Setting
A retrospective case-control study was conducted at the Sleep-Wake Disorders Center and Imaging Institute at Soroka University Medical Center, in which all patients were enrollees of Clalit Health Care Services (CHS). According to the Israeli National Health Insurance Law,42 all enrollees have free access to medical services, and physicians have no economic incentives to prevent or deter patients from PSG study and/or CT examination.3,4,18
This was a retrospective study and a waiver for informed consent was obtained from the Institutional Review Committee of Soroka University Medical Center. All methods were performed in accordance with the Israeli regulations. The Institutional Review Committee of Soroka University Medical Center approved the study protocol (protocol number SOR-20-0250).
Study groups
From June 2010 until September 2020, we retrospectively recruited the entire cohort of adults (n = 259, aged 24 through 85 years), who were referred to overnight PSG diagnosis of OSA and who performed at least two ambulatory abdominal CT examinations (at a constant peak voltage of 120 kV) (
Supplementary Fig. 2). All subjects had “typical” symptoms of OSA and had been referred by an otolaryngology surgeon, pulmonologist, or neurologist. Subjects who underwent PSG were matched with a comparison group (non-OSA) that was selected randomly from the general population in the Soroka University Medical Center database. According to this database, none of the comparison group had a history of sleep problems or had had a PSG study.
Excluded from both groups were patients with any disease that influenced bone metabolism or who were receiving medicine that could influence bone metabolism (n = 41, Fig. 1) such as: chronic obstructive pulmonary disease, genetic disorders, cancer, autoimmune disorders, chronic liver disease, chronic renal insufficiency, musculoskeletal and connective tissue disorders, lumbar surgery, malabsorption disease, fibromatosis, unspecified anticonvulsants, and endocrine disorders, or who were wheelchair bound. We excluded patients receiving the following medications: oral corticoid, hormone replacement, or osteoporosis therapy; proton pump inhibitors; and anticonvulsant and anticoagulant drugs. Of the remaining 280 patients, two subjects had missing PSG data, and in 15 patients CT data could not be analyzed (the scan was not performed at 120 kV). 201 patients were included in the analysis, and of these, 27 did not have OSA (AHI <5 events/hr). Therefore, the OSA group comprised 174 patients.
CT examination of 124 non-OSA patients were analyzed and, of these, data of 10 CTs could not be analyzed (examination not performed at 120 kV). We grouped 27 PSG patients who tested negative to OSA (AHI < 5 events/h) with the 114 subjects in comparison group; the non-OSA group included 141 subjects.
Medical diagnoses retrieved from the Soroka University Medical Center database are documented only by physicians using the International Classification of Diseases, Ninth Revision (ICD-9) code. This database contains >99% of all patient diagnoses. We reviewed the following diagnosed cardiovascular diseases [codes 410–414, 426–438, 443] and hypertension [code 401–405]. A self-administered questionnaire assessed the Epworth sleepiness scale (ESS), where a higher the score indicates a higher level of sleepiness.45
PSG study: Data were acquired using a commercially available sleep monitoring system (Viasys, SomnoStar Pro; Yorba Linda, CA, USA or SomniPro 19 PSG; Deymed Diagnostic, Hronov, Czech Republic), as previously described by our laboratory.46,47 On the study day, participants were advised to maintain their sleep–wake routine and avoid consumption of caffeine and soft drinks. The overnight PSG included recordings of an EEG (C3/A2, C4/A1, and O2/A1, O1/A2), electrooculogram (right and left outer cantHU), electromyogram, and electrocardiogram. Scoring was done by a trained technologist and reviewed by one of the investigators (A.T.). Arousals and awakenings were scored using the American Sleep Disorders Association (ASDA) assessment.47 The apnea–hypopnea index was defined as the sum of all obstructive and mixed apneas, plus hypopneas associated with a ≥30% reduction in airflow and either ≥ 4% oxygen desaturation or electroencephalographic arousal, divided by the hours of total sleep time.46–48 The percent of sleeping time in which oxygen saturation was below 90% (T90) was calculated. OSA severity was defined as AHI 1–4.9 events/h, AHI 5.0–14.9 events/h, or AHI ≥ 15 events/h were considered as no OSA and mild and moderate/severe OSA, respectively.
Measurement of BMD of vertebral bone
We used 1-mm thick abdominal sections from CT examinations obtained during ambulatory or emergency room visits using a Siemens SOMATOM Definition AS+ Scanner (Siemens Healthcare GmbH, Erlangen Germany) or a Philips Brilliance ICT scanner (Haifa, Israel). We retrospectively accessed the CT examinations at a constant peak voltage of 120 kV with a variable mAs tube. We included CT scans with or without a contrast agent and evaluated vertebral BMD on a standard radiology picture archiving and communication (PACS) system workstation, with images viewed in bone windows, i.e., gray-scale assignment of the image display, to emphasize bone without the influence of attenuation/BMD values (Supplementary Fig. 2).16,49 We included examinations either with or without an intravenous enhancement agent. We assessed vertebral BMD by placing a single oval click-and-drag region of interest (ROI) in an axial and sagittal slice over an area of vertebral body trabecular bone and then measured CT attenuation in HU, with a lower HU (lower attenuation) representing less-dense bone, at each of the T12 and L1 levels. On the axial images of the selected slice—the superior part of the vertebra—the elliptical ROI was encompassed manually as the largest possible area at the anterior portion of each vertebral body, and in the sagittal plane, we focused on the upper anterior part of the vertebra in order to avoid the Dense Tracecular zone. The mean CT scan density of the ROI was measured. We avoided placing the ROI near areas that would distort the BMD measurement (focal heterogeneity or lesion, posterior venous plexus, compression fracture, and artifacts). A CHS engineer from the Biomedical Engineering Department calibrated the CT scanners routinely according to the manufacturer’s instructions using an American College of Radiology-accredited phantom. BMD was expressed in milligrams per milliliter of hydroxyapatite.16
Statistical Analysis
Statistical analyses were performed using R Statistical Software, version 3.5.2 (Foundation for Statistical Computing, Vienna, Austria). We compared the proportion of cardiovascular diseases and hypertension between patients with and without OSA using a Pearson chi-square test. Age, body mass index, the Epworth sleepiness scale, the arousal and awakening index, sleep efficiency, oxygen desaturation index (> 4%), and the percent of sleeping time in which oxygen saturation was below 90% were compared between patients with mild and moderate/severe OSA using a Student's t-test. The first and second BMD measurements and the difference between the two BMD (i.e., BMD on the second scan minus BMD on the first scan) measurements were compared between the OSA and non-OSA groups using a Student's t-test. We further evaluated the difference within the two measurements between the OSA and non-OSA groups using a two-ways repeated measurements ANOVA. To assess the independent association between OSA and the first measurement BMD and the BMD difference between the two measurements, we used a multivariate linear regression adjusted for patient age, gender, and diagnosis of cardiovascular diseases. The beta values, including the 95% Confidence Interval (CI) and p values for each variable, were calculated. Null hypotheses were rejected at the 5% level.