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Preliminary study of the association of serum irisin levels with poor sleep quality in rheumatoid arthritis patients
Journal Pre-proof Preliminary Study of the Association of Serum Irisin Levels with Poor Sleep Quality in Rheumatoid Arthritis Patients Rania M. Gamal, Mona Embarek Mohamed, Nevin Hammam, Noha Abo El Fetoh, Ahmed M. Rashed, Daniel E. Furst PII:
S1389-9457(18)30858-X
DOI:
https://doi.org/10.1016/j.sleep.2019.10.021
Reference:
SLEEP 4230
To appear in:
Sleep Medicine
Received Date: 24 October 2018 Revised Date:
22 July 2019
Accepted Date: 17 October 2019
Please cite this article as: Gamal RM, Mohamed ME, Hammam N, El Fetoh NA, Rashed AM, Furst DE, Preliminary Study of the Association of Serum Irisin Levels with Poor Sleep Quality in Rheumatoid Arthritis Patients, Sleep Medicine, https://doi.org/10.1016/j.sleep.2019.10.021. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. 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. © 2019 Elsevier B.V. All rights reserved.
Preliminary Study of the Association of Serum Irisin Levels with Poor Sleep Quality in Rheumatoid Arthritis Patients
Short title: Irisin in RA patients
Rania M. Gamal1, Mona Embarek Mohamed2, Nevin Hammam1, Noha Abo El Fetoh3, Ahmed M. Rashed1, Daniel E. Furst4,5&6 1 Rheumatology& Rehabilitation Department, Assuit University Hospitals, Egypt. 71515. 2 Microbiology and Immunology Department, Faculty of Medicine, Assiut University, Assiut, Egypt.71515 3 Neurology and Psychiatry Department, Faculty of Medicine, Assiut University, Egypt. 4 Department of Medicine, Division of Rheumatology, University of California, Los Angeles (emeritus), Los Angeles, California. 5 Department of Rheumatology, Division of Rheumatology, University of Washington, Seattle Washington 6 Division of Rheumatology and Experimental Medicine, University of Florence, Florence, Italy.
Corresponding Author and offprint requests: Rania M. Gamal (MD) Associate Professor. Rheumatology & Rehabilitation Department, Assiut University, Assiut, Egypt. Postal code 71515 Fax: 02-0882333327 Mobile: 0020-1006289729 E-mail: [email protected]
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Abstract Study Objectives: Sleep disorders are significant problems in patients with rheumatoid arthritis (RA) and are associated with poor quality of life. Irisin is myokine which may have antiinflammatory and energy regulatory roles. This study assessed the association of serum irisin levels with the quality of sleep and disease activity in RA patients. Methods: Fifty-eight RA patients and 30 matched healthy controls were included. Disease activity score in 28 joints (DAS28-ESR) and the patients’ global score were calculated. RA patients were grouped according to the Pittsburgh Sleep Quality Index score (PSQI) into goodsleepers (group 1) defined as a PQSI score <=5 and poor sleepers (group 2) with a PSQI > 5. Serum irisin levels were measured for both patients and controls by commercially available enzyme-linked immunosorbent assay kits. Results: Poor sleep quality was found in 26 (45%) of the RA patients. Irisin levels were significantly lower in RA patients with poor sleep compared to those with good sleep and healthy controls (p<0.001). Serum irisin levels correlated inversely with disease duration, morning stiffness duration, DAS28-ESR, global score, and total PSQI score (r= -0.722 to -0.263 & p values <=0.001-0.04) indicating a possible anti-inflammatory role of irisin in RA patients. The analysis employed Student's t-test, ANOVA, and Pearson correlation. Conclusions: Irisin levels were decreased in RA patients with poor sleep quality compared to RA patients with good sleep quality and healthy controls, indicating a possible association of decreased serum irisin with sleep impairment in RA patients
Keywords: Irisin, Rheumatoid arthritis, Sleep, Disease activity.
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Introduction Rheumatoid arthritis (RA) is a chronic disease characterized by inflammation in the synovial joints, leading to pain, swelling, morning stiffness, and joint deformities [1]. Patients with RA experience disturbed sleep that affects their health-related quality of life and physical or psychological function [2]. Sleep disturbances were reported in 54-70% of RA patients [3]; RA patients with poor sleep quality suffered from night-time awakenings, short sleep duration, and daytime dysfunction [1]. Sleep quality was affected by many factors in RA, including disease activity, degree of pain, socioeconomic status, and psychological disorders [4]. Previous data explored the role of specific cytokines in the regulation of sleep, including interleukin-1ß (IL-1ß) and tumor necrosis factor-α (TNF-α) [5]; however, the underlying mechanism for poor sleep is not fully known. Irisin was first described by Boström and others [6] as a myokine secreted from skeletal muscles after exercise’ it drives the conversion of brown fat from white adipose tissues. It has a beneficial role in energy expenditure and regulation of metabolism. Irisin is also secreted from other tissues like the liver, nerves, the heart, kidneys, and skin [7]. Recent reports described lower circulating irisin in obese persons and patients suffering from obstructive sleep apnea syndrome (OSAS) [8]. Serum irisin levels were also low in type II diabetes mellitus (DM) patients compared to nondiabetic controls [9]. Irisin is proteolytically cleaved from a fibronectin type III domain containing 5 (FNDC5) and is released into the circulation with translation and cleavage of FNDC5/irisin greatly enhanced by exercise in humans [10]. Irisin exerts its action through different pathways, including; anti-inflammatory, anti-apoptotic, and anti-oxidative mechanisms [11]. The anti-inflammatory role of irisin (which has relevance to combat the inflammatory process in RA) is achieved by many mechanisms including; suppressing secretion of necrosis
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factor ⱪß (NF-ⱪß), TNF-α, IL-6, and other pro-inflammatory cytokines from macrophages and adipocytes [12], decreasing recruitment of T lymphocytes and macrophages to atherosclerotic lesions, and preserving integrity of the endothelial cells [13]. To date, no study has reported the association between serum irisin levels and sleep quality in RA patients. In this preliminary study, we explore the association of serum irisin levels with sleep disorders and/or disease activity in RA patients. Methods The study was approved by the Medical Ethical Committee of Assiut University and included 58 RA patients and 30 age- and sex-matched healthy controls. RA patients fulfilled the American College of Rheumatology (ACR) classification criteria for RA [14] and attended the rheumatology and rehabilitation department at Assiut University Hospitals. Patients participated in the study after signing voluntary, fully-informed consent. Eligibility criteria included: age ≥ 18 years, stable medication usage in the last three months. The exclusion criteria included: neurological disorders that may affect sleep quality (including obstructive sleep apnea), patients on sleep-inducing medication, unstable thyroid disease, diabetes, unstable congestive heart failure, renal disease, and other connective tissue or autoimmune diseases. Demographic data were collected from RA patients and controls. The height, weight, and blood pressure of the participants were measured. In RA patients, disease activity status was assessed using the Disease Activity Score, including 28 joint counts (DAS28-ESR) [15]. Sleep quality was measured using the Pittsburgh Sleep Quality Index (PQSI) that measures the pattern and quality of sleep in adults by measuring seven domains: subjective sleep quality, sleep latency, sleep duration, habitual sleep efficiency, sleep disturbances, use of sleep medication, and daytime dysfunction over the last month. According to PQSI, “good-sleepers” are defined as a PQSI
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score=<5 and “poor sleepers” as a PSQI > 5 [16]. Healthy controls were chosen as "goodsleepers." Treatment with anti-rheumatic medications was reported in RA patients. Blood samples were collected at the time of clinic visits, centrifuged, and sera were stored at −20°C until examined. Laboratory investigations were performed for each participant and included: hemoglobin levels, white blood cells (WBCs) count, platelets count, serum glucose, erythrocyte sedimentation rate (ESR), and C-reactive protein (CRP). Routine liver, kidney, and thyroid function tests were performed six months apart from Irisin measurement. Serum irisin levels were measured by an enzyme-linked immunosorbent assay (Biovender, Czech Republic) with a detection range of 1-5000 ng/ml. The minimum concentration was one ng/ml. Data were analyzed using the Statistical Package for the Social Sciences (SPSS) version 20.0. Descriptive data included number and percent, mean ± SD. Student’s T-test and analysis of variance (ANOVA) were used for comparing means between different groups; Pearson correlation coefficient (r) was used to test the association between quantitative variables. A pvalue=<0.05 was considered statistically significant. No compensation was made for repeated analyses. Results RA patients were 54 females and four males; their mean age was 44.12±11.78 years (range 1871), and mean disease duration was 7.6±5.28 years (range 1-22). Healthy controls were 28 females (93%) and two males (7%), aged 41.37±10.9 years (range 23-67), with no significant differences between RA patients and controls regarding sex or age. RA patients were divided according to the PSQI score into two groups; group 1 were poor-sleepers (26 RA patients), and group 2 (32 RA patients) were good sleepers. RA patients with poor sleep had significantly longer morning stiffness (MS), worse global scores, and higher DAS28-ESR compared to good
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sleeper RA patients (p=<0.04 for all). Medications used were statistically different only for methotrexate and leflunomide (p<0.05) between the two RA groups (table 1). The serum levels of irisin were numerically but not statistically higher for RA patients receiving methotrexate treatment than RA patients not receiving methotrexate (139.5±59.6 vs. 127±78.4 ng/ml).
The total PSQI score was significantly worse (higher numbers) in both RA poor and good sleepers compared to the healthy control group (p=<0.001 for both). Sleep duration, sleep disturbance, sleep latency, day dysfunctions, sleep efficiency, and overall sleep quality were significantly higher (worse) in RA poor sleepers compared to RA good sleepers and healthy controls (p<0.001 for all). Also, RA good sleepers experienced more sleep disturbance, sleep latency, and day dysfunctions than healthy controls ((p =<0.007 for all) (table 2) ESR levels were significantly higher in RA patients (Groups 1 and 2) in contrast to controls as expected (p=0.006 and p=0.001, respectively), with no significant difference between RA groups (p=0.896). Platelet count was significantly higher in Group 2 (good sleepers) versus the healthy controls (p=0.042) (table 3). RA poor sleepers had significantly lower serum irisin levels than RA good sleepers (79.7±35.5 versus 173.34±67.06) (p=0.001). Levels of irisin in both RA groups were significantly lower than those in healthy controls (326.3±148.1) (p=<0.001 for all), all of whom were good sleepers (figure 1).
In a correlation analysis among RA patients, irisin concentrations were associated with total Pittsburgh Sleep Quality Index scores (Pearson correlation; r=-0.722, p=<0.001),. Circulating irisin levels were also negatively associated with disease duration (r=-0.263, p=.04), morning
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stiffness duration (r=-0.348, p=.006), DAS28-ESR (itself a composite representing inflammation/disease activity) (r=-0.298, p=0.02) and Global score (r=-0.263, p=0.04) (table 4)
Discussion Our preliminary study is the first to show that irisin levels are lower in RA than normals and lower in RA "poor sleepers" compared RA "good sleepers." In addition, irisin levels correlated with RA sleep quality, disease duration, and disease activity. Improving the quality of life is an important goal in RA, as RA patients have significant pain, fatigue, functional disabilities, depression, and poor sleep quality [1]. They have decreased general health, affecting physical and psychological activities [17]. Surrogate markers are needed that can be measured and tell if change/improvement has occurred or is needed in this aspect of RA patients’ lives. Based on our data, irisin could be a surrogate for RA inflammatory activity. Since its first description in 2012, irisin has attracted attention for its effects on muscles, cardiac cells, and brain [18, 19]. Irisin is an adipomyokine that regulates the metabolic rate of adipocytes and myocytes and acts as an exercise-induced insulin sensitizer [20]. Irisin may be anti-inflammatory [21, 22]. A report by Mazur-Bialy and colleagues [12, 23] demonstrated that pretreatment of LPS-stimulated macrophages with a low concentration of irisin reduced their inflammatory activity. They observed reduced release and expression of proinflammatory cytokines, such as IL-1ß, TNF-α, and IL-6. Further, they found that pretreatment with irisin significantly decreased the Toll-like receptor 4 (TLR4) expression and MyD88 protein levels in macrophages that consequently reduced the release of pro-inflammatory cytokines by
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macrophages in a dose- and time-dependent manner. Also, Dong et al. [24] reported that irisin inhibited IL-1ß, TNF-α, and IL-6. Irisin’s effect on inflammation may also be indirect. It is released into the circulation after exercise [6], and previous data documented the role of exercise in downregulation of the inflammatory process. For example, Wang et al. [25] reported increased expression of circulating T regulatory (Treg) cells, which are potent IL-10 producers and have an anti-inflammatory role, after exercise. Other studies reported that stimulated cytokine production by T helper 1 (Th1) lymphocytes is inhibited following exercise [26, 27] representing a decrease of cell-mediated immunity. Irisin may also be anti-inflammatory as it reduces reactive oxygen species (ROS) production, thereby modulating macrophage activity [28]. Exogenous irisin has been considered for treating inflammation [11]. Our study showed decreased serum irisin levels in RA patients with poor sleep quality compared to RA patients with good sleep quality. These findings could be mediated by the brain-derived neurotrophic factor (BDNF), as irisin enhances BDNF expression in brain tissues [19, 29-32], and exercise increases BDNF concentration in the brain [33, 34]. Exercise promoted a significant up-regulation of BDNF levels in older women with significant depression [35]. Kushikata et al. [36] studied BDNF and showed that BDNF increased the time spent in non-rapid eye movement sleep in rats and rabbits and rapid eye movement sleep in rabbits. Faraguna et al. [37] demonstrated a causal relationship between BDNF and sleep regulation. They infused BDNF into the cerebral cortex of awake rats; this induced a significant increase in sleep and increased slow-wave activity (SWA) in the injected side compared to the non-injected side. We did not measure serum BDNF. Future studies could include both irisin and BDNF to clarify the role of the irisin/BDNF cascade in sleep regulation in both the general population and in RA. Another
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possible explanation for the association of irisin with good sleep could be its role in alleviating the inflammatory process that is crucial in decreasing pain in RA patients. [38]. Irisin decreased inflammatory cell infiltration and counteracted inflammatory cytokines shown in a recent paper [39]. Researchers reported the effects of myokines including irisin that are secreted during exercise, in alleviating inflammation in some rheumatic diseases such as systemic sclerosis [40]. Previous data revealed inflammation to be associated with sleep disorders and quality of sleep in healthy individuals [41]. Reports showed alteration in the immune response after experimental deprivation of sleep with increased levels of inflammatory mediators [42]. Clearly, there are several factors to consider when interpreting data associating insufficient or disturbed sleep with inflammation. Methotrexate and leflunomide treatment did not affect irisin levels nor sleep structure in our RA patients, although RA patients treated with methotrexate showed numerically (not statistically) higher irisin levels than those not treated; a study with more patients might clarify this relationship. High dose corticosteroids may affect sleep. In our study, no patients used higher than 7.5 mg daily prednisone or its equivalent; thus, it is unlikely that corticosteroids had any effect on sleep in this study. As previously noted, the role of irisin in the sleep cascade needs further clarification, although its associations seem to make it an excellent candidate to be a surrogate marker for sleep quality. In a correlation analysis among our RA patients, the level of serum irisin was associated with disease duration, morning stiffness, DAS28-ESR, global score, sleep latency, and total PQSI score. These correlations were only modest but are very encouraging in this preliminary
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investigation, indicating that this surrogate marker was associated with increased inflammation (negatively correlated with global score and DAS28-ESR) [15, 21]. The present study has limitations. As noted above, BDNF was not measured, which was not the purpose of this study; nonetheless, it could have been useful data. The relatively small number of participants limits its generalizability, but the findings certainly are encouraging to prompt further study. Although neurological causes for sleep disturbance were excluded, we did not specifically inquire about RLS. The use of a specific insomnia questionnaire would also have been of interest, although the PSQI deals with this indirectly in several questions. This study was cross-sectional, therefore a longitudinal follow-up would be desirable. Also, the reproducibility of the irisin quantification has to be verified in such work. Although we employed a commonly used measure of irisin, the specificity of this methodology has been questioned by some [43, 44]; one author even questioned the existence of irisin [45], although we believe that is not really in doubt. In conclusion, we found that irisin concentrations were low in patients with RA, especially in those who sleep poorly. Our data revealed an association of irisin with sleep quality and disease activity in RA. We believe that further work clarifying irisin’s place in the sleep cascade is warranted for its potential to be a surrogate marker and, even, a potential therapeutic modality.
Acknowledgments We want to thank all the patients and their families, and the healthy controls, for their enthusiastic support during this research study.
Contributors
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RMG contributed to the study's concept and design, cases recruitment, acquisition and interpretation of data, statistical analyses, writing, and final revision of the article. MEM contributed to the study's concept and design, acquisition and interpretation of data, statistical analyses, writing, and final revision of the article. NH contributed to the study's design, cases recruitment, acquisition, and interpretation of data. NAF contributed to the draft and revision of the article. AMR contributed to cases recruitment, acquisition, and interpretation of data. DEF contributed to data acquisition and interpretation, statistical analysis, writing, and revision of the article. All authors revised and approved the final version of the manuscript . Funding None Declared
Competing interests The authors declare that there are no conflicts of interest to disclose.
Figure legends Figure 1. Serum irisin levels in RA patients (poor and good sleepers) and healthy controls p= 0.001 for RA poor sleepers vs. RA good sleepers, p<0.001 for RA poor sleepers vs. healthy controls and p<0.001 for RA good sleepers vs. healthy controls.
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Table 3. Laboratory data of RA patients and healthy controls
ESR value (mm) (normal<20 mm/h)
RA patients (no=60)
Healthy controls
Gr.1(Poor-sleepers) Gr.2=Good-sleepers
(all Good
P value
(N=28; 47%)
(N=32; 53%)
sleepers)(N=30)
35.23±15.75
37.25±18.7
20.8±16.6
0.896*
(16-70)
(3-81)
(3-68)
0.006§ 0.001¶
Hemoglobin level
12.58±1.35
12.34±1.77
11.65±1.7
0.848*
(g/dl)(Normal range:
(10.2-15.4)
(8.5-15.7)
(9.4-17)
0.091§ 0.222¶
male=14-16, female=12-14) WBC count
5.94±2.8
6.44±2.94
7.2±2.73
0.783*
(Normal=3.9-10.9 x 109/L)
(2.8-15.6)
(3.6-18.4)
(3.7-15)
0.204§ 0.505¶
Platelet count
298.4±94
303.8±66
256.8±67
0.960*
(Normal=150-400 x 109/L)
(144-551)
(197-464)
(169-410)
0.105§ 0.043¶
* represents p value for difference between group 1 and 2, § represents p value for difference between group 1 and controls, and ¶ represents p value for difference between group 2 and controls
Table 2. Sleep data of RA patients and healthy controls
Total PSQI score
RA patients (no=58)
Healthy controls
Gr.1(Poor-sleepers) Gr.2=Good-sleepers
(all Good
(N=26; 45%)
(N=32; 55%)
sleepers)(N=30)
8.5±2.44 (6-13)
2.97±1.23 (1-5)
0.5±0.94 (0-3)
P value <0.001* <0.001§
(range=0-13)
<0.001¶ Sleep duration
1±1.06
0.13±0.55
0.07±0.25
<0.001* <0.001§ 0.939¶
Sleep disturbance
1.77±0.43
1.19±0.4
0.13±0.35
<0.001* <0.001§ <0.001¶
Sleep latency
2.04±0.824
0.94±0.91
0.2±41
<0.001* <0.001§ 0.001¶
Day dysfunctions
1.54±0.86
0.44±0.5
0.0±0.0
<0.001* <0.001§ 0.007¶
Sleep efficiency
0.69±0.93
0.03±0.18
0.1±0.3
<0.001* <0.001§ 0.873¶
Overall sleep quality
1.38±0.94
0.25±0.51
0.0±0.0
<0.001* <0.001§ 0.230¶
Need med. to sleep:
2/26
0/32
0/30
0.085
yes/no * represents p value for difference between group 1 and 2, § represents p value for difference between group 1 and controls, and ¶ represents p value for difference between group 2 and controls
Table 1. Clinical data of RA patients and healthy controls RA patients (N=58)
Healthy controls P
Gr.1=Poor-sleepers
Gr.2=Good-sleepers
(all Good
(N=26; 45%)
(N=32; 55%)
sleepers)(N=30)
16 (61.5%)
7 (22%)
---
0.003
35 ± 42.7
9.37 ± 19.33
---
0.004
N of swollen joints
2.65 ± 3.46
2.47 ± 2.46
---
0.332
No of tender joints
8.65 ± 6.08
5.9 ± 5.5
---
0.288
23.27 ± 16.18
13.13 ± 11.7
---
0.008
4.63 ± 1.01
4.16 ± 1.11
---
0.101
0 (0%)
4 (12.5%)
Low activity (>2.6 ≤ 3.2)
2 (7.5%)
1 (3%)
---
0.04
Moderate (> 3.2 ≤ 5.1)
16 (61.5%)
24 (75%)
High activity (> 5.1)
8 (31%)
3 (9.5%)
Hypertension(BP>140)
5 (18%)
4 (12.5%)
4 (13%)
0.867
Methotrexate
5 (18%)
15 (47%)
---
0.015
Hydroxychloroquine
20 (77%)
25 (78%)
---
1.00
Leflunomide
19 (68%)
13 (41%)
---
0.042
Sulfasalazine
1 (3.6%)
4 (12.5%)
---
0.359
Combined
18 (64%)
23 (72%)
---
0.586
Steroids
8 (28.6%)
10 (31%)
---
1.00
NSAIDs
15 (57.7%)
10 (31%)
---
0.063
Morning stiffness (MS) N (%) MS duration (min)
Global score DAS-28 Remission (≤ 2.6)
Treatment:
Table 4. Correlation of serum irisin levels with clinical, PSQI score, and lab. parameters in RA patients (no=58) Item
r
p
Disease duration
-.263*
.042
MS duration
-.348**
.006
DAS-28
-.298*
.021
Global score
-.366**
.004
ESR
-.074
.573
Sleep latency
-.561**
<0.001
Total PSQI score
-.722**
<0.001
Correlation is significant at the 0.05 level (2-tailed)*
Abbreviations: DAS-28= Disease Activity Score including 28 joint counts; ESR=erythrocyte sedimentation rate; MS=morning stiffness; PSQI=Pittsburgh Sleep Quality Index
Figure 1. Serum irisin levels in RA patients (poor and good sleepers) and healthy controls p= 0.001 for RA poor sleepers vs RA good sleepers, p<0.001 for RA poor sleepers vs healthy controls and p<0.001 for RA good sleepers vs healthy controls.
Highlights • Sleep disorders are significant problems in patients with rheumatoid arthritis (RA. • Irisin is myokine and adipokine, which may be antiinflammatory and may help regulate energy expenditure. • This study evaluated the association of serum irisin levels with quality of sleep and disease activity in RA patients. • RA patients with poor sleep quality had decreased serum irisin levels compared to RA patients with good sleep quality.
• In patients with RA, the correlations of serum Irisin level with disease duration, morning stiffness, RA disease activity index and Pittsburgh Sleep Quality Index were evaluated.
S1389-9457(18)30858-X
DOI:
https://doi.org/10.1016/j.sleep.2019.10.021
Reference:
SLEEP 4230
To appear in:
Sleep Medicine
Received Date: 24 October 2018 Revised Date:
22 July 2019
Accepted Date: 17 October 2019
Please cite this article as: Gamal RM, Mohamed ME, Hammam N, El Fetoh NA, Rashed AM, Furst DE, Preliminary Study of the Association of Serum Irisin Levels with Poor Sleep Quality in Rheumatoid Arthritis Patients, Sleep Medicine, https://doi.org/10.1016/j.sleep.2019.10.021. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. 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. © 2019 Elsevier B.V. All rights reserved.
Preliminary Study of the Association of Serum Irisin Levels with Poor Sleep Quality in Rheumatoid Arthritis Patients
Short title: Irisin in RA patients
Rania M. Gamal1, Mona Embarek Mohamed2, Nevin Hammam1, Noha Abo El Fetoh3, Ahmed M. Rashed1, Daniel E. Furst4,5&6 1 Rheumatology& Rehabilitation Department, Assuit University Hospitals, Egypt. 71515. 2 Microbiology and Immunology Department, Faculty of Medicine, Assiut University, Assiut, Egypt.71515 3 Neurology and Psychiatry Department, Faculty of Medicine, Assiut University, Egypt. 4 Department of Medicine, Division of Rheumatology, University of California, Los Angeles (emeritus), Los Angeles, California. 5 Department of Rheumatology, Division of Rheumatology, University of Washington, Seattle Washington 6 Division of Rheumatology and Experimental Medicine, University of Florence, Florence, Italy.
Corresponding Author and offprint requests: Rania M. Gamal (MD) Associate Professor. Rheumatology & Rehabilitation Department, Assiut University, Assiut, Egypt. Postal code 71515 Fax: 02-0882333327 Mobile: 0020-1006289729 E-mail: [email protected]
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Abstract Study Objectives: Sleep disorders are significant problems in patients with rheumatoid arthritis (RA) and are associated with poor quality of life. Irisin is myokine which may have antiinflammatory and energy regulatory roles. This study assessed the association of serum irisin levels with the quality of sleep and disease activity in RA patients. Methods: Fifty-eight RA patients and 30 matched healthy controls were included. Disease activity score in 28 joints (DAS28-ESR) and the patients’ global score were calculated. RA patients were grouped according to the Pittsburgh Sleep Quality Index score (PSQI) into goodsleepers (group 1) defined as a PQSI score <=5 and poor sleepers (group 2) with a PSQI > 5. Serum irisin levels were measured for both patients and controls by commercially available enzyme-linked immunosorbent assay kits. Results: Poor sleep quality was found in 26 (45%) of the RA patients. Irisin levels were significantly lower in RA patients with poor sleep compared to those with good sleep and healthy controls (p<0.001). Serum irisin levels correlated inversely with disease duration, morning stiffness duration, DAS28-ESR, global score, and total PSQI score (r= -0.722 to -0.263 & p values <=0.001-0.04) indicating a possible anti-inflammatory role of irisin in RA patients. The analysis employed Student's t-test, ANOVA, and Pearson correlation. Conclusions: Irisin levels were decreased in RA patients with poor sleep quality compared to RA patients with good sleep quality and healthy controls, indicating a possible association of decreased serum irisin with sleep impairment in RA patients
Keywords: Irisin, Rheumatoid arthritis, Sleep, Disease activity.
.
2
Introduction Rheumatoid arthritis (RA) is a chronic disease characterized by inflammation in the synovial joints, leading to pain, swelling, morning stiffness, and joint deformities [1]. Patients with RA experience disturbed sleep that affects their health-related quality of life and physical or psychological function [2]. Sleep disturbances were reported in 54-70% of RA patients [3]; RA patients with poor sleep quality suffered from night-time awakenings, short sleep duration, and daytime dysfunction [1]. Sleep quality was affected by many factors in RA, including disease activity, degree of pain, socioeconomic status, and psychological disorders [4]. Previous data explored the role of specific cytokines in the regulation of sleep, including interleukin-1ß (IL-1ß) and tumor necrosis factor-α (TNF-α) [5]; however, the underlying mechanism for poor sleep is not fully known. Irisin was first described by Boström and others [6] as a myokine secreted from skeletal muscles after exercise’ it drives the conversion of brown fat from white adipose tissues. It has a beneficial role in energy expenditure and regulation of metabolism. Irisin is also secreted from other tissues like the liver, nerves, the heart, kidneys, and skin [7]. Recent reports described lower circulating irisin in obese persons and patients suffering from obstructive sleep apnea syndrome (OSAS) [8]. Serum irisin levels were also low in type II diabetes mellitus (DM) patients compared to nondiabetic controls [9]. Irisin is proteolytically cleaved from a fibronectin type III domain containing 5 (FNDC5) and is released into the circulation with translation and cleavage of FNDC5/irisin greatly enhanced by exercise in humans [10]. Irisin exerts its action through different pathways, including; anti-inflammatory, anti-apoptotic, and anti-oxidative mechanisms [11]. The anti-inflammatory role of irisin (which has relevance to combat the inflammatory process in RA) is achieved by many mechanisms including; suppressing secretion of necrosis
3
factor ⱪß (NF-ⱪß), TNF-α, IL-6, and other pro-inflammatory cytokines from macrophages and adipocytes [12], decreasing recruitment of T lymphocytes and macrophages to atherosclerotic lesions, and preserving integrity of the endothelial cells [13]. To date, no study has reported the association between serum irisin levels and sleep quality in RA patients. In this preliminary study, we explore the association of serum irisin levels with sleep disorders and/or disease activity in RA patients. Methods The study was approved by the Medical Ethical Committee of Assiut University and included 58 RA patients and 30 age- and sex-matched healthy controls. RA patients fulfilled the American College of Rheumatology (ACR) classification criteria for RA [14] and attended the rheumatology and rehabilitation department at Assiut University Hospitals. Patients participated in the study after signing voluntary, fully-informed consent. Eligibility criteria included: age ≥ 18 years, stable medication usage in the last three months. The exclusion criteria included: neurological disorders that may affect sleep quality (including obstructive sleep apnea), patients on sleep-inducing medication, unstable thyroid disease, diabetes, unstable congestive heart failure, renal disease, and other connective tissue or autoimmune diseases. Demographic data were collected from RA patients and controls. The height, weight, and blood pressure of the participants were measured. In RA patients, disease activity status was assessed using the Disease Activity Score, including 28 joint counts (DAS28-ESR) [15]. Sleep quality was measured using the Pittsburgh Sleep Quality Index (PQSI) that measures the pattern and quality of sleep in adults by measuring seven domains: subjective sleep quality, sleep latency, sleep duration, habitual sleep efficiency, sleep disturbances, use of sleep medication, and daytime dysfunction over the last month. According to PQSI, “good-sleepers” are defined as a PQSI
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score=<5 and “poor sleepers” as a PSQI > 5 [16]. Healthy controls were chosen as "goodsleepers." Treatment with anti-rheumatic medications was reported in RA patients. Blood samples were collected at the time of clinic visits, centrifuged, and sera were stored at −20°C until examined. Laboratory investigations were performed for each participant and included: hemoglobin levels, white blood cells (WBCs) count, platelets count, serum glucose, erythrocyte sedimentation rate (ESR), and C-reactive protein (CRP). Routine liver, kidney, and thyroid function tests were performed six months apart from Irisin measurement. Serum irisin levels were measured by an enzyme-linked immunosorbent assay (Biovender, Czech Republic) with a detection range of 1-5000 ng/ml. The minimum concentration was one ng/ml. Data were analyzed using the Statistical Package for the Social Sciences (SPSS) version 20.0. Descriptive data included number and percent, mean ± SD. Student’s T-test and analysis of variance (ANOVA) were used for comparing means between different groups; Pearson correlation coefficient (r) was used to test the association between quantitative variables. A pvalue=<0.05 was considered statistically significant. No compensation was made for repeated analyses. Results RA patients were 54 females and four males; their mean age was 44.12±11.78 years (range 1871), and mean disease duration was 7.6±5.28 years (range 1-22). Healthy controls were 28 females (93%) and two males (7%), aged 41.37±10.9 years (range 23-67), with no significant differences between RA patients and controls regarding sex or age. RA patients were divided according to the PSQI score into two groups; group 1 were poor-sleepers (26 RA patients), and group 2 (32 RA patients) were good sleepers. RA patients with poor sleep had significantly longer morning stiffness (MS), worse global scores, and higher DAS28-ESR compared to good
5
sleeper RA patients (p=<0.04 for all). Medications used were statistically different only for methotrexate and leflunomide (p<0.05) between the two RA groups (table 1). The serum levels of irisin were numerically but not statistically higher for RA patients receiving methotrexate treatment than RA patients not receiving methotrexate (139.5±59.6 vs. 127±78.4 ng/ml).
The total PSQI score was significantly worse (higher numbers) in both RA poor and good sleepers compared to the healthy control group (p=<0.001 for both). Sleep duration, sleep disturbance, sleep latency, day dysfunctions, sleep efficiency, and overall sleep quality were significantly higher (worse) in RA poor sleepers compared to RA good sleepers and healthy controls (p<0.001 for all). Also, RA good sleepers experienced more sleep disturbance, sleep latency, and day dysfunctions than healthy controls ((p =<0.007 for all) (table 2) ESR levels were significantly higher in RA patients (Groups 1 and 2) in contrast to controls as expected (p=0.006 and p=0.001, respectively), with no significant difference between RA groups (p=0.896). Platelet count was significantly higher in Group 2 (good sleepers) versus the healthy controls (p=0.042) (table 3). RA poor sleepers had significantly lower serum irisin levels than RA good sleepers (79.7±35.5 versus 173.34±67.06) (p=0.001). Levels of irisin in both RA groups were significantly lower than those in healthy controls (326.3±148.1) (p=<0.001 for all), all of whom were good sleepers (figure 1).
In a correlation analysis among RA patients, irisin concentrations were associated with total Pittsburgh Sleep Quality Index scores (Pearson correlation; r=-0.722, p=<0.001),. Circulating irisin levels were also negatively associated with disease duration (r=-0.263, p=.04), morning
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stiffness duration (r=-0.348, p=.006), DAS28-ESR (itself a composite representing inflammation/disease activity) (r=-0.298, p=0.02) and Global score (r=-0.263, p=0.04) (table 4)
Discussion Our preliminary study is the first to show that irisin levels are lower in RA than normals and lower in RA "poor sleepers" compared RA "good sleepers." In addition, irisin levels correlated with RA sleep quality, disease duration, and disease activity. Improving the quality of life is an important goal in RA, as RA patients have significant pain, fatigue, functional disabilities, depression, and poor sleep quality [1]. They have decreased general health, affecting physical and psychological activities [17]. Surrogate markers are needed that can be measured and tell if change/improvement has occurred or is needed in this aspect of RA patients’ lives. Based on our data, irisin could be a surrogate for RA inflammatory activity. Since its first description in 2012, irisin has attracted attention for its effects on muscles, cardiac cells, and brain [18, 19]. Irisin is an adipomyokine that regulates the metabolic rate of adipocytes and myocytes and acts as an exercise-induced insulin sensitizer [20]. Irisin may be anti-inflammatory [21, 22]. A report by Mazur-Bialy and colleagues [12, 23] demonstrated that pretreatment of LPS-stimulated macrophages with a low concentration of irisin reduced their inflammatory activity. They observed reduced release and expression of proinflammatory cytokines, such as IL-1ß, TNF-α, and IL-6. Further, they found that pretreatment with irisin significantly decreased the Toll-like receptor 4 (TLR4) expression and MyD88 protein levels in macrophages that consequently reduced the release of pro-inflammatory cytokines by
7
macrophages in a dose- and time-dependent manner. Also, Dong et al. [24] reported that irisin inhibited IL-1ß, TNF-α, and IL-6. Irisin’s effect on inflammation may also be indirect. It is released into the circulation after exercise [6], and previous data documented the role of exercise in downregulation of the inflammatory process. For example, Wang et al. [25] reported increased expression of circulating T regulatory (Treg) cells, which are potent IL-10 producers and have an anti-inflammatory role, after exercise. Other studies reported that stimulated cytokine production by T helper 1 (Th1) lymphocytes is inhibited following exercise [26, 27] representing a decrease of cell-mediated immunity. Irisin may also be anti-inflammatory as it reduces reactive oxygen species (ROS) production, thereby modulating macrophage activity [28]. Exogenous irisin has been considered for treating inflammation [11]. Our study showed decreased serum irisin levels in RA patients with poor sleep quality compared to RA patients with good sleep quality. These findings could be mediated by the brain-derived neurotrophic factor (BDNF), as irisin enhances BDNF expression in brain tissues [19, 29-32], and exercise increases BDNF concentration in the brain [33, 34]. Exercise promoted a significant up-regulation of BDNF levels in older women with significant depression [35]. Kushikata et al. [36] studied BDNF and showed that BDNF increased the time spent in non-rapid eye movement sleep in rats and rabbits and rapid eye movement sleep in rabbits. Faraguna et al. [37] demonstrated a causal relationship between BDNF and sleep regulation. They infused BDNF into the cerebral cortex of awake rats; this induced a significant increase in sleep and increased slow-wave activity (SWA) in the injected side compared to the non-injected side. We did not measure serum BDNF. Future studies could include both irisin and BDNF to clarify the role of the irisin/BDNF cascade in sleep regulation in both the general population and in RA. Another
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possible explanation for the association of irisin with good sleep could be its role in alleviating the inflammatory process that is crucial in decreasing pain in RA patients. [38]. Irisin decreased inflammatory cell infiltration and counteracted inflammatory cytokines shown in a recent paper [39]. Researchers reported the effects of myokines including irisin that are secreted during exercise, in alleviating inflammation in some rheumatic diseases such as systemic sclerosis [40]. Previous data revealed inflammation to be associated with sleep disorders and quality of sleep in healthy individuals [41]. Reports showed alteration in the immune response after experimental deprivation of sleep with increased levels of inflammatory mediators [42]. Clearly, there are several factors to consider when interpreting data associating insufficient or disturbed sleep with inflammation. Methotrexate and leflunomide treatment did not affect irisin levels nor sleep structure in our RA patients, although RA patients treated with methotrexate showed numerically (not statistically) higher irisin levels than those not treated; a study with more patients might clarify this relationship. High dose corticosteroids may affect sleep. In our study, no patients used higher than 7.5 mg daily prednisone or its equivalent; thus, it is unlikely that corticosteroids had any effect on sleep in this study. As previously noted, the role of irisin in the sleep cascade needs further clarification, although its associations seem to make it an excellent candidate to be a surrogate marker for sleep quality. In a correlation analysis among our RA patients, the level of serum irisin was associated with disease duration, morning stiffness, DAS28-ESR, global score, sleep latency, and total PQSI score. These correlations were only modest but are very encouraging in this preliminary
9
investigation, indicating that this surrogate marker was associated with increased inflammation (negatively correlated with global score and DAS28-ESR) [15, 21]. The present study has limitations. As noted above, BDNF was not measured, which was not the purpose of this study; nonetheless, it could have been useful data. The relatively small number of participants limits its generalizability, but the findings certainly are encouraging to prompt further study. Although neurological causes for sleep disturbance were excluded, we did not specifically inquire about RLS. The use of a specific insomnia questionnaire would also have been of interest, although the PSQI deals with this indirectly in several questions. This study was cross-sectional, therefore a longitudinal follow-up would be desirable. Also, the reproducibility of the irisin quantification has to be verified in such work. Although we employed a commonly used measure of irisin, the specificity of this methodology has been questioned by some [43, 44]; one author even questioned the existence of irisin [45], although we believe that is not really in doubt. In conclusion, we found that irisin concentrations were low in patients with RA, especially in those who sleep poorly. Our data revealed an association of irisin with sleep quality and disease activity in RA. We believe that further work clarifying irisin’s place in the sleep cascade is warranted for its potential to be a surrogate marker and, even, a potential therapeutic modality.
Acknowledgments We want to thank all the patients and their families, and the healthy controls, for their enthusiastic support during this research study.
Contributors
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RMG contributed to the study's concept and design, cases recruitment, acquisition and interpretation of data, statistical analyses, writing, and final revision of the article. MEM contributed to the study's concept and design, acquisition and interpretation of data, statistical analyses, writing, and final revision of the article. NH contributed to the study's design, cases recruitment, acquisition, and interpretation of data. NAF contributed to the draft and revision of the article. AMR contributed to cases recruitment, acquisition, and interpretation of data. DEF contributed to data acquisition and interpretation, statistical analysis, writing, and revision of the article. All authors revised and approved the final version of the manuscript . Funding None Declared
Competing interests The authors declare that there are no conflicts of interest to disclose.
Figure legends Figure 1. Serum irisin levels in RA patients (poor and good sleepers) and healthy controls p= 0.001 for RA poor sleepers vs. RA good sleepers, p<0.001 for RA poor sleepers vs. healthy controls and p<0.001 for RA good sleepers vs. healthy controls.
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[40] Lunetta C, Lizio A, Tremolizzo L, Ruscica M, Macchi C, Riva N, et al. Serum irisin is upregulated in patients affected by amyotrophic lateral sclerosis and correlates with functional and metabolic status. Journal of Neurology. 2018:1-8. [41] Mills PJ, von Känel R, Norman D, Natarajan L, Ziegler MG, Dimsdale JE. Inflammation and sleep in healthy individuals. Sleep. 2007;30:729-35. [42] Irwin MR, Wang M, Campomayor CO, Collado-Hidalgo A, Cole S. Sleep deprivation and activation of morning levels of cellular and genomic markers of inflammation. Archives of internal medicine. 2006;166:1756-62. [43] Timmons JA, Baar K, Davidsen PK, Atherton PJ. Is irisin a human exercise gene? Nature. 2012;488:E9-E10. [44] Spiegelman BM, Wrann C. Response to Comment on Wu and Spiegelman. Irisin ERKs the fat. Diabetes 2014;63:381-383. Diabetes. 2014;63:e17. [45] Albrecht E, Norheim F, Thiede B, Holen T, Ohashi T, Schering L, et al. Irisin – a myth rather than an exercise-inducible myokine. Scientific Reports. 2015;5:8889.
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Table 3. Laboratory data of RA patients and healthy controls
ESR value (mm) (normal<20 mm/h)
RA patients (no=60)
Healthy controls
Gr.1(Poor-sleepers) Gr.2=Good-sleepers
(all Good
P value
(N=28; 47%)
(N=32; 53%)
sleepers)(N=30)
35.23±15.75
37.25±18.7
20.8±16.6
0.896*
(16-70)
(3-81)
(3-68)
0.006§ 0.001¶
Hemoglobin level
12.58±1.35
12.34±1.77
11.65±1.7
0.848*
(g/dl)(Normal range:
(10.2-15.4)
(8.5-15.7)
(9.4-17)
0.091§ 0.222¶
male=14-16, female=12-14) WBC count
5.94±2.8
6.44±2.94
7.2±2.73
0.783*
(Normal=3.9-10.9 x 109/L)
(2.8-15.6)
(3.6-18.4)
(3.7-15)
0.204§ 0.505¶
Platelet count
298.4±94
303.8±66
256.8±67
0.960*
(Normal=150-400 x 109/L)
(144-551)
(197-464)
(169-410)
0.105§ 0.043¶
* represents p value for difference between group 1 and 2, § represents p value for difference between group 1 and controls, and ¶ represents p value for difference between group 2 and controls
Table 2. Sleep data of RA patients and healthy controls
Total PSQI score
RA patients (no=58)
Healthy controls
Gr.1(Poor-sleepers) Gr.2=Good-sleepers
(all Good
(N=26; 45%)
(N=32; 55%)
sleepers)(N=30)
8.5±2.44 (6-13)
2.97±1.23 (1-5)
0.5±0.94 (0-3)
P value <0.001* <0.001§
(range=0-13)
<0.001¶ Sleep duration
1±1.06
0.13±0.55
0.07±0.25
<0.001* <0.001§ 0.939¶
Sleep disturbance
1.77±0.43
1.19±0.4
0.13±0.35
<0.001* <0.001§ <0.001¶
Sleep latency
2.04±0.824
0.94±0.91
0.2±41
<0.001* <0.001§ 0.001¶
Day dysfunctions
1.54±0.86
0.44±0.5
0.0±0.0
<0.001* <0.001§ 0.007¶
Sleep efficiency
0.69±0.93
0.03±0.18
0.1±0.3
<0.001* <0.001§ 0.873¶
Overall sleep quality
1.38±0.94
0.25±0.51
0.0±0.0
<0.001* <0.001§ 0.230¶
Need med. to sleep:
2/26
0/32
0/30
0.085
yes/no * represents p value for difference between group 1 and 2, § represents p value for difference between group 1 and controls, and ¶ represents p value for difference between group 2 and controls
Table 1. Clinical data of RA patients and healthy controls RA patients (N=58)
Healthy controls P
Gr.1=Poor-sleepers
Gr.2=Good-sleepers
(all Good
(N=26; 45%)
(N=32; 55%)
sleepers)(N=30)
16 (61.5%)
7 (22%)
---
0.003
35 ± 42.7
9.37 ± 19.33
---
0.004
N of swollen joints
2.65 ± 3.46
2.47 ± 2.46
---
0.332
No of tender joints
8.65 ± 6.08
5.9 ± 5.5
---
0.288
23.27 ± 16.18
13.13 ± 11.7
---
0.008
4.63 ± 1.01
4.16 ± 1.11
---
0.101
0 (0%)
4 (12.5%)
Low activity (>2.6 ≤ 3.2)
2 (7.5%)
1 (3%)
---
0.04
Moderate (> 3.2 ≤ 5.1)
16 (61.5%)
24 (75%)
High activity (> 5.1)
8 (31%)
3 (9.5%)
Hypertension(BP>140)
5 (18%)
4 (12.5%)
4 (13%)
0.867
Methotrexate
5 (18%)
15 (47%)
---
0.015
Hydroxychloroquine
20 (77%)
25 (78%)
---
1.00
Leflunomide
19 (68%)
13 (41%)
---
0.042
Sulfasalazine
1 (3.6%)
4 (12.5%)
---
0.359
Combined
18 (64%)
23 (72%)
---
0.586
Steroids
8 (28.6%)
10 (31%)
---
1.00
NSAIDs
15 (57.7%)
10 (31%)
---
0.063
Morning stiffness (MS) N (%) MS duration (min)
Global score DAS-28 Remission (≤ 2.6)
Treatment:
Table 4. Correlation of serum irisin levels with clinical, PSQI score, and lab. parameters in RA patients (no=58) Item
r
p
Disease duration
-.263*
.042
MS duration
-.348**
.006
DAS-28
-.298*
.021
Global score
-.366**
.004
ESR
-.074
.573
Sleep latency
-.561**
<0.001
Total PSQI score
-.722**
<0.001
Correlation is significant at the 0.05 level (2-tailed)*
Abbreviations: DAS-28= Disease Activity Score including 28 joint counts; ESR=erythrocyte sedimentation rate; MS=morning stiffness; PSQI=Pittsburgh Sleep Quality Index
Figure 1. Serum irisin levels in RA patients (poor and good sleepers) and healthy controls p= 0.001 for RA poor sleepers vs RA good sleepers, p<0.001 for RA poor sleepers vs healthy controls and p<0.001 for RA good sleepers vs healthy controls.
Highlights • Sleep disorders are significant problems in patients with rheumatoid arthritis (RA. • Irisin is myokine and adipokine, which may be antiinflammatory and may help regulate energy expenditure. • This study evaluated the association of serum irisin levels with quality of sleep and disease activity in RA patients. • RA patients with poor sleep quality had decreased serum irisin levels compared to RA patients with good sleep quality.
• In patients with RA, the correlations of serum Irisin level with disease duration, morning stiffness, RA disease activity index and Pittsburgh Sleep Quality Index were evaluated.