Background
Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a debilitating disease characterized by persistent or relapsing unexplained fatigue of at least 6 months’ duration that is not relieved by rest and that causes a substantial reduction in daily activities [
1,
2]. Currently, the number of effective treatment options for ME/CFS is limited. Therefore, it is important for patients to engage in self-help strategies, such as pacing, to manage their symptoms.
Yoga is a representative mind-body therapy [
3] that has been recommended as a self-help tool for coping with ME/CFS [
2]. However, there was limited evidence on its efficacy [
4] until we investigated the effects of isometric yoga on patients with ME/CFS [
5,
6]. Isometric yoga is a yoga program that was developed by Oka et al. specifically for patients with ME/CFS. To date, we have developed two types of isometric yoga programs, enabling patients to practice poses with a self-adjustable isometric load in a seated [
5] or recumbent [
6] position. The recumbent isometric yoga program was designed for patients who cannot sit for a long period of time and who were reluctant to practice a seated isometric yoga program. We demonstrated that regular practice of both seated [
5] and recumbent [
6] isometric yoga reduced the fatigue of these patients, according to subjective self-reported measures.
The reason for developing these isometric yoga programs was based on findings that showed the positive effects of yoga on breast cancer patients and survivors. Although these yoga programs differed from that used in our studies, the combined results demonstrated that regular yoga practice can reduce fatigue and improve psychological health outcomes, e.g. reduction of perceived stress, anxiety, depression, and insomnia (for a review, see [
7,
8]). This suggested that these yoga-induced effects might also be beneficial for patients with ME/CFS. However, based on our clinical experience, it seemed that patients with ME/CFS would have difficulty practicing yoga programs similar to those designed for patients with breast cancer or healthy individuals. In fact, some patients actually reported experiencing exacerbated fatigue and pain after attending a public yoga class and attempting the same poses as healthy participants. Therefore, we discussed these issues with yoga instructors of the Japan Yoga Therapy Society and with them developed two isometric yoga programs for patients with ME/CFS, taking into consideration the pathophysiological characteristics of the disease and the limitations of the patients. First, given that the patients have severe fatigue and their condition changes moment to moment, we selected easy isometric poses so that patients could adjust the muscular strength they apply depending on their present condition. Second, as patients have orthostatic intolerance, we intentionally did not include any standing postures. Third, given that the patients have severe pain, we avoided poses that would require flexibility and active stretching. Fourth, given that the patients have impaired concentration and short-term memory, we kept the programs as simple as possible. Fifth, because some patients are deconditioned, we designed the programs to help in the prevention of disuse muscular atrophy by including isometric loading. Most importantly, because the patients experience post-exertional malaise, we also designed the programs to enhance the awareness of inner sensations (interoception and proprioception), so that the patients can be more sensitive and aware of the thresholds that induce post-exertional malaise and to keep their daily activities within their personal limits. Furthermore, we asked the yoga instructor to pay careful attention to the patient’s physical condition to avoid exacerbation of symptoms or post-exertion malaise and permitted the instructor to divide and modify the program if necessary. We also asked the instructor to adjust the brightness of ceiling lights and to not play music, use perfume, or touch participants when giving instruction because they may have hyper-sensitivity to light, sound, or smell, and may have allodynia. Given that the severity of symptoms differed from person to person, the yoga instruction was not performed in a group setting but instead on a one-to-one basis. It should be noted that this yoga program was not developed as another option for graded exercise therapy but rather as an exercise to induce post-isometric relaxation [
9,
10] and to facilitate awareness for more efficient pacing.
With this newly developed program, we sought to investigate the underlying mechanisms for the fatigue-improving effects of isometric yoga. While the pathophysiological mechanisms of ME/CFS are not yet fully understood, previous studies have suggested several abnormalities in patients with ME/CFS, including dysfunction of the autonomic nervous system, hypothalamic-pituitary-adrenocortical (HPA) axis, and immune system [
11]. Autonomic dysfunction is characterized by low vagal tone and sympathetic overactivity [
12,
13]. Dysfunctions of the HPA axis include attenuated diurnal changes in cortisol [
14] and decreased levels of dehydroepiandrosterone sulfate (DHEA-S) [
15]. Dysfunctions of the immune system include increased blood levels of transforming growth factor (TGF)-β1 [
16] and proinflammatory cytokines such as tumor necrosis factor (TNF)-α [
17,
18]. In our previous study, we demonstrated that seated isometric yoga resulted in improvements in some of these abnormalities, including increased DHEA-S serum levels and the high frequency component of heart rate variability, suggesting an increase in parasympathetic function,and reduced cortisol and TNF-α serum levels in patients with ME/CFS [
19]. Therefore, these changes might be associated with the therapeutic mechanisms of isometric yoga on ME/CFS. However, recent findings on the biological abnormalities of patients with ME/CFS, such as peripheral cytokines ([
20]; for review, see [
21,
22]); are not necessarily consistent. Thus, caution should be taken when interpreting the role of isometric yoga on inducing these fatigue-relieving effects.
To further investigate the mechanisms behind the fatigue-relieving effects of practicing isometric yoga, we extended our study to assess changes in a new class of biomarkers, microribonucleic acids (miRNAs). Recent studies have suggested that miRNAs are involved in ME/CFS [
23‐
25]. miRNAs are single-stranded, non-coding small RNAs, 18–25 nucleotides in length, that bind to the 3’UTRs of target messenger RNAs (mRNAs) and contribute to gene silencing by suppressing protein production or facilitating mRNA degradation [
26,
27]. miRNAs play a critical role in various biological processes, such as cell differentiation, development and homeostasis [
28‐
30], and circulating miRNAs have been proposed as biomarkers for some medical conditions [
31‐
33]. Several studies have demonstrated differences between patients with ME/CFS and non-fatigued control subjects in the levels of miRNA expression in peripheral blood mononuclear cells [
25], cytotoxic lymphocytes [
23], and plasma [
24]. These studies proposed the use of miRNAs as potential biomarkers of ME/CFS [
24,
25].
To date, however, it is not known if yoga affects the miRNA expression of patients with ME/CFS. Therefore, we sought to undertake this exploratory pilot study to investigate if the regular practice of isometric yoga affects the serum miRNA expression of patients with ME/CFS.
Discussion
To our knowledge, this is the first report to suggest that recumbent isometric yoga alters the serum miRNAs of patients with ME/CFS. We demonstrated that regular practice of recumbent isometric yoga reduced the CFQ 11 scores of patients with ME/CFS. Several studies have already suggested the beneficial effects of yoga, although the type of yoga was different from that used in our studies, in at least a subset of patients with ME/CFS [
4‐
6,
19] [
46]. In a case report of one patient with CFS, yoga-based lifestyle intervention improved their clinical profile, positive aspects of personality, and subjective well-being, in addition to reducing anxiety [
4]. In our previous study we conducted a randomized, controlled trial to assess the effect of isometric yoga on patients with ME/CFS and found that isometric yoga practiced in a seated position improved fatigue and pain in conventional treatment-resistant patients with ME/CFS who were able to sit for at least 30 min [
5]. We also developed a recumbent isometric yoga program for patients who found it difficult to sit for long periods of time and were reluctant to practice seated isometric yoga. We found that practicing recumbent yoga for 3 months decreased the CFQ 11 scores of these patients [
6]. Participants in the present study were a different cohort than those in our previous study [
6]; however, we obtained the same results. These studies suggest that regular practice of recumbent isometric yoga reduces the fatigue of patients with ME/CFS. However, the present results should be interpreted carefully because, as with patient 5, it is possible that the questionnaire may have been misunderstood. Furthermore, as was previously pointed out [
47], when evaluating the changes of patients with a high score (near 33), the potential for a ceiling effect must also be taken into consideration.
The present study also demonstrated changes in several serum miRNA levels of patients with ME/CFS after practicing recumbent isometric yoga for 3 months. Serum miRNAs, which are known to be stably encapsulated in vesicles and secreted or excreted from cells or tissues, have been proposed as clinical biomarkers, such as in cancer detection [
31‐
33]. In the present study, we found that 4 miRNAs were upregulated, and 42 miRNAs were downregulated in serum samples after practicing recumbent isometric yoga. However, these miRNAs do not correspond with the miRNAs reported to be differentially expressed in patients with ME/CFS compared to healthy subjects. For example, Brenu et al. demonstrated that plasma levels of miR-127-3p, miR-142-5p and miR143-3p were upregulated in patients with ME/CFS in comparison to non-fatigued control subjects [
24]. They also found that miR-21 and several other miRNAs were decreased in the natural killer (NK) and CD8
+ T cells of ME/CFS patients [
23]. Another study reported that 34 miRNAs were upregulated in peripheral blood mononuclear cells of patients with ME/CFS, and that four miRNAs, miR-99b, miR-330, miR-126, and miR-30c, may be useful biomarkers of ME/CFS [
25]. Our study failed to replicate the results of these previous studies in that we did not observe changes in these miRNAs after recumbent isometric yoga. Therefore, we must be cautious in our interpretation of the miRNAs that were changed after recumbent isometric yoga and whether they actually affect the pathophysiological mechanisms of ME/CFS.
The reasons for this discrepancy are unclear. However, there are several possible explanations. One is that isometric yoga improves fatigue not by reversing the mechanisms that are involved in the development of ME/CFS but by acting on other mechanisms, e.g. enhancing fatigue-relieving mechanisms, which are not directly responsible for the pathophysiology of ME/CFS. Another reason could be related to the different sample types that were used in the different studies [
23‐
25]. These various sample types, i.e. plasma, serum or NK cells, could have different miRNA profiles, thus the findings might not be readily comparable. Additionally, the observed differences might also result from varied dietary habits. Recent studies have reported that different dietary habits affect the plasma miRNA expression of healthy individuals [
48]. Because this study was conducted in Japan and previous studies were conducted in Australia [
23,
24] and the United Kingdom [
25], it is possible that Japanese dietary habits affected the outcomes.
Among the upregulated miRNAs, miR-19a, the second most upregulated miRNA, is of particular interest because it is known to target TNF-α [
49]. Although the findings are inconsistent [
21,
22], immunological abnormalities, including increased TNF-α [
50], have been documented in patients with ME/CFS [
50]. In general, practicing yoga is known to shift autonomic nervous system function from a sympathetic nerve-predominant state to a parasympathetic nerve-predominant state and to reduce inflammatory markers, such as TNF-α, interleukin-1 (IL-1), or nuclear factor kappa B (NF-κB), as well as reduce stress hormones [
51‐
53]. Increased TNF-α and IL-1 are reported to be correlated with fatigue, sadness, autonomic symptoms, and a flu-like malaise in these patients [
18]. We previously observed that seated isometric yoga significantly reduced the TNF-α serum level [
19]. Therefore, the upregulation of miR-19a expression by isometric yoga might be associated with a reduction of fatigue through the decreased production of TNF-α. Additionally, yoga has been shown to reduce methylation of the TNF region of DNA in chronically stressed women [
54]. Therefore, yoga-induced changes in DNA methylation might also contribute to a decrease in TNF-α. Furthermore, vagal nerve activation by yoga [
19], which is impaired in ME/CFS patients [
55,
56], might also contribute to a decrease in the serum TNF-α level via vagal anti-inflammatory pathways [
57].
miR-485 [
58] as well as miR-19a [
59] have inhibitory effects on TGF-β signaling and the expression levels of TGF-β1. Several studies, but not all [
60], have reported that the blood level of TGF-β1 of patients with ME/CFS was higher than in healthy subjects [
16]. We have previously observed a correlation between the reduction in fatigue by seated isometric yoga with a reduction in the plasma level of TGF-β1 [
39]. Therefore, miR-485 might play a beneficial role via the inhibition of TGF-β signaling. The role of miR-668, which was the most upregulated after yoga, in ME/CFS remains uncertain. Previous studies have indicated that it plays a protective role in acute kidney injury by repressing mitochondrial fission-associated protein [
61,
62] and enhances resistance in breast cancer cells [
63].
In contrast to the upregulated miRNAs, miR-4273, which was the most downregulated in this study, has been shown to inhibit prostate cancer growth through regulation of Abelson kinase [
64]. miR-940, another downregulated miRNA, can promote angiogenic abilities of cerebral microvascular endothelial cells after cerebral infarction [
65] and induce osteoblastic phenotypes in the bone metastatic microenvironment [
66]. Although the functions of most of the downregulated miRNAs remain unknown, these miRNAs could also be related to the fatigue-relieving mechanisms in ME/CFS.
There are several limitations in the present study. First, we were unable to blind the treatment and did not have a control group for comparison. Because miRNA expression can change due to fluctuations in lifestyle factors such as diet [
48], exercise [
67], or sleep [
68,
69], it is undeniable that the present results could be affected by these or other unknown factors. Second, we did not perform corrections for multiple comparisons. Therefore, the data in the current study should be considered exploratory pilot data. Third, the sample size was small. Because it was not certain if recumbent isometric yoga affects circulating miRNAs, this study was conducted as a pilot study to assess if recumbent isometric yoga actually affects circulating miRNAs. Because this study suggests that isometric yoga can change the serum miRNA level, future studies should attempt to replicate the present results with a larger sample size. Fourth, we did not validate the expression of miRNAs using quantitative real-time polymerase chain reaction. Therefore, the possibility remains that there may be slight deviations from the expression levels observed in the blood. Finally, we could not investigate the actual targets of the affected miRNAs in this clinical study. To identify the targets of these miRNAs, further basic research is needed. There is, of course, a limitation on using the CFQ 11, a subjective measure, to evaluate fatigue because it is possible that the scores could be affected by an expectancy effect or a ceiling effect. Therefore, future studies should include objective measures in addition to the CFQ 11 to assess the fatigue-relieving effects of recumbent isometric yoga.
Despite these limitations, this is the first study to show that isometric yoga impacts the expression of miRNAs. It also suggests that these altered miRNAs may have roles in improving ME/CFS and could represent biomarkers for improvement of ME/CFS symptoms by isometric yoga.
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