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Showing posts with label The Trivedi Effect®. Show all posts

Wednesday, August 10, 2022

Effect of Biofield Treatment on Blood, Hepatic, and Cardiac Biomarkers in Vitamin D3 Deficiency Diet (VDD) Induced Sprague Dawley Rats - Juniper Publishers

Pharmacy & Pharmaceutical Sciences - Juniper Publishers

Abstract

A novel proprietary formulation was designed that consist of minerals (magnesium, zinc, copper, calcium, selenium, and iron), vitamins (ascorbic acid, pyridoxine HCl, alpha tocopherol, cyanocobalamin, and cholecalciferol), Panax ginseng extract, β-carotene, and cannabidiol isolate. The study objective was to evaluate the impact of Consciousness Energy Healing Treatment (the Trivedi Effect®) on a novel test formulation in male Sprague Dawley (SD) rats, fed with vitamin D3 deficiency diet (VDD) for immunomodulatory activity. The test formulation was divided into two parts. One part was denoted as the untreated test formulation without any Biofield Energy Treatment, while the other part was defined as the Biofield Energy Treated sample, which received the Biofield Energy Healing Treatment by renowned Biofield Energy Healer, Mr. Mahendra Kumar Trivedi. The level of total leukocytes count (TLC) were significantly increased by 23.1% and 17.44% in the Biofield Energy Treated test formulation (G5) and Biofield Energy Treated test formulation from day -15 (G7) groups, respectively compared with the disease control group (G2) induced by VDD. Moreover, the level of neutrophils was significantly increased by 71.68%, 92.92% (p≤0.01), 42.48%, and 49.56% in the G5, G7, Biofield Energy Treatment per se plus Biofield Energy Treated test formulation from day -15 (G8), and Biofield Energy Treatment per se animals plus untreated test formulation (G9) groups, respectively compared with the G2 group. Monocyte level was also increased by 23.53%, 17.65%, and 29.41% in the G5, Biofield Energy Treatment per se to animals from day -15 (G6), and G9 groups, respectively compared to the G2 group. Hepatic biomarker like alkaline phosphatase (ALP) was significantly reduced by 14.98% and 23.07% (p≤0.05) in the G7 and G9 groups, respectively as compared to the G2 group. Besides, cardiac biomarker like creatine kinase myocardium band (CK-MB) was also significantly (p≤0.05) reduced by 20.99% in the G6 group compared with the G2. Altogether, results suggested that the Biofield Treated test formulation and Biofield Energy Treatment per se significantly increased immune-related parameters, which might be beneficial for the management of immune-compromised patients as well as to boost-up the immunity in healthy peoples. The results showed a significant slowdown of disease progression and all other disease-related complications/symptoms in the preventive Biofield Energy Treatment group per se and the Biofield Energy Treated Test formulation groups (viz. G6, G7, G8, and G9) as compared to the disease control group.

Keywords: Biofield treatment; Immunomodulation; The Trivedi effect®; Hematology; Biochemistry; Vitamin D3 deficiency diet; Calcitriol

Abbreviations: CAM: Complementary and Alternative Medicine; IBS: Irritable Bowel Syndrome; QoL: Quality of Life; TLC: Total Leukocytes Count; SGOT: Serum Glutamate Oxaloacetate Transaminase; SGPT: Serum Glutamate Pyruvate Transaminase; ALP: Alkaline Phosphatase; SEM: Standard Error of Mean; ANOVA: One-Way Analysis of Variance; CK-MB: Creatine Kinase Myocardium Band; DLC: Differential Leukocyte Counts; SD: Sprague Dawley; API: Active Pharmaceutical Ingredients

Introduction

Lack of vitamin D3 has been directly linked to various health problems like cognitive decline, osteoporosis, depression, diabetes, cardiovascular disease, hypertension, and cancer [1,2]. It is also very essential for bone health in children and adults. The processes by which intake of vitamin D3 like synthesis through skin via UV-rays and absorption from foods become less efficient with age [3]. That is why, hypovitaminosis of vitamin D3 is more common worldwide [4]. Based on this situation authors constructed the current research work to assess the effect ofBiofield Treatment on hematology and biochemical parameters with special reference to hepatic and cardiac biomarkers after induction of vitamin D3 deficiency diet (VDD) in male Sprague Dawley rats. The novel test formulation, which is a combination of different minerals (selenium, zinc, iron, calcium, copper, and magnesium), vitamins (cyanocobalamin, ascorbic acid, pyridoxine HCl, alpha tocopherol, and cholecalciferol), cannabidiol isolate and Panax ginseng extract. The active pharmaceutical ingredients (API) used in this test formulation already has been extensively applicable as nutraceutical supplement [5-8].

Biofield Therapy is one of the approaches of Complementary and Alternative Medicine (CAM) therapies. At present, it is considering as the first-line model for the management of numerous chronic, life-style oriented and metabolic disorders. According to National Health Interview Survey (NHIS) 2012, reported that most of the Americans used the dietary supplement as complementary health approaches than conventional medicine therapy. Apart from this, The National Center of Complementary and Integrative Health (NCCIH) has recognized and accepted the Biofield Therapy as a CAM health care approach in addition to other therapies, medicines and practices such as Tai Chi, Ayurvedic medicine, Qi Gong, deep breathing, Rolfing structural integration, yoga, chiropractic/osteopathic manipulation, natural products, massage, relaxation techniques, meditation, aromatherapy, progressive relaxation, hypnotherapy, Pilates, acupuncture, mindfulness, healing touch, naturopathy, special diets, homeopathy, acupressure, cranial sacral therapy traditional Chinese herbs and medicines, movement therapy, guided imagery, Reiki, essential oils, and applied prayer (as is common in other religions, like Judaism, Buddhism, Christianity, and Hinduism).

Human Biofield Energy has certain kind energy that can work effectively [9]. CAM therapies have been extensively used all over the world for the benefits aspect in the healthcare system [10]. Biofield Energy can be harnessed and transmitted by individuals into both living and non-living things via the process of unique thought transmission process. Biofield Energy Healing Treatment (the Trivedi Effect®) has been published in numerous peerreviewed science journals with significant outcomes in many scientific fields such as cancer research [11,12], microbiology and biotechnology [13-15], pharmaceutical science [16- 19], agricultural science [20-22], materials science [23-25], nutraceuticals [26,27], skin health [28,29], human health and wellness. In this context, authors have planned to investigate the impact of the Consciousness Energy Healing Treatment (the Trivedi Effect®) on the test formulation for immunomodulatory activity concerning hematology and biochemical parameters.

Materials and Methods

Chemicals and reagents

Pyridoxine hydrochloride (vitamin B6), calcitriol, zinc chloride, magnesium (II) gluconate, and β-carotene (retinol, Provit A) were purchased from TCI, Japan. Copper chloride, cyanocobalamin (vitamin B12), calcium chloride, vitamin E (Alpha-Tocopherol), cholecalciferol (vitamin D3), iron (II) sulfate, and sodium carboxymethyl cellulose (Na-CMC) were procured from Sigma- Aldrich, USA. Ascorbic acid (vitamin C) and sodium selenate were obtained from Alfa Aesar, India. Cannabidiol isolate and Panax ginseng extract were obtained from Panacea Phytoextracts, India and Standard Hemp Company, USA, respectively.

Maintenance of animal

Randomly breed male Sprague Dawley (SD) rats with body weight ranges from 200 to 300 gm were used in this study. The animals were purchased from M/s. Vivo Bio Tech, Hyderabad, India. Animals were randomly divided into nine groups based on their body weights consist of 6 animals of each group. They were kept individually in sterilized polypropylene cages with stainless steel top grill having provision for holding pellet feed and drinking water bottle fitted with stainless steel sipper tube. The animals were maintained as per standard protocol throughout the experiment.

Consciousness energy healing strategies

The test formulation was distributed into two parts. One part of each ingredient was considered as the untreated test formulation, where no Biofield Energy Treatment was provided. Another part of each ingredient was received Biofield Energy Treatment by Mr. Mahendra Kumar Trivedi (the Trivedi Effect®) for ~3 minutes under laboratory conditions. Besides, three group of animals were also received Biofield Energy Treatment under laboratory conditions for ~3 minutes. The blessing/treatment was given to the test items by his physical presence without touching in the laboratory of Dabur Research Foundation, near New Delhi, India. Similar way, the control samples were also treated by a “sham” healer for ~3 minutes under the same laboratory conditions. The “sham” healer did not know about the Biofield Treatment. After that, the Biofield Energy Treated and untreated test formulations were kept in the similar sealed condition and used as per the study plan. The Biofield Energy Treated animals were taken back to experimental room for further proceedings.

Experimental procedure

Seven days after acclimatization, animals were randomized and grouped based on the body weight. Dosing for groups G7 and G8 were initiated on day -15 and continued till end of the experiment. However, G1 to G6 and G9 groups were dosed from day 1 till the end of experiment. All the animals except G1 group received vitamin D3 deficient diet (VDD) daily to the end of the experiment. Three weeks after the initiation of induction of VDD, all the groups were dosed with respective formulations. At the end of 8th weeks after bleeding, blood from all the animals was collected from the retro-orbital plexus using capillary tube for hematology analysis. From one portion of blood, serum was isolated for the analysis of biochemical parameters. The selective vital organs were collected and weighed.

Assessment of hematology parameters

Hematological parameters such as total leukocyte count (TLC), and differential leukocyte counts (DLC) were analyzed using Hematology analyzer (Abbott Model-CD-3700) in blood samples [30]

Assessment of cardiac and hepatic enzymes

Creatine kinase myocardium band (CK-MB), alkaline phosphatase (ALP), serum glutamic oxaloacetic transaminase (SGOT), and serum glutamate-pyruvate transaminase (SGPT) were analyzed using serum by Biochemistry Analyzer, Spectra lab A– plus, Italy [31,32].

Sample preparation for commercial products

The samples of excipients like Avicel, Microcrystalline cellulose, Sodium CMC and pharmaceutical dosage forms like Ophthalmic suspension and Tablet are prepared as per the procedure mentioned under sample preparation for nasal spray suspension for the determination of Avicel content.

Determination of body weight, feed Intake, and organ weight parameters

All the experimental animals were daily analyzed for their change in body weight, feed intake, and organ weight parameters, which was calculated by weighing the daily feed supply and the left-over amount that evaluate the average daily feed intake. The average intake of feed was recorded in every three days interval throughout the experimental period. After terminal bleeding, the animals were sacrificed and the following organs such as liver, lungs, kidney, brain, heart, eyes, pancreas, spleen, thymus, adrenal gland, intestine, and reproductive organs, i.e., testis, prostate, epididymis, and vas deferens were collected. These organs were trimmed off any adherent tissue and fat, as appropriate and weighed. The organ to body weight ratio percentage was identified by comparing the weight of each organ with the final body weight of individual rat [33]. All the data were reported through the study treatment regimen. Relative organ weight was calculated as per Equation 1.

Clinical sign and symptoms

All the animals in different test groups were analyzed for various clinical sign and symptoms in accordance with in-house protocol. Abnormal behavior in animals was recorded with the time of onset and disappearance.

Statistical analysis

The data were represented as mean ± standard error of mean (SEM) and subjected to statistical analysis using Sigma-Plot statistical software (Version 11.0). For multiple comparison Oneway analysis of variance (ANOVA) followed by post-hoc analysis by Dunnett’s test and for between two groups comparison Student’s t-test was performed. The p≤0.05 was considered as statistically significant.

Results and Discussion

Hematology parameters

The experimental data of hematology parameters in various groups (G1 to G9) are summarized in Table 1. The results suggested an improved animal hematology profile as compared with the disease control group (G2). The tested hematology parameters like total leukocytes count (TLC) were significantly increased by 39.53% (p≤0.01), 16.67%, 23.1%, and 17.44% in the G3, G4, G5, and G7 groups, respectively as compared with the disease control group (G2) induced by vitamin D3 deficient diet. Similarly, the level of neutrophils was significantly increased by 110.62% (p ≤ 0.01), 86.73% (p ≤ 0.01), 71.68%, 92.92% (p ≤ 0.01), 42.48%, and 49.56% in the G3, G4, G5, G7, G8, and G9 groups, respectively as compared with the G2 group. Moreover, the level of lymphocytes was increased by 29.42% (p≤0.01), 1.53%, 15.14%, and 5.78% in the G3, G4, G5, and G7 groups as compared with the G2 group. Further, the level of eosinophils was significantly increased by 41.18%, 23.53%, 17.65%, and 29.41% in the G4, G5, G6, and G9 groups, respectively as compared with the G2 group. Monocyte level was also increased by 41.18%, 23.53%, 17.65%, and 29.41% in the G4, G5, G6, and G9 groups, respectively compared to the G2 group.

Altogether, study data suggested that the Biofield Treatment has the significant capacity to improve the blood immunity-related parameters. The altered hematology profile might be used in many acute infection, gout, rheumatoid arthritis, chronic inflammatory diseases, rheumatic fever, etc. However, minerals and vitamins play a vital role to control the hematology profile [34-36]. Increase the level of TLC and neutrophils count directly supports to the improvement of cell mediated immunity [37]. The study data concluded that the Biofield Energy Treated (the Trivedi Effect®) test formulation significantly improved the concentrations of TLC, lymphocytes, neutrophils, and monocytes in hematology profile assay, which suggest immunomodulatory potential of the test formulation with respect to altered hematological animal profile.

G: Group; G1: Normal control (0.5% CMC); G2: Disease control (VDD: Vitamin D3 deficient diet + 0.5% CMC); G3: Reference item (VDD + Calcitriol); G4: (VDD + Untreated test formulation); G5: (VDD + Biofield Energy Treated test formulation); G6: (VDD + Biofield Energy Treatment per se to animals from day -15; G7: (VDD + Biofield Energy Treated test formulation from day -15); G8: (VDD + Biofield Energy Treatment per se plus Biofield Energy Treated test formulation from day -15), and G9: (VDD + Biofield Energy Treatment per se animals plus untreated test formulation). Values are presented as mean ± SEM (n=6). **p≤0.01 vs. G2.

Measurement of hepatic and cardiac biomarkers

Hepatic and cardiac biochemical markers were tested for the test formulation and the results are tabulated in Table 2. The parameters used are serum glutamate oxaloacetate transaminase (SGOT), serum glutamate pyruvate transaminase (SGPT), alkaline phosphatase (ALP) and cardiac enzyme creatine kinase myocardium band (CK-MB), and other biomarkers such as, total bilirubin, albumin, and globulin of different groups (G1 to G9) are summarized and compared with their respective controls. The level of ALP was also found to be significantly reduced by 35.41% (p≤0.05), 22.9%, 7.72%, 7.89%, 14.98%, and 23.07% (p≤0.05) in the G3, G4, G5, G6, G7, and G9 groups, respectively compared to the G2 group. Further, CK-MB was significantly reduced by 6.18%, 13.99%, and 20.99% (p≤0.05) in the G3, G4, and G6 groups, respectively compared with the G2.

Liver toxicity was measured by the hepatic biomarkers, and any high alteration in these enzymes results in infection and liver damage [38]. The study data suggested that the Biofield Treated test formulation showed an improved liver and cardiac health as reflected by many improved levels of hepatic enzymes. Therefore, it can be concluded that the Trivedi Effect®-Biofield Energy Healing significantly improved the liver health and its immunity profile.

Estimation of animal weight parameters, feed intake, and relative organ weight

The test formulation in whole experimental study was calculated and defined with respect to weight parameters, feed intake, and relative organ weight. The results of animal tested organ weight parameters are summarized in the Table 3. The change in animal weights is reported as per the normal physiological process. Thus, the relative organ weight parameters did not show any significant change in the tested organ weight throughout the experiment suggested that the test formulation was found to be safe for the treatment. Organ to body weight ratio is the valuable index for any experimental test procedure with respect to the documentation of swelling, atrophy, or hypertrophy after exposure of test samples. Overall, the animal weight data, relative organ weight, and feed intake data suggested no significant abnormal change than the disease control group (G2). It suggests that Biofield Energy Treated test formulation and Biofield Energy Treatment per se were found as safe in all the tested animals.

this research plan, four groups were considered as preventive maintenance groups. These groups were G6 (Biofield Energy Treatment per se to animals at -15 days), G7 (Biofield Energy Treated test formulation from day -15), G8 (Biofield Energy Treatment per se to animals along with Biofield Treated test formulation from day -15), and G9 (Biofield treatment per se at -15 days to animals with untreated test formulation). Based on the overall data, it suggests that the Biofield Energy Healing Therapy was found to be most effective and beneficial to prevent and protect from the occurrence of any type of disease in the rat model. The data indicated that this therapy could act as a preventive maintenance therapy to prevent the occurrence of disease, slowdown the disease progression when disease-related complications are present which will ultimately improve the overall health and quality of life.

Conclusion

Blood profile data showed that the total leukocytes count (TLC) was significantly increased by 39.53% (p≤0.01) and 23.1% in the G3 and G5 groups, respectively compared to the disease control group (G2) induced by vitamin D3 deficient diet. Additionally, the level of neutrophils was increased by 110.62%, 86.73%, 71.68%, 92.92%, 42.48%, and 49.56% in the G3, G4, G5, G7, G8, and G9 groups, respectively as compared to the G2. Monocyte level was also increased by 41.18%, 23.53%, 17.65%, and 29.41% in the G4, G5, G6, and G9 groups, respectively compared to the G2 group. Besides, ALP level was reduced by 35.41%, 22.9%, 14.98%, and 23.07% in the G3, G4, G7, and G9 groups, respectively compared to the G2 group. Further, creatine kinase myocardium band (CKMB) was also reduced by 13.99% and 20.99% in the G4 and G6 groups, respectively compared with the G2. An experimental weight parameter such as body weight, organ weight, and feed intake data suggested normal changes, which suggest no toxicity profile of the test formulation. Thus, the present experiment suggested that the Trivedi Effect®-Biofield Energy Healing based novel test formulation have some impact on the immune system. The Biofield Energy Healing Treatment (the Trivedi Effect®) per se showed the best results with respect to different beneficial efficacy and biomarker parameters in the preventive maintenance group, G6, as compared to the other preventive maintenance groups (G7, G8, and G9) in the rat model study.

The Biofield Energy Healing Treatment also helped to slowdown the disease progression and disease-related complications impacting the overall animals’ health. These data suggested that Biofield Energy Treatment per se and Biofield Energy Treated Test formulation in combination would be the best treatment strategy to prevent and protect from the occurrence of any type of disease. Therefore, the Biofield Energy Healing Treatment (the Trivedi Effect®) per se might be an effective in healthy humans, when used as a preventive maintenance therapy to sustain good health, to boost overall health, promote healthy aging and increase quality of life.

In the presence of disease, the Biofield Energy therapy might reduce the severity of any acute/chronic disease (such as autoimmune related and inflammatory disorders) and / or slow the disease progression. Therefore, the Biofield Energy Treated test formulation and Biofield Energy Treatment per se may act as an effective anti-inflammatory and immunomodulatory product, and it can be used as a Complementary and Alternative Medicine (CAM) with a safe therapeutic index for various autoimmune disorders such as Systemic Lupus Erythematosus, Fibromyalgia, Addison Disease, Hashimoto Thyroiditis, Celiac Disease (glutensensitive enteropathy), Myasthenia Gravis, Pernicious Anemia, Aplastic Anemia, Dermatomyositis, Multiple Sclerosis, Chronic Fatigue Syndrome Graves’ Disease, Scleroderma, Psoriasis, Sjogren Syndrome, Crohn’s Disease, Vasculitis, Rheumatoid Arthritis, Reactive Arthritis, Type 1 Diabetes`, Vitiligo, and Alopecia Areata, as well as inflammatory disorders such as Irritable Bowel Syndrome (IBS), Asthma, Ulcerative Colitis, Alzheimer’s Disease, Parkinson’s Disease, Atherosclerosis, Dermatitis, Hepatitis, and Diverticulitis. Further, the Biofield Energy Healing Treated test formulation can also be used in the prevention of immunemediated tissue damage and can be used as a stress prevention and management which include overall health and improved Quality of Life (QoL).

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Monday, April 18, 2022

In vitro Assessment of Biofield Energy Treated DMEM on Thermogenesis Using Myoblasts Cell Line (C2C12) - Juniper Publishers

Cell Science & Molecular Biology - Juniper Publishers

Abstract

Mitochondrial dysfunction lead to various serious disorders, which are considered as one of the important components related with the aging, such as type-2 diabetes and Alzheimer’s disease. The aim of the present study was to examine the effect of Consciousness Energy Healing based DMEM medium on murine myoblasts (C2C12) cells to evaluate the mitochondrial mass content using 10-N-nonyl acridine orange (NAO) dye assay. The test item (DMEM medium) was divided into two parts, one part received Consciousness Energy Healing Treatment by a renowned Biofield Energy Healer, Dahryn Trivedi and was labeled as the Biofield Energy Treated DMEM group, while the other part did not receive any kind of Treatment and denoted as the untreated DMEM group. The level of mitochondrial mass content was assessed using 10-N-nonyl acridine orange (NAO) dye method. Cell viability of the test items using MTT assay showed 72.32% and 125.32% viable cells in the untreated DMEM and Biofield Energy Treated DMEM groups, respectively suggested a safe and nontoxic profile of the test items. Besides, the mitochondrial mass content in terms of Fluorescence Unit (FU) was significantly (p≤0.05) increased by 81.78% in the Biofield Energy Treated DMEM group compared to the untreated DMEM group. Overall, the experimental data suggested that the Consciousness Energy Healing Based DMEM showed a significant improvement of mitochondrial mass content and results in better thermogenesis with respect to naive DMEM. Thus, an increased level of NAO dye accumulation in muscle cells indicated increased mitochondrial mass content and hence, better thermogenesis. In the present study, results demonstrated that an increased mitochondrial mass content in the cells when treated with The Trivedi Effect®. This indicates that the test sample has the potential to improve thermogenesis, which can be used against various metabolic diseases, such as insulin resistance, type-2 diabetes, and cardiovascular diseases, etc.

Keywords: Biofield energy; The Trivedi Effect®; Thermogenesis; Mitochondrial biogenesis; Metabolic disorders; Murine myoblast cell; DMEM

Abbreviations: CAM: Complementary and Alternative Medicine, NCCAM: National Center for Complementary and Alternative Medicine; DMEM: Dulbecco’s Modified Eagle’s Medium; FBS: Fetal Bovine Serum

Introduction

Mitochondria (also known as power generator of the cell) produce most of the vital energy required for the cellular function through oxidative phosphorylation involved in electron transport and ATP synthesis. They produce ATP through the process of cellular respiration mainly aerobic respiration, which requires oxygen. Number and amount of mitochondria in a cell be governed by the energy requirement of the cell [1]. For example, the muscle cells have found comparatively more number of mitochondria since, they need to produce energy to move the body. On the other hand, red blood cells carry oxygen to other cells, do not need to produce energy as compared with the muscle cells. Mitochondria are the powerhouse in the cell, which produce energy from basic components. They undergo fusion, fission, transport, and degradation, all of the process is vital to maintain a healthy mitochondrial population [1]. However, the mitochondrial biogenesis process is involved an increased and controlled mitochondrial mass with number that helped to produce greater production of ATP as a response to greater energy expenditure [2]. Physiologic, metabolic, and pathologic changes along with morphological and functional adaptability are the vital factor to regulate the process of mitochondrial biogenesis. In addition, proteins and transcription factors, upstream regulatory proteins and secondary mechanisms are also involved in the biogenesis process, which also stabilizes the new mitochondrial DNA [3].

Mitochondrial biogenesis regulates and control various therapeutic interference in wide number of diseases such as metabolic syndrome, neurodegenerative disorders, sarcopenia, cardiac pathophysiology and physiological processes like aging [4]. Nonyl-acridine orange (NAO) is a non-fluorescent dye that converts into fluorescent dye in the presence of oxidative species [5]. NAO assay is one of the gold standard assays to detect the mitochondrial mass alteration, which is a metachromatic dye that binds to cardiolipin, an inner mitochondrial membrane lipid, regardless of the energetic state of the cell. Therefore, mitochondrial mass of the cells could be estimated by studying accumulation of the fluorescent dye in the mitochondria. Furthermore, an alternative therapies such as nuclear gene was reported to regulate total mitochondrial mass in response to mitochondrial dysfunction [6]. In order to improve the mitochondrial mass content via thermogenesis process, some alternative treatment approach without any associated side-effect is needed.

Biofield Energy Healing is a categorized as one of the Complementary and Alternative Medicine (CAM) accepted worldwide for the various treatment. Biofield Energy Therapy was accepted by National Center for Complementary and Alternative Medicine (NCCAM). Biofield Energy Healing is one of the emerging frontier in medicine, which has been increased in order to promote wellness by uncovering the root cause of diseases with universal solutions. CAM therapies have shown various significant clinical benefits. Over the past few decades, many energy healing practices have been reported a significant outcomes in various clinical and non-clinical fields. The effects of the CAM therapies have great potential, which include external qigong, Johrei, Reiki, therapeutic touch, yoga, Qi Gong, polarity therapy, Tai Chi, pranic healing, deep breathing, chiropractic/osteopathic manipulation, guided imagery, meditation, massage, homeopathy, hypnotherapy, progressive relaxation, acupressure, acupuncture, special diets, relaxation techniques, Rolfing structural integration, healing touch, movement therapy, pilates, mindfulness, Ayurvedic medicine, traditional Chinese herbs and medicines in biological systems both In vitro and in vivo [7]. Every living organisms possess some kind of unique energy known as Biofield Energy, which is infinite, para-dimensional and electromagnetic field surrounding the human body. Biofield (Putative Energy Fields) based Energy Healing Therapies have been reported to have significant outcomes against various disease conditions. Biofield Energy Healing Treatment (The Trivedi Effect®) contain a putative bioenergy, which is channeled by a renowned practitioner from a distance. Biofield Energy Healing as a CAM showed a significant results in biological studies [8]. However, the National Center for Complementary and Alternative Medicine (NCCAM), well-defined Biofield Therapies in the subcategory of Energy Therapies [9]. The Trivedi Effect®- Consciousness Energy Healing Treatment has been reported with significant revolution in the physicochemical properties of metals, chemicals, ceramics and polymers [10- 12], improved agricultural crop yield, productivity, and quality [13,14], transformed antimicrobial characteristics [15-17], biotechnology [18,19], improved bioavailability [20-22], skin health [23, 24], nutraceuticals [25,26], cancer research [27,28], bone health [29-31], human health and wellness. On the basis of outstanding benefits of Biofield Energy Treatment, the present study was aimed to evaluate the impact of the Biofield Energy Treatment (The Trivedi Effect®) on DMEM as test sample to alter the mitochondrial mass content on thermogenesis using NAO dye staining using standard in vitro assay in murine myoblasts (C2C12) cells.

Material and Methods

Chemicals and reagents

Fetal bovine serum (FBS) and Dulbecco’s Modified Eagle’s Medium (DMEM) were purchased from Life Technology, USA. Antibiotics solution (penicillin-streptomycin) were procured from HiMedia, India, and ethylenediaminetetraacetic acid (EDTA) was purchased from Sigma, USA. All the other chemicals used in this experiment were analytical grade procured from India.

Cell culture

C2C12 (murine myoblasts) was used as a test system in the present study. The C2C12 cell line was maintained in DMEM growth medium for routine culture supplemented with 10% FBS. Growth conditions were maintained at 37°C, 5% CO2, and 95% humidity and subcultured by trypsinisation followed by splitting the cell suspension into fresh flasks and supplementing with fresh cell growth medium. Before initiation of the experiment, cells were incubated in DMEM+2% horse serum (HS) for 3 days to allow the cells to differentiate into myotubes.

Experimental design

The experimental groups consisted of group 1 (G-I) with untreated DMEM and group 2 (G-II) included the Biofield Energy Treated DMEM.

Consciousness energy healing treatment strategies

The test item, DMEM was divided into two parts. One part of the test item was treated with the Biofield Energy by a renowned Biofield Energy Healer, Dahryn Trivedi remotely for ~5 minutes and coded as the Biofield Energy Treated DMEM, while the second part did not receive any sort of treatment and denoted as the untreated DMEM group. Biofield Energy Healer was located in the USA, while the test items were located in the research laboratory of Dabur Research Foundation, New Delhi, India. This Biofield Energy Treatment was administered through Healer’s unique Energy Transmission process to the test sample under laboratory conditions. Dahryn Trivedi in this study never visited the laboratory in person, nor had any contact with the test item (DMEM). Further, the untreated DMEM group was treated with a “sham” healer for comparative purposes. The “sham” healer did not have any knowledge about the Biofield Energy Treatment. After that, the Biofield Energy Treated and untreated samples were kept in similar sealed conditions for experimental study.

Assessment of cell viability using MTT assay

The cell viability was performed by MTT assay in C2C12 cell line. The cells were counted and plated in 96-well plates at the density corresponding to 10 X 10³ cells/well/180μL of cell growth medium (DMEM+2% HS). The above cells were incubated overnight under growth conditions and allowed the cell recovery and exponential growth, which were subjected to serum stripping or starvation. The cells were treated with the Untreated and Biofield Energy Treated test item. The cells in the above plate(s) were incubated for 24 to 72 hours in a CO₂ incubator at 37°C, 5% CO₂, and 95% humidity. Following incubation, the plates were taken out and 20μL of 5 mg/mL of MTT solution were added to all the wells followed by additional incubation for 3 hours at 37°C. The supernatant was aspirated and 150μL of DMSO was added to each well to dissolve formazan crystals. The absorbance of each well was read at 540nm using Synergy HT microplate reader, Bio Tek, USA [29]. The percentage cytotoxicity at each tested concentrations of the test items were calculated using the following equation (1):

%cytotoxicity = (1− X / R)*100...........(1)

Where, X = Absorbance of the Biofield Treated cells; R = Absorbance of untreated cells

The percentage cell viability corresponding to each treatment was obtained using the following equation (2):

% Cell Viability = (100- % Cytotoxicity).........(2)

The concentrations exhibiting ≥70% cell viability was considered as non-cytotoxic.

Assessment of mitochondrial content

For the assessment of mitochondrial mass, the cells were counted using an hemocytometer and plated at 4500 cells/well in dark walled 96-well plates in DMEM supplemented with 2% HS. The cells were incubated overnight under standard growth conditions to allow the cell recovery and exponential growth, which were treated by the test items in different groups followed by incubation with the test items for 72 hours. After incubation with the test items, mitochondrial content was determined by 10-N-nonyl acridine orange (NAO) dye. 50nM dye was added to each well and the cells were incubated for 30 minutes at 37°C and 5%CO2. After 30 minutes of incubation, media was discarded and cells were washed with phosphate buffer saline (PBS). 150μL of PBS was added to each well and fluorescence was read at 485/20 excitation, 528/20 emission filter using synergy HT microplate reader. The percentage increase in mitochondrial content was calculated using following equation

Where, FU denotes Fluorescence unit

Statistical analysis

All the values were represented as Mean ± SEM (standard error of mean) of three independent experiments. The statistical analysis was performed using Sigma Plot statistical software (v11.0). For two groups comparison student’s t-test was used. Statistically significant values were set at the level of p≤0.05.

Results and Discussion

Cell viability using MTT assay

Cell viability data of the untreated and Biofield Energy Treated DMEM groups in C2C12 cells using MTT assay is shown in Figure 1. The percentage of cell viability in the untreated DMEM group was 72.32%, while it was 125.32% in the Biofield Energy Treated DMEM group (Figure 1). Overall, data suggest that all the test samples were found safe against the tested C2C12 cells, which were used for the estimation of mitochondrial mass content, which indicate extend of thermogenesis.

Extend of mitochondrial mass content in C2C12 cells

Mitochondrial mass content or biogenesis showed a significant effects against various metabolic diseases. The increased cell capacity to control and maintain the cell metabolism, signal transduction, and regulation of mitochondrial ROS production [2]. Alteration or decrease in the mitochondrial biogenesis are related with the mitochondrial dysfunction and mitochondrial oxidative stress, which leads to various diseases [32]. Mitochondrial mass content results in the improved production of ATP as a response to greater energy expenditure [33]. Various factors such as physical exercise, nutritional factors, etc. reported to have an improved mitochondrial mass content, which results in greater glucose uptake by muscles, along with an increased metabolic enzymes level for glycolysis, oxidative phosphorylation and ultimately a greater mitochondrial metabolic capacity [34]. Aging process leads to decrease level of mitochondrial mass content and results in various diseases such as enhanced aging, insulin resistance, type-2 diabetes, cardiovascular diseases, obesity, etc. [35]. The experiment was conducted to study the influence of Biofield Energy Healing Treatment on mitochondrial content in C2C12 cells using NAO dye assay. The results of mitochondrial mass content in terms of increase number of fluorescence unit (FU) among different groups in C2C12 cells using NAO dye assay are presented in Figure 2. The untreated DMEM group showed 272.3 ± 12.14 FU. Besides, the Biofield Energy Treated DMEM group showed 81.78% increase the level of mitochondrial mass in terms of FU, compared to the untreated DMEM group (Figure 2).

Thus, the data suggested that the Consciousness Energy Blessed DMEM showed a significant improvement of thermogenesis, which results in mitochondrial mass content. This phenomenon can be significantly used against various metabolic diseases, such as insulin resistance, type 2 diabetes, and cardiovascular diseases. Overall, the Biofield Energy Healing Treatment (The Trivedi Effect®) has the significant capacity to improve the overall Quality of life with an improved thermogenesis and mitochondrial content.

Conclusion

MTT data showed a significant cell viability with ranges from 72.32% to 125.32% in different test item groups, which indicated that the test items were safe and non-toxic in nature. Correspondingly, The Trivedi Effect® showed a significant change in mitochondrial mass among different groups. The Biofield Energy Treated DMEM group demonstrated significant increase in the mitochondrial mass by 81.78% as compared to the untreated DMEM group. Thus, Biofield Energy Healing based DMEM can be significantly used to improve the energy level, endurance, body energy, which can be utilized against many diseases such aging, diabetes, cancer, depression, hypertension, cardiovascular disease, aging, and physical strength. The Biofield Energy Treated (The Trivedi Effect®) DMEM were found to have a significant impact on thermogenesis, which might significantly improve the mitochondrial content in muscle cells. Therefore, the Consciousness Energy Healing based DMEM might be a suitable alternative media for cell growth. It can be useful for the management of various disorders such as Lupus, Systemic Lupus Erythematosus, Fibromyalgia, Addison Disease, Hashimoto Thyroiditis, Celiac Disease (gluten-sensitive enteropathy), Multiple Sclerosis, Dermatomyositis, Graves’ Disease, Myasthenia Gravis, Pernicious Anemia, Aplastic Anemia, Scleroderma, Psoriasis, Rheumatoid Arthritis, Reactive Arthritis, Diabetes, Sjogren Syndrome, Crohn’s Disease, Vasculitis, Vitiligo, Chronic Fatigue Syndrome and Alopecia Areata, as well as inflammatory disorders such as Irritable Bowel Syndrome (IBS), Asthma, Ulcerative Colitis, Alzheimer’s Disease, Parkinson’s Disease, Atherosclerosis, Dermatitis, Hepatitis, and Diverticulitis. Further, the Biofield Energy Healing Treatment can also be used in the prevention of immune-mediated tissue damage in cases of organ transplants (for example heart transplants, kidney transplants and liver transplants), for anti-aging, stress prevention and management, and in the improvement of overall health and Quality of Life.

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Tuesday, November 16, 2021

Evaluation of Biofield Energy Healing Treatment Based Proprietary Test Formulation on Gut Health Potential in Colon Cancer Cell Line (HT-29) - Juniper Publishers

Pharmacology & Clinical Research - Juniper Publishers

Abstract

The present study was aimed to evaluate the anti-inflammatory potential of Biofield Energy Healing (the Trivedi Effect®-Consciousness Energy Healing) on the test formulation in colon cancer cell line (HT-29). Each ingredient of the test formulation was divided into two parts, one part was denoted as the untreated test formulation and the other part was demarcated as the Biofield Energy Treated test formulation, which received Biofield Energy Healing Treatment by a renowned Biofield Energy Healer, Mr. Mahendra Kumar Trivedi. MTT assay showed that the test formulation was found safe and non-toxic upto 122 μg/mL in HT-29 cells with more than 78% cell viability. The level of interleukin-6 (IL-6) expression was significantly reduced by 34.09% and 59.41% (p≤0.001) at 3 and 15 μM, respectively compared to the vehicle control (VC) group under the stimulation of tumor necrosis factor - alpha (TNF-α). Moreover, IL-8 level was significantly suppressed in the Biofield Energy Treated test formulation by 57.09% and 42.88% at 0.1 and 3 μM, respectively compared with the VC group. However, the Biofield Energy Treated test formulation further substantial altered the level of interferon gamma compared to the VC group. The Trivedi Effect®-Consciousness Energy Healing significantly regulate the inflammatory condition after treatment with the test formulation in colon cancer cell line (HT-29). This experimental data suggested that the Biofield Treated test formulation can be utilized for many inflammatory disease conditions such as rheumatoid arthritis, multiple sclerosis, psoriasis, inflammatory bowel diseases, scleroderma, and type 1 diabetes mellitus.

Keywords: Biofield Energy Healing; The Trivedi Effect®; Inflammation; Colon cancer cell line (HT-29); Pro-inflammatory cytokines

Abbreviations: FBS: Fetal bovine serum; MTT: 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide; ELISA: Enzyme-linked immunosorbent assay; NCCAM: National Center for Complementary and Alternative Medicine; CAM: Complementary and Alternative Medicine

Introduction

The relation among of the gut health, micro biota, and cytokines have been well studies and reported in past two decades against various inflammatory bowel diseases (IBD) and associated mucosal inflammations. The cytokines play an important role in the mechanism in inflammation especially in Crohn’s disease and ulcerative colitis. The pathogenesis of IBD is not completely understood, but the role of cytokines in the intestinal immune system has significant impact in disruption of normal state of controlled gut inflammations [1,2]. Innate immune response is the major response in gut inflammation and its related diseases. Most of the cytokines are secreted by activated dendritic cells and the macrophages, which regulates the inflammatory response in gut inflammatory diseases. Once, these cytokines are secreted by antigen presenting cells, they triggers and differentiate various T cells by activating adaptive immune response. Gut inflammation dysregulates the T-cells, and manage the over-reactive and auto-reactive cells. T-cell regulation or its overproduction leads to the development of gut inflammatory diseases [3]. These cells along with various types of cytokines play a complex role in inflammatory gut diseases [4, 5]. Thus, there is the need of some novel formulation which alters the level of cytokines to improve the gut health. The present study was aimed to test the impact of the Biofield Energy Treated test formulation comprised of zinc chloride, ferrous sulphate, copper chloride (II-cupric), vitamin B6 (pyridoxine HCl), vitamin B12 (cyanocobalamin), magnesium (II) gluconate, and cholecalciferol (vit. D3) against the colon cytokines. The novel test formulation was treated with Biofield Energy Healing Treatment, as one of the best CAM approach with significant therapeutic outcomes. Biofield Energy Healing is one of the emerging frontiers aspect to CAM and various clinical approach has been used with significant results [6-9]. CAM therapies have been recommended by The National Center for Complementary/Alternative Medicine (NCCAM) and there therapies exist in various forms such as external qigong, Johrei, Reiki, therapeutic touch, yoga, Qi Gong, polarity therapy, Tai Chi, pranic healing, deep breathing, chiropractic/osteopathic manipulation, guided imagery, meditation, massage, homeopathy, hypnotherapy, progressive relaxation, acupressure, acupuncture, special diets, relaxation techniques, Rolfing structural integration, healing touch, movement therapy, pilates, mindfulness, Ayurvedic medicine, traditional Chinese herbs and medicines in biological systems both in vitro and in vivo. Human Biofield Energy has subtle energy that has the capacity to work in an effective manner [10] with its various clinical benefits [11]. This energy can be harness and transmit it into living and non-living things by the process of Biofield Energy Healing Treatment. Biofield Energy Treatment (the Trivedi Effect®- Consciousness Energy Healing Treatment) has been extensively studied with significant outcomes in the field of pharmaceuticals [12-14], nutraceuticals [15,16], metals and ceramics [17-19], microbiology [20-22], microbial genetics [23, 24], cancer research [25,26], livestock, agriculture science [27-29], improved bioavailability of many compounds [30-32], improved skin health [33, 34], improved properties of nutraceuticals [35, 36], improved overall bone health [37-39], human health and wellness. Thus, the study was planned on colon cytokines estimation that could significantly helped to improve the prevalence of gut inflammatory diseases using novel test formulation consisting of minerals such as Mg, Zn, Fe, Cu and vitamins including B6, B12, D3 in colon cancer cell line (HT-29).

Materials and Methods

Chemicals and Reagents

Antibiotics solution (Penicillin-Streptomycin) was purchased from HiMedia, India. 3-(4, 5-dimethyl-2-thiazolyl) 2, 5 diphenyl-2 H-tetrazolium) (MTT), Dulbecco’s Modified Eagle’s Medium (DMEM), NaHCO3, and EDTA were purchased from Sigma Chemical Corp. (St. Louis, MO), a subsidiary of Sigma-Aldrich Corporation. ELISA (enzyme-link immunosorbent assay) assay kits for all cytokines tumor necrosis factor alpha (TNF-α), macrophage inflammatory protein-1α (MIP-1α), and interleukin-1 beta (IL-1β) were purchased from R&D Systems, USA. Fetal bovine serum (FBS) was purchased from GIBCO, USA. Iron sulfate, copper chloride, and cholecalciferol (vitamin D3) were obtained from Sigma Chemical Co. (St. Louis, MO). Zinc chloride and magnesium (II) gluconate hydrate were obtained from TCI, Japan. Pyridoxine- HCL (vit-B6), cyanocobalamin (vit-B12) were procured from Alfa Aesar, USA. All other chemicals used in this study were analytical grade available in India.

Test Formulation and Reference Standard

The test formulation contained a combination of vitamins with minerals viz. iron sulfate, copper chloride, zinc chloride and magnesium (II) gluconate hydrate, cholecalciferol (vitamin D3), pyridoxine-HCL (Vit-B6), and cyanocobalamin (Vit-B12). Tumor necrosis factor alpha (TNF-α) was used as an inflammatory stimulant, while epigallocatechin-3-gallate (EGCG) was used as a reference standard (positive control) for immunomodulatory action in colon cancer cell line (HT-29).

Biofield Energy Healing Strategies

One part of each ingredient of the test formulation did not receive any sort of treatment and was defined as the untreated test formulation group, while another part received Biofield Energy Treatment known as Biofield Treated Test formulation by Mr. Mahendra Kumar Trivedi, a renowned Biofield Energy Healer under standard laboratory conditions for ~3 minutes. This treatment was provided through the Biofield Energy Healer unique Energy Transmission process (the Trivedi Effect®) to the test formulation. Further, the untreated test formulation was treated with a “sham” healer for comparison purposes. The “sham” healer did not have any knowledge about the Biofield Energy Treatment. After that, the Biofield Energy treated and untreated test formulations were kept in similar sealed conditions and used for the in vitro study on colon cancer cell line (HT-29) for cytokines estimation.

Experimental Design

The colon cancer cell line (HT-29) was divided into four different groups. Group 1 comprised of the HT-29 cells vehicle was denoted as the vehicle control, group 2 included cells with epigallocatechin-3-gallate (EGCG) as positive control at various concentrations. Group 3 and 4 included the cells with the untreated and Biofield Energy Treated test formulation, respectively at concentration range 0.1 to 15 μg/mL in presence of tumor necrosis factor - alpha (TNF-α).

Cytotoxicity by MTT Assay

The effect of the Biofield Energy Treated and untreated test formulations at a wide range concentration were tested for cell viability using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. The number of viable cells were determined by the ability of mitochondria to convert MTT to formazan dye. The details procedure was followed as per Plikerd et al. 2017 [40]. The effect of the test formulation on cell viability of HT-29 cells was determined as:

%Cell viability=100-%cytotoxicity (1)

Where; % cytotoxicity = [(O.D. of control cells – O.D. of cells treated with the test formulation)/O.D. of control cells]*100.

The results of the concentrations that showed >75% viability were selected subsequently for cytokine estimation.

Determination of Cytokine levels by ELISA

The HT-29 cell suspension in DMEM medium containing 10% FBS was plated at a density of 0.5 X 106 in 12-well cell culture plates. Cells were incubated at 37 °C for 24 hours. Cells were sera starved by replacing medium with DMEM containing 0% FBS and again incubated at 37 °C for another 24 hours. Cells were treated with proprietary test formulation at selected noncytotoxic concentrations and stimulated with Hu-TNF-α. Cells treated with TNF-α + EGCG were included as the positive control. After treatment, cells were incubated in a 5% CO2 incubator for 72 hours. After incubation, culture supernatants were collected from each well and stored at -20 °C until analysis. The level of cytokines (IL-6, IL-8, and IFN-γ) in culture supernatants of HT-29 cells were determined using ELISA as per manufacturer’s instructions.

Statistical Analysis

All the data were expressed as mean of three replicates ± SEM and were subjected to one-way analysis of variance (ANOVA) followed by Dunnett’s test and Student’s t-test for two groups comparison. Statistical significance was considered at p≤0.05.

Results & Discussion

MTT Assay

The cell viability results are summarized in the Figure 1. The percent cell viability in the vehicle control (VC) group was found as 117.5%. Moreover, the positive control, epigallocatechin-3- gallate (EGCG) showed 119.2%, 83%, and 101.2% cell viability at the concentration of 10, 50, and 100 μM, respectively. Further, the untreated test formulation showed more than 97% cell viability upto 122 μM; while the Biofield Treated test formulation showed more than 78% cell viability upto 122 μM. Based on the MTT cell viability assay the test formulation was found as safe and nontoxic upto the concentration of 122 μM. MTT assay is widely used for the cell toxicity against any test formulations. In addition, this assay was found as more rapid, less costly, less time consuming, and non-radioactive method as compared with the other assays. This assay display cell proliferation results on the basis of the cell growth and metabolic activity [41]. MTT assay suggest that the concentrations of the test formulation were found safe up to 122 μg/mL with respect to the viability in the colon cancer cell line (HT-29).

Estimation of Interleukin-6 (IL-6) Expression

The level of interleukin-6 (IL-6) expression in colon cancer (HT-29) cells is represented in the Figure 2. The positive control, epigallocatechin-3-gallate (EGCG) was significantly reduced the level of IL-6 by 38.71%, 92.43%, 91.74%, and 98.20% (p≤0.001) at the concentrations of 1, 10, 50, and 100 μM, respectively as compared to the vehicle control (VC) group. Moreover, the untreated test formulation showed significant reduction of IL-6 by 11.96%, 35.94%, and 52.3% (p≤0.001) at 0.1, 3, and 15 μM, respectively as compared to the VC group. Further, the Biofield Energy Treated test formulation group showed 14.19%, 34.09%, and 59.41% (p≤0.001) reduction of IL-6 at 0.1, 3, and 15 μM, respectively as compared to the VC group under the stimulation of TNF-α stimulation. Besides, the Biofield Treated test formulation also significantly reduced the level of IL-6 by 14.87% as compared to the untreated test formulation group. Overall, the minerals and vitamin-based Biofield Energy Treated test formulation showed an anti-inflammatory activity by reducing the level of IL-6 under the stimulation of TNF-α as compared with the vehicle control as well as untreated test formulation groups. Hence, the Biofield Energy Treated test formulation could be used a major role in immune-related disorders and also defined as controlling factor for many diseases [42]. Thus, it can be suggested that the Biofield Energy Treated test formulation can be used in many inflammatory disorders.

Estimation of IL-8 Expression

The level of interleukin-8 (IL-8) expression in colon cancer (HT-29) cells is represented in the Figure 3. The positive control, epigallocatechin-3-gallate (EGCG) was significantly reduced the level of IL-8 by 41.49%, 36.77%, 82.89% (p≤0.001), and 91.21% (p≤0.001) at the concentrations of 1,10,50, and 100 μM, respectively as compared to the vehicle control (VC) group. Moreover, the untreated test formulation showed significant reduction of IL-6 by 27.34% (p≤0.001), 21.03% ( ≤0.001), and 9.08% at 0.1, 3, and 15 μM, respectively as compared to the VC group. Further, the Biofield Energy Treated test formulation group showed significant (p≤0.001) reduction of IL-8 by 57.09% and 42.88% at 0.1 and 3 μM, respectively as compared to the VC group under TNF-α stimulation. Besides, the Biofield Treated test formulation also significantly reduced the level of IL-8 by 40.94% and 38.25% at 0.1 and 3 μg/mL, respectively as compared to the untreated test formulation group. Overall, the minerals and vitamin-based Biofield Energy Treated test formulation showed an anti-inflammatory activity by reducing the level of IL-8 under the stimulation of TNF-α as compared with the vehicle control as well as untreated test formulation group. Chronic inflammatory conditions leads to the massive production of proinflammatory factors such as chemokines. IL-8 is one of the chemokine in chronic inflammation and it is initially act as a neutrophil chemotactic and activating factor [43,44]. Overall, the experimental data suggested that Biofield Energy Healing Treatment has the significant capacity to reduce the level of IL-8 with respect to vehicle control and untreated test formulation.

Estimation of IFN-γ Expression

The level of interferon gamma (IFN-γ) expression in colon cancer (HT-29) cells is shown in the Figure 4. The positive control, epigallocatechin-3-gallate (EGCG) was significantly reduced the level of IFN-γ by 13.64%, 12.64%, 37.64%, and 22.99% at the concentrations of 1, 10, 50, and 100 μM, respectively as compared to the vehicle control (VC) group. Moreover, the untreated test formulation showed significant reduction of IFN-γ by 4.55% and 2.78% at 0.1 and 3 μM, respectively as compared to the VC group. Further, the Biofield Energy Treated test formulation significantly altered the level of IFN-γ as compared to the VC group under TNF-α stimulation. This suggests that the Biofield Energy Treated test formulation has significant immunomodulatory activity. Scientific reports suggest that various immunological and inflammatory functions of chemokines play significant role in controlling the immune response during infections. Overall, the immunomodulatory effect might be the result of specific modulation of NF-κB, a transcription factor involved in the activation of many inflammatory mediator genes [45,46].

Worldwide scope of alternative medicine and its outcomes have been increased significantly. However, an important phytoconstituents along with minerals and vitamins are reported to have beneficial role against many diseases such as diabetes, indigestion, inflammation of intestine, osteomalacia, blood disorders, infertility, potent revitalizer, etc. [47]. Due to high safety profile with the wide therapeutic action of alternative medicines, the scope has been increased worldwide [48]. Besides, the individual constituents of the novel proprietary test formulation has been reported to have substantial immunomodulatory action, and Biofield Energy Healing Treatment significantly alters the action of cytokines. Overall, the Biofield Energy Healing Treatment on the test formulation can be a novel approach in supports of the use of Biofield Treated test formulation for various types of autoimmune disorders in colon cancer cell line (HT-29).

Conclusion

On the basis of current study findings, it is concluded that the novel proprietary test formulation showed significant antiinflammatory action on the tested cytokines (IL-6, IL-8, and IFN-γ) in colon cancer cell line (HT-29) after administration of the Biofield Energy Treated formulation. MTT assay in the Biofield Energy Treated colon cancer cell line (HT-29) suggest that the test formulation showed more than 78% cell viability and found as safe and non-toxic. In addition, the levels of cytokine, interleukin-6 (IL-6) was significantly reduced by 34.09% and 59.41% (p≤0.001) at 3 and 15 μM, respectively as compared to the vehicle control (VC) group. Moreover, IL-8 level was reported to be significantly suppressed in the Biofield Energy Treated test formulation by 57.09% and 42.88% at 0.1 and 3 μM, respectively as compared with the VC group. On the basis of experimental results of various tested cytokines and their expression, significant anti-inflammatory activity in colon cancer cells was reported in the new test formulation after treated with the Trivedi Effect®- Biofield Energy Healing. Biofield Energy Treated test formulation can be used as a Complementary and Alternative Medicine (CAM) to prevent the immune-mediated diseases such as Irritable Bowel Syndrome, Rheumatoid arthritis, Ulcerative colitis and Crohn’s disease, Stress, Asthma, and many more with safe therapeutic index. Besides, it can also be utilized in organ transplants (for example kidney transplants, liver transplants and heart transplants), various autoimmune disorders such as Lupus, Addison Disease, Celiac Disease (gluten-sensitive enteropathy), Dermatomyositis, Graves’ Disease, Hashimoto Thyroiditis, Multiple Sclerosis, Myasthenia Gravis, Pernicious Anemia, Aplastic Anemia, Sjogren Syndrome, Systemic Lupus Erythematosus, Diabetes, Alopecia Areata, Fibromyalgia, Vitiligo, Psoriasis, Scleroderma, Chronic Fatigue Syndrome and Vasculitis, Type 1 to improve the overall health and quality of life.

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