The Potential Benefits of Biofield Energy Treated Vitamin D3 on Bone Mineralization in Human Bone Osteosarcoma Cells (MG-63)

2018·0.9 MB·English

by  Aileen Carol Lee

International Journal of Nutrition and Food Sciences
2018; 7(1): 30-38
http://www.sciencepublishinggroup.com/j/ijnfs
doi: 10.11648/j.ijnfs.20180701.15

ISSN: 2327-2694 (Print); ISSN: 2327-2716 (Online)

The Potential Benefits of Biofield Energy Treated Vitamin
D on Bone Mineralization in Human Bone Osteosarcoma
3
Cells (MG-63)
Aileen Carol Lee1, Mahendra Kumar Trivedi1, Alice Branton1, Dahryn Trivedi1, Gopal Nayak1,
Sambhu Charan Mondal2, Snehasis Jana2, *
1Trivedi Global, Inc., Henderson, USA
2Trivedi Science Research Laboratory Pvt. Ltd., Bhopal, India
Email address:

*Corresponding author
To cite this article:
Aileen Carol Lee, Mahendra Kumar Trivedi, Alice Branton, Dahryn Trivedi, Gopal Nayak, Sambhu Charan Mondal, Snehasis Jana. The
Potential Benefits of Biofield Energy Treated Vitamin D on Bone Mineralization in Human Bone Osteosarcoma Cells (MG-63).
3
International Journal of Nutrition and Food Sciences. Vol. 7, No. 1, 2018, pp. 30-38. doi: 10.11648/j.ijnfs.20180701.15
Received: November 28, 2017; Accepted: December 14, 2017; Published: January 17, 2018

Abstract: The study was aimed to evaluate the potential of Consciousness Energy Healing based vitamin D3 and DMEM
medium on bone health. The test items, were divided into two parts. One part of each sample received the Consciousness Energy
Healing Treatment by Aileen Carol Lee and those samples were labeled as the Biofield Energy Treated (BT) samples, while the
other parts of each sample were denoted as the untreated test items (UT). ALP, collagen, and bone mineralization activities were
performed to assess bone health in human bone osteosarcoma cells (MG-63). The cell viability assay (MTT) data showed that the
test samples were found as safe in the tested concentrations. The level of ALP was significantly increased by 251.16%, 103.75%,
and 119.55% in the UT-DMEM + BT-Test item, BT-DMEM + UT-Test item, and BT-DMEM + BT-Test item, respectively at 50
µg/mL, while 123.37% increased in the BT-DMEM + UT-Test item at 0.1 µg/mL compared to the UT-DMEM + UT-Test item
group. Further, the ALP level was significantly elevated by 194.1%, 72.34%, and 65.31% in the UT-DMEM + BT-Test item, BT-
DMEM + UT-Test item, and BT-DMEM + BT-Test item groups, respectively at 100 µg/mL compared to the UT-DMEM + UT-
Test item group. Collagen was significantly increased by 83.02%, 145.67%, and 191.41% in the UT-DMEM + BT-Test item, BT-
DMEM + UT-Test item, and BT-DMEM + BT-Test item groups, respectively at 0.1 µg/mL compared to the untreated group.
Further, the collagen level was significantly increased by 50.29% and 52.48% in the BT-DMEM + UT-Test item and BT-DMEM
+ BT-Test item, respectively at 50 µg/mL compared to the untreated group. Additionally, the percent of bone mineralization was
distinctly increased by 64.46% and 94.16% in the UT-DMEM + BT-Test item and BT-DMEM + UT-Test item groups,
respectively at 100 µg/mL compared to the untreated group. The percent of bone mineralization was distinctly increased by
26.59% and 28.33% in the BT-DMEM + UT-Test item group at 10 and 50 µg/mL, respectively compared to the UT-DMEM +
UT-Test item group. Overall, the Biofield Energy Treated vitamin D3 was remarkably improved the bone health parameters and it
could be a powerful alternative nutraceutical supplement to combat vitamin D3 deficiency and fight against various bone related
problems including low bone density, osteoporosis, osteogenesis imperfecta, rickets, osteomalacia, bone and joint pain, bone
fractures, deformed bones, osteoma, chondrodystrophia fetalis, autoimmune and inflammatory diseases, stress management and
prevention, and anti-aging by improving overall health.
Keywords: The Trivedi Effect®, Osteosarcoma Cells (MG-63), Alizarin Red S Staining, ALP, Collagen,
Bone Mineralization, Vitamin D Deficiency, Low Bone Density
3
International Journal of Nutrition and Food Sciences 2018; 7(1): 30-38 31

1. Introduction
(CAM) therapies are now rising as preferred models of
treatment, among which Biofield Therapy (or Healing
Vitamin D has multiple effects, which regulate the
Modalities) is one approach that has been reported to have
functions in different organs viz. brain, liver, lungs, heart,
several benefits to enhance physical, mental and emotional
kidneys, skeletal, immune and reproductive systems.
human wellness. However, as per the data of 2012 from the
Moreover, it has significant anti-inflammatory, anti-aging,
National Health Interview Survey (NHIS), which indicated that
anti-stress, anti-arthritic, anti-osteoporosis, anti-apoptotic,
the highest percentage (17.7%) of the Americans used dietary
wound healing, anti-cancer, anti-psychotic and anti-fibrotic
supplements as a complementary health approach as compared
actions [1]. Vitamin D receptors are widely distributed in
with other practices in past years. The National Center of
most of the body organs viz. brain, liver, heart, lungs, kidney,
Complementary and Integrative Health (NCCIH) has recognized
pancreas, large and small intestines, muscles, reproductive,
and accepted Biofield Energy Healing as a CAM health care
nervous system, etc. Vitamin D receptors influence cell-to-
approach in addition to other therapies, medicines and practices
cell communication, normal cell growth, cell differentiation,
such as natural products, deep breathing, yoga, Tai Chi, Qi
cell cycling and proliferation, hormonal balance,
Gong, chiropractic/osteopathic manipulation, meditation,
neurotransmission process, skin health, immune and
massage, special diets, homeopathy, progressive relaxation,
cardiovascular functions. In any living vertebrates, vitamin D
guided imagery, acupressure, acupuncture, relaxation
plays an important role in maintaining a healthy skeletal
techniques, hypnotherapy, healing touch, movement therapy,
structure and is essential for bone health. Naturally, it is
pilates, rolfing structural integration, mindfulness, Ayurvedic
synthesized in the presence of sunlight in the skin [2]. Most
medicine, traditional Chinese herbs and medicines, naturopathy,
foods do not contain any vitamin D, additionally now-a-days
essential oils, aromatherapy, Reiki, and cranial sacral therapy.
due to aging, use of sunscreen, and change of zenith angle of
Human Biofield Energy has subtle energy that has the capacity
sun the production of vitamin D has reduced [3]. Increasing
3 to work in an effective manner [14]. CAM therapies have been
age is not only related to a decrease in bone marrow
practiced worldwide with reported clinical benefits in different
depression and muscle strength but is also associated with
health disease profiles [15]. This energy can be harnessed and
marked changes in the immune and inflammatory responses
transmitted by the experts into living and non-living things via
[4]. Deficiency of vitamin D causes metabolic bone diseases
3 the process of Biofield Energy Healing. Biofield Energy
like osteomalacia and exacerbate osteoporosis, etc. [5]. The Treatment (The Trivedi Effect®) has been published in numerous
quality of life for menopausal women is one of the most
peer-reviewed science journals with significant outcomes in
critical health problem in the today world. Metabolic bone
many scientific fields such as cancer research [16, 17],
disorders like osteoporosis are mainly prevalent in post-
microbiology [18-21], biotechnology [22, 23], pharmaceutical
menopausal women. Hormonal factors and rapid bone loss in
science [24-27], agricultural science [28-31], materials science
post-menopausal women leads to an increased risk of
[32-35], nutraceuticals [36, 37], skin health, human health and
fractures [6]. Hence, the serum calcium and alkaline
wellness.
phosphatase (ALP) levels in post-menopausal women are the
Based on the literature information and importance of
main two vital biochemical markers of bone metabolism.
vitamin D on bone health, the authors sought to evaluate the
3
However, bone-specific ALP is the most important marker
impact of the Biofield Energy Treatment (The Trivedi
for osteoblast differentiation [7]. Further, it is generally Effect®) on the test samples (vitamin D and DMEM
3
accepted that an increased calcium intake along with an
medium) for bone health activity with respect to the
adequate source of vitamin D is important for maintaining
assessment of different bone health parameters like ALP,
good bone health. Vitamin D also plays an important role in
collagen content, and bone mineralization using standard
maintaining an adequate level of serum calcium and
assays in MG-63 cells.
phosphorus. Therefore, vitamin D has a great impact in
forming and maintaining strong bones [8, 9]. Bone strength
2. Materials and Methods
depends on the quality, geometry, shape, microarchitecture,
turnover, mineral content, and the collagen content. Collagen
2.1. Chemicals and Reagents
is the major structural protein responsible for bone
calcification. In the aging state, the mechanical properties of Fetal bovine serum (FBS) and Dulbecco's Modified
the bones become impaired and the bones get fragile, that Eagle's Medium (DMEM) were purchased from Life
causes various clinical disorders associated with bone Technology, USA. Rutin hydrate was purchased from TCI,
collagen abnormalities and bone fragility, such as Japan, while vitamin D (denoted as test item) and L-ascorbic
3
osteogenesis imperfecta and osteoporosis [10, 11]. acid were obtained from Sigma-Aldrich, USA. Antibiotic
In recent years, several scientific reports and clinical trials solution (penicillin-streptomycin) was procured from
have revealed the useful effects of Biofield Energy Treatments, HiMedia, India, while 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-
which have shown to enhance immune function in cases of diphenyl-2H-tetrazolium) (MTT), Direct Red 80, and
cervical cancer patients via therapeutic touch [12], massage ethylene diamine tetra acetic acid (EDTA) were purchased
therapy [13], etc. Complementary and Alternative Medicine from Sigma, USA. All the other chemicals used in this
32 Aileen Carol Lee et al.: The Potential Benefits of Biofield Energy Treated Vitamin D on Bone Mineralization in
3
Human Bone Osteosarcoma Cells (MG-63)
experiment were analytical grade procured from India. corresponding to 5 X 103 to 10 X 103 cells/well/180 µL of
cell growth medium. The above cells were incubated
2.2. Cell Culture
overnight under growth conditions and allowed cell recovery
and exponential growth, then they were subjected to serum
The human bone osteosarcoma cell line, MG-63, was used
stripping or starvation. The cells were treated with the test
as the test system in the present study. The MG-63 cell line
item, DMEM, and the positive control. The untreated cells
was maintained under the DMEM growth medium for routine
served as baseline control. The cells in the above plate (s)
culture and supplemented with 10% FBS. Growth conditions
were incubated for a time point ranging from 24 to 72 hours
were maintained as 37°C, 5% CO and 95% humidity and
2
in CO incubator at 37°C, 5% CO and 95% humidity.
subcultured by trypsinisation followed by splitting the cell 2 2
Following incubation, the plates were taken out and 20 µL of
suspension into fresh flasks and supplementing with fresh
5 mg/mL of MTT solution was added to all the wells
cell growth medium. Three days before the start of the
followed by an additional incubation for 3 hours at 37°C. The
experiment (i.e., day -3), the growth medium of near-
supernatant was aspirated and 150 µL of DMSO and was
confluent cells was replaced with fresh phenol-free DMEM,
added to each well to dissolve formazan crystals. The
supplemented with 10% charcoal dextran stripped FBS (CD-
absorbance of each well was read at 540 nm using a Synergy
FBS) and 1% penicillin-streptomycin [38].
HT micro plate reader, BioTek, USA. The percentage
2.3. Experimental Design cytotoxicity at each tested concentration of the test substance
was calculated using the following Equation 1:
The experimental groups consisted of cells in baseline
control, vehicle control groups (0.05% DMSO with Biofield % Cytotoxicity = {(1-X)/R}*100 (1)
Energy Treated and untreated DMEM), a positive control
Where, X = Absorbance of treated cells; R = Absorbance
group (rutin hydrate) and experimental test groups.
of untreated cells
Experimental groups included the combination of the
The percentage cell viability corresponding to each
Biofield Energy Treated and untreated vitamin D /DMEM. It
3
treatment was then be obtained using the following Equation
consisted of four major treatment groups on specified cells
2:
with UT-DMEM + UT-Test item, UT-DMEM + Biofield
Energy Treated test item (BT-Test item), BT-DMEM + UT- % Cell Viability = 100 -% Cytotoxicity (2)
Test item, and BT-DMEM + BT-Test item.
The concentrations exhibiting ≥70% Cell viability was
2.4. Consciousness Energy Healing Treatment Strategies
considered as non-cytotoxic [39].
The test item (vitamin D3) and DMEM were divided into 2.6. Assessment of Alkaline Phosphatase (ALP) Activity
two parts. One part each of the test item and DMEM were
treated with the Biofield Energy (also known as The Trivedi The cells were counted using an hemocytometer and plated
Effect®) and coded as the Biofield Energy Treated items, in a 24-well plate at the density corresponding 1 x 104
while the second part did not receive any sort of treatment cells/well in phenol free DMEM supplemented with 10%
and was defined as the untreated samples. This Biofield CD-FBS. Following the respective treatments, the cells in the
Energy Healing Treatment was provided by Aileen Carol above plate were incubated for 48 hours in CO incubator at
2
Lee, who participated in this study and performed the 37°C, 5% CO and 95% humidity. After 48 hours of
2
Biofield Energy Treatment remotely for ~5 minutes. Aileen incubation, the plate was taken out and processed for the
Carol Lee was remotely located in the USA, while the test measurement of ALP enzyme activity. The cells were washed
samples were located in the research laboratory of Dabur with 1X PBS and lysed by freeze thaw method i.e.,
Research Foundation, New Delhi, India. The Biofield Energy incubation at -80°C for 20 minutes followed by incubation at
Treatment was administered for 5 minutes through the 37°C for 10 minutes. To the lysed cells, 50 µL of substrate
healer’s unique Energy Transmission process remotely to the solution i.e., 5 mM of p-nitrophenyl phosphate (pNPP) in 1M
test samples under laboratory conditions. Aileen Carol Lee in diethanolamine and 0.24 mM magnesium chloride (MgCl )
2
this study, never visited the laboratory in person, nor had any solution (pH 10.4) was added to all the wells followed by
contact with the test item and medium. Further, the control incubation for 1 hour at 37°C. The absorbance of the above
group was treated with a sham healer for comparative solution was read at 405 nm using Synergy HT micro plate
purposes. The sham healer did not have any knowledge about reader (Biotek, USA). The absorbance values obtained were
the Biofield Energy Treatment. After that, the Biofield normalized with substrate blank (pNPP solution alone)
Energy Treated and untreated samples were kept in similar absorbance values. The percentage increase in ALP enzyme
sealed conditions for experimental study. activity with respect to the untreated cells (baseline group)
was calculated using Equation 3:
2.5. Determination of Non-Cytotoxic Concentration
% Increase in ALP = {(X-R)/R}*100 (3)
The cell viability test was performed by MTT assay in the
human bone osteosarcoma cell line (MG-63). The cells were Where, X = Absorbance of cells corresponding to positive
counted and plated in 96 well plates at the density control and test groups
International Journal of Nutrition and Food Sciences 2018; 7(1): 30-38 33

R = Absorbance of cells corresponding to baseline group growth. Following overnight incubation, the above cells were
(untreated cells) subjected to serum stripping for 24 hours. The cells were then
treated with non-cytotoxic concentrations of the test samples and
2.7. Assessment of Collagen Synthesis
positive control. Following 3-7 days of incubation with the test
samples and positive control, the plates were taken out, cell
The MG-63 cells were counted using an hemocytometer and
plated in 24-well plate at the density corresponding to 10 x 103 layers processed further by staining with Alizarin Red S dye.
The cells were fixed in 70% ethanol for 1 hour, after which
cells/well in phenol free DMEM supplemented with 10% CD-
Alizarin Red solution (40 µm; pH 4.2) was added to the samples
FBS. Following the respective treatments, the cells in the above
for 20 minutes with shaking. The cells were washed with
plate were incubated for 48 hours in CO incubator at 37°C, 5%
2
distilled water to remove unbound dye. For quantitative analysis
CO and 95% humidity. After 48 hours of incubation, the plate
2
by absorbance evaluation, nodules were solubilized with 10%
was taken out and the amount of collagen accumulated in MG-
cetylpyridinium chloride for 15 minutes with shaking.
63 cells corresponding to each treatment was measured by
Absorbance was measured at 562 nm using Biotek Synergy HT
Direct Sirius red dye binding assay. In brief, the cell layers were
micro plate reader. The percentage increase in bone
washed with PBS and fixed in Bouin’s solution (5% acetic acid,
mineralization with respect to the untreated cells (baseline
9% formaldehyde and 0.9% picric acid) for 1 hour at room
group) was calculated using the following Equation 5:
temperature (RT). After 1 hour of incubation, the above wells
were washed with milliQ water and air dried. The cells were
% Increase = {(X-R)/R}*100 (5)
then stained with Sirius red dye solution for 1 hour at RT
followed by washing in 0.01 N HCl to remove unbound dye. Where, X = Absorbance in cells corresponding to positive
The collagen dye complex obtained in the above step was control or test groups; R = Absorbance in cells corresponding
dissolved in 0.1 N NaOH and absorbance was read at 540 nm to baseline (untreated) group.
using Biotek Synergy HT micro plate reader. The level of
2.9. Statistical Analysis
collagen was extrapolated using standard curve obtained from
purified Calf Collagen Bornstein and Traub Type I (Sigma Type
All the values were represented as percentage of the
III). The percentage increase in collagen level with respect to the
respective parameters. For statistical analysis Sigma-Plot
untreated cells (baseline group) was calculated using Equation 4:
(version 11.0) was used as a statistical tool. Statistically
significant values were set at the level of p≤0.05.
% Increase in collagen levels = {(X-R)/R}*100 (4)
Where, X = Collagen levels in cells corresponding to 3. Results and Discussion
positive control and test groups
R = Collagen levels in cells corresponding to baseline 3.1. MTT Assay
group (untreated cells)
The cell viability data of the Biofield Energy Treated
2.8. Assessment of Bone Mineralization by Alizarin Red S vitamin D and DMEM by MTT assay in MG-63 cells are
3
Staining depicted in Figure 1. The data showed that the test samples in
combination did not exhibit any cytotoxicity (as evidence of
The MG-63 cells were counted using an hemocytometer and
cell viability approximately greater than 73%) across all the
plated in 24-well plate at the density corresponding to 10 x103
tested concentrations up to 100 µg/mL. Hence, the same
cells/well in phenol free DMEM supplemented with 10% CD-
concentrations were assessed further to see the effect of the test
FBS. Following the respective treatments, the cells in the above
samples on the levels of alkaline phosphatase (ALP) activity,
plate were incubated for 48 hours in CO incubator at 37°C, 5%
2 collagen synthesis, and bone mineralization in MG-63 cells.
CO and 95% humidity to allow cell recovery and exponential
2

Figure 1. The cell viability expressed as percentage of the test samples (vitamin D3 and DMEM medium) in different concentrations in MG-63 cells after 72
hours of treatment. VC: Vehicle control (0.05% DMSO); UT: Untreated; BT: Biofield Energy Treated, TI: Test Item.
34 Aileen Carol Lee et al.: The Potential Benefits of Biofield Energy Treated Vitamin D on Bone Mineralization in
3
Human Bone Osteosarcoma Cells (MG-63)

3.2. Alkaline Phosphatase (ALP) Activity was significantly elevated by 194.1%, 72.34%, and 65.31%
in the UT-DMEM + BT-Test item, BT-DMEM + UT-Test
The effect of the Biofield Energy Treated test items on the
item, and BT-DMEM + BT-Test item groups, respectively at
level of alkaline phosphatase (ALP) in human bone
100 µg/mL compared to the UT-DMEM + UT-Test item
osteosarcoma cells is presented in Figure 2. The level of ALP
group. Overall, the Consciousness Energy Healing Treated
was found as 7.5% in the vehicle control (VC) group
(The Trivedi Effect®) test item group (i.e., vitamin D3)
compared to the baseline control group. The ALP activity
showed an improved synthesis of ALP level in the human
was significantly increased in a dose dependent manner by
osteosarcoma cells with respect to the untreated item items
34.31%, 51.47%, and 189.39% in the positive control group
group. For bone formation and bone resorption, various
at the concentration of 0.01, 0.1, and 1 µg/mL, respectively
biomarkers are now available for specific and sensitive
compared to the untreated cells group. The level of ALP was
assessment. For example, the bone formation biomarkers are
significantly increased by 23.38%, 123.37%, and 111.43% in
total alkaline phosphatase (ALP), bone-specific alkaline
the UT-DMEM + BT-Test item, BT-DMEM + UT-Test item,
phosphatase (BALP), osteocalcin (OC), procollagen type 1
and BT-DMEM + BT-Test item groups, respectively at the
N-terminal propeptide (P1NP) and procollagen type 1 C-
concentration of 0.1 µg/mL compared to the UT-DMEM +
terminal propeptide (P1CP) [40, 6]. ALP is a group of cell
UT-Test item group. Moreover, ALP level was significantly
membrane metalloenzymes that catalyze the hydrolysis of
increased by 85.51% and 87.61% in the BT-DMEM + UT-
phosphate esters in an alkaline environment, generating an
Test item and BT-DMEM + BT-Test item groups,
organic radical and inorganic phosphate. It is expressed
respectively at the concentration of 1 µg/mL compared to the
mainly in bone, liver, intestines, proximal convoluted tubules
untreated group. The level of ALP was increased by 2.2% in
of the kidney, and in the placenta and released from these
the BT-DMEM + BT-Test item group, at the concentration of
tissues in the blood [41]. In this experiment, it was also
10 µg/mL compared to the UT-DMEM + UT-Test item
evident that the Biofield Energy Treated vitamin D3
group. Further, the level of ALP was significant increased by
significantly increased the level of ALP expression, which
251.16%, 103.75%, and 119.55% in the UT-DMEM + BT-
might be very advantageous to maintain a healthy skeletal
Test item, BT-DMEM + UT-Test item, and BT-DMEM +
structure for the patients suffering from various bone related
BT-Test item groups, respectively at 50 µg/mL compared to
disorders.
the UT-DMEM + UT-Test item group. Further, the ALP level

Figure 2. The effect of the Biofield Energy Treated test samples on alkaline phosphatase enzyme activity in human bone osteosarcoma cell. VC: Vehicle control
(0.05% DMSO), UT: Untreated; BT: Biofield Energy Treated, TI: Test Item.
3.3. Assessment of Collagen Activity DMEM + BT-Test item group at 1 µg/mL compared to the
UT-DMEM + UT-Test item group. The collagen synthesis
The effect of the test samples on the collagen content in
was significantly increased by 14.12% and 31.79% in the
human bone osteosarcoma cells is shown in Figure 3.
BT-DMEM + UT-Test item and BT-DMEM + BT-Test item
Collagen level in the VC group was found as 13%. The level
groups, respectively at 10 µg/mL compared to the UT-
of collagen synthesis was significantly increased by 63.44%,
DMEM + UT-Test item group. Moreover, the collagen level
63.83%, and 79.45% at 0.01, 0.1, and 1 µg/mL, respectively
was significantly increased by 8.76%, 50.29%, and 52.48%
in the positive control group compared to the untreated cells
in the UT-DMEM + BT-Test item, BT-DMEM + UT-Test
group. The level of collagen was significantly increased by
item, and BT-DMEM + BT-Test item groups, respectively at
83.02%, 145.67%, and 191.41% in the UT-DMEM + BT-
50 µg/mL compared to the UT-DMEM + UT-Test item
Test item, BT-DMEM + UT-Test item, and BT-DMEM +
group. Additionally, at 100 µg/mL the level of collagen was
BT-Test item groups, respectively at 0.1 µg/mL compared to
also significantly increased by 12.88%, 64.51%, and 71.50%
the UT-DMEM + UT-Test item group. Further, collagen
in the UT-DMEM + BT-Test item, BT-DMEM + UT-Test
level was significantly enhanced by 40.40% in the BT-
item, and BT-DMEM + BT-Test item groups, respectively
International Journal of Nutrition and Food Sciences 2018; 7(1): 30-38 35

with respect to the UT-DMEM + UT-Test item group (Figure differentiation [42]. From literature it was reported that the
3). Overall, the Consciousness Energy Healing based test dietary collagen peptides would effectively prevent age-
item group (i.e., vitamin D3) showed an improved synthesis related bone loss like osteogenesis imperfecta and
of collagen content in the human osteosarcoma cells with osteoporosis, there was an abnormality of collagen synthesis,
respect to all the treatment groups. Bones with sufficient stability and maturation [43-47]. Overall, The Trivedi
collagen act as a steel like strong and elastic. Bones that are Effect® - Consciousness Energy Healing Treatment modality
lack of collagen are like dry, brittle wood, easily broken. showed a significant improvement of the collagen level in
Type I collagen is the major structural protein in extra human osteosarcoma cells. Thus, it is assumed that The
cellular matrix responsible for bone calcification. It also Trivedi effect® has the potential to improve the bone health
plays a vital role in further promoting osteoblast in various skeletal disorders.

Figure 3. The effect of the test samples on collagen activity in human bone osteosarcoma cells. VC: Vehicle control (0.05% DMSO), UT: Untreated; BT:
Biofield Energy Treated, TI: Test Item.
3.4. Assessment of Bone Mineralization by Alizarin Red S was distinctly increased by 26.59% in the BT-DMEM + UT-
(ARS) Staining Test item group at 10 µg/mL compared to the UT-DMEM +
UT-Test item group. Further, a noticeably increased percentage
The Alizarin red S (ARS) staining is widely utilized for the
of bone mineralization by 28.33% in the BT-DMEM + UT-Test
assessment of calcium-rich deposits or calcium crystals in the
item group was observed at 50 µg/mL with respect to the UT-
cell culture study. Endocrine factors can strongly influenced to
DMEM + UT-Test item group. In addition, the data showed a
restore the balance between bone formation and resorption.
significant increase of bone mineralization by 64.46% and
Bone formation involves the mineralization of the extracellular
94.16% in the UT-DMEM + BT-Test item and BT-DMEM +
matrix formed by osteoblasts. In this process vitamin D
UT-Test item groups, respectively than the UT-DMEM + UT-
stimulates the bone mineralization of human osteoblasts but is
Test item group (Figure 4) at 100 µg/mL. Thus, based on the
often found inhibitory for mineralization of murine osteoblasts
above findings it is hypothesized that the Consciousness Energy
[48, 49]. The bone mineralization in human bone osteosarcoma Healing Treatment (The Trivedi Effect®) based test item groups
cells is shown in Figure 4. The percentage of bone
(i.e., vitamin D) showed a remarkable improvement of bone
3
mineralization was significantly increased in a concentration
mineralization content assessed by in vitro in the human
dependent manner by 75%, 105.94%, and 135.15% at 5, 10, and
osteosarcoma cells (MG-63) with respect to the all others
25 µg/mL, respectively in the positive control group compared
treatment groups.
to the untreated cells group. The percent of bone mineralization

Figure 4. The effect of the Biofield Energy Treated test samples on human bone osteosarcoma cells for the assessment of bone mineralization activity. VC:
Vehicle control (0.05% DMSO), UT: Untreated; BT: Biofield Energy Treated, TI: Test Item.
36 Aileen Carol Lee et al.: The Potential Benefits of Biofield Energy Treated Vitamin D on Bone Mineralization in
3
Human Bone Osteosarcoma Cells (MG-63)

4. Conclusions
National Center of Complementary and Integrative Health,
DMEM: Dulbecco's modified eagle's medium, FBS: Fetal
The MTT cell viability assay data showed more than 73%
bovine serum, ATCC: American type culture collection, UT:
cells were viable, which indicated that the test samples were
Untreated, BT: Biofield Energy Treated, AGE: Advanced
safe and nontoxic in all the tested concentrations. ALP was
glycation end products.
significantly increased by 251.16%, 103.75%, and 119.55%
in the UT-DMEM + BT-Test item, BT-DMEM + UT-Test
Acknowledgements
item, and BT-DMEM + BT-Test item, respectively at 50
µg/mL, while 123.37% increased in the BT-DMEM + UT-
Authors are grateful to Dabur Research Foundation,
Test item at 0.1 µg/mL compared to the UT-DMEM + UT-
Trivedi Global, Inc., Trivedi Science, Trivedi Testimonials
Test item group. Collagen was significantly increased by
and Trivedi Master Wellness for their support throughout the
83.02%, 145.67%, and 191.41% in the UT-DMEM + BT-
work.
Test item, BT-DMEM + UT-Test item, and BT-DMEM +
BT-Test item group at 0.1 µg/mL, respectively compared to
the untreated group. Further, the percent of bone References
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[3] Matsuoka LY, Ide L, Wortsman J, Mac Laughlin JA, Holick
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