Qigong is wellness practice, a system of disciplines. Coordinated physical postures, movements, breathing techniques, and meditations used for rehabilitative purposes, health cultivation, martial arts training and in some cases as a spiritual practice. Qigong takes roots in ancient Chinese culture, Traditional Chinese Medicine and martial arts dating back more than 4,000 years.

Qigong ((/ˈtʃiːˈɡɒŋ/), qi gong, chi kung, or chi gung) means "The Practice of balancing and cultivating Qi".


Qi = Essence/Energy

Qi = Yin (female essence) + Yang (male essence)

Qi stands for the energy in everything. The life-force that everything and every person has.

Qi is the central underlying principle in Traditional Chinese Medicine and Acupuncture.

The practice of balancing and cultivating Qi is called Qigong.

My interest in Qigong has always been for clinical purposes, for its wide range of clinical applications, and self maintenance, health and stamina cultivation. During last few decades a strong body of research has been emerging regarding the therapeutic effects of Qigong.

Medical Qigong is used in Traditional Chinese Medicine for preventative, rehabilitative and healing acceleration. It is used as a physical rehabilitation modality, manual therapy applications to address physical body and physical energies; and a somatic/meditative therapy to address the mental/emotional aspect of the person.

It is believed that strong essence lives only in a strong body and strong body always manifests strong essence. It makes total sense. This is why in Qigong we train and discipline the body in tendon with training and disciplining of the mind.

Through diligent training, serious students achieve high levels of physical and mental awareness, concentration and ability to balance the body and bodily processes with the mind and mental processes. This position is known as the sweet spot of qigong. This is where stress, anxieties, panic attacks, depression, fatigue can not exist. This sweet spot is where you are in optimal state. This is where the body enters into total homeostasis. This is where health lives and where health is cultivated.

Qigong sessions incorporate a wide range of breathing exercises coupled with sitting or standing meditation postures as well as either gentle or vigorous body shaking, physical movements, including slow, meditative, flowing, dance-like motions. All done with the purposeful regulation of both breath and mind coordinated with the regulation of the body.

2-1. Introduction

Before you start your qigong training, you must first understand the three treasures of life: essence (jing, 精), internal energy (qi, 氣), and spirit (shen, 神)—as well as their interrelationship. If you lack this understanding, you are missing the root of qigong train- ing, as well as the basic idea of qigong theory. The main goals of qigong training are to learn how to retain your jing, strengthen and smooth your qi flow, and enlighten your shen. To reach these goals you must learn how to regulate the body (tiao shen, 調身), regulate the mind (tiao xin, 調心), regulate the breathing (tiao xi, 調息), regulate the qi (tiao qi, 調氣), and regulate the shen (tiao shen, 調神).

Regulating the body includes understanding how to find and build the root of the body as well as of the individual forms you are practicing. To build a firm root, you must know how to keep your center, how to balance your body, and most important of all, how to relax so that the qi can flow.

Regulating the mind involves learning how to keep your mind calm, peaceful, and centered, so that you can judge situations objectively and lead qi to the desired places. The mind is the main key to success in qigong practice.

To regulate your breathing, you must learn how to breathe so that your breathing and your mind mutually correspond and cooperate. When you breathe this way, your mind will be able to attain peace more quickly, and therefore concentrate more easily on leading the qi.

Regulating the qi is one of the ultimate goals of qigong practice. In order to regulate your qi effectively, you must first have regulated your body, mind, and breathing. Only then will your mind be clear enough to sense how the qi is distributed in your body and understand how to adjust it.

For Buddhist priests, who seek the enlightenment of the Buddha, regulating the shen is the final goal of qigong. This enables them to maintain a neutral, objective perspective of life, and this perspective is the eternal life of the Buddha. The average qigong practitio- ner has lower goals. He raises his shen in order to increase his concentration and enhance his vitality. This makes it possible for him to lead qi effectively to his entire body so that it carries out the managing and guarding duties. This maintains his health and slows down the aging process.


30 Simple Qigong Exercises for Health

If you understand these few things you will be able to quickly enter into the field of qigong. Without all of these important elements, your training will be ineffective and your time will be wasted.

2-2. Three Treasures (Jing, Qi, Shen, 三寶-精,氣,神)

Before you start any qigong training, you must first understand the three treasures (san bao, 三寶): essence (jing, 精), internal energy (qi, 氣), and spirit (shen, 神). They are also called the three origins or the three roots (san yuan, 三元) because they are con- sidered the origins and roots of your life. Jing means the essence, the most original and refined part of every thing. Jing exists in everything. It represents the most basic part of anything that shows its characteristics. Sperm is called jing zi (精子), which means “essence of the son,” because it contains the jing of the father, which is passed on to his son (or daughter) and becomes the child’s jing. Jing is the original source of every living thing, and it determines the nature and characteristics of that thing. It is the root of life.

Qi is the internal energy of your body. It is like the electricity that passes through a machine to keep it running. Qi comes either from the conversion of the jing that you have received from your parents, or from the food you eat and the air you breathe.

Shen is the center of your mind, the spirit of your being. It is what makes you human because animals do not have a shen. The shen in your body must be nourished by your qi or energy. When your qi is full, your shen will be enlivened.

These three elements are interrelated in a number of ways. Chinese meditators and qigong practitioners believe that the body contains two general types of qi. The first type is called original qi (yuan qi, 元氣) or pre-birth qi (xian tian qi, 先天氣), and it comes from converted original jing (yuan jing, 元精), which you get from your parents at con- ception. The second type, which is called post-birth qi (hou tian qi, 後天氣), is drawn from the jing of the food and air we take in. When this qi flows or is led to the brain, it can energize the shen and soul. This energized and raised shen is able to lead the qi to the entire body.

Each one of these three elements or treasures has its own root. You must know the roots so that you can strengthen and protect your three treasures:

1. There are many kinds of jing that your body requires. Except for the jing that you inherit from your parents, which is called original jing (yuan jing, 元精), all other jings must be obtained from food, air, or even the energy surrounding you. Among all of these jings, original jing is the most important one. It is the root and the seed of your life, and your basic strength. If your parents were strong and healthy, your original jing will be strong and healthy, and you will have a strong foundation on which to grow. The Chinese people believe that in order to stay healthy and live a long life, you must protect and maintain this jing. It is like money that you have

Chapter 2. Qigong Training Theory 31invested in a bank. You can live off the interest for a long time, but if you withdraw

the principal and spend it, you will have nothing left.

The root of original jing before your birth was in your parents. After birth this origi- nal jing stays in its residence—the kidneys, which are considered the root of your jing. When you keep this root strong, you will have sufficient original jing to supply to your body. Although you cannot increase the amount of jing you have, qigong training can improve the quality of your jing. Qigong can also teach you how to convert your jing into original qi more efficiently and how to use this qi effectively.

  1. Qi is converted both from the jing that you have inherited from your parents and from the jing that you draw from the food and air you take in. Qi which is con- verted from the original jing, which you have inherited, is called original qi (yuan qi, 元氣).1 Just as original jing is the most important type of jing, original qi is the most important type of qi. It is pure and of high quality, while the qi from food and air may make your body too positive or too negative, depending on how and where you absorb it. When you retain and protect your original jing, you will be able to generate original qi in a pure, continuous stream. As a qigong practitioner, you must know how to convert your original jing into original qi in a smooth, steady stream.

    Since your original qi comes from your original jing, they both have the kidneys for their root. When your kidneys are strong, the original jing is strong, and the origi- nal qi converted from this original jing will also be full and strong. This qi resides in the lower dan tian in your abdomen. Once you learn how to convert your original jing, you will be able to supply your body with all the qi it needs.

  2. Shen is the force that keeps you alive. It has no substance, but it gives expression and appearance to your jing. Shen is also the control tower for the qi. When your shen is strong, your qi is strong and you can lead it efficiently. The root of shen (spirit) is your mind (yi, 意) or intention. When your brain is energized and stimulated, your mind will be more aware and you will be able to concentrate more intensely. Also, your shen will be raised. Advanced qigong practitioners believe that your brain must always be sufficiently nourished by your qi. It is the qi that keeps your mind clear and concentrated. With an abundant qi supply, the mind can be energized and can raise the shen and enhance your vitality.

    The deeper levels of qigong training include the conversion of jing into qi, which is then led to the brain to raise the shen. This process is called “huan jing bu nao” (還精補腦) and means “return the jing to nourish the brain.” When qi is led to the head, it stays at the upper dan tian (center of the forehead). The upper dan tian is the residence of shen. Qi and shen are mutually related. When your shen is weak,


Simple Qigong Exercises for Health

your qi is weak, and your body will degenerate rapidly. Shen is the headquarters of qi. Likewise, qi supports the shen, energizing it and keeping it sharp, clear, and strong. If the qi in your body is weak, your shen will also be weak.

2-3. Qigong Training Theory

In qigong training, you must understand the principle behind everything you are doing. The principle is the root of your practice, and it is this root that brings forth the results you want. The root gives life, while the branches and flowers (results) give only temporary beauty. If you keep the root, you can regrow. If you have just branches and flowers, they will die in a short time.

Every qigong form or practice has its special purpose and theory. If you do not know the purpose and theory, you have lost the root (meaning) of the practice. Therefore, as a qigong practitioner, you must continue to ponder and practice until you understand the root of every set or form.

Before you start training, you must first understand that all of the training originates in your mind. You must have a clear idea of what you are doing, and your mind must be calm, centered, and balanced. This also implies that your feeling, sensing, and judgment must be objective and accurate. This requires emotional balance and a clear mind. This takes a lot of hard work, but once you have reached this level, you will have built the root of your physical training, and your yi will be able to lead your qi throughout your physical body.

As mentioned previously, qigong training includes five important elements: regulating the body, regulating the breath, regulating the mind, regulating the qi, and regulating the spirit (shen). These elements are the foundation of successful qigong practice. Without this foundation, your understanding of qigong and your practice will remain superficial.

1. Regulating the Body (Tiao Shen, 調身)
Regulating the body is called “tiao shen” (調身) in Chinese. This means to adjust

your body until it is in the most comfortable and relaxed state. This implies that your body must be centered and balanced. If it is not, you will be tense and uneasy, and this will affect the judgment of your yi and the circulation of your qi. In Chinese medical society it is said: “(When) shape (body’s posture) is not correct, then the qi will not be smooth. (When) the qi is not smooth, the yi (mind) will not be peaceful. (When) the yi is not peaceful, then the qi is disordered.”2 You should understand that the relaxation of your body originates with your yi. Therefore, before you can relax your body, you must first relax or regulate your mind (yi). This is called “shen xin ping heng” (身心平衡), which means “body and heart (mind) balanced.” The body and the mind are mutually related. A relaxed and balanced body helps your yi to relax and concentrate. When your

While some of the studies compared Qigong or Tai Chi to other forms of exercise (n=13), many compared to a non-exercise treatment control group such as education or usual care (n=43) and some used both exercise and non-exercise comparison groups to evaluate effects of Qigong or Tai Chi interventions (n=11). Most studies included healthy adults (n=16 studies), but several studies included participants based on specific risk factors or diagnosis of disease including: arthritis (n=5); heart disease (n=6); hypertension (n=5); osteoporosis risk [e.g., peri-menopausal (n=3)]; fall risk determined by age and sedentary lifestyle or poor physical function and balance (n=18); breast cancer (n=1); depression (n=2); fibromyalgia (n=2); immune dysfunction including HIV/AIDS and varicella history or vaccine response (n=3); muscular dystrophy (n=1); Parkinson’s disease (n=1); neck pain (n=1); sleep complaints (n=1); chronic disease (n=1); and traumatic brain injury (n=1). Some of the studies (n=9) monitored adverse effects during the interventions and none reported an adverse event.

The studies originated from 13 countries (USA, n=34; China (including Hong Kong), n=9; Korea, n=4; Australia and New Zealand, n=5; Sweden, n=4; Great Britain, n=3; Italy and Taiwan, each n=2; Netherlands, Israel, Poland, and Spain, each n=1).

Resistance training and other weight bearing exercises are known to increase bone formation and have been recommended for post-menopausal women for that purpose. Interestingly, most Qigong practices involve no resistance and only minimal weight bearing (such as gentle knee bends), yet the four RCTs (total sample size=427) included in this review reported positive effects on bone health. One study examined the effect of Qigong40 and three examined Tai Chi.4143 Bone loss was retarded and numbers of fractures were less among post-menopausal women practicing Tai Chi compared to usual care.41 In another study, bone loss was less pronounced for post-menopausal females practicing Tai Chi or resistance training compared to no-exercise controls, but this effect was not found in the older men participating in the study.43 Shen and colleagues42 compared Tai Chi to resistance training and reported significant changes in biomarkers of bone health in both groups. Bone mineral density increased for women following Qigong exercises as compared to no-exercise controls.40 In summary, current research suggests a favorable effect on bone health for those practicing Tai Chi or Qigong.


Nineteen studies [Qigong (n=7) and Tai Chi (n=12)] reported favorable cardiovascular and/or pulmonary outcomes. Participants in this grouping of studies were generally older adults (mean age=61.02) and inclusion criteria varied from history of disease to reported sedentary behavior. Measures of cardiopulmonary function were representative of cardiopulmonary fitness and cardiovascular disease risk and included blood pressure, heart rate, ejection fraction rates, blood lipids, 6-minute walk distance, ventilatory function, and body mass index (BMI).

One of the most consistent findings was the significant reduction in blood pressure reported in multiple studies, especially when Qigong4445 or Tai Chi4647 were compared to inactive control groups such as usual care, educational classes, or wait-list controls. Even when compared to active control groups such as aerobic exercise or balance training, Tai Chi showed a significant reduction in blood pressure in two studies.4849 Other studies, however, that utilized active control interventions (low to moderate levels of physical activity) expected to reduce blood pressure showed positive changes for both groups, but without significant differences between Qigong28 or Tai Chi5051 and the comparison group; thus, providing preliminary evidence that these meditative movement practices achieve similar results to conventional exercise.

Other indicators of cardiac health have been evaluated. Reduced heart rate is reported495152as well as increases in heart rate variability.53 These reported changes in blood pressure, heart rate, and heart rate variability suggest that one or several of the key components of Tai Chi and Qigong, that is body, breath, and mind, may affect sympathetic and parasympathetic balance and activity.

Biomarkers of heart-health have been shown to improve in response to Qigong or Tai Chi practice. Yeh and colleagues34 reported significantly improved Serum B-type natriuretic peptide levels in response to Tai Chi compared to usual care controls, indicating improved left ventricular function. Lipid profiles improved in two studies4446 comparing Qigong and Tai Chi to inactive controls while another study of Qigong54 reported no change in cholesterol levels compared to inactive (wait-list) controls. Pippa and colleagues54 also reported no change in ejection fraction rates following a 16-week study of Qigong among participants with a history of chronic atrial fibrillation. Urine catecholamine levels were significantly decreased in participants practicing Tai Chi compared to wait-list controls45 but a similar trend did not reach significance in another study with only 15 participants per treatment condition.34

A variety of cardiopulmonary fitness indicators have been examined for both Qigong and Tai Chi. Participants with a history of heart failure reported significant improvements in the incremental shuttle walk following a combined Tai Chi/Qigong intervention implemented in two studies incorporating inactive control groups.3455 Women treated for breast cancer achieved significantly increased distances in the 6-minute walk test in response to Tai Chi compared to a psychosocial support control intervention56 and VO2 max increased significantly more following a Tai Chi intervention compared to resistance training and usual care control groups.53 In contrast to these consistent findings for cardiopulmonary benefits, one study found no significant improvement in response to Qigong, while aerobic training did achieve significant changes. In this small (n=11 in each arm of study) cross-over study of patients with Parkinson’s disease, participants practiced Qigong or aerobic training in random order for 7 weeks (with 8 weeks rest in between intervention periods), results on the 6-minute walk test, VO2peak and VO2/Kg ratio were significantly improved for those who completed the aerobic exercise protocol, but no significant effects were found for those practicing Qigong.57

Most of the non-significant findings have been found in studies with participants with some form of chronic illness or recovery from cancer at study entry. For example, respiratory function improved clinically, but not significantly, for patients with chronic heart failure practicing Tai Chi compared to usual care,34 and as described above, was relatively unchanged for the Qigong group with a history of Parkinson’s disease compared to an aerobic training control group.57 A group of patients with muscular dystrophy58 showed a trend for improvement that did not reach significance compared to a wait-list control. Further, no change in cardiovascular function was reported for sedentary participants with a history of osteoarthritis.59 Aerobic capacity was shown to improve with Tai Chi, though not significantly more so than with inactive controls in a small study of breast cancer survivors.525356 It is important to point out that of these five studies that failed to demonstrate significant improvements following Qigong or Tai Chi, four had 31 or fewer participants. It is difficult to discern if non-significant findings in cardiopulmonary fitness are due to some pattern of ineffectiveness with chronic and debilitating illness or if they are a result of the limited statistical power.

One of the key risk factors for cardiac disease is obesity. Qigong has demonstrated a greater reduction in BMI as compared to an exercise control group in two studies,2847 but this difference was not significant. Another study demonstrated a marked but non-significant reduction in waist circumference with Tai Chi compared to usual care for older adults.52Conversely, one study using Qigong and two with Tai Chi (respectively)485459 reported no change in BMI compared to usual care and another implementing a Qigong intervention60failed to maintain weight loss, suggesting the data are inconclusive at this point as to whether or not these practices may consistently affect weight.

A few studies of both Qigong and Tai Chi have examined level of intensity, indicating that some forms of these practices fall within the moderate intensity level,1161 but for the most part, level of exercise intensity is not reported. Cardiopulmonary benefits of Qigong and Tai Chi may partially be explained as a response to aerobic exercise, but with the wide range of speeds with which these exercises are executed, it would be important to assess this factor for a better understanding of the elements that contribute to outcomes. Regardless of the mechanisms, the preponderance of studies on cardiopulmonary outcomes show that Qigong and Tai Chi are effective compared to inactive controls, or at least approximately equal to the expected benefits of conventional exercise.

Physical Function

Decreased physical activity is related to declining physical function in all populations and that decline is compounded by the natural process of aging.6263 Changes in physical function were assessed in 16 studies (Qigong, n=2; Tai Chi, n=14). Most of the studies were conducted with older adults (i.e., studies in which mean age = 55 years or older, n=13) and several recruited specifically for participants with chronic pain (e.g., osteoarthritis, neck pain, or fibromyalgia, n = 5). A number of behavioral measures of physical function performance were included in this category of outcomes which also includes self-reported responses on scales representing physical function. Although fitness outcomes, such as the 6-minute walk test, might also be seen as assessing overall physical function, we did not include tests already discussed in the cardiopulmonary fitness category, but rather focused on functional tests that are usually used to assess capacity for daily living. Studies that assessed changes in overall physical activity levels are also included as an outcome pertaining to physical function.

Physical function measured with a wide variety of performance indicators, including chair rise, 50-ft walk, gait speed, muscle contraction strength, hand grip, flexibility, and function as measured on the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC, an osteoarthritis-specific assessment for function, stiffness, and pain), were variously found to be significantly improved in 5 studies comparing Tai Chi to minimal activity (usual or stretching activity, psychosocial support, or education) comparison groups566466 and one study of Tai Chi compared to an exercise therapy control intervention.64 One of these studies combined functional walking with Tai Chi to achieve significant improvements with pre-frail elders compared to usual care.67

In contrast, in 7 studies including participants with osteoarthritis or multiple co-morbidities, some of the physical function measures were not significantly different for Tai Chi or Qigong in comparison to inactive controls. This was the case for gait speed,47 timed up and go, 50-ft. walk and stair climb68 and 50-ft. walk and chair stand.6970 In one study of 30 patients with osteoarthritis practicing Tai Chi twice per week69 and another with 36 participants with fibromyalgia that utilized hand grip and chair stand to test a 20 minutes per week Qigong intervention,71 neither achieved significant improvements compared to usual care. In one exception to this trend, one measure of functional performance, time to complete chair rise, was significantly improved in transitionally frail elders in the Tai Chi group compared to a wellness education control group.47

Studies using self-report measures consistently show positive results for Tai Chi. Self-reported improvement in physical function for sedentary older adults was demonstrated for Tai Chi compared to wait-list controls,7072 and a stretching exercise control.33

Results in this category of outcomes are inconsistent, with a preponderance of studies recruiting sedentary, or chronically ill or frail elder participants. Even so, a handful of these studies successfully demonstrated potential for Qigong and Tai Chi to build performance, even with health compromised individuals. Further studies are needed to examine the factors that are important to more critically evaluate these interventions (such as power considerations or dose and frequency of the interventions), or learn if there are particular states of ill health that are less likely to respond to this form of exercise.

Falls and Balance

Another large grouping of studies focused primarily on falls prevention, balance, and physical function tests related to falls and balance (such as one-leg stance). Although there may be some crossover of implied benefits to the more general physical function measures reported above, this separate category was established to report on the studies of interventions primarily targeting falls and related measures. Fear of falling is reported with the psychological outcomes and fall self-efficacy is reported in the self-efficacy outcomes rather than in this category of falls and balance.

Outcomes related to falls such as balance, fall rates, and improved strength and flexibility were reported in 24 articles (Qigong, n=2, Tai Chi, n=20, and 2 studies that included both practices). Scores directly assessing balance (such as one-leg stance) or other closely related measures were consistently, significantly improved in 16 Tai Chi studies that only included participants who were sedentary or deemed at risk for falls at baseline.3343495359657383

Qigong has been less studied in relationship to balance-related outcomes; however, results suggest that there was a trend to maintain balance using Qigong in a population of patients with muscular dystrophy.58 In two studies that used both Qigong and Tai Chi, several measures of balance were significantly improved with sedentary women84 and with elderly healthy adults (mean age 80.4 years) compared to wait list controls.85

Another set of studies shows the effect of Tai Chi on balance to be similar to conventional exercise or physical therapy control interventions aimed at improving physical function related to balance;537486 or vestibular rehabilitation.8788 On the other hand, in a study of stroke survivors comparing Tai Chi to balance exercises, significant improvements in balance were achieved in the exercise control group, but not for Tai Chi.89 While knee extension was significantly improved, balance was not improved significantly in a Tai Chi intervention with sedentary women compared to a flexibility training control group.76

Mechanisms of gait performance which are important to understanding how Tai Chi affects balance were also studied. Reported improvements were found in 4 studies.82878890Strength and flexibility are also important to fall prevention. Four studies found significant improvements in these factors when Tai Chi was compared to an active control (brisk walking)3353597583 or inactive controls.597583

Eight studies directly monitored fall rates. Studies that incorporate educational or less active control interventions (e.g., stretching), variously demonstrated significant falls reduction for Tai Chi67778081 or non-significant reductions compared to control.4349 In a study comparing Tai Chi to an active physical therapy intervention designed to improve balance, results were similar (non-significant differences) between the two groups.86 The results are difficult to interpret as some participants may fall more because their level of activity has increased and some interventions are not monitored long enough to detect changes in fall rates.75

This category of outcomes has a large body of research supporting the efficacy of Tai Chi on improving factors related to falls, and growing evidence that falls may be reduced. Longer term studies to examine fall rates, and parallel studies that utilize Qigong as the intervention may further clarify the potential of these forms of exercise to affect falls and balance.

Quality of Life

Quality of life (QOL) outcomes were reported in 17 articles (Qigong, n=4 and Tai Chi, n=13). QOL is a broad ranging concept derived in a complex process from measures of a person’s perceived physical health, psychological state, personal beliefs, social relationships and relationship to relevant features of one’s environment.91 In 13 studies of a wide range of participants (including healthy adults, patients with cancer, post-stroke, arthritis, etc.) at least one of the components of QOL was reported to be significantly improved by Tai Chi compared to inactive346669739295 or active controls,89 and by Qigong compared to inactive9697 or active control groups.74 Qigong showed improvements in QOL compared to an exercise intervention, but not significantly so.74

Conversely, two studies reported no change in QOL, both with severely health-compromised individuals. One was of short duration (6 weeks) conducted with patients with traumatic brain injury.98 Some improvement in coping was shown with muscular dystrophy patients in response to a Qigong intervention,58 however, this finding was not significant, and direct QOL measures remained unchanged. One study reported no change in QOL when Tai Chi was compared to balance training and an education control among healthy older adults.99

With a few exceptions, the preponderance of studies indicate that Qigong and Tai Chi hold great potential for improving QOL in both healthy and chronically ill patients.


Self-efficacy is the confidence a person feels in performing one or several behaviors and the perceived ability to overcome the barriers associated with the performance of those behaviors.100 Although this is not a health outcome itself, it is often associated directly with health behaviors and benefits (e.g., falls self-efficacy associated with reduced falls), or with psychological health. Significant improvements in this outcome were reported in 8 studies (Qigong, n=2 and Tai Chi, n=6). Self-efficacy was generally assessed in the RCTs as a secondary outcome and reflected the “problem” area under investigation, such as falls self-efficacy (i.e., feeling confident that one will not fall) or efficacy to manage a disease (arthritis, fibromyalgia) or symptom (pain). Self-efficacy for falls was significantly increased as a result of participation in Tai Chi in 3 studies with adults at risk for falls compared to wait-list or usual care, sedentary control groups.7075101 In studies with clinical populations, persons with arthritis experienced improvements in arthritis self-efficacy69 and fibromyalgia patients experienced improvements in the ability to manage pain102 after participating in Tai chi as compared to inactive control groups that provided social interaction (telephone calls and relaxation therapy respectively). Lastly, the perceived ability to handle stress or novel experiences97103 and exercise self-efficacy99103 were enhanced relative to inactive control groups as a function of participation in Qigong and Tai Chi.

Patient Reported Outcomes

Patient reported outcomes (PROs) include reports of symptoms related to disease as perceived by the patient. The definition of PROs as “a measurement of any aspect of a patient’s health status that comes directly from the patient, without the interpretation of the patient’s responses by a physician or anyone else,”104 has developed over the past decade as an important indicator of treatment outcomes that matter to the patient, including an array of symptoms such as pain, fatigue, and nausea. Although PRO lists often include factors such as anxiety and depression, these are not included here, but rather in a separate section to address a range of psychological effects.

Thirteen studies are included in this category (Qigong, n=3 and Tai Chi, n=10). Arthritic pain6873105106 decreased significantly in response to Tai Chi compared to inactive (health education or usual care) controls. Self-reported neck pain and disability64 improved to a similar degree for Qigong and an exercise comparison intervention, but the difference between groups was not significant. Fibromyalgia symptoms improved significantly in one study comparing Tai Chi to a relaxation intervention,102 while another study reported slight improvements in symptoms for both Qigong and a usual care control group with no significant difference between the groups.71 Perceived symptoms of heart failure,55 disability,67 and sickness impact scores107 decreased in response to Tai Chi interventions as compared to inactive controls (either usual care or educational interventions) and sleep quality improved for Tai Chi even as compared to an exercise intervention.108 With Tai Chi, dissociative experiences and symptoms improved clinically, but were not statistically different from gains achieved by a support group among male veterans.109 Parkinson’s disease symptoms and disability were not significantly changed following a 7-week session of Qigong compared to aerobic training sessions.57

With the wide range of symptoms, and irregular outcomes of these PROs studies, it is difficult to draw meaningful conclusions about this category. Pain consistently responded to Tai Chi in four studies, while other symptoms were not uniformly assessed.


Twenty-seven articles (Qigong, n=7 and Tai Chi, n=19 and one study using both Qigong and Tai Chi) reported on psychological factors such as anxiety, depression, stress, mood, fear of falling, and self-esteem. Most of these studies examined psychological factors as secondary goals of the study, and consequently, they often did not intentionally recruit participants with appreciable psychological distress. Nevertheless, a number of substantial findings dominate this category.

Anxiety decreased significantly for participants practicing Qigong compared to an active exercise group.2846110 Depression was shown to improve significantly in studies comparing Qigong to an inactive control, newspaper reading97 and for Tai Chi compared to usual care, psychosocial support or stretching/education controls.5673111 General measures of mood (e.g, Profile of Mood States) were improved significantly for participants practicing Tai Chi compared to usual care controls.6698103112

Depression improved, but not significantly, for both Qigong and exercise comparison groups2896 and for Tai Chi compared to an educational intervention.113 One study reported improved depression, anxiety, and stress among patients with osteoarthritis for both Tai Chi and hydrotherapy groups compared to a wait-list control, but only significantly so for hydrotherapy.68

Non-significant changes in anxiety were reported in a study of Tai Chi compared to a relaxation intervention102 and two other studies did not detect significant differences in depression in response to Tai Chi55102 or Qigong58 compared to usual care or inactive controls. Fear of falling decreased significantly in most studies498283101114 except for one that showed no change.84 Reports of self-esteem significantly improved in tests of Tai Chi compared to usual care93115 and psychosocial support,95 but the increase in self-esteem compared to exercise and education controls was not significant.99

Jin112 specifically created a stressful situation and measured the response in mood, self-reported stress levels, and BP, across 4 interventions, including Tai Chi, meditation, brisk walking and neutral reading. Significant improvements were shown in adrenaline, heart rate, and noradrenaline in Tai Chi compared to a neutral reading intervention, while all groups showed improvements in cortisol. In another study examining blood markers related to stress response, norepinephrine, epinephrine and cortisol blood levels were significantly decreased in response to Qigong compared to a wait-list control group.110

This category of symptoms, particularly anxiety and depression, shows fairly consistent responses to both Tai Chi and Qigong, especially when the control intervention does not include active interventions such as exercise. In particular, with a few studies indicating that there may be changes in biomarkers associated with anxiety and/or depression in response to the interventions, this category shows promise for examining potential mechanisms of action for the change in psychological state.

Immune Function and Inflammation

Immune-related responses have also been reported in response to Qigong (n=3) and Tai Chi (n=3) studies. Manzaneque et al.116 reported improvements in a number of immune-related blood markers, including total number of leukocytes, number of eosinphils, and number and percentage of monocytes, as well as the complement C3 levels following a 1-month Qigong intervention compared to usual care. Antibody levels in response to flu vaccinations were significantly increased among a Qigong group compared to usual care.117 Varicella zoster virus titers and T-cells increased in response to vaccine among Tai Chi practitioners.113 An earlier study conducted by Irwin and colleagues92 reported an increase in varicella zoster virus specific cell-mediated immunity among those practicing Tai Chi compared to wait-list controls.

Immune function and inflammation are closely related, and are often assessed using a variety of blood markers, particularly certain cytokines and C-reactive protein. Interleukin-6, an important marker of inflammation, was found to be significantly modulated in response to practicing Qigong, compared to a no-exercise control group.40 On the other hand, C-reactive protein and erythrocyte sedimentation rates remained unchanged among a group of rheumatoid arthritic patients who participated in a Tai Chi class compared to stretching and wellness education.73

A number of studies not utilizing an RCT design have examined blood markers prior to and after Tai Chi or Qigong interventions, providing some indication of factors that might be important to explore in future RCTs (and not reported in the table). For example, improvements in thyroid-stimulating hormone, follicle-stimulating hormone, triiodothyronine,118 and lymphocyte production119 have been noted in response to Tai Chi compared to matched controls. Pre-post Tai Chi intervention designs have also shown an improvement in immunoglobulin G (IgG)120 and natural killer (NK) cells121 and similar non-RCTs have suggested that Qigong improves immune function and reduces inflammation profiles as indicated by cytokine and T-lymphocyte subset proportions.122124

As with the category of psychological outcomes, these immune and inflammation related parameters fairly consistently respond to Tai Chi and Qigong, while also providing potential for examining mechanisms of action.

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In answering Research Question 1, we have identified 9 categories of health benefits related to Tai Chi and Qigong interventions, with varying levels of support. Six domains of health-related benefits have dominated the research with sixteen or more RCTs published for each of these outcomes: psychological effects (27), falls/balance (23), cardiopulmonary fitness (19), QOL (17), PROs (18), and physical function (16). These areas represent most of the RCTs reviewed, with many of the studies including multiple measured outcomes spanning across several categories (n=42). Substantially fewer RCTs have been completed in the other three categories, including bone density (4), self-efficacy (8), and studies examining markers of immune function or inflammation (6).

The preponderance of studies showed significant, positive results on the tested health outcomes, especially when comparisons were made with minimally active or inactive controls (n=52). For some of the outcomes addressed in this review, there were studies that did not demonstrate significant improvements for the Tai Chi or Qigong intervention as compared to the control condition. For the most part, however, these non-significant findings occurred in studies in which the control design was actually a treatment type of control expected to produce similar benefits, such as an educational control group intervention producing similar outcomes as Tai Chi for self-esteem,99 aerobic exercise showing similar results to Qigong in reducing depression,2857 an acupressure group successfully maintaining weight loss compared to no intervention effect for Qigong,60 or resistance training producing similar (nonsignificant) effects as Tai Chi for muscle strength, balance, and falls.4366 It is important to note that although the Tai Chi and Qigong interventions did not produce larger benefits than these active treatment controls, in most cases substantial improvements in the outcome were observed for both treatment groups.

Other studies in which the improvements did not significantly differ between the treatment group and the control group suffered from: (a) study designs of shorter duration (4 to 8 weeks, rather than the usual 12 or more weeks)5198 although there were some exceptional studies with significant results after only 8 weeks;4483103 (b) selection of very health-compromised participants or individuals with conditions that do not generally respond to other conventional treatments or medicines such as muscular dystrophy,58 multiple morbidities,47 fibromyalgia,71arthritis;73 or (c) the outcome measured was not noted as particularly problematic nor set as an eligibility criteria for poor starting levels at baseline (n=5).2896

On the other hand, in the areas of research that address outcomes typically associated with physical exercise, such as cardiopulmonary health or physical function, results are fairly consistent in showing that positive, significantly larger effects are observed for both Tai Chi and Qigong when compared to no-exercise control groups and similar health outcomes are found when compared to exercise controls. Even with the very wide range of study design types, strength of control interventions, and the entry level of the health status of study participants, there remain a number of remarkable and persistent findings of health benefits in response to both Qigong and Tai Chi.

In response to Research Question 2, we have noted in earlier sections the ways in which Qigong and Tai Chi are considered equivalent, and now address how studies identifying similar outcomes in response to these practices may provide additional evidence for equivalence. On the surface, research that examines the effects of Qigong on health outcomes appears to be of lesser magnitude than the research on what is typically called Tai Chi. For each category of outcomes described above, we noted how many RCTs had been conducted for each, Tai Chi and Qigong, and for the most part, there were many fewer reports on Qigong than for what is named Tai Chi for any given outcome examined. Nevertheless, across the outcomes examined in RCTs, the findings are often similar, with no particular trends indicating that one has different effects than the other.

As noted earlier, however, it is not unusual for the intervention used in a study or trial to be named Tai Chi, but to actually apply a set of activities which is more a form of Qigong, that is, easy-to-learn movements that are simple and repeatable rather than the long complex sequences of traditional Tai Chi movements that can take a long time to learn. For example, a large number of studies examining Tai Chi effects on balance use a modified, repetitive form of Tai Chi which is more like Qigong. Thus, while it appears that fewer studies have been conducted to test what is called Qigong, it is also clear that when a practice called Tai Chi is modified to focus, especially on balance enhancement, for example, it actually may be Tai Chi in name only.

Given the apparent similarity of practice forms utilized in research, the discussion of equivalence of Tai Chi and Qigong extends beyond the earlier observation that they are similar in practice and philosophy. Since research designs often incorporate blended aspects of both Qigong and Tai Chi, it is unreasonable to claim that the evidence is lacking for one or the other and it becomes inappropriate not to claim their equivalence. We suggest that the combined current research provides a wider base of growing evidence indicating that these two forms produce a wide range of health-related benefits.

The problem with claiming equivalence, then, does not lie within the smaller number of studies using a form called Qigong, but rather in the lack of detail reported across the studies regarding whether or not the interventions contain the key elements philosophically and operationally thought to define meditative movement practices such as Tai Chi and Qigong. In previous publications, and in this review, we note that the roots of both of these TCM-based wellness practices require that the key elements of meditative movement be implemented: focus on regulating the body (movement/posture); focus on regulating the breath; and focus on regulating the mind (consciousness) to achieve a meditative state. Given the equivalence noted in foundational principles and practice, the differences among interventions and resultant effects on outcomes would perhaps more purposefully be assessed for intervention fidelity (i.e., adherence to the criteria of meditative movement).

Beyond the meditative movement factors that tie the practices and expected outcomes together, other, more conventional factors would be important to assess, each potentially contributing to variations in outcomes achieved. For example, dosing (i.e., frequency, duration and level of intensity, including estimate of aerobic level or metabolic equivalents) may be important in whether or not benefits accrue. Or, a focus on particular muscle groups may be critical to understanding changes relative to certain goals (e.g., how many of the exercises chosen for a study protocol develop quadricep strength likely to produce results for specific physical function tests?). Beyond the important similarities of movement and a focus on breath and mind to achieve meditative states, there are other aspects that vary greatly within the wide variety of both Tai Chi and Qigong exercises, including speed of execution, muscle groups used and range of motion, all of which may provide differences in the physiologically-oriented outcomes (similar to the differences that could be noted in the wide variety of exercises considered under the aerobic “umbrella”).

The question of the equivalence Tai Chi and Qigong, then, may be helpful if the focus is on similarity in philosophy and practice. With consistent reporting on adherence to the above mentioned aspects of practice, not only could a level of standardization be implemented, but also measures that control for variation of interventions could be used to better understand differences and similarities in effects.1

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For purposes of this review, a study was selected if it was designed as an RCT and compared the effects of either Tai Chi or Qigong to those of a control condition on a physical or psychological health outcome. However, there was no further grading of the quality of the research design. As a result of this relatively broad inclusion criterion, the studies represent a wide variety in methods of controlling for balanced randomization and intent to treat analyses, in the specific methods of implementing Tai Chi and Qigong, in the outcomes assessed, in the measurement tools used to ascertain the outcomes, and in the populations being studied.

One difficulty in examining such a broad scope of studies is that the large number of studies required that we logically, but artificially, construct categories within which to discuss each group of outcomes. However, by choosing to categorize by health outcomes, rather than participant, patient or disease types, we have provided one particular view of the data, and may have obscured other aspects. For example, in a recently published review, the authors analyzed studies that were conducted with community-dwelling adults over the age of 55.125 Results showed that interventions utilizing Tai Chi and Qigong may help older adults improve physical function and reduce blood pressure, fall risk, and depression and anxiety. Another view of these data may emerge if only studies of chronically ill participants are evaluated. Thus, there may be other ways to examine the RCTs reported in the current review such that specific diseases or selected study populations may reveal more consistent findings (positive or negative) for certain outcomes that are clearly tied to entry level values.

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A compelling body of research emerges when Tai Chi studies and the growing body of Qigong studies are combined. The evidence suggests that a wide range of health benefits accrue in response to these meditative movement forms, some consistently so, and some with limitations in the findings thus far. This review has identified numerous outcomes with varying levels of evidence for the efficacy for Qigong and Tai Chi, including bone health, cardiopulmonary fitness and related biomarkers, physical function, falls prevention and balance, general quality of life and patient reported outcomes, immunity, and psychological factors such as anxiety, depression and self-efficacy. A substantial number RCTs have demonstrated consistent, positive results especially when the studies are designed with limited activity for controls. When both Tai Chi and Qigong are investigated together, as two approaches to a single category of practice, meditative movement, the magnitude of the body of research is quite impressive.

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So What?

Application to research

The current state of research splinters these TCM-based wellness practices by identifying them with different names, and treating them as separate and different methodologies. Our intent has been to recognize the common critical elements of Qigong and Tai Chi, based on the similarities in philosophy and practice and the range of findings for similar health outcomes, and to treat the two as equivalent forms. Studies in the future should acknowledge these elements, and even test for intervention fidelity, to assure that the practices do, in fact, reflect the guiding principles of Tai Chi and Qigong. Beyond this we assert that it is critical to begin delineating the practice characteristics that actually do differ both between and within these practices, so that a more specific body of knowledge can begin to accumulate about the types of practices, the component features of the practices and their effects on health-related outcomes.

Some studies of these forms of meditative movement indicate that study participants with severe, chronic, progressive illnesses may be slower to respond or not respond at all to the practices. Interestingly, however, other studies suggest that these practices may improve or slow the progression of such illnesses. This may be especially likely when the practices are implemented early as an aspect of wellness, prevention or disease management in a proactive, risk reduction context. These findings suggest that continued research on these and other forms of meditative movement is warranted for a broad number of conditions and across populations.

The wide variations in populations and outcomes studied, descriptions of interventions (or the lack of such description), reports of dose, and the extreme variety in the sorts of tools used to assess outcomes, point to the need to develop more standardized protocols and trends in measurement for the field of meditative movement research. Application in health promotion

The preponderance of findings are positive for a wide range of health benefits in response to Tai Chi, and a growing evidence base for similar benefits for Qigong. As described, the foundational similarities and the often adapted Tai Chi protocols which more closely resemble forms of Qigong, allow us to suggest that outcomes can be counted across both types of studies, further supporting claims of equivalence.

In a recent review addressing Tai Chi and Qigong research among older adults, it was pointed out that no adverse events were reported across studies.125 The substantial potential for achieving health benefits, the minimal cost incurred by this form of self-care, and the apparent safety of implementation across populations, points to the importance of wider implementation and dissemination. The health promotion challenge is that both Tai Chi and Qigong are still often considered novel forms of exercise and adopted by a small market segment of our population. On the positive side, however, there is a rapid increase of visibility of what is popularly referenced as Tai Chi which is known as an effective intervention for balance enhancement and falls prevention among the elderly, and there is a growing interest in safe, alternative forms of exercise across all age groups. Tai Chi and Qigong interventions provide an accessible alternative option for individuals who may prefer these activities over more conventional or vigorous forms of exercise. The growing interest in these forms of exercise that include a mindful focus on the breath and meditation provides an opportunity for changing the landscape of personal choice making and shifts the motivations that people have to exercise, while presenting an entirely new set of exercise research opportunities. This suggests that Tai Chi and Qigong (or more generally, meditative movement types of exercise) may provide an attractive and effective exercise alternatives for the large populations of people at risk for preventable disease, sedentary, and lacking the motivation to engage in more conventional exercise.

21st Century Breakthrough: Researching the Benefits of Mind-Body Practice by Investigating Genetic Expression

By Roger Jahnke, OMD

Founder, Director of the Institute of Integral Qigong and Tai Chi

Research on Mind-Body approaches is accelerating. One of a number of characteristics of Mind-Body practice is the purposeful elicitation of the Relaxation Response (RR). The various forms of practices which elicit the RR include Tai Chi, Qi Gong, Yoga, meditation, repetitive prayer, breathing exercises, progressive muscle relaxation, biofeedback, guided visualization, affirmation, etc. These methods tend to trigger physiological and perhaps energetic mechanisms that move the body into a state of deep rest. It appears that this can literally change how genes behave in response to stress.

Mind-Body practices that produce the Relaxation Response have been used by people across cultures for thousands of years to prevent and treat disease and generate states of mind that foster greater performance and intuitive insight. Recently, a number of studies have turned toward investigation of the effect that Mind-Body practice can have on genetic expression. In research on natural healing, functional maximization and holistic, complementary and integrative medicine there has been a growing trend away from simply studying disease mechanisms and outcomes, toward the study of the subtle factors that predispose individuals for sustainable wellbeing as well as for disease. Gene expression is an emerging arena wherein the total continuum – from wellbeing to disease – can be effectively investigated.

The RR is characterized by reduction in oxygen intake, increase in exhalation of nitric oxide, and lower psychological distress. It has been proposed as the


counterpoint to the "flight or fight" state (FF) – the stress response. FF and RR are like a Yin-Yang. Numerous studies have shown that both RR and the FF have distinct profiles of physiological and gene expression changes.

Recently - July, 2008 - a breakthrough study was completed exploring the extent to which Mind-Body practices that trigger the RR and have an influence on gene expression: Genomic Counter-Stress Changes Induced by the Relaxation Response. Dr. Herbert Benson, who was the lead developer of the RR concept was among the researchers on this study.

Earlier -- February 2005 - a similar study was completed which focused specifically on gene expression in Qigong: Genomic Profiling of Neutrophil Transcripts in Asian Qigong Practitioners: A Pilot Study in Gene Regulation by Mind–Body Interaction.

A less widely known study on a Yoga/Pranayama method – East Indian Qigong – also explored the effects of a RR method on gene expression: Gene expression profiling in practitioners of Sudarshan Kriya.

The abstracts and links to PDFs of these studies are presented below. Genomic counter-stress changes induced by the relaxation response is included in full at the end of this article.

Due to the magnitude of the credibility of Dr. Herbert Benson, his role as the “father” of the relaxation response and the refined design of the study, theGenomic counter-stress changes induced by the relaxation response article has gotten a greater amount of press. The highest number of participants, however, was involved in the Gene expression profiling in practitioners of Sudarshan Kriya article. The Benson study, n=57 (RCT, 19 long term, 20 short term & 19 control), Qigong study, n=12 (6 Qigong and 6 control), Pranayama study, n=84 (42 Pranayama and 42 control).


In a number of press releases the authors of Genomic counter-stress changes induced by the relaxation response made a number of comments that are easily applicable to all three studies. They state that:

"This study provides the first compelling evidence that the RR [relaxation response] elicits specific gene expression changes in short-term and long- term practitioners."

Actually the other studies were earlier and they all suggest this.

The Genomic Counter-stress authors wrote that their findings suggest: "Consistent and constitutive changes in gene expression resulting from RR may relate to long term physiological effects," and that "Our study may stimulate new investigations into applying transcriptional profiling for accurately measuring RR and stress related responses in multiple disease settings."

It is likely that these studies portend a “sea change” in research and will trigger an outpouring of similar research. Dr. Herbert Benson, professor emeritus of Harvard University and director emeritus of the Benson-Henry Institute and co- senior author of the study said:

"Now we've found how changing the activity of the mind can alter the way basic genetic instructions are implemented," said Benson.

Dr. Towia Libermann, director of the BIDMC Genomics Center and also co-senior author of the study added:

"This is the first comprehensive study of how the mind can affect gene expression, linking what has been looked on as a 'soft' science with the 'hard' science of genomics.” "It is also important because of its focus on gene


expression in healthy individuals, rather than in disease states," explained Libermann.

The authors said their study showed that the relaxation response changed the expression of genes involved with inflammation, programmed cell death and the handling of free radicals. Free radicals are normal byproducts of metabolism that the body neutralizes in order to stop damage to cells and tissues.

Co-lead author of the study Dr. Jeffery Dusek formerly of the Benson-Henry Institute and now with the Abbott Northwestern Hospital in Minneapolis said:

"Changes in the activation of these same genes have previously been seen in conditions such as post-traumatic stress disorder; but the relaxation-response- associated changes were the opposite of stress-associated changes and were much more pronounced in the long-term practitioners."

Dr. Benson reflected that people across different cultures have been using Mind- Body techniques for thousands of years. They found that it didn't particularly matter which techniques was used, whether it was Tai Chi, Qigong, meditation, Yoga, breathing, or repetitive praying, they all act through the same underlying mechanism.

"Now, we need to see if similar changes occur in patients who use the relaxation response to help treat stress-related disorders, and those studies are underway now".

Probably the most compelling statement from the article on the findings of the study was “It is becoming increasingly clear that psychosocial stress can manifest as system-wide perturbations of cellular processes, generally increasing oxidative stress and promoting a pro-inflammatory milieu. Stress associated changes in peripheral blood leukocyte expression of single genes have been


identified. More recently, chronic psychosocial stress has been associated with accelerated aging at the cellular level. Specifically, shortened telomeres, low telomerase activity, decreased anti-oxidant capacity and increased oxidative stress are correlated with increased psychosocial stress and with increased vulnerability to a variety of disease states.”

These 3 studies strongly suggest that Mind-Body practices, especially those that trigger a sustained and accumulative RR effect – a counter stress effect – can prevent and ameliorate disease. This effect of Mind-Body practice on gene expression transforms the landscape of scientific exploration and launches an entirely new direction for the investigation for the emerging field of health maximization based integrative medicine.


Sea Change Studies -- The Abstracts

I. Genomic counter-stress changes induced by the relaxation response

Dusek JA, Otu HH, Wohlhueter AL, Bhasin M, Zerbini LF, Joseph MG, Benson H, Libermann TA. PLoS ONE – Online Journal of Medicine 2008 Jul 2;3(7):e2576

To review the article in full:


This is included at the end of this article.

Benson-Henry Institute for Mind Body Medicine at Massachusetts General Hospital, Chestnut Hill, Massachusetts, United States of America.

BACKGROUND: Mind-body practices that elicit the relaxation response (RR) have been used worldwide for millennia to prevent and treat disease. The RR is characterized by decreased oxygen consumption, increased exhaled nitric oxide, and reduced psychological distress. It is believed to be the counterpart of the stress response that exhibits a distinct pattern of physiology and transcriptional profile. We hypothesized that RR elicitation results in characteristic gene expression changes that can be used to measure physiological responses elicited by the RR in an unbiased fashion.

METHODS/PRINCIPAL FINDINGS: We assessed whole blood transcriptional profiles in 19 healthy, long-term practitioners of daily RR practice (group M), 19 healthy controls (group N(1)), and 20 N(1) individuals who completed 8 weeks of RR training (group N(2)). 2209 genes were differentially expressed in group M relative to group N(1) (p<0.05) and 1561 genes in group N(2) compared to group N(1) (p<0.05). Importantly, 433 (p<10(-10)) of 2209 and 1561 differentially expressed genes were shared among long-term (M) and short-term practitioners


(N(2)). Gene ontology and gene set enrichment analyses revealed significant alterations in cellular metabolism, oxidative phosphorylation, generation of reactive oxygen species and response to oxidative stress in long-term and short- term practitioners of daily RR practice that may counteract cellular damage related to chronic psychological stress. A significant number of genes and pathways were confirmed in an independent validation set containing 5 N(1) controls, 5 N(2) short-term and 6 M long-term practitioners.

CONCLUSIONS/SIGNIFICANCE: This study provides the first compelling evidence that the RR elicits specific gene expression changes in short-term and long-term practitioners. Our results suggest consistent and constitutive changes in gene expression resulting from RR may relate to long term physiological effects. Our study may stimulate new investigations into applying transcriptional profiling for accurately measuring RR and stress related responses in multiple disease settings.

II. Genomic profiling of neutrophil transcripts in Asian Qigong practitioners: a pilot study in gene regulation by mind-body interaction.

Li QZ, Li P, Garcia GE, Johnson RJ, Feng L. Journal of Alternative and Complement Medicine 2005 Feb;11(1):29-39

The full article can be reviewed at:


Microarray Core, Center for Immunology, University of Texas Southwestern Medical Center, Dallas, TX, USA.

BACKGROUND AND OBJECTIVES: The great similarity of the genomes of humans and other species stimulated us to search for genes regulated by


elements associated with human uniqueness, such as the mind-body interaction. DNA microarray technology offers the advantage of analyzing thousands of genes simultaneously, with the potential to determine healthy phenotypic changes in gene expression. The aim of this study was to determine the genomic profile and function of neutrophils in Falun Gong (FLG, an ancient Chinese Qigong) practitioners, with healthy subjects as controls.

SUBJECTS AND DESIGN: Six (6) Asian FLG practitioners and 6 Asian normal healthy controls were recruited for our study. The practitioners have practiced FLG for at least 1 year (range, 1-5 years). The practice includes daily reading of FLG books and daily practice of exercises lasting 1-2 hours. Selected normal healthy controls did not perform Qigong, yoga, t'ai chi, or any other type of mind- body practice, and had not followed any conventional physical exercise program for at least 1 year. Neutrophils were isolated from fresh blood and assayed for gene expression, using microarrays and RNase protection assay (RPA), as well as for function (phagocytosis) and survival (apoptosis).

RESULTS: The changes in gene expression of FLG practitioners in contrast to normal healthy controls were characterized by enhanced immunity, downregulation of cellular metabolism, and alteration of apoptotic genes in favor of a rapid resolution of inflammation. The lifespan of normal neutrophils was prolonged, while the inflammatory neutrophils displayed accelerated cell death in FLG practitioners as determined by enzyme-linked immunosorbent assay. Correlating with enhanced immunity reflected by microarray data, neutrophil phagocytosis was significantly increased in Qigong practitioners. Some of the altered genes observed by microarray were confirmed by RPA.

CONCLUSION: Qigong practice may regulate immunity, metabolic rate, and cell death, possibly at the transcriptional level. Our pilot study provides the first evidence that Qigong practice may exert transcriptional regulation at a genomic level. New approaches are needed to study how genes are regulated by


elements associated with human uniqueness, such as consciousness, cognition, and spirituality.

III. Gene expression profiling in practitioners of Sudarshan Kriya

Sharma H, Datta P, Singh A, Sen S, Bhardwaj NK, Kochupillai V, Singh N.Journal of Psychosomatic Research 2008 Feb;64(2):213-8

To review a more comprehensive version of the article:

http://www.jbtdrc.org/National%20Symposium%20- %202006/Proc%20pages/CMB/CMB1.pdf

Department of Biochemistry, All India Institute of Medical Sciences, Ansari Nagar, New Delhi, India.

BACKGROUND: The rapid pace of life, eating habits, and environmental pollution have increased stress levels and its related disorders. Stress genes and a variety of regulatory pathways mediate the complex molecular response to stress. Oxidative stress is internal damage caused by reactive oxygen species. Increasing evidence suggests that chronic psychosocial stress may increase oxidative stress, which in turn may contribute to aging, and etiology of coronary diseases, cancer, arthritis, etc. Psychophysiological concomitants of meditation have been extensively researched, but there are very little data available on biochemical activity leading to relieving stress by causing a relaxation response by Sudarshan Kriya (SK). SK is a breathing technique that involves breathing in three different rhythms. It is preceded by Ujjayi Pranayam (long and deep breaths with constriction at the base of throat) and Bhastrika (fast and forceful breaths through nose along with arm movements).

METHODS: Forty-two SK practitioners and 42 normal healthy controls were recruited for our study. The practitioners had practiced SK for at least 1 year.


Selected normal healthy controls did not perform any conventional physical exercise or any formal stress management technique. Whole blood was used for glutathione peroxidase estimation and red blood cell lysate was used for superoxide dismutase activity assay and for glutathione estimation. White blood cells were isolated from fresh blood and assayed for gene expression using reverse transcriptase-polymerase chain reaction. The parameters studied are antioxidant enzymes, genes involved in oxidative stress, DNA damage, cell cycle control, aging, and apoptosis.

RESULTS: A better antioxidant status both at the enzyme activity and RNA level was seen in SK practitioners. This was accompanied by better stress regulation and better immune status due to prolonged life span of lymphocytes by up- regulation of antiapoptotic genes and prosurvival genes in these subjects.

CONCLUSIONS: Our pilot study provides the first evidence suggesting that SK practice may exert effects on immunity, aging, cell death, and stress regulation through transcriptional regulation.

The methods of the Genomic counter-stress changes induced by the relaxation response study are also somewhat exemplary and shed some light on how such research is done. The researchers recruited three groups of people. In the first group (called the M group) there were 19 long-term practitioners who had been practicing various ways of producing the relaxation response every day for a long time (for instance with daily yoga, repeated prayer, Tai Chi or meditation practice).

In the second group were another 19 people who they called the "healthy controls" (group N1), who were not daily practitioners, and the third group was like the healthy controls group, except these 20 people completed 8 weeks of relaxation response training (this group was N2).


The researchers assessed transcriptional profiles of the people in all three groups from blood samples.

They found the expressions of a total of 2,209 genes were significantly different between groups M and N1, and a total of 1,561 genes were similarly significantly different between groups N2 and N1.

More importantly, however, was the fact 433 of the genes were common to both sets of comparisons: the same ones were different between M and N1 and between M and N2, so even short term practice of the relaxation response appeared to produce changes in these 433 gene expressions.

Further analysis using techniques called gene ontology and gene set enrichment, showed that groups M and N1 (the long term and the short term practitioners of the relaxation response) exhibited similar physiological changes such as in "cell metabolism, oxidative phosphorylation, generation of reactive oxygen species and response to oxidative stress".

A second phase of the study involving 5 N1 healthy controls, 5 N2 short term practitioners, and 6 M long term practitioners, was done to validate a significant number of genes and pathways.

This is clearly a very well designed study, and extremely thorough. The findings of these three studies points the way to investigating gene expression profiles in well, at risk and diseased populations to demonstrate the benefits of Mind-Body practices which elicit the RR to both prevent and cure disease. Given the low cost of learning and practicing Mind-Body practice, it is clear that proliferation of these practices in school, at work and at home can potentiate a nationwide and culture wide response to eliminate the health cost crisis.


Details on the Dusek, Benson, Libermann study follow in full with an excellent references section at the conclusion.

Genomic Counter-Stress Changes Induced by the Relaxation Response

Jeffery A. Dusek1,2,3,6#, Hasan H. Otu3,4#, Ann L. Wohlhueter1, Manoj Bhasin3,4, Luiz F. Zerbini3,4, Marie G. Joseph4, Herbert Benson1,3,5*, Towia A. Libermann3,4*

Benson-Henry Institute for Mind Body Medicine at Massachusetts General Hospital, Chestnut Hill, Massachusetts, United States of America
Department of Psychiatry, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, United States of America

Department of Medicine, Division of Interdisciplinary Medicine and Biotechnology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts, United States of America
BIDMC Genomics Center, Beth Israel Deaconess Medical Center, Boston, Massachusetts, United States of America

Department of Medicine, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, United States of AmericaInstitute for Health and Healing, Abbott Northwestern Hospital, Minneapolis, Minnesota, United States of America

For full text of research article



Mind-body practices that elicit the relaxation response (RR) have been used worldwide for millennia to prevent and treat disease. The RR is characterized by decreased oxygen consumption, increased exhaled nitric oxide, and reduced


psychological distress. It is believed to be the counterpart of the stress response that exhibits a distinct pattern of physiology and transcriptional profile. We hypothesized that RR elicitation results in characteristic gene expression changes that can be used to measure physiological responses elicited by the RR in an unbiased fashion.

Methods/Principal Findings

We assessed whole blood transcriptional profiles in 19 healthy, long-term practitioners of daily RR practice (group M), 19 healthy controls (group N1), and 20 N1 individuals who completed 8 weeks of RR training (group N2). 2209 genes were differentially expressed in group M relative to group N1 (p<0.05) and 1561 genes in group N2 compared to group N1 (p<0.05). Importantly, 433 (p<10−10) of 2209 and 1561 differentially expressed genes were shared among long-term (M) and short-term practitioners (N2). Gene ontology and gene set enrichmentanalyses revealed significant alterations in cellular metabolism, oxidative phosphorylation, generation of reactive oxygen species and response to oxidative stress in long-term and short-term practitioners of daily RR practice that may counteract cellular damage related to chronic psychological stress. A significant number of genes and pathways were confirmed in an independent validation set containing 5 N1 controls, 5 N2 short-term and 6 M long-term practitioners.


This study provides the first compelling evidence that the RR elicits specific gene expression changes in short-term and long-term practitioners. Our results suggest consistent and constitutive changes in gene expression resulting from RR may relate to long-term physiological effects. Our study may stimulate new investigations into applying transcriptional profiling for accurately measuring RR and stress related responses in multiple disease settings.



The relaxation response (RR) has been defined as a mind-body intervention that offsets the physiological effects caused by stress [1], [2]. The RR has been reported to be useful therapeutically (often as an adjunct to medical treatment) in numerous conditions that are caused or exacerbated by stress [3]–[6].

Mind-body approaches that elicit the RR include: various forms of meditation, repetitive prayer, yoga, tai chi, breathing exercises, progressive muscle relaxation, biofeedback, guided imagery and Qi Gong [7]. One way that the RR can be elicited is when individuals repeat a word, sound, phrase, prayer or focus on their breathing with a disregard of intrusive everyday thoughts [2]. The non- pharmacological benefit of the RR on stress reduction and other physiological as well as pathological parameters has attracted significant interest in recent years to decipher the physiological effects of the RR. In addition to decreased oxygen consumption [8]–[10], other consistent physiologic changes observed in long- term practitioners of RR techniques include decreased carbon dioxide elimination, reduced blood pressure, heart and respiration rate [1], [2], [11], prominent low frequency heart rate oscillations [12] and alterations in cortical and subcortical brain regions [13], [14].

Despite these observations and the well-established clinical effects of RR- eliciting practices [15], [16], the mechanisms underlying the RR have not been identified. Similarly, the impact of the RR on gene expression and signaling pathways has not yet been explored in detail, although a transcriptional profiling study of Qi Gong [17] practitioners, another RR method, revealed apparent distinct gene expression differences between Qi Gong practitioners and age matched controls. It is likely that differences in gene expression may be an underlying factor in the physiologic and psychologic changes noted above. Toward that end, we conducted a study to explore the gene expression profile of healthy long-term practitioners versus healthy age and gender matched controls. As a further evaluation, we provided 8-weeks of RR training to the control


subjects and re-assessed their gene expression.

Patient characteristics

This study includes both cross sectional and an 8-week prospective design. Healthy adults were enrolled, comprising 2 groups: individuals with a long-term RR practice (group M; n = 19) or those with no prior RR experience (novice; group N1; n = 19). Group N1 novices, furthermore, underwent 8-weeks of RR training (Group N2; n = 20) for the prospective analysis. In the cross sectional study, we compare gene expression profiles (GEP) in whole blood between groups M and N1, whereas in the prospective study GEP is compared for each individual novice subject before and after RR experience, matched individuals of groups N1 versus N2 respectively.

Gene expression changes associated with the RR

Transcriptional differences between the different groups and within individuals before and after the RR are assessed by microarray analysis using Affymetrix HG-U133 Plus 2.0 genechips (www.affymetrix.com). This technology is a well established and reliable method to assess global gene expression differences[18]. Comparing group M (subjects with long term RR practice) to group N1(subjects prior to RR training), we find statistically significant differential expression of 2209 genes; 1275 significantly up-regulated and 934 significantly down-regulated in M vs. N1. Additionally, 1561 genes are differentially expressed in novices after RR experience, N2 vs. N1; 874 significantly up-regulated and 687 significantly down-regulated. Comparison of gene lists from M vs. N1, N2 vs. N1and M vs. N2 with Venn diagrams reveals significant overlap (Fig. 1a). Significance of overlaps is calculated using hypergeometric distributional assumption [19] and p-values are adjusted using Bonferroni correction for multiple comparisons [20]. Heatmaps were generated from genes in the intersecting areas of the Venn diagrams (Fig. 1b). We find 316 up-regulated and


279 down-regulated genes are differentially expressed in group M compared to both group N1 and N2; these changes in GEP are only observed in long-term RR practitioners. Similarly, 260 genes are up-regulated and 168 genes are down- regulated in both groups M and N2 compared to N1; they represent GEP changes characteristic of RR practice over at least 8 weeks.

Figure 1. Gene Ontology Analysis.
Analysis of differentially expressed genes: a) Venn diagrams: * indicates significant overlaps (p<106); b) Heatmaps of the 595 differentially regulated genes in both M vs. N1 and M vs. N2 (left) and the 418 differentially regulated genes in both M vs. N1 and N2 vs. N1; c) Heatmap of 15 genes in the intersection of all three groups (gene symbols listed on the right). In heatmaps, rows represent genes and columns represent samples from N1, N2, and M groups. Genes are clustered using row-normalized signals and mapped to the [−1,1] interval (shown in scales beneath each heatmap). Red and green represent high and low expression values, respectively.

Heatmaps generated using these genes exhibit consistent GEP changes across the three groups with a few samples in each group resembling the GEP of another group. To determine if any demographic characteristics (e.g. age, ethnicity, etc.) influences this observation, we clustered each group separately using the same set of genes. For each cluster analysis, we calculated the significance of observing a characteristic among the samples in the subgroups formed. We found that number of times M subjects reported eliciting the RR per week was significantly associated with the subgroups formed when M samples were clustered using genes differentially expressed in long term RR practitioners only. Specifically, there were 316 up-regulated and 279 down-regulated genes differentially expressed in group M compared to both group N1 and N2; (Fig. 1b). All remaining cluster analyses revealed no such significant influence of demographic characteristics (see online supplementary data).


Finally, the intersection of all 3 areas (M vs. N1, N2 vs. N1 and M vs. N2) identifies genes with expression behavior that is monotonically changed between N1 to N2to M (Fig. 1c). These results clearly demonstrate that short term as well as long term RR practice lead to distinct and consistent gene expression changes in hematopoietic cells.

Signaling pathways modulated by the RR

We performed Expression Analysis Systematic Explorer (EASE) analysis [21]using M vs. N1, and N2 vs. N1 data-sets, to identify Gene Ontology (GO) categories where specific genes in these data occur more often than would be expected by random distribution of genes. These findings (and those of the validation data-set below) are summarized in Table 1, where select over- represented GO categories are listed along with specific genes differentially expressed in our data-sets. These categories include oxidative phosphorylation, ubiquitin-dependent protein catabolism, nuclear messenger RNA (mRNA) splicing, ribosomes, metabolic processes, regulation of apoptosis, NF-κB pathways, cysteine-type endo-peptidase activity and antigen processing. Most are significant in both long-term (M vs. N1) and short-term (N2 vs. N1) practitioners of daily RR practice (see Table).

Table 1. Gene Ontology Categories
Even though our analyses of differentially expressed genes and GO categories associated with RR practice meet widely accepted criteria for statistical significance, we were concerned about the relatively small fold changes that were observed (see Supplementary Methods). To address this issue we employed Gene Set Enrichment Analysis (GSEA). GSEA has proven to be useful for capturing subtle expression changes in complex gene signatures based on predefined gene sets or pathways [22]. As described above, we examined expression data for 2 comparisons, M vs. N1 and N2 vs. N1. The selected


pathways or gene sets that are significantly enriched (False Discovery Rate (FDR.)<50%, nominal p-value (NPV)< = 0.02) are shown in Figure 2, with gene sets for N2 vs. N1 and M vs. N1 in Fig 2A and Fig 2B respectively.

Figure 2. GSEA Analysis.
The analysis has been performed for >1200 predefined datasets using GSEA 2.0 software. Signal values for each gene are obtained by collapsing the probe values using max_probe algorithm. Representative datasets, significantly enriched (FDR.<50%, or NPV< = 0.01) between any two groups and corresponding heatmaps (depicting relative gene expression changes of core enrichment) are shown in a) N2 vs. N1 and b) M vs. N1. Datasets that are enriched in both the original and validation analyses are marked with *. c) Heatmaps of ribosomal proteins and ubiquitin mediated proteolysis illustrate transitional trends in gene expression across the N1, N2 and M groups. doi:10.1371/journal.pone.0002576.g002

GSEA analysis of N2 vs. N1 showed highly significant enrichment in gene sets related to various cellular stressors/stress responses and metabolism. To a pronounced degree these observations complement the results of GO analysis presented in the Table, also depicting significant alterations in cellular response to stress, oxidative and primary metabolism. The transition effect of the RR from novice to short term (8 weeks) to long term RR practitioners has been denoted through a colorgram of ribosomal proteins and ubiquitin mediated proteolysis gene sets (Fig. 2C). Whereas expression of ribosomal genes is significantly upregulated in RR practitioners at 8 weeks and more pronounced in long term practitioners, ubiquitin mediated proteolysis gene expression in general shows an opposite trend. Closer inspection of the colorgrams for ribosomal proteins and Ub proteolysis gene sets shows some variation in the GEP in each subgroup (N1, N2M). The GEP of a few N1 or N2 subjects resembles the GEP of M subjects and


vice versa. To elucidate the association between GEP and subject characteristics (Race, Age, etc), we performed clustering of each subgroup separately (N1, M) using the enriched gene sets (Ribosomal and Ub Proteolysis). This analysis identified a subcluster in the N2 subgroup that has significant over-representation of Asian subjects (P value <0.05) when the clustering was performed using the ribosomal protein gene set. This observation needs further validation on a larger dataset as the current study contains only five Asian subjects. No other characteristic exhibited significant association with the ribosomal protein gene set. No significant association between the GEP profiles and subject characteristic was found when clustering was performed using the Ub proteolysis gene set. This analysis provides further insight into the stress response related genes that are influenced by RR practice.

Independent validation set analysis

As a validation of our results, we repeated the experimental and analysis procedures defined in the “Methods” section on a new set of samples consisting of 5 N1, 5 N2 and 6 M subjects. We found 1846 and 2390 probe sets differentially expressed between M vs. N1, and N2 vs. N1 groups. The validation data-set showed a significant (p<10−5) number of genes in common with the original analysis of 58 samples. We also found that 70–75% of all GO categories from the original analysis were retained in the validation set (p ~0), and 30–65% of significantly over-represented GO categories were shared. Of note, biologically relevant GO categories such as oxidative phosphorylation, regulation of apoptosis, and antigen presentation, come up as significantly over-represented in both the original and validation analyses. Results of the validation set and comparison analyses can be found in the online supplementary data. In addition, validation GSEA analysis on N2 vs. N1 subjects shows enrichment of ribosomal proteins and platelet expressed gene sets and enrichment of ribosomal proteins, oxidative phosphorylation and electron transport chain gene sets in M vs. N1subjects (Fig. 2A and 2B). The similarities between the original and validation


results from GSEA analysis argues against random chance accounting for the observed enrichment of these gene sets.


Results from our study indicate that there are distinct differences in the GEPs between individuals with many years of RR practice (group M) and those without such experience (group N1). Furthermore we find significant GEP changes within the same individuals before (N1) and after 8 weeks of RR training (N2). Finally, the changes in GEP found in M vs. N1, and those of N2 vs. N1, are to a great degree similar when assessed by analysis of differentially expressed genes, GO analysis and GSEA.

It is becoming increasingly clear that psychosocial stress can manifest as system-wide perturbations of cellular processes, generally increasing oxidative stress and promoting a pro-inflammatory milieu [23]–[25]. Stress associated changes in peripheral blood leukocyte expression of single genes have been identified [26]–[28]. More recently, chronic psychosocial stress has been associated with accelerated aging at the cellular level. Specifically, shortened telomeres, low telomerase activity, decreased anti-oxidant capacity and increased oxidative stress are correlated with increased psychosocial stress [29]and with increased vulnerability to a variety of disease states [30]. Stress-related changes in GEP have been demonstrated by microarray analysis in healthy subjects, including up-regulation of several cytokines/chemokines and their receptors [31], and in individuals suffering from post-traumatic stress disorder, including inflammation, apoptosis and stress response [32] as well as metabolism and RNA processing pathways [33]. The pro-inflammatory transcription factor NF-kappa B (NF-κB) which is activated by psychosocial stress has been identified as a potential link between stress and oxidative cellular activation [34].

The RR is clinically effective for ameliorating symptoms in a variety of stress- related disorders including cardiovascular, autoimmune and other inflammatory


conditions and pain [15]. We hypothesize that RR elicitation is associated with systemic gene expression changes in molecular and biochemical pathways involved in cellular metabolism, oxidative phosphorylation/generation of reactive oxygen species and response to oxidative stress and that these changes to some degree serve to ameliorate the negative impact of stress. Genome-wide evaluation of PBL GEP is a reasonable approach to survey the transcriptional changes that are involved in elicitation of the RR. The GEP of RR practitioners presented here reveals altered gene expression in specific functional groups which suggest a greater capacity to respond to oxidative stress and the associated cellular damage. Genes including COX7B, UQCRB and CASP2 change in opposite direction from that in the stress response [31], [32].

Our findings are relatively consistent with those found in a study of Qi Gong [17], a practice that elicits the RR. In their study of 6 Qi Gong practitioners and 6 aged matched controls, practitioners had down-regulation of ubiquitin, proteasome, ribosomal protein and stress response genes and mixed up- and down-regulation of genes involved in apoptosis and immune function. We find a similar pattern of GO categories that are significantly over-represented in GO or enriched in GSEA in our cross sectional comparison, M vs. N1. However, in our data-set ribosomal proteins were up-regulated.

Overall, similar genomic pattern changes occurred in practitioners of a specific mind body technique (Qi Gong) as well as in our long-term practitioners who utilized different RR practices including Vipassana, mantra, mindfulness or transcendental meditation, breath focus, Kripalu or Kundalini Yoga, and repetitive prayer. This indicates there is a common RR state regardless of the techniques used to elicit it.

Our study is the first to prospectively evaluate GEP changes in individuals before and after a short-term (8 week) RR training which consequently enables an appreciation of the parallel GEP changes that occur with short- and/or long-term RR practice. Replications in larger cohorts are warranted. Future investigations


could better define the therapeutic value and required duration of RR training to counter stress-related disorders.

Materials and Methods Participants

Nineteen healthy practitioners of various RR eliciting techniques (including several types of meditation, Yoga, and repetitive prayer) participated (M group; n = 19). Years of practice averaged 9.4 years (5.0 sd) and ranged from 4 to 20 years. Twenty individuals without any prior RR eliciting experience served as controls (N group; n = 20).. As shown in Table 2, the M and N groups are matched with respect to age, gender, race, height, weight, and marital status, which do not exhibit significant difference between the groups (p>0.05, t- and chi- square test).

Table 2. Demographicsdoi:10.1371/journal.pone.0002576.t002


The study protocol was approved by the Committee on Clinical Investigations at the Beth Israel Deaconess Medical Center (BIDMC), Boston MA. All subjects provided written informed consent and the study was conducted in the General Clinical Research Center (GCRC) of the BIDMC. After providing written informed consent, participants were screened by a physician and had blood drawn to ensure good health. All participants completed a testing session in the GCRC. N1(novice) subjects had 8-weeks of RR training, listened to a 20-minute RR-eliciting CD daily and returned to the GCRC for a repeat testing session (hereafter classified as the N2 group).


Relaxation-Response Training

N subjects received 8 weeks of RR training. Training included information about reducing daily stress, and a 20-minute elicitation of the RR [35]. Subjects randomized to the RR group received 8 weekly individual RR-training sessions from an experienced clinician as per our manualized research protocol [35]. The first session provided an educational overview of the stress response and the RR, instructions on how to elicit the RR, and a 20-minute guided RR experience. Sessions 2 through 8 consisted of a review of the subject's home practice card for compliance and a 20-minute guided RR experience.

During the RR elicitation in the weekly session, the subject was guided through a RR sequence including: diaphragmatic breathing, body scan, as well as mantra and mindfulness meditation, while subjects passively ignored intrusive thoughts. The specific CD guided the subject through the same sequence and has our clinical research studies and clinical practice for more than 15 years [35]. Subjects were asked to listen to the RR-eliciting CD once a day for 20 minutes at home.

To measure compliance, participants' daily home practice logs were reviewed each week and at the end of the 8 week training. These logs indicate that N subjects listened for an average of 17.5 minutes per day (3.7 sd) over 8-weeks.

Microarray Analysis

Following previously described protocols, the transcriptional profile of samples were probed using Affymetrix HG-U 133 Plus 2.0 chips representing over 47,000 transcripts and variants using more than 54,000 probesets. Scanned image output files were visually examined for major chip defects and hybridization artifacts and then analyzed with Affymetrix GeneChip Microarray Analysis Suite 5.0 (MAS5) software (Affymetrix). The image from each chip was scaled such that the 2% trimmed mean intensity value for all arrays was adjusted to target


intensity and reported as a non-negative quantity. Chips used for subsequent analysis consisted of 19 M, 19 N1 and 20 N2 samples (one chip from the N1group had insufficient signal values). A hierarchical clustering technique was used to construct an Unweighted Pair Group Method with Arithmetic-mean (UPGMA) tree using Pearson's correlation as the metric of similarity [36]. The expression data matrix was row-normalized for each gene prior to the application of average linkage clustering. When comparing 2 groups of samples to identify genes enriched in a given group, we used combination of three criteria. We considered genes with significantly different expression across the two groups using t-test (p<0.05) that further remained significant at a 5% false discovery rate (FDR.) using permutation testing with 1,000 permutations [37], [38]. In order to finalize a set of genes significantly up-regulated in a given group compared to another group, among the genes that passed the aforementioned steps, we filtered the ones that are “present” in at least half of the samples in the enriched group using Affymetrix' MAS5 Presence/Absence (P/A) calls. We used a paired t- test when comparing samples in groups N1 and N2.

Data deposition: All data sets have been deposited in the Gene Expression Omnibus, www.ncbi.nlm.nih.gov/geo (accession nos. GSE10041 and GSM253663-253734).

Gene Ontology and Gene Set Enrichment Analyses

Differentially expressed genes between the 3 groups (N2 vs. N1, M vs. N1 and M vs. N2) were separately analyzed using EASE to identify biologically relevant categories that are over-represented in the input set [21]. EASE analyses tested each list against all genes on the chip and overrepresentation describes a group of genes belonging to a certain GO category that appear more often in the given input list than expected to occur if the distribution were random. GO categories that had EASE scores of 0.05 or lower were selected as significantly over- represented. Gene Set Enrichment Analysis (GSEA 2.0 packagehttp://www.broad.mit.edu/gsea/) was used to determine whether an a priori


defined set of genes showed statistically significant, concordant differences between 2 groups (N2 vs. N1, and M vs. N1) in the context of known biological pathways. We tested expression values of all the genes in the relevant sample groups against 1687 gene sets obtained from the MSigDB2.0 for enrichment belonging to various metabolic pathways, chromosomal locations and functional sets (gene sets related to cancer/cancer cells are not included). The enriched gene sets have nominal p-value (NPV) less than 1% and False Discovery Rate (FDR.) <50% after 100 random permutations. These criteria ensure that there is minimal chance of identifying false positives.


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