yoga and DNA dAMAGE

1. REGULAR ARTICLE
Role of Yoga and Its Plausible Mechanism in the Mitigation of
DNA Damage in Type-2 Diabetes: A Randomized Clinical Trial
Rajesh G. Nair, M.S.1
∙ Mithila M Vasudev, PhD1
∙ Ramesh Mavathur, PhD1
Published online: 28 June 2021
© Society of Behavioral Medicine 2021. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.
Abstract
Background Although yoga is found to be beneficial in
the management of type 2 diabetes (T2D), its mechanism
of action is poorly understood. T2D is also known to be
associated with increased oxidative stress (OS) and DNA
damage.
Purpose This study examines how yoga modulates
OS-induced DNA damage and the efficiency of DNA
repair in T2D conditions.
Methods In this assessor-masked randomized clinical
trial, T2D subjects (n = 61), aged (Mean ± SD, 50.3 ±
4.2) were randomly allocated into Yoga group (31) that
received 10 weeks of yoga intervention and Control (30)
with routine exercises. Molecular and biochemical as-
sessments were done before and after the intervention
period. Structural Equation Modeling using “R” was
used for mediation analysis.
Results At the end of the 10th week, Yoga group
showed significant reduction in DNA damage indica-
tors like Tail Moment (−5.88[95%CI: −10.47 to −1.30];
P = .013) and Olive Tail Moment (−2.93[95%CI: −4.87
to −1.00]; P < .01), oxidative DNA damage marker
8-OHdG (−60.39[95%CI: −92.55 to −28.23]; P < .001)
and Fasting Blood Sugar (-22.58[95%CI: −44.33 to
−0.83]; P = .042) compared to Control. OGG1 protein
expression indicating DNA repair, improved signifi-
cantly (17.55[95%CI:1.37 to 33.73]; P = .034) whereas
Total Antioxidant Capacity did not (5.80[95%CI: -0.86
to 12.47]; P = 0.086). Mediation analysis indicated that
improvements in oxidative DNA damage and DNA re-
pair together played a major mediatory role (97.4%) in
carrying the effect of yoga.
Conclusion The beneficial effect of yoga on DNA damage
in T2D subjects was found to be mediated by mitigation
of oxidative DNA damage and enhancement of DNA
repair.
Clinical Trial information (www.ctri.nic.in)
CTRI/2018/07/014825
Keywords: Yoga ∙ Type 2 diabetes ∙ DNA damage and
Repair ∙ Mechanism ∙ Oxidative stress ∙ 8-OHdG
Introduction
Type-2 diabetes (T2D) is becoming one of the fastest-
growing major non-communicable diseases (NCDs) ad-
versely affecting human life and well-being globally, in a
multitude of ways. The latest epidemiological data shows
its worldwide prevalence as 463 million and is projected
to reach 700 million by 2045 [1]. Owing to its treatment
costs and loss of productive years, the global burden of
diabetes, including direct and indirect costs for 2015 was
estimated to be US$1.31 trillion and is predicted to reach
$2.1 trillion by 2030 [2]. Besides incurring huge expend-
iture and stretching the existing medical infrastructure,
the cost of treatment and ongoing management is a major
issue, particularly in low-and middle-income countries [3].
People with T2D are susceptible to micro and
macrovascular diseases, peripheral neuropathy, cogni-
tive decline, and depression [4] and it is considered as
a significant risk factor for cardiovascular (CVD) and
chronic kidney diseases [5]. T2D also adversely affects
the immune system, making them prone to infections
[6–8] and at increased risk of illness from infectious dis-
eases, such as flu or Covid-19 [9]. Besides being one of
the top 10 causes of death among adults globally, studies
indicate that T2D incidence is rising even in young adults
because of unhealthy lifestyle [1, 10].
Rajesh G Nair
rajeshakanair@gmail.com
1
Molecular Bioscience Lab, Anvesana Research Labs,
S-VYASA, Bangalore, India
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DOI: 10.1093/abm/kaab043
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2. T2D – Oxidative stress, DNA damage, and repair
Oxidative stress (OS) refers to the imbalance between
the generation of reactive oxygen species (ROS) and
their scavenging by the inherent antioxidant defenses
of the cell [11, 12]. The abnormal accumulation of ROS
is the underlying pathology in a variety of micro and
macrovascular diseases [13–15], endothelial dysfunction,
atherosclerotic plaques, and subsequent development
of CVDs and stroke [13, 16]. The mechanism by which
abnormal accumulation of ROS contributes to patho-
logical conditions involves damage or oxidative modifi-
cation of biomolecules, such as nucleic acids, lipids, and
proteins. One of the most common targets of ROS is the
DNA, the biomolecule carrying genetic information,
which is fundamental for the functioning and survival of
all living organisms. The innumerable types of damage
that can occur due to endogenous and exogenous causes
pose a serious threat to the transmission of the exact
complement of genetic material [17]. On average, each
human cell experiences more than 10,000 DNA lesions
per day [18], which are typically caused by normal cel-
lular processes. Most of these damages are repaired by
the organism’s inherent repair mechanisms [19]. But any
prolonged inability to repair the damages becomes the
basis of genomic instability and possible pathological
outcomes [20, 21]. At the molecular level, T2D con-
dition is associated with a higher level of OS, reduced
antioxidant capacity, increased oxidative DNA damage
[13], and impaired DNA damage-repair capability [14].
Among the DNA nucleobases, guanine is the most oxi-
dized because of its low redox potential [14, 22]. In this
premise, 8-hydroxy-2’-deoxyguanosine (8-OHdG) is con-
sidered as a key biomarker for oxidative DNA damage
induced by ROS and is found to be the most prolific oxi-
dative DNA repair product in the human system [16, 23].
8-Oxoguanine glycosylase 1 (OGG1), a short patch gene
as part of the Base Excision Repair (BER) machinery
is mainly involved in the repair of oxidatively induced
DNA damage [24, 25]. Inability or improper repair of
oxidative DNA damage can lead to single or double-
strand breaks and possibly to mutations, cell senescence,
or apoptosis [14, 26], which are associated with the eti-
ology and pathophysiology of many diseases including
cancer [16, 23, 26].
Yoga – T2D, OS, DNA damage and repair
Besides non-modifiable factors, an unhealthy lifestyle is
considered to be a major contributing factor in the de-
velopment of T2D and its complications [27]. Hence,
apart from the medical management, diet and physical
activity-based health behavior modifications are also
strongly recommended by experts and governing bodies
for T2D management [4, 10, 28]. Yoga, a mind-body
practice that originated in India and being practiced
worldwide, is considered one of the effective therapeutic
approaches based on health behavior modifications
for the prevention and management of T2D [29, 30].
Research evidence indicates that yoga practices benefited
T2D patients by improving blood sugar and lipid levels,
body composition, pulmonary and autonomic functions,
mood, sleep, quality of life, and reduced medications [28,
31]. Studies found yoga interventions feasible with high
acceptability [32, 33], and also highly cost-effective com-
pared to the conventional treatment [34]. Although cer-
tain studies have looked into the effect of yoga on DNA
damage, OS, and cellular aging [35] and some reported
that yoga reduced OS [31, 36] in T2D patients and DNA
damage in sperm cells [37], hitherto, no study has looked
into the mechanism of yoga at the molecular level in
T2D-associated DNA damage and repair.
Considering this evidence, we hypothesize that
the effect of yoga is mediated through alleviation of
OS-induced DNA damage and improvement in DNA
repair capability in T2D conditions. Hence, this study
aims to elucidate the mechanism of action of yoga on
T2D related DNA damage in terms of its effect on oxi-
dative DNA damage and DNA repair markers. In add-
ition to that, the study also assesses the effect of yoga on
the anthropometric and biochemical parameters in T2D
subjects.
Methods
Study Design
This study followed an assessor-masked randomized
clinical trial design. (Figure 1). T2D subjects (n = 61)
meeting study criteria were recruited and randomly al-
located in a 1:1 ratio using permuted random blocks
of variable size (maximum, 6) to Yoga group (31) and
Control group (30). An independent researcher from an-
other lab generated a randomization sequence and did
the allocation concealment using sequentially numbered,
sealed, opaque envelopes. Participants were not blinded
from the intervention because of its interactive nature,
though for the lab assessments and data analysis, par-
ticipant identity was masked by coding. This trial was
approved by the Institutional Ethics Committee of
S-VYASA (Reference No: RES/IEC-SVYASA/119/2017)
and registered at the Clinical Trial Registry of India
(Trial Registration No: CTRI/2018/07/014825). All par-
ticipants of this study, which was conducted from Jan 3,
2019, to May 27, 2019, were service personnel or their im-
mediate family members from Adugodi Police Quarters,
Bangalore, Karnataka, India. Written informed consent
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3. was obtained from all the participants before the com-
mencement of the study.
Study Participants
Subjects identified with T2D (fasting plasma glucose
above 126 mg/dL, as per American Diabetes Association
guidelines), age ranging from 35 to 60 from both genders
(Male: 33, Female: 28) and having no prior yoga experi-
ence were enrolled using convenience sampling for this
study. Key exclusion criteria were people having type-1
diabetes, uncontrolled T2D (FBS 200 mg/dL), cancer,
CVDs, neurological or psychological disorders (eating,
attention disorders), mobility restrictions, or any other
serious conditions that can confound the study outcome.
Sample size (70), was arrived at (ES = 0.6, α = 0.05,
power = 0.8) based on a pilot study conducted earlier at
the S-VYASA yoga clinic. Though 72 subjects assented
for the study, 11 did not appear for baseline data col-
lection and were excluded. All participants were using
medicines for glycemic control. Few participants were
using blood pressure 11(18%) and cholesterol-lowering
6(9.8%) medications. None of the participants had
any serious co-morbid medical conditions in the past.
Participants were not appreciably different in terms of
demographic, socioeconomic status, and clinical char-
acteristics. Baseline characteristics of study participants
are given in Table 1.
Intervention
The Yoga group underwent 10 weeks of yoga sessions
including Asana (specific postures) and Pranayama (spe-
cific breathing) practices as given in the intervention
protocol (Supplement 1 Intervention Protocol), 1 hour
per day for 4 days a week, by a certified and trained
yoga professional at a designated place. On average, a
participant in the Yoga group attended 37 hours of in-
structed yoga practices. This study used an adapted
version of the yoga module, developed, validated, and
used by AYUSH, Ministry of Health, Govt. of India,
for a national level program for T2D management [38].
Yoga practices were provided as adjuvant therapy to
their already existing course of treatment. The Control
group did physical exercises like walking, jogging, and
stretching 4 hours/week for 10 weeks, though they were
not instructor-led sessions. Compliance with the exercise
routine was around 81% as obtained by self-reporting.
None from the Control group practiced yoga during the
study period. Study participants were not asked to drop
or change any medication, nor any specific diet plans
were recommended as part of the study.
Outcome Measures
Primary outcome measures include Fasting Blood Sugar
(FBS) for glycemic status; Comet assay parameters: Tail
Fig. 1. CONSORT flow diagram.
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4. Moment (TM) and Olive Tail Moment (OTM) as indices
of DNA damage (higher values indicate more damage),
where TM is defined as the product of the tail length
and the fraction of total DNA in the tail (TM = Tail
length x Tail DNA%), OTM is defined as the product
of distance between the intensity-centroids of the comet
head and tail, and the fraction of total DNA in the tail
(OTM = Distance between the comet head and tail cen-
troids x Tail DNA%); 8-hydroxy-2’-deoxyguanosine
(8-OHdG), a marker for oxidative DNA damage;
8-Oxoguanine glycosylase 1 (OGG1) protein expression,
a primary enzyme responsible for the excision of OS in-
duced DNA damage, for DNA repair activity; and Total
Antioxidant Capacity (TAC) for free radical scaven-
ging ability. Secondary outcome measures include Low
Density Lipoprotein (LDL), High Density Lipoprotein
(HDL), Triglycerides (TG), Total cholesterol (TC),
Blood pressure, Waist-Hip Ratio (WHR), and Body
Mass Index (BMI).
The authors would like to state that certain outcome
variables enlisted in the trial registry are different from
those assessed in the current study. The details are listed
in the supplement (Supplement 5 Changed outcome meas-
ures) along with the justification.
Assessments
Anthropometric data and venous blood samples
(fasting) were collected at the baseline and the end of
the 10th week. All the biochemical variables were as-
sessed using an automated serum chemistry analyzer
(Mindray BS390). Alkaline comet assay was performed
using peripheral blood mononuclear cells (PBMCs) to
assess the DNA damage, which is based on the prin-
ciple that fragmented or the damaged DNA will mi-
grate out of the cell during agarose gel electrophoresis
creating a comet-like structure with damaged DNA
fragments forming the tail, while the intact DNA re-
mains in the head. OGG1 protein expression was as-
sessed using Western Blot technique. Serum 8-OHdG
was assessed using ELISA kit (Cat No. CEA660Ge,
Cloud-clone Corp, Texas). Total Antioxidant
Capacity (TAC) was assessed by ABTS (2,2’-azino-
bis (3-ethylbenzothiazoline-6-sulfonic acid) method.
The optical density measurements for both ELISA
and TAC were obtained using a plate reader (Enspire,
Perkin Elmer, Massachusetts). Detailed protocol de-
scriptions are given in the lab protocols (Supplement 2
Lab protocols).
Statistical Analysis
Data were analyzed for outliers and normality and were
presentedasMean(SD).Statisticalcomparisons(2-sided)
between groups were done using Student’s Independent
t-test for parametric data and Wilcoxon rank-sum test
for non-parametric data. Confidence Intervals (CI) were
reported with 95% confidence. P-value 0.05 was con-
sidered statistically significant. Effect Size (ES) (Cohen’s
d) also was calculated. All tests were done using “R”stat-
istical software (version 3.6.1).
Mediation Analysis
This study employed a mediation model based on mul-
tiple regression to examine and analyze any mediatory
role played by oxidative DNA damage (8-OHdG) and
DNA repair activity (OGG1) in carrying the effect
of yoga on resultant DNA damage (TM). Mediation
modeling and analysis were done using Structural
Equation Modeling (SEM) technique provided by
“R” statistical package “lavaan” (version 0.6–5) [39].
Model fit measures, direct and indirect effects, their
strength and significance were assessed using SEM
Table 1. Baseline characteristics of study participants
Yoga Control
(n = 31) (n = 30)
Characteristic
Age, mean (SD), y 49.8(4.6) 50.8(3.8)
Sex, No. (%)
Male 16(51.6) 17(56.7)
Female 15(48.4) 13(43.3)
Education, no. (%)
1 ≤ High school 5(16) 6(20)
2 = Pre-University 13(42) 14(47)
3 = Graduate 9(29) 8(27)
4 = Post-Graduate 4(13) 2(7)
Employed, No. (%) 22(71) 24(80)
Household income/year, no. (%)
1 ≤ $ 5000 5(16) 2(7)
2 = $ 5000 - $ 6999 14(45) 18(60)
3 = $ 7000 - $ 8999 11(35) 5(17)
4 ≥ $ 9000 1(3) 5(17)
Blood pressure, mean (SD)
Systolic (mm Hg) 126.2(9.7) 129.1(9.5)
Diastolic (mm Hg) 86.7(8.4) 88.6(8.1)
Body Mass Index, mean (SD) 26.3(1.2) 26.5(1.7)
Waist-Hip Ratio, mean (SD) 0.93(0.04) 0.95(0.05)
T2D duration, mean (SD), y 5.5(1.3) 5.6(1.4)
Medications, No. (%)
T2D 31(100) 30(100)
Hypertension 5(16) 6(20)
Cholesterol 4(13) 2(6.7)
Demographic, socioeconomic, and clinical characteristics of the
participants.
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5. analysis. As the bootstrapping method is employed to
evaluate standard error, values of “Std.Err,” “z-value,”
and “p-value” vary with repeated executions, though
the “Estimates” (regression coefficients) remain con-
stant. Basic assumptions for the multiple regression
model were tested, including the causative relation-
ship between variables and multicollinearity between
predictors.
Results
As seen in the CONSORT Flow Diagram (Figure 1), out
of 61 participants enrolled, 45 remained until the end of
this study and all received intended treatment. Dropouts
were almost equal for both groups and were at random.
Mean ± SD of age and duration of T2D for the Yoga
group (n = 22, Male/Female: 11/11) was 49.5 ± 4.7 and
5.44 ± 1.49 and that of the Control group (n = 23, Male/
Female: 13/10) were 50.6 ± 3.9 and 5.73 ± 1.44 respect-
ively. Baseline outcome measures showed no significant
difference between groups (Supplement 3 Baseline meas-
ures). No medication changes were reported by any par-
ticipant during the study.
Adverse events
No adverse events were reported by any participant.
A few reported muscular pain in the first week of inter-
vention, which faded away in subsequent days without
any medication.
Primary outcome variables
Among primary outcome variables (Table 2), at the end
of 10th week, participants in the Yoga group showed
significant reduction (expressed as between-group mean
difference (ΔG
) with 95% CI, effect size and significance)
in DNA damage markers, TM (ΔG
= −5.88[95%CI:
−10.47 to −1.30]; d = 0.77, P = .013) and OTM (ΔG
=
−2.93[95%CI: −4.87 to −1.00]; d = 0.91, P .01) com-
pared to the Control. Though FBS (ΔG
= −22.58[95%CI:
−44.33 to −0.83]; d = 0.62, P = .042) reduced signifi-
cantly for Yoga group, improvement observed in TAC
(ΔG
= 5.80[95%CI: −0.86 to 12.47]; d = 0.52, P = 0.086)
was not significant. While OGG1 protein expression
(ΔG
= 17.55[95%CI:1.37 to 33.73]; d = 0.65, P = .034),
representing DNA repair improved significantly,
8-OHdG (ΔG
= −60.39[95%CI: −92.55 to −28.23];
d = 1.13, P .001), the marker for oxidative DNA
Table 2. Primary and secondary outcome measures
Yoga (n = 22) Control (n = 23)
Mean (SD) Mean (SD) 95% CI ES P-value
Primary Outcomes
FBS (mg/dL) 162.7 (34.8) 185.3 (37.4) −44.33 to −0.83 0.62 .042
TAC (TE/mL) 34.0 (10.2) 28.2 (11.9) −0.86 to 12.47 0.52 .086
8-OHdG (pg/dL) 207.1 (48.3) 267.5 (58.0) −92.55 to −28.23 1.13 .001
OGG1 (AU) 79.3 (24.2) 61.8 (29.3) 1.37 to 33.73 0.65 .034
TM (AU) 11.8 (6.0) 17.7 (8.9) −10.47 to −1.3 0.77 .013
OTM (AU) 6.2 (2.5) 9.2 (3.8) −4.87 to −1 0.91 .004
Secondary Outcomes
BMI (kg/m2
) 26.1 (1.4) 26.8 (2.1) −1.84 to 0.27 0.45 .14
WHR 0.92 (0.05) 0.96 (0.06) −0.07 to −0.01 0.77 .013
SBP (mmHg) 123.8 (8.2) 130.8 (11.6) −13.05 to −0.97 0.7 .024
DBP (mmHg) 84.6 (8.1) 89.4 (9.4) −10.08 to 0.48 0.55 .074
LDL (mg/dL) 134.4 (17.4) 146.7 (22.5) −24.51 to −0.24 0.61 .046
HDL (mg/dL) 43.0 (4.2) 39.7 (3.7) 0.98 to 5.71 0.85 .007
TG (mg/dL) 130.7 (45.2) 178.9 (69.8) −83.75 to −12.71 0.82 .009
TC (mg/dL) 203.5 (18.1) 222.2 (27.9) −32.88 to −4.46 0.79 .011
All values are expressed as Mean (SD). FBS: Fasting Blood Sugar; TAC: Total Antioxidant Capacity;
8-OHdG: 8-hydroxy-2’-deoxyguanosine; OGG1: 8-Oxoguanine glycosylase; TM: Tail Moment; OTM: Olive Tail Moment; LDL/HDL:
Low/High Density Lipoprotein; TG: Triglycerides; TC: Total Cholesterol; BMI: Body Mass Index; WHR: Waist to hip ratio; SBP/DBP:
Systolic/Diastolic Blood Pressure; TE: Trolox Equivalents; AU: Arbitrary Units; CI: Confidence Interval; ES: Effect Size (Cohen’s d). P
.05 considered as statistically significant. SI conversion: To convert glucose, cholesterol, and triglycerides to mmol/L, multiply values by
0.055, 0.0259, and 0.0113 respectively. To convert 8-OHdG to pmol/L, multiply values by 0.0353
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6. damage showed highly significant reduction in Yoga
group compared to the Control.
Results from the mediation analysis
Asforassumptiontesting,thelinearregression-basedana-
lysis showed a significant causative relationship between
“group” and the outcome variable (TM) (P = .013) and
with mediators [8-OHdG (P .001), OGG1(P = .03)].
Also, 8-OHdG (P .001) and OGG1(P = .024)
showed a significant causative relationship with TM.
Multicollinearity, a measure of the correlation be-
tween the predictors (8-OHdG and OGG1 predicting
TM), as indicated by the variance inflation factor (VIF)
(8-OHdG = 1.002166 and OGG1 = 1.002166), were
found to be within the acceptable levels ( 5).
Figure 2 represents the proposed mediation model with
details of regression coefficients and their significance for
each path: Direct path (Group → TM) is represented by
ć, and Indirect paths by Group→ 8-OHdG → TM (a1, b1)
and Group → OGG1 → TM (a2, b2). SEM-based medi-
ation analysis showed that the mediatory effect of 8-OHdG
(a1*b1 = −4.174[95%CI: −8.412 to −1.395]; P = .02) as a
proportion of total effect was 70.9% and that of OGG1
(a2*b2 = −1.563[95%CI: −3.338 to −0.104]; P = .063) was
26.6%, while that of direct path (ć = −0.150[95%CI: −4.23
to4.32];P = .944)wasonly2.5%.Proportionalcontribution
of total indirect effect (a1*b1 + a2*b2 = −5.737[95%CI:
−10.173 to −2.558]; P = .005) was found to be highly sig-
nificant at97.4%.Detailedresultsof SEM-basedmediation
analysis are given in SEM analysis results (Supplement 4
SEM results).
Fit measures obtained for the present model
showed that they were within acceptable levels [40].
Comparative Fit Index = 1.000 [CFI ≥.95], Tucker-Lewis
Index = 1.027 [TLI ≥.95], Root Mean Square Error of
Approximation = 0.000 [RMSEA 0.08], Standardized
Root Mean Square Residual = 0.036 [SRMR 0.08] and
Chi-square test statistic = 0.841, (P = 0.359) [P 0.05].
Discussion
In this randomized clinical trial involving 10 weeks of
yoga intervention for T2D patients compared to a con-
trol group with routine exercises, the study results showed
significant improvements in various outcome measures
for the Yoga group. Yoga group showed a significant
reduction in DNA damage, as indicated by changes in
Tail Moment and Olive Tail Moment compared to the
Control group. This may have been due to consistent
improvement observed in causative parameters like
blood sugar, lipid levels, and oxidative DNA damage
(8-OHdG) observed in the Yoga group, though the im-
provement observed in the antioxidant status (TAC) was
not significant (Table 2). Previous studies also have re-
ported that yoga practices reduce blood sugar [41], oxi-
dative stress, and improve antioxidant status and thus
may have therapeutic effects on metabolic disorders [31,
36, 42]. Tolahunase et al., reported a significant reduc-
tion in cellular aging rates, represented by lower levels of
8-OHdG, ROS, cortisol, IL-6, increased TAC, and tel-
omerase activity in healthy population practicing yoga
[35].
On the DNA repair front, results from the pre-
sent study showed significant improvement in OGG1
protein expression. It is evident from the results that
DNA damage is diminishing in the Yoga group post-
intervention as indicated by the slopes of the regression
equation compared to baseline (Figure 3), with lower
8-OHdG and higher OGG1 levels. This shows, that oxi-
dative DNA damage was reduced and the efficiency of
DNA repair was enhanced in the Yoga group. None
of these improvements were observed in the Control
group (Figure 4). Similar observations were made by
studies employing exercise and yoga training in people
with T2D and a healthy population. While exercise
training increased genomic stability with better regu-
lation of DNA damage response in T2D patients [43,
44], others reported regular exercises increased OGG1
activity and promote DNA repair [22] and yogic prac-
tices were found to upregulate DNA repair genes like
PARP and OGG1 in sperm cells leading to reduced
DNA damage [45].
Interestingly, oxidative stress was also found to di-
minish OGG1 protein stability by downregulating p300
mediated acetylation [14] apart from its down-regulation
in hyperglycemic conditions [46]. Increased DNA damage
found in the Control group with lower OGG1 levels in
the present study is in tune with the above observation.
Hence, lower DNA damage observed in the Yoga group
Fig. 2. Mediation model. Path diagram showing mediation effect
with regression coefficients and their significance for the pro-
posed mediation model. Mediatory paths (indirect) are Group→
8-OHdG → TM (a1, b1) and Group → OGG1 → TM (a2, b2).
Direct path (Group → TM) is represented by ć. Regression coeffi-
cients are a1, b1, a2, b2, and ć.
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7. in the post scenario could be due to the combined effect
of lower systemic oxidative stress, better scavenging of
oxidative radical by the antioxidant mechanism, and en-
hanced DNA repair among the practitioners. A scien-
tific review observed that yoga and meditative practices
conserve DNA integrity by reducing OS-induced DNA
damage and better regulation of genes involved in the
DNA repair process, cell-cycle control, and anti-inflam-
matory effects [37].
Among secondary parameters, study results showed
significant improvements in biochemical and anthropo-
metric measures in the Yoga group compared to the
Control. Along with significant improvements observed
in lipid profile which is reported to decrease free fatty
acids and thereby improve insulin sensitivity [47], im-
provements observed in body composition indicators
like WHR and BMI might have contributed to better
glucose uptake and reduced hyperglycemia-induced oxi-
dative stress. Previous studies involving yoga interven-
tion for T2D also reported similar benefits [31, 41], while
others reported improvements in weight, waist circum-
ference, and BMI except for blood sugar [33]. Systematic
reviews and meta-analyses on the effect of yoga on T2D
reported significant improvements in glycemic control,
lipid profile, blood pressure, BMI, WHR, and cortisol
levels for yoga practitioners compared to exercise [30,
48]. Our study findings are consistent with the conclu-
sions of various studies involving yoga intervention for
T2D [30, 48–50].
Mechanism of action of yoga as obtained from the medi-
ation analysis
Apart from assessing the effect of yoga on DNA damage
in T2D patients, we also elucidated its mechanism of
action by employing a statistical mediation analysis.
The SEM-based analysis showed that (Figure 2) there
exists a causative relationship between Group (binary
variable; presence/absence of yoga) and DNA damage
reduction in T2D patients, and that effect was carried
forward by its influence on oxidative DNA damage
and DNA repair, acting as mediators. Further, the ana-
lysis showed, the bulk of that effect (70.9%) was medi-
ated through a reduction in 8-OHdG and less (26.5%)
through OGG1 enhancement. Differential results from
the analysis showed that the path Group → 8-OHdG
→ TM (a1, b1) was more significant than the Group →
OGG1 → TM (a2, b2) path. This significance should be
understood in the light of their proportional contribu-
tion while considering the total effect. Individually con-
sidering, all regression coefficients (a1, b1, a2, and b2)
of both paths were significant. While the combined con-
tribution of mediators or total indirect effect (a1b1 +
a2b2) was found to be highly significant at 97.5%, that
of the direct path (ć) was insignificant at 2.5%, which
means that majority of the yoga-effect was carried by the
mediators. Though studies with similar settings and ana-
lysis were not observed, few showed that yoga practices re-
duce OS and enhance DNA repair gene activities [37, 45].
Fig. 3. Causative relationship between mediators and DNA damage. (A–D): Causative relationship between the mediatory variables’
Oxidative DNA damage (8-OHdG) and DNA repair (OGG1) with DNA damage (Tail Moment) in the light of a 3rd variable (indicated
by dot / triangle size) for both pre-and post-intervention
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8. Mediation results indicate that yoga’s effect is mainly
mediated through reduced oxidative DNA damage and
enhanced DNA repair in T2D related DNA damage,
though the former was more prominent than the latter.
Studies also observed that medication-based inten-
sive glucose control may lower the risk of cardiovascular
events but may not yield long-term mortality benefit [51].
But when medication is combined with behavioral changes,
long term benefits like reduction of vascular complications
[52] and all-cause mortality [27] were observed, though the
“LOOK AHEAD” study [53] observed non-significance,
nevertheless reported benefits like better glycemic control,
physical fitness, quality of life and reduced health expend-
iture as other studies. These evidences make a case for
integrating holistic and mind-body practice like yoga, with
its empowering nature that promotes positive health, which
is also cost-effective and relatively safe, in the management
of a lifestyle- disease like T2D.
Strengths and limitations of the study
To our understanding, this is the first study to eluci-
date yoga’s mechanism of action in T2D related DNA
damage and employ statistical mediation analysis for
the same. The study also followed an RCT design along
with assessments on anthropometric, biochemical, and
specific molecular markers. As for limitations, the study
had dropouts, a relatively smaller sample size and dur-
ation, and did not assess diet, sleep quality, and stress
levels of participants that could have an impact on
physiology. Though the study assessed OGG1 activity for
DNA repair, DNA damage response is a complex pro-
cess involving many players.
Conclusions
The findings of the study demonstrate that the bene-
ficial effect of yoga on DNA damage in T2D subjects
was found to be primarily mediated by mitigation
of oxidative DNA damage and enhanced DNA re-
pair. In general, yoga was found to be a potent ad-
junct therapy in the management of T2D by way of
its efficacy seen on anthropometric, biochemical, and
molecular markers in T2D patients. But, generaliza-
tion and interpretation of the study results across the
Figure 4. Group comparison of primary outcome measures with its distribution profile. (A–F): Distribution profile of primary outcome
measures with group comparison for both pre-and post-scenario. P-values were obtained using independent t-test. (A) Fasting Blood
Sugar; (B) Total Antioxidant Capacity; (C) OHdG (8-hydroxy-2’-deoxyguanosine); (D) OGG1 (8-Oxoguanine glycosylase); (E) Tail
Moment; (F) Olive Tail Moment.
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9. population should be done cautiously in the light of
socio-cultural, ethnic, and genetic differences, and
may entail more studies with larger sample size and
broader measures, including psychological assess-
ments like stress, anxiety, depression, and quality-of-
life in a diverse population.
Acknowledgment
We thank the lab technicians of Anvesana Research Labs for
their technical assistance and service in sample collection and
testing.
Funding: This work was supported by University Grants
Commission (UGC), India. (Research Grant Number: RA-2016-
18-GE-KAR-7161). UGC had no role in the design and conduct
of the study; collection, analysis, and interpretation of the data; in
the writing, review, or approval of the manuscript; and decision to
submit the manuscript for publication.
Compliance with Ethical Standards
Authors’ Statement of Conflict of Interest and Adherence to Ethical
Standards Authors Authors (Mithila M Vasudev, Rajesh G Nair,
and Ramesh Mavathur) declare that we have no conflicts of
interest.
Author Contributions: RM and RN were responsible for the con-
cept, design, and subject recruitment of the study. Experiments,
acquisition, analysis, or interpretation of data done by MV and
RN. While statistical analysis and drafting of the manuscript
done by RN, critical revision of the manuscript for important in-
tellectual content and project supervision were done by MV and
RM along with administrative, technical, and material support.
Funding was obtained by RM. All authors contributed with their
specific skills in the conduct of the study and approved the final
manuscript.
Human Rights: All procedures performed in studies involving
human participants were in accordance with the ethical standards
of the institutional and/or national research committee and with
the 1964 Helsinki declaration and its later amendments or compar-
able ethical standards.
Informed Consent: Informed consent was obtained from all partici-
pants included in the study.
Welfare of Animals: This study does not involve any animal
experiments.
Appendix A. Supplementary material
Supplement 1 Intervention Protocol
Supplement 2 Lab protocols
Supplement 3 Baseline measures
Supplement 4 SEM results
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