nervous system. For this protocol, the 
test stimulus will be the TS sequence at highest tolerable temperature, applied to the outer 
volar surface of the non-dominant forearm, along C8-T1 innervations. Immersion of the 
contralateral foot into a painfully hot water bath (47 °C, Techne® water baths, Bibby Scientific 
US, Burlington, NJ) will be the conditioning pain stimulus that is intended to activate the pain-
inhibitory circuits and thereby decrease the perceived pain of the forearm test stimulus (see 
Figure 2 for setup).  
In total, four CPM trials will be performed: two trials using a hot water bath (47 °C) and two 
baseline trials without a water bath. For each hot water bath trial, the participant’s foot will be 
first submerged in the water bath. After 20sec of immersion, the 10-pulse temporal summation 
sequence will be applied to the forearm and the participant will be prompted to rate the pain 
intensity of the 1st, 4th, 7th, and 10th stimulus using the VAS. There will be a two-minute rest 
period between all trials during which the thermode will be systematically moved from the 
distal to proximal sites along the C8-T1 innervated skin in order to avoid re-stimulation of the 
previously sensitized skin. For statistical analysis, ratings of the two 10-pulse sequences 
under the conditioning stimulus (hot water) will be averaged, and contrasted against ratings 
from the baseline (no-water) condition. A decrease in pain intensity ratings under the hot water 
condition compared to the no-water condition will be used as an index of the pain-inhibitory 
capacity.   
 
 
 

(CCI) Committee on Clinical Investigations 
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Study Description – Part B 
CCI Form:  9-2015 
 
 
PI Revision 
Date:02/26/2016 
 
B. 
Statistical Considerations 
Sample Size Justification:  For sufficient statistical power, we will need to have 20 participants with a 
diagnosis of insomnia disorder (based on DSM-V criteria), and 20 healthy control sleepers complete 
the study protocol (60% women, see power calculation below). 
Power calculation 
Power calculations were performed for the primary objective of this proposal. With respect to the 
proposed stronger inflammatory response reactivity in insomnia compared to healthy controls, 
power calculations were based on findings from Heffner et al.16, reporting a difference in the IL-6 
response between poor and good sleepers to an acute challenge of 1.6 pg/ml (SD 1.71 pg/ml, 
estimated from graph). Thus, we need to study 20 participants in each group to reach optimal 
power of 80% (unpaired t-test, alpha = 0.05). 
With respect to the proposed stronger cortisol response reactivity in insomnia compared to healthy 
controls, power calculations were based on findings from Goodin et al.14, reporting a difference in the 
cortisol response between poor and good sleepers to an acute challenge (CPT) of 10 units (area 
under the curve, SD 11 units, estimated from graph). Thus, 80% power will be reached with a sample 
size of N=20 per group 
 
Statistics:  Primary objective: Testing response reactivity to a physical challenge. The overall 
statistical analysis will be performed within the frame of mixed linear models. Primary and secondary 
outcome variables pertaining to the primary objective (see above) will constitute the independent 
variable, time (baseline, 1st CPT post-testing) and group (insomnia vs. healthy controls) will be 
entered as fixed factors, while subject number will serve as random factor. Simple contrasts will be 
performed between groups to identify differences at single time points. 
Secondary objective 1: Testing response habituation to a repeated physical challenge. Data 
analysis will be similar as for the primary objective, but the time element will be presented by 
measures at baseline, 1st CPT post-testing, 2nd CPT post-testing, and 3rd CPT post-testing. 
Secondary objective 2: GC sensitivity as a predictor of inflammatory response reactivity. 
Univariate analysis will be used to identify differences between insomnia and healthy controls in GC 
sensitivity at baseline. Regression analysis will be used to predict inflammatory response reactivity.  
Secondary objective 3: Actigraphy-based sleep characteristics as predictors of inflammatory, 
autonomic, and HPA response reactivity. Univariate analysis will be performed to identify differences 
in sleep variables (as mentioned under secondary outcome variables above). Regression analysis will 
be used to predict response reactivity.  
For all analysis, alpha will be set to 0.05. 
 
 

(CCI) Committee on Clinical Investigations 
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Study Description – Part B 
CCI Form:  9-2015 
 
 
PI Revision 
Date:02/26/2016 
 
C. 
Subject Selection 
Human Subjects Involvement and Characteristics 
A total of 40 healthy adults (18-55 years, not including attrition rate of 10%) will be enrolled. 20 
participants with the diagnosis of insomnia disorder (DSM-V), 20 age- and gender matched healthy 
controls.  60% women (equivalent to an insomnia risk ratio of 1.4 in women versus men) Subjects of 
all racial/ethnic backgrounds will be eligible to participate. Participants will be accepted into the study 
if they are in good health, do not have a history of any medical disorders (including sleep disorders), 
as determined by a medical history, physical screening, and overnight sleep study (PSG).  
Participants from the Boston area, who meet these strict medical eligibility criteria, will be recruited to 
participate in the study. The study physician will also perform an in-person visit with each participant on 
day that they are in-hospital, in order to ensure patient’s well-being.  
 
Inclusion Criteria:  
- 
Women and men between the ages 18-55 years. 
- 
Body mass index (BMI) <=35. 
- 
Blood chemistry in the normal range 
 
  Specific to insomnia group: 
      Diagnosis of insomnia based on DSM-V criteria1: 
A. A predominant complaint of dissatisfaction with sleep quantity or quality, associated 
with one (or more) of the following symptoms: difficulty initiating sleep, difficulty 
maintaining sleep, early-morning awakening with inability to return to sleep. 
B. The sleep disturbance causes clinically significant distress or impairment of functioning 
(social, occupational etc.). 
C. The sleep difficulty occurs at least 3 nights per week. 
D. The sleep difficulty occurs despite adequate opportunity for sleep. 
E. The insomnia is not better explained by and does not occur exclusively during the 
course of another sleep-wake disorder. 
F. The insomnia is not attributable to the physiological effects of a substance (e.g., a drug 
of abuse). 
G. Coexisting mental disorders and medical conditions do not adequately explain the 
predominant complaint of insomnia. 
 
Specific for control group: 
- 
Good quality and quantity sleep (as determined by sleep diary and actigraphy: 7 to 9 
hours of sleep duration/night, sleep onset latency (SOL) and wake after sleep onset 
(WASO) <20min/night.  
- 
 
Exclusion Criteria: 
- 
Active infection/disease. 
- 
History of neurological, chronic pain, immune, cardiovascular, liver/kidney, or metabolic 
disorder within the last 6 months prior to study start. 
- 
History of psychiatric disorders in the last 6 months prior to study start, including major 
depressive disorders, bipolar disorders, panic disorders, post-traumatic stress disorders 
(PTSD), thought disorders, and substance abuse/dependence disorders. 
- 
Sleep disorders other than insomnia:  Apnea Hypopnea Index of >15 events/hour on 
polysomnographic sleep study, periodic leg movement index (PLMI) >10/hour; sleep efficiency 
<80% (only for control participants) based on polysomnographic screening night; restless legs 
syndrome, circadian rhythm disorders, and nightmare disorders determined by diagnostic 
interview. 
- 
Reynaud’s disease. 
- 
Psychotropic, sleep, or any other medications or herbs interfering with the inflammatory, 

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Study Description – Part B 
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PI Revision 
Date:02/26/2016 
 
autonomic, or HPA system in the last 2 weeks prior to study start, except oral contraceptives.  
Note 1: Individuals who are currently using medication will be invited to undergo a taper 
with permission from, and supervised by, their physician and will be eligible for 
participation 2 weeks following cessation of all medication use. 
Note 2: Information on historical medication use (prior to the 2-week medication-free 
period before study start) will be collected and incorporated into the statistical analysis 
plan if group differences exist. 
- 
In psychotherapy or any other behavioral interventions at study start (e.g., acupuncture for 
insomnia). 
- 
Pregnant/nursing. 
- 
History of psychiatric, neurological, pain-related, immune, gastrointestinal, or cardiovascular 
disease; liver/kidney or metabolic disorder within the last 6  months prior to the study start.. 
- 
Esophageal reflux; gastric or duodenal ulcers; or asthma 
- 
Pregnant/nursing. 
- 
Regular medication use other than oral contraceptives. 
- 
Donation of blood or platelets 3 month prior to or in-between in-hospital visits. 
- 
Substance abuse. 
Participants with slightly abnormal blood values will be included if they are not clinically meaningful.  
Results from the toxicology screen will not be used as eligibility criteria for study continuation, but will be 
used for data quality control purposes. 
 
Sources of Materials: 
Blood and urine samples from participants will be assayed for markers involved in the stress 
response. 
 
Inclusion of Women:  
All women of child bearing age who do or do not take oral contraceptives and who are not pregnant 
or nursing are eligible for the study. 
 
  
 
 
B4. 
POSSIBLE BENEFITS 
This research will not benefit participants directly.  However it will provide important information regarding 
our understanding of the biological consequences on insomnia disorder with respect to cardiovascular, 
metabolic and mood disorders.   
 
 

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Study Description – Part B 
CCI Form:  9-2015 
 
 
PI Revision 
Date:02/26/2016 
 
 
B5. 
POSSIBLE RISKS AND ANALYSIS OF RISK/BENEFIT RATIO 
There is a minimal risk of infection associated with drawing blood from a vein. On rare occasions, 
pressure pain testing can cause bruising at the site where the pressure has been applied. The 
application of heat pain can lead to sensitization of the skin for up to 48 hours. There are no known 
risks of cold pressor test. There are no known risks of wearing BP or PSG equipment.  
 
Participants will be extensively screened before being accepted into the study. We will exclude 
participants with a history of immune, cardiovascular, pain-related, gastrointestinal, significant 
psychiatric disease, significant allergies, or conditions where extreme vasoconstriction can occur, 
such as Raynaud’s syndrome. Participant screening will include blood tests for CBC and differentials, 
as well as thyroid hormone, and blood glucose; urinary toxicology screens will also be conducted. We 
will exclude participants who smoke or take regular medications other than hormonal birth control 
medication. The medical history and physical will be conducted by the study physician. The study 
physician will check on all participants while they are admitted to the CRC, and serve as the doctor on-
call throughout each participant’s stay. Should participants need medical assistance, the monitoring 
research assistant will notify the ward nurses, who will address any immediate medical needs and also 
immediately notify the study physician and the PI. In the event of a medical emergency, the CRC is 
located in a critical care hospital, and a code team is available at all times.  
 
Risk/benefit ratio: This work has the potential to understand how insomnia disorder may affect the 
reactivity of biological systems to a stressful challenge, which may serve as an important indicator of 
the system’s dysregulation and associated disease risk. 
 
 
B6. 
RECRUITMENT AND CONSENT PROCEDURES 
Recruitment 
Participants will be recruited via internet postings, flyers, bulletin advertisements, and local 
newspapers.  When participants contact the research office, they will participate in preliminary 
screening via telephone and/or email.  If it appears that an individual may qualify to participate in the 
study, s/he will be invited to come to the Clinical Research Center (CRC) where s/he will go through 
the informed consent process and complete questionnaires designed to assist in determining 
eligibility. During the initial visit to the CRC, nurses will also take vital signs and collect a blood 
sample.  These results will be reviewed by the study physician prior to the potential participants’ 2nd 
CRC visit, during which participants will undergo a medical history/physical and overnight sleep 
screening.  If an eligible individual is still interested in study participation, s/he will be scheduled to 
participate in the in-hospital portion of the study.  
 
Consent 
The PI and/or senior research team member will interview each subject and obtain informed consent 
from interested participants, after the participant has the study explained to them in detail and has had 
any questions answered. 
      
 
Subject Protection 
While none of the subjects in this study will be drawn from a population vulnerable to coercion or 
undue influence, the following measures to ensure subject protection will remain in place.  
Participants will be extensively screened before being accepted into the study. We will exclude 
participants with a history of immune, cardiovascular, pain-related, gastrointestinal, or significant 
psychiatric disease, or with significant allergies or Raynaud’s Syndrome or a history of substance 
abuse. Participant screening will include blood tests for CBC and differentials, as well as thyroid 
hormone, and blood glucose; urinary toxicology screens will also be conducted. We will exclude 

(CCI) Committee on Clinical Investigations 
Page 12 of 13 
 
Study Description – Part B 
CCI Form:  9-2015 
 
 
PI Revision 
Date:02/26/2016 
 
participants who smoke or take regular medications other than hormonal birth control medication. The 
medical history and physical will be conducted by the study physician. Should participants need 
medical assistance, the monitoring research assistant will notify the ward nurses, who will address any 
immediate medical needs and also immediately notify the study physician and the PI. In the event of a 
medical emergency, the CRC is located in a critical care hospital, and a code team is available at all 
times.  
 
If a participant reports to have suicidal ideation (positive answer to item 9 of the Becks Depression 
Inventory), the physician will be called to evaluate the condition of the subject which may result in 
bringing the subject to the emergency room.       
 
B7. 
STUDY LOCATION 
 Privacy and Physical Setting 
Every effort to maintain subject privacy will be made.  When interested participants contact the study 
office, a research assistant will return their call from a private office, in order to maintain 
confidentiality.  Emails sent to and received by study participants will be based on a study email 
address, and direct mail will only be sent with the participant’s express permission and provision of a 
physical mailing address.  Participants will only be contacted by phone at the number they provide to 
study staff. 
 
For their screening visits, participants will present to the CRC, a central shared clinical research floor 
in the main part of the hospital.  Presence in this area of the hospital does not provide any specific 
information for an outside observer who witnesses the participant presenting for screening.  All in-
person screening, including collection of physical data from participants, will take place in a private 
room, behind closed doors, in the CRC.  The in-hospital portion of the study will be conducted in a 
private or shared room within the CRC).  
 
Whenever possible, participant information will be identified only with a ‘participant identification 
number’ and not with actual names.  The exception to this is clinical data acquired in the CRC, which, 
in accordance with hospital policy, is identified with first and last name.  As soon as it is feasible, this 
name identification is converted to a numeric identifier to maximize confidentiality. 
 
 
 

(CCI) Committee on Clinical Investigations 
Page 13 of 13 
 
Study Description – Part B 
CCI Form:  9-2015 
 
 
PI Revision 
Date:02/26/2016 
 
B8. 
DATA SECURITY 
All possible precautions will be taken to ensure data security.  Data collected from participants will be 
identified using a subject number, rather than a name, whenever possible.  All patient hard copies of 
materials containing patient information will be stored in a locked file cabinet in a locked office on a 
secured (swipe-card access) research floor of the hospital.  Electronic data will be maintained on a 
secure server behind the BIDMC firewall.  All computers in the laboratory require use of a login 
before accessing any information, and data are encrypted.  Similar to hard copy materials, laboratory 
computers are located in locked rooms on a secure (locked) research floor of the hospital.      
 
B9 
Multi-Site Studies 
 
Is the BIDMC the coordinating site?       
 Yes    
 No  
 
Is the BIDMC PI the lead investigator of the multi-site study?     
 Yes    
 No  
 
B10 
Dissemination of Research Results 
 
Research subjects who express the desire to receive information about their individual study results 
or study progress will be provided with the following information: (1) Blood screening results provided 
by the study physician. (2) Information on sleep screening night provided and explained by PI or 
senior co-investigator. (3) Copy of journal publication. In addition, participation of research subjects 
will be acknowledged in data presentations and journal publications.