Impact of Biofield Energy Treatment Based Test Formulation on Vital Organ Health Specific Biomarkers Using Cell Line Study

Multiple organ dysfunction syndrome or failure is one of the major concerns against healthcare services in order to maintain the normal function. The present study aimed to explore the impact of the Biofield Energy Treated test formulation on the function of vital organs such as bones, heart, liver, lungs, and brain using standard activity parameters in specific cell-based assays. The test formulation and cells medium was divided into two parts, one untreated (UT) and other part received the Biofield Energy Treatment remotely by a renowned Biofield Energy Healer, Ariadne Esmene Afaganis, Canada and was labeled as the Biofield Treated (BT) test formulation/media. The test formulation was tested for cell viability, and the data suggested that the test formulation was found safe and non-toxic against all the cell lines. Cytoprotective activity among the experimental groups showed a significant improved activity by 94.4% at 1 μg/mL in untreated medium (UT-Med) + Biofield Treated Test Item (BT-TI) group in human cardiac fibroblasts cells (HCF) cells, while 84.4% at 10 μg/mL in BT-Med + BT-TI groups in human hepatoma cells (HepG2), and 124% increased cytoprotective action at 1 μg/mL in UT-Med + BT-TI group in adenocarcinomic human alveolar basal epithelial cells (A549) cells as compared with the untreated test group. ALP activity in MG-63 cells was significantly increased by 85.9% at 10 μg/mL in the UT-Med + BT-TI group, while in Ishikawa cells showed maximum increased ALP activity by 59.2% at 0.1 μg/mL in BT-Med + BT-TI groups as compared to the untreated group. The percent protection of HCF (heart) cells (decreased of LDH activity) was significantly increased by 53% and 40.5% at 1 and 10 μg/mL concentrations respectively, in UT-Med + BT-TI group, while BT-Med + UT-TI group showed increased protection by 68.5%, 70.7%, and 16.8% at 0.1, 1, and 10 μg/mL respectively, and 86.5%, 62.5%, and 34.2% improved cellular protection at 0.1, 1, and 10 μg/mL respectively, in BT-Med + BT-TI group as compared to the untreated test group. The percent protection of HepG2 (liver) cells (decreased of ALT activity) was reported by 33.5%, 63.2%, and 99.2% at 10 μg/mL in the UT-Med + BT-TI, BT-Med + UT-TI, and BT-Med + BT-TI groups, respectively compared to the untreated group. Cellular protection of A549 (lungs) cells (increased of SOD activity) in terms of percentage was increased by increased by 39.8% (at 10 μg/mL), 44% (at 25.5 μg/mL), and 59.7% (at 25.5 μg/mL) in the UT-Med + BT-TI, BT-Med + UT-TI, and BT-Med + BT-TI groups, respectively compared to untreated group. Serotonin level was significantly increased by 59.2% (at 0.1 μg/mL), 190.3% (at 0.1 μg/mL), and 201% (at 1 μg/mL) in the UT-Med + BT-TI, BT-Med + UT-TI, and BT-Med + BT-TI groups, respectively compared to untreated in human neuroblastoma cells (SH-SY5Y). However, the relative quantification (RQ) of vitamin D receptor (VDR) was significantly increased by 159.1% (at 50 μg/mL), 212.7% (at 1 μg/mL), and 278.3% (at 10 μg/mL) in the UT-Med + BT-TI, BT-Med + UT-TI, and BT-Med + BT-TI groups, respectively as compared to the untreated in MG-63 cells. Thus, the present data concluded that the overall multiple organ health using various standard biomarkers in specific cell lines were significantly improved with Freely Available Online www.openaccesspub.org JTRR CC-license DOI: 10.14302/issn.2640-6403.jtrr-19-2946 Vol-1 Issue 2 Pg. no.– 23 Introduction Herbal product based test formulations using vital phytonutrients or nutraceuticals are showing immense importance around the world due to its high acceptance rate against various health care challenges with respect to health of major vital organs such as bones, heart, liver, lungs, and, brain [1]. More than 80% of the world populations (~ 4 billion peoples) rely on herbal product for their respective health challenge in the developing world [2, 3]. Thus, for measuring the biomarkers for overall health, specific cell lines were selected for various activities such as for bone health MG-63 cells (Human Osteoblasts like Cells) were used for estimation of ALP activity (phenotypic marker for the early differentiation and maturation of osteoblasts). The study data suggested that the response of MG-63 cells for 1, 25-dihydroxyvitamin D3 (1,25(OH)2D3) administration has been found to be very similar to the normal human osteoblast cells [4]. MG-63 cell line has been widely reported for studying the potential of any test compounds to improve bone health [5]. For estimation of menstrual pain dysmenorrheal, Ishikawa (Human Endometrial cancer cell line) were used for estimation of ALP activity (represents estrogenic activity). These cells are derived from human endometrium that are found to be very significant in female reproductive functions along with fertility-determining factor [6-8]. Similarly, lung health was determined using estimation of cytoprotection SOD activity in A549 (Human lung adenocarcinoma cell line) [9]. Liver health was determined using HepG2 (Human liver cell line) by estimation of cytoprotection ALT action [10-12]. Heart health was determined using estimation of cytoprotection LDH in Human Cardiac fibroblasts cells [13, 14]. Memory and cognitive health was detected using estimation of serotonin release using SH-SY5Y (Human Neuroblastoma Cells) [15, 16]. Similarly, Vitamin D metabolism was detected using estimation of VDR expression in MG-63 (Human Osteoblasts like Cells), as these are the widely used biomarkers to study VDR expression as an in-vitro model [17, 18]. The scientific data suggested that the death of more than 17 million people worldwide was due to cardiovascular disorders, multiple organ failure, and all are somehow related with the generation of free radicals, which are responsible for the onset of these health issues [19, 20]. Thus, the present study was aimed to detect the overall health status using novel test formulation, which included a combination of 11 ingredients, calcium chloride, panax ginseng extract, vitamin B12, beta carotene, vitamin D3, zinc chloride, magnesium gluconate, sodium selenate, ferrous sulfate, ascorbic acid and vitamin B6. Further, the effect of test formulation was tested on Ishikawa, MG-63, A549, SH-SY5Y, HepG2, and Human Cardiac Fibroblast (HCF) for respective activities after Biofield Energy Healing Treatment. Biofield Energy Healing therapies are accepted worldwide as one of the best Complementary and Alternative Medicine (CAM) concept for altering overall quality of life. Various forms of Energy Healing Therapies have been reported with significant clinical and non-clinical outcomes [21, 22]. Energy Healing Therapies have been practiced and accepted by the U.S. population and is well defined by National Center for Complementary and Alternative Medicine Corresponding author: Snehasis Jana, Trivedi Science Research Laboratory Pvt. Ltd., Thane-West, Maharashtra, India, Email: publication@trivedieffect.com


Introduction
Herbal product based test formulations using vital phytonutrients or nutraceuticals are showing immense importance around the world due to its high acceptance rate against various health care challenges with respect to health of major vital organs such as bones, heart, liver, lungs, and, brain [1]. More than 80% of the world populations (~ 4 billion peoples) rely on herbal product for their respective health challenge in the developing world [2,3]. Thus, for measuring the biomarkers for overall health, specific cell lines were administration has been found to be very similar to the normal human osteoblast cells [4]. MG-63 cell line has been widely reported for studying the potential of any test compounds to improve bone health [5]. For estimation of menstrual pain dysmenorrheal, Ishikawa (Human Endometrial cancer cell line) were used for estimation of ALP activity (represents estrogenic activity). These cells are derived from human endometrium that are found to be very significant in female reproductive functions along with fertility-determining factor [6][7][8]. Similarly, lung health was determined using estimation of cytoprotection SOD activity in A549 (Human lung adenocarcinoma cell line) [9]. Liver health was determined using HepG2 (Human liver cell line) by estimation of cytoprotection ALT action [10][11][12]. Heart health was determined using estimation of cytoprotection LDH in Human Cardiac fibroblasts cells [13,14]. Memory and cognitive health was detected using estimation of serotonin release using SH-SY5Y (Human Neuroblastoma Cells) [15,16].
Similarly, Vitamin D metabolism was detected using estimation of VDR expression in MG-63 (Human Osteoblasts like Cells), as these are the widely used biomarkers to study VDR expression as an in-vitro model [17,18]. The scientific data suggested that the death of more than 17 million people worldwide was due to cardiovascular disorders, multiple organ failure, and all are somehow related with the generation of free radicals, which are responsible for the onset of these health issues [19,20].
Thus, the present study was aimed to detect the overall health status using novel test formulation, which included a combination of 11  Consciousness Energy Healing Treatment found to be significant to improve the metal physicochemical properties [25,26], improved crop yield in agriculture science [27], microbiology [28,29], biotechnology [30,31], improved bioavailability of many compounds [32,33], improved skin health [34,35], improved properties of nutraceuticals [36], cancer science research [37], improved overall bone health [38][39][40], human health and wellness. Due to the continued outcomes and wide applications of Biofield Energy Healing Treatments, the test formulation was studied for impact of the Biofield Energy Healing Treated test formulation on the function of vital organs such as bones, heart, liver, lungs, and brain specific biomarkers in different cell-lines.    [(X-R)/R]*100…………… (7) Where, X = Serotonin levels corresponding to test item or positive control, R = Serotonin levels corresponding to control group.

Effect of Test Formulation on Vitamin D Receptor (VDR) in Bone (MG-63) Cells
The effect of test formulation on vitamin D receptor (VDR) activity in bone (MG-63) cells were counted using the hemocytometer at density 2 X 10 5 cells/well in 6-well plates followed by overnight  (8) Where, N is the relative Threshold Cycle (C T ) value of treated sample with respect to the untreated sample.

Statistical Analysis
All the values were represented as percentage.

Results and Discussion
Cell Viability Using MTT Assay  respectively as compared to the UT-Med + UT-TI group [43]. Oxidative stress has been reported as the major contributor for cellular damage and high incidence has been found with respect to the initiation of many liver

Estimation of Alanine Amino Transferase (ALT) Activity in HepG2 Cells
Alanine amino transferase (ALT) activity of the test formulation was evaluated on the protection of  HepG2 cells in terms of decrease ALT activity is shown in However, it has been reported that various factors such as endogenously or exogenously such as air pollutants, cigarette smoke, etc. were exposed to the lungs causing respiratory illness and other inflammatory disorders.
SOD is one of the important antioxidant defense mechanism and also a powerful anti-inflammatory.
However, their SOD enzyme based supplement significantly reduced the ROS generation, oxidative stress and also inhibits the endothelial activation [55].
The experimental data showed that Biofield Energy Treatment has the significant capacity to improve the

Effect of Test Formulation on Serotonin in Human Neuroblastoma (SH-SY5Y) Cells
The effect of test formulation on serotonin level was assessed in SH-SY5Y cells after 24 hours of treatment by ELISA and the results are presented in  Figure 6). 5-HT has been found in the brain, bowels,