Biological markers for the effects of yoga as a complementary and alternative medicine

Antioxidant nature of yoga

Oxidative stress is a result of increased levels of reactive oxygen species (ROS) and reactive nitrogen species (RNS) over antioxidants and leads to the damage of different biomolecules like DNA, protein, lipids which further contribute to diseases including cancer, cardiovascular disease, neurodegenerative disorders and aging [12, 13, 14, 15]. For instance, generation of ROS leads to the overexpression of JUN, an oncogene which is involved in lung cancer [16]. To respond to deleterious effects of oxidative stress it is very crucial to maintain the required level of antioxidants in the body. For this cells produce various antioxidant enzymes like superoxide dismutase (SOD), catalase, peroxiredoxins (PRXs), malondialdehyde (MDA) etc. (Table 1).

Level of glutathione (GSH) is a major non-enzymatic intracellular marker of antioxidant status [4]. The level of Glutathione increased significantly from 235.3 + 16.9 nmol/L to 331.7 + 37.6 nmol/L among male volunteers of Indian navy who practiced yoga [17]. A Pilot study among pre-diabetics showed no significant difference between the baseline value 7.8 ± 2.5 and the final value 8.2 ± 2.4 after 3-months of yoga practice [18]. Whereas yoga practices (Yogasana, pranayama and meditation) for three months resulted in 2.1 fold increase in GSH among healthy university students [19].

GSH can be oxidized to glutathione disulfide (GSSG) with the action of glutathione peroxidase (GPX) in response to oxidative stress and Glutathione reductase (GR) recycle GSSG to its reduced state using reduced nicotinamide adenine dinucleotide phosphate (NADPH) as hydrogen donor. The ratio of GSH/GSSG, also known as the glutathione redox ratio (GRR) is a sensitive indicator of oxidative stress and has been shown to increase significantly (p<0.001) after 3 months of yoga practice (Yogasana, pranayama, and meditation) [17, 20].

Glutathione peroxidase (GPx) glutathione reductase (GR) and glutathione S-transferase (GST), SOD and catalase are the enzymatic antioxidant components that protect against oxidative stress. Studies on healthy individuals showed that activities of GPx, and GST were significantly increased after 3 months yoga practice (p<0.05). Whereas no significant change was observed in glutathione reductase activity (p<0.05) [19].

In another study conducted on physically active males from Indian Air Force, activity of glutathione S-transferase increased significantly (p<0.001), while GPx activity decreased significantly (p<0.001). GR activity also increased significantly (p<0.05) following 3 months yogic practice [17].

Superoxide radical (O₂^−•) is highly active oxidant however, SOD detoxifies superoxide radical (O₂^−•) by converting it to hydrogen peroxide and O₂; further, catalase and GSH detoxifies hydrogen peroxide into water and alcohol [21]. The activity of SOD increases significantly after following 3 months yoga practice among healthy individuals [17] whereas, Hegde et al documented significant decrease in SOD activity (unit/gmHb) after 3 months of yogic practices (Yogasana, shavasana and pranayama) 4721.0 ± 1263.0 (3992.0-5450) vs. 4340.0 ± 978.0 (3776.0-4905.0) among pre-diabetes patients [18]. There was significant 4.65% increase in SOD activity and 0.09% in catalase among patients with end-stage renal disease on hemodialysis who practiced hatha yoga for 4-months [22].

Vitamin C and vitamin E both are potent components of an antioxidant defense system. The levels of vitamin C and vitamin E were increased significantly (p<0.001) following yogic practices [17].

MDA is the end product of lipid peroxidation, which is stimulated by increased level of ROS. Moreover, increased level of MDA induces various cellular reactions which results in destruction of DNA and protein [23, 24]. A study done among the healthy yoga practitioners has shown significant reduction in the level of MDA as compared with the control group (p<0.01) [25]. In another study done among prediabetic patients, 3-month practice of yogasanas and pranayama and shavasana resulted in significant reduction in malondialdehyde level (p<0.001) [18].

A study conducted among hypertensive individuals, has shown that the level of MDA was significantly reduced (p<0.05) among patients who performed yoga (Yogasana and Pranayama), the decrease was by 4.0% after four months of intervention (p=0.096) [25]. Pal et al also observed Non significant decrease in both the control group (8.56 ± 0.63 to 8.38 ± 0.60) and yoga group (7.78 ± 0.50 to 6.01 ± 0.46) among healthy males [17]. This decrease in the MDA level may be due to decrease of lipid peroxidation via increased antioxidant level [26].

Total antioxidant status (TAS) is also an important antioxidant marker. The level of TAS increased significantly (p<0.001) following 3 months of yogic practice which indicates a marked improvement in the overall cellular antioxidant level [17].

The above mentioned studies involved different types of participants; healthy as well as individual suffering from various health conditions. The yoga interventions and the duration of the program were also varied. Majority of studies reported that practicing yoga was helpful in increasing the level of GSH, GSH/GSSH, total antioxidants status (TAS), vitamin C, vitamin E, glutathione reductase (GR) activity, catalase and GST activity and reducing the level of GSSG and MDA (Table 2).

Improvement of cardiovascular health due to yogic practices

People suffering from mood disorders have higher risk of cardiovascular disease due to less cardiorespiratory coupling and autonomic dysfunction [27, 28, 29]. Anxiety which is common in mood disorders is known to change the breathing patterns which evokes increase in tidal volume, respiratory rate and decrease in respiratory time amongst healthy people [30, 31]. Yoga has been shown to help in improved breathing function and coronary artery calcium (CAC), increased adaptation to hypoxia and improved mood [32, 33, 34]. Sudarshan Kriya yoga (SKY) which is known to be helpful in depression, anxiety and stress has now also been reported to increase the spontaneous respiratory coupling and cardiac autonomous control in patients with anxiety and stress disorders which decreases the risk of cardiovascular disease in such patients (Table 3) [39, 40, 41].

A study done among the heart failure (HF) patients has shown that 12 weeks of yoga therapy (Meditation and pranayama) along with standard medical therapy resulted in a significant decrease in Heart Rate (HR) (p<0.001), Rate pressure product (RPP) (p<0.001), systolic blood pressure (p<0.01), diastolic blood pressure (p<0.001), normalized Low Frequency power (LFnu) (p<0.001), low freq., high freq. ratio (LF/HF ratio) (p<0.001) and a significant increase in normalized High Frequency power (HFnu) (p<0.001) [35].

Another study among heart failure patients has shown significant improvement in left ventrical ejection fraction (LVEF) and Tei index after 12 weeks of yoga therapy (Meditation and pranayama). LVEF was increased from 38.93 ± 5.1 to 52.96 ± 6.01 (36.88% in the yoga group (YG) and 16.9% in the control group (CG) (p<0.01) and Tei index was reduced from 0.54 ± 0.85 to 0.38 ± 0.03 (27.87% in the YG) and 2.79% in the CG (p<0.01). A significant decrease 63.75% in the YG and 10.77% in the CG (p<0.01) in N-terminal pro b-type natriuretic peptide (NT-proBNP) level, which was also reported in Yoga Group (pre: 3965.48 ± 1365.08, post: 1395 ± 997.08) [36]. These results indicate that 12-week yoga therapy offered additional benefits to standard medical therapy for Heart Failure patients by improving cardiac function, parasympathetic activity while reducing the sympathetic activity and myocardial stress [36, 42, 43].

Pro-inflammatory markers such as interleukin-6 (IL-6), high sensitivity C-reactive protein (hs-CRP) and extra cellular superoxide dismutase (EcSOD) have been associated with unfavorable cardiovascular outcomes in heart failure [42]. Increased levels of IL-6 and CRP have been adversely associated with HF [43, 44] and ECSOD activity has been correlated with endothelium-mediated, flow-dependent vasodilatation [45]. Pullen et al showed that after yogic intervention (Yogasanas and pranayama) there was a significant reduction in serum level of IL-6 (19.6 ± 2.5 to 16.0 ± 2.1 mg. dL⁻¹: p<0.001) and hs-CRP (2.4 ± 0.58 to 1.9 ± 0.4 mg. dL⁻¹: p<0.001). Levels of EC-SOD increased from 509 ± 71.9 to 610 ± 86.2 U.mL⁻¹: p<0.001) [37] and the results showed consistency with the earlier studies of Pullen et al which showed statistically significant reductions in serum levels of inflammatory markers: IL-6 and hs-CRP and an increase in EC-SOD in the Yoga (Yogasanas and pranayama) treatment group (all p<0.005) [38].

Anti-ageing impacts of yoga

Aging is sequential change in an organism, or a decline of physiological goodness that leads to an increased threat of disease, debility, organism’s inability to habituate to metabolic stress and ultimately death [46]. Ageing at genomic level is largely the result of DNA damage caused by ROS, chemicals like benzo[a]pyrene, UV/IR radiations, spontaneous hydrolytic reactions, DNA replication errors which leads to various genetic lesions which includes point mutations, gene disruption, telomere shortening, translocations etc. Damage caused by these lesions is repaired by DNA repair mechanisms for example base excision repair (BER), nucleotide excision repair (NER), non-homologous end joining (NHEJ) [47, 48, 49, 50, 51]. Excessive DNA damage and insufficient DNA repair mechanism favors the aging process.

Recently published studies demonstrated that aging is associated with telomerase activity and telomere length and maintaining telomere length is important to prevent cellular senescence [46]. Telomere is repetitive nucleotide sequence at each end of the chromosome which protects chromosome from damage and prevents the fusion with adjoining chromosomes and these ends of chromosomes are vulnerable to age-related decay [52, 53, 54]. The length of telomere gets shorten with ageing and age-related diseases. Telomeres are found to be prone to inflammation and oxidative stress which can further promote telomere shortening, hence ageing. Loss of telomere-protective sequences due to deprivation of human telomerase reverse transcriptase (hTERT) activity lead to aging in humans and mice, whereas aging can be delayed in mice by reactivation of telomerase [46, 55, 56]. Telomeric DNA is protected by a six-subunit protein called Shelterin which bounds telomere prevents the ingression of DNA repair proteins to the telomeres [57]. Lack of shelterin induces telomere uncapping, non-homologous end joining, senescence and/or apoptosis [58, 59].

Several studies have provided significant evidence of the impact of yoga-intervention on telomerase activity and telomere length.

Tolahunase et al studied the effect of a 12 week Yoga (Yogasana, pranayama, and meditation) Based Lifestyle Intervention on both cardinal and metabotropic biomarkers associated with cellular aging. The findings showed a significant reduction in the mean levels of 8-hydroxy 2ʹ deoxyguanosine (8-OH₂dG) and ROS while enhancement in the mean levels of total antioxidant capacity (TAC) and telomerase activity (all values p<0.05). The mean level of telomere length was increased, but this result was not significant (p=0.069). The metabotrophic blood biomarkers associated with cellular aging are cortisol, β-endorphin, IL-6, Brain-derived neurotrophic factor (BDNF), and sirtuin-1. The mean levels of cortisol and IL-6 were significantly reduced and mean levels of β-endorphin, BDNF, and sirtuin-1were significantly increased (all values p<0.05) [60] (Table 4).

A study by Kumar et al showed there was an enhancement in telomerase activity and decrease in oxidative stress (ROS) and DNA damage marker (8-OH₂dG) by yoga (Asnas, pranayama, shavasana) lifestyle intervention. The yoga program included asanas (postures), pranayama (breathing exercises), stress management, group discussions, lectures, and individualized advice [61] (Table 4).

Lavretsky et al measured the effect of 12 min kirtan kriya and listening to relaxation music on telomerase activity using telomeric repeat amplification protocol (TRAPeze) telomerase detection kit, and observed a significant 43% increase in telomerase activity which leads to increase in length of telomeres and ultimately delay aging [62] (Table 4).

A similar study by Krishna et al reported that leukocyte telomere length (LTL) was significantly increased (p<0.001) in the yoga group by yogic actions like Asanas (bodily/tangible positions), Pranayama, Dhyana (meditation) and was measured by quantitative PCR [63] (Table 4).

Limitations

In many case studies the control group was missing, so it is very difficult to suggest whether the results are due to the yogic intervention or not [60, 61, 62]. In some studies, yogic intervention along with medication, physical exercise, or relaxing music were employed to analyze the changes in biological indicators and so it is difficult to assess the impact of yoga alone [19, 22, 25, 37, 38, 62, 63]. While in some studies the focus was to establish cause and effect relationship without going into the details of mechanism.