|Year : 2021 | Volume
| Issue : 1 | Page : 55-69
Focused-pulsed electromagnetic field treatment reverses lipopolysaccharide-induced alterations in gene expression profile in human gastrointestinal epithelial cells
Asit Panja1, Rolf Binder2, Silvia Binder3
1 AlfaGene Bioscience Inc. Fords, New Jersey, USA
2 ONDAMED GmbH, Schwanau, Germany
3 The Binder Institute for Personalized Medicine, Schwanau, Germany
|Date of Submission||04-Jul-2020|
|Date of Decision||16-Jul-2020|
|Date of Acceptance||31-Aug-2020|
|Date of Web Publication||2-Feb-2021|
Alfagene Bioscience Inc., 20 Corrielle Street, Fords, New Jersey 08863
Source of Support: None, Conflict of Interest: None
BACKGROUND: Although the therapeutic effects of pulsed electromagnetic field (PEMF) treatment are well documented, underlying mechanisms of PEMF in treating pathological conditions are incompletely understood.
METHODS: We utilized a human gastrointestinal epithelial cell system to investigate the influence of a low-frequency electromagnetic field generated from a focused PEMF (f-PEMF) device on the expression of human genes. We simulated an inflammatory condition by stimulating the cells with lipopolysaccharide (LPS). A set of LPS-activated cells were then subjected to 100-Hz f-PEMF for 30 seconds to observe the therapeutic effect of f-PEMF. We determined the therapeutic effect by analyzing the reversal of LPS-induced alterations in gene expression using RNA-seq analysis. The results were compared to the changes between untreated controls versus LPS treated cells, defining the homeostatic alteration of changes in gene expression profile caused by LPS stimulation alone. We further compared LPS treated versus LPS + f-PEMF treated cells to examine the effect of f-PEMF in the reversal of the LPS-induced alterations in gene expression patterns.
RESULTS: A total of 38,162 genes (of 60,448 tested) were constitutively expressed in the untreated control cells. Stimulation with LPS altered the expression profile through de novo-induction of >1950 genes that were originally unexpressed and silencing 2486 constitutively expressed genes. LPS treatment also altered expression levels in a large panel of genes. Exposing LPS-treated cells to 100 Hz of f-PEMF for 30 seconds (s) showed reversals of LPS treatment-induced altered gene expression. In this paper, we emphasize the f-PEMF regulation of genes associated with inflammatory processes.
CONCLUSION: Our data indicates for possibility of developing new nonchemical alternative therapeutic approaches for treatment of inflammation and pain.
Keywords: Gastrointestinal epithelial cell, gene expression, inflammation, lipopolysaccharide, pulsed electro-magnetic field
|How to cite this article:|
Panja A, Binder R, Binder S. Focused-pulsed electromagnetic field treatment reverses lipopolysaccharide-induced alterations in gene expression profile in human gastrointestinal epithelial cells. Int J Health Allied Sci 2021;10:55-69
|How to cite this URL:|
Panja A, Binder R, Binder S. Focused-pulsed electromagnetic field treatment reverses lipopolysaccharide-induced alterations in gene expression profile in human gastrointestinal epithelial cells. Int J Health Allied Sci [serial online] 2021 [cited 2023 Mar 29];10:55-69. Available from: https://www.ijhas.in/text.asp?2021/10/1/55/308584
| Introduction|| |
Advances in medical sciences during the past century have meaningfully shifted the nature of disease landscape and treatment modalities for many diseases. Acute illnesses caused by specific infectious microorganisms are being replaced increasingly by chronic disorders with multiple etio-pathophysiological factors. This emerging trend of chronic inflammatory health conditions arises from complex interactions among cellular, molecular, genetic components of various organs and tissues in the body, which are further influenced by environmental factors.,,,, Inflammation is also caused by direct injuries such as trauma or infection to the tissues and associated organs.,, Inflammation remains the major cause of many chronic illnesses.,, In many cases, inflammation-mediated illnesses are diagnosed and treated based on symptoms and signs within a single system or organ rather than the variables affecting the disease pathogenesis. For example, nonsteroidal anti-inflammatory drugs (NSAIDs) are often used to block the effects of inflammatory mediator prostaglandin produced by vascular components in response to inflammation in the localized tissues.,,, While such symptomatic treatments alleviate symptoms, they do not target homeostatic dysbalances in the inner core of the cellular components of the affected tissues, organs, and/or the systems. Many of such anti-inflammatory drugs used for treating these conditions also cause severe side effects.,, In addition, the cost of these medications is expensive, especially when the treatment is required for a long period.,,, Therefore, health care providers have an obligation to overcome these challenges by exploring alternative therapeutic approaches which would be safe, effective, rapid, and without any invasive risks.
Electromagnetic (EM) therapy has been known to stimulate the body's own healing mechanisms by regulating intrinsic mechanisms for repair and regeneration of the pathological state of selected body areas or weakened tissues,,,,,, possibly, through restoration and/or realignment of the subcellular components and/or the quantum energy field inside the cells (Cyto-Quantum-Energy). Thus, EM and other alternative therapy are increasingly in demand by physicians and patients alike. However, despite its historical use and modern popularity, a considerable level of wariness exists in the integration of such concepts into modern medicine. This is largely in part due to the lack of understanding of how the low-frequency EM impulses interact with the cellular, molecular, genetic components and regulate the function of specific organs and/or tissues in the human body.
Therefore, we have recently initiated a multi-disciplinary collaborative program to study bio-interactive mechanisms of focused EM fields with a human gastrointestinal (GI) stem cell-like primary epithelial cell system and a pulsed electromagnetic field (PEMF) device using focused EM fields. The results from a pilot study demonstrating the effect of pulsed EM stimulation on gene expression profile and its relevance to potential mechanisms of therapeutic effects on chronic inflammatory conditions are discussed in this paper.
According to the results, LPS induced changes in the inflammation associated gene expression profile in human cells is reversible by optimum focused PEMF treatment, perhaps, through restoration and/or reorganization of the quantum energy field inside the cells (cyto-quantum-energy-homeostasis) used in this study. The reported concept of cyto-quantum-energy-homeostasis for gene expression regulation by focused PEMF opens new possibilities for the development and utility of devices that may re-establish the physiological environment in cells, organs, and tissues of inflamed or chronically damaged areas in the body.
| Materials and Methods|| |
The aim of this study is to determine the mechanisms of action of focused PEMF (f-PEMF) treatment in cellular, molecular, and genetic levels in the human body. Emphasis has been on the inflammatory panel of genes. An in vitro model of an inflammatory scenario was established by stimulating a GI stem cell-like human intestinal primary epithelial cell population, with lipopolysaccharides (LPS) (Sigma, St Louis, MO). LPS is a cell wall protein from Gram-negative bacteria and is known for its inflammatory effects in humans.,,, In many acute and/or chronic inflammatory conditions of the GI tract, epithelial barrier function is thought to be disrupted by LPS. This further leads to the activation of immunoresponsive cells in the gut, causing overproduction of inflammatory cytokines (cytokine storm). To examine whether and how focused PEMF could influence the LPS altered molecular biology of the cells, in a separate condition, the same LPS stimulated cells were further exposed to a focused EM field of 100 Hz for 30s by using the ONDAMED device [Figure 1]. Cells were then cultured for an additional 14 h in a humidified CO2 incubator. Gene expression profile in all three conditions (untreated, LPS treated, and LPS + ONDAMED' focused PEMF) were analyzed using RNAseq analysis through a third-party laboratory.
|Figure 1: Pulsed electro-magnetic frequency exposure to lipopolysaccharide treated gastrointestinal epithelial cells in culture|
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A frozen vial of 500,000 GI stem cell-like primary epithelial cells (from AlfaGene Bioscience Inc, Fords, NJ, USA) was defrosted, washed, and cultured in 3 ml of HIPEC medium, in a vented-capped 25 cm2 sterile tissue culture flask (Corning, NY, USA) at 37°C temperature in a humidified (90%) CO2 (5%) incubator. Upon 80% confluency, cells were trypsinized and dissociated from the culture flask, washed three times in F12 medium. Cells were then re-seeded in 6-well tissue culture plates (7.5 × 105 cells/2 ml of medium/well) for the experiment, as described above in [Figure 1]. Three separate plates were used for each of the three conditions (1. medium alone, 2. medium + LPS 1 mg/ml, and 3. medium + LPS + PEMF) and placed in three separate racks in the incubator for 14 h after the addition of LPS (conditions 2 and 3) and PEMF (condition 3 only) treatment.
Ondamed's focused-pulsed electromagnetic field exposure
PEMF exposure to the cultured cells was carried out by using a focused PEMF treatment device developed by the Ondamed Company (Schwanau, Germany). The device, invented in Germany in 1993 by electronics engineer Rolf Binder, employs the mechanism of action of pulsed EM fields emitted through various applicators. Four differently sized applicators deliver targeted stimulation to a localized area while a fifth available applicator delivers a wide field stimulation providing a systemic therapeutic effect such as the lymph system, bone, and joints. Frequencies range between 0.1 and 31,835 Hz with an available intensity setting between 0.5 and 55 mT (milli Tesla). The field density varies for each applicator.
An EM field was created by placing an applicator with a 2.4” (6 cm) distance for stimulation connected to the PEMF generating device on a flat surface of a wooden countertop at room temperature. A bridge of approximately 2.4” (6 cm) height was created by placing a plastic box on both horizontal sides of the EM matrix applicator. Then, a holding bridge was created by placing another container on the top of the surface, as shown in [Figure 1].
The 6-well cell culture plate containing cultured human intestinal primary epithelial cells (750K cells/2 ml of medium/well) were treated with LPS (1 μg/ml) 10 min before the EM exposure was placed on the top of the bridge surface. This set up allowed vertical transmission of EM waves (100 Hz) to the center top of the designed bridge. The device contains control switches for an EM frequency intensity as well as exposure time. LPS-stimulated cells were subjected to a single exposure of 100 Hz PEMF for 30s followed by their placement in the incubator as described above in the cell culture section.
After 14 h of (posttreatment) incubation period, medium from the cultured cells was collected, centrifuged to obtain cell-free supernatants and stored at −20C for future analysis for secreted proteins. At the same time, the adhered cells in the wells of the culture plates were gently washed with PBS, followed by lysis in RNA later solution. Total RNA was extracted from these cell lysate samples from each of the three experimental conditions by using the RNAeasy kit (Qiagen, Germantown, MD) following the manufacturer's instructions. The final RNA pellet was dissolved in RNAse-free water. The yield of total RNA from each of three conditions (with the equal number of seeded cells) was 13 μg (medium only), 2 μg (medium + LPS), and 9 μg (medium + LPS + f-PEMF), respectively. These results from total RNA yield itself indicated that the reduction in the amount of total RNA from the LPS treated cells might have been due to inhibition of cell growth or induction of cell death by LPS treatment to these primary epithelial cells, which was overcome by f-PEMF treatment.
Extracted total RNA samples were labeled in a coded fashion and sent for further quality assessment and subsequent RNAseq analysis to an FDA compliant Next Generation Sequence (third party contract research organization) laboratory.
RNAseq and gene expression analysis
RNAseq analysis was performed using CLC Genomics Workbench v. 10.0.1. (Qiagen, Redwood City, CA). Illumina reads were trimmed to remove low-quality ends and adapter sequences. Sequences of at least 50 bp in length were then mapped to the GRCh38 reference genome using the RNA-Seq Analysis tool in CLC. Total gene and transcript hit counts were measured, and RPKM (read per kilobase million) values were calculated. Gene and transcript values were analyzed separately. Kal's Z-test was used to compare the expression values in different conditions. Genes with P ≤ 0.05 and absolute proportions fold change ≥2 were called as differentially expressed genes for each comparison. Transcripts with P ≤ 0.05 and absolute proportions fold change ≥2 were called differentially expressed transcripts for each comparison. Gene results were annotated with the Gene Ontology Biological Process information. Then, Hyper-G tests for each comparison were conducted using the genes with a fold change >2 and P < 0.05.
| Results|| |
To gain insights on how f-PEMF may act in cellular constituents of tissue in various pathological settings, we took a simple approach by asking the question of what changes occur at the genomic level of a cell during the initiation and perpetuation of an inflammatory process and then whether PEMF treatment can have any role in reversing the changes if there are any [Table 1] and [Table 2]. Hence, the first step to address this question was to assess the changes in the total number of expressed versus unexpressed genes caused by LPS alone or LPS + PEMF compared to the untreated (medium only) cells. As can be seen in [Table 1], out of total 60,448 genes tested, only 38,162 genes were constitutively expressed in the untreated cells (medium only). This number of expressed genes remained in balance (almost unchanged) after LPS stimulation (38,164 genes), which was however, reduced by 2,932 genes on exposure to focused PEMF treatment (35,232 genes). The observed balance in the total number expressed or unexpressed genes in control (medium only) versus LPS treated cells raised two possibilities: (i) LPS may not have any effect in turning on or off any genes in these cells but may have regulatory effects only on the expression levels or (ii) The number of genes induced (turned on expression) by signals from LPS stimulation was balanced by silencing the equal number of genes (turning off the expression) along with its effects on the expression levels.
|Table 1: Changes in total number of expressed versus unexpressed genes in LPS treated versus LPS + PEMF treated cells|
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|Table 2: Total number of genes turned-on (induced) or turned-off (silenced) by LPS treatment or LPS + PEMF treatment compared to baseline (unstimulated/untreated) control condition|
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Therefore, to rule in or rule out any of these possibilities, we counted the total number de novo expressed genes (that were not expressed in the untreated) and the number silenced (that were originally expressed in untreated cells) in both LPS treated and LPS + PEMF treated cells compared to the control condition (medium only). De novo expression of at least 1,950 genes in LPS treated and 1,371 genes in LPS + focused PEMF treated cells were observed. Similarly, there were 2,486 and 4,303 genes were turned off by LPS alone and LPS + PEMF treatment, respectively [Table 2]. Similar analytical observation was made in the transcriptome profiles as well. However, as mentioned above in this paper, our focus is only on the gene expression profile with special emphasis on the inflammation-related genes. Transcriptome data, as well as the analysis of other functional groups of genes, will be reported in subsequent papers from our group.
Our next goal was to further explore whether and how PEMF regulates actual levels of expression of various functional groups of genes that are altered by disease initiating processes. For this purpose, we created at least 12 functional groups of genes filtered from the genome database by using CLC software. Then we further filtered and organized the list of genes in two groups. One with the list of genes of which the expression levels were increased by LPS treatment. Similarly, the second group included the genes that were decreased by LPS stimulation compared to the baseline control. We display and compared side-by-side the RPKM values (measure of expression level) of all the genes from each of the three experimental conditions (medium only, LPS stimulated, LPS stimulated + focused PEMF treated) for all the 12 functional groups (not shown in this paper). Results from the functional module of inflammation-related genes are shown in [Table 3] and [Table 4]. As can be seen in both of these tables, PEMF treatment had a profound effect in reversing the altered expression levels irrespective of the signaling directions (up-regulatory or down-regulatory) by LPS. Similar pattern of genes expression regulation by PEMF was seen in the other functional groups (e.g. cell growth, differentiation, apoptosis, sensory, immunoregulatory, cytokines, chemokines, etc.). These observations suggest that an optimized level of EM energy may have a significant impact on biological systems.
|Table 3: Pulsed electro-magnetic frequency treatment reverses upregulated expression of inflammation associated genes in LPS -stimulated human gastrointestinal stem cell-Derived epithelial cells|
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|Table 4: Pulsed electro-magnetic field treatment reverses downregulated expression of inflammation-associated genes in LPS-stimulated human gastrointestinal epithelial cells|
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Most interestingly, we have noted that 23 genes were regulated with statistical significance (P = 0.05). Our findings not only substantiate the therapeutic potential of PEMF through genomic homeostasis regulation but also provide valuable information that PEMF treatment could potentially offer an alternative solution to safety concerns of currently marketed anti-inflammatory drugs such as NSAIDs, which acts through inhibition of PSTG gene but bears a wide range of side effects. However, before considering PEMF as an alternative to NSAIDS, it is important to determine if there is any potential toxicity caused by PEMF exposure in our experiments. Therefore, similar to the analysis of inflammation-associated genes described above, by using the same CLC software, we filtered a panel of toxicology associated genes from the entire list of 60,446 annotated genes used for our RNAseq analysis. We then examined the influence of focused PEMF on the expression regulation of this toxicology associated panel genes. We again sub-divided the toxicology gene panel by two groups i) those of which the expression levels were increased by LPS and ii) those of which were decreased by LPS stimulation in adult human intestinal primary epithelial cells. As can be seen in [Table 5] and [Table 6], consistent with the observation in inflammatory genes panel, the expression level of most of the toxicology genes that were either upregulated [Table 5] or downregulated [Table 6] by LPS were put into the reverse trend by focused PEMF treatment. Although this was a preliminary study with one single dose of 100 Hz EM, these results provide important evidence that exposure to the low frequency of PEMF may play a role in the restoration of physiologic homeostasis without any overt toxic effect. Of course, further studies are necessary to characterize the safety and efficacy ranges in biological cell, tissue, organ, and/or system-specific manner.
|Table 5: Pulsed electro-magnetic field treatment reverses upregulated expression of toxicology pathway associated genes in LPS-Stimulated human gastrointestinal epithelial cells|
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|Table 6: Pulsed electro-magnetic frequency treatment reverses downregulated expression of toxicology pathway associated genes in lipopolysaccharide -stimulated human gastrointestinal epithelial cells|
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| Discussion|| |
Inflammation is considered to be the underlying cause of most of the major chronic organ system clinical diseases., For example, pathophysiological processes of cancer, heart attack, Alzheimer, arthritis, and many other chronic conditions are associated with the initiation and perpetuation of inflammation in the respective tissues or organs.,,, Such chronic diseases involve multiple structural and functional alterations within the inner core of cells.,,,,, All of these alterations are fundamentally associated with changes in gene expression profile, which in turn, impair physiological properties in specific cells, tissues, and organs in the human body. For many of these diseases, currently, no adequately safe treatment exists.
In the present study, we examined the effect of a single exposure of 100 Hz on gene expression regulation in LPS stimulated human intestinal epithelial cells. The adult human GI stem cell-like primary epithelial cells used in our study serves as an interesting model to study inflammatory responses as well as to study the biological effects of the EM field because there exists a large body of information in the literature for their interactions with bacterial toxins as well as transmission of electromagnetically coupled energy through these cells. Bacterial endotoxin induces a potent inflammatory response by these cells involving a variety of mechanisms., Previous studies have shown in the past that LPS induces proinflammatory cytokine production as well modulates surface antigen expression in these cells., EM transmission is the basic principle used in many clinical investigational procedures. Visualization of the epithelial surface inside the GI tract by battery-less wireless capsule endoscopy is based on EM transmission. Thus, these cells suited us well for our experimental design.
Not surprisingly, our experimental results demonstrated that LPS caused massive alterations in the human genome profile both in expression and transcription levels. An interesting phenomenon was observed that the number of genes that were turned on (de novo induced) by LPS (1 μg/ml) stimulation was almost equal to the number of genes that were silenced during the inflammatory response to LPS by these cells [Table 1] and [Table 2]. More interestingly, our experimental results demonstrated that a single treatment of low frequency focused PEMF (100 Hz) for 30s reverse directed the LPS actions on gene expression regulation. A large panel of inflammation-associated genes that were upregulated by LPS stimulation was downregulated by PEMF treatment and vice versa [Table 3] and [Table 4]. We expect to identify valuable information from this simple and single pilot study of human genome expression regulation conducted in that may guide for drawing new strategies for discovery and development of new, safer, more effective, and economical treatments for chronic inflammatory diseases. In this regard, we highlight one critical observation that an exposure of focused PEMF (100 Hz for 30 s) inhibits prostaglandin-endoperoxide synthase (PTGS2) by ~ 40% [Table 3]. PTGS2 is known as cyclooxygenase-2 (Cox2), which is the key enzyme in the prostaglandin biosynthesis pathway and as is normally targeted by NSAID. It is worth mentioning that NSAIDs represent one of the most widely prescribed drugs for treatments of inflammation, pain, and fever. Their effects are largely attributed to the suppression of prostaglandin production by inhibiting cyclooxygenases. However, NSAIDs are also known to cause huge health burdens when taken for longer periods. Chronic use of NSAIDs causes GI ulcers, hemorrhage, liver toxicity, cardiovascular conditions, and many adverse effects in human health. Therefore in addition to general homeostatic restoration, focused PEMF may offer a solution for safety and efficacy concerns of currently used anti-inflammatory drugs. Finally, our analysis on toxicological panel shows a similar trend of reversal of the LPS effect without any overt induction of toxicological genes [Table 5] and [Table 6].
Taken together our results provide new insights toward the understanding of potential mechanisms by which focused PEMF offering effects on subtlest constituents (genes) of the human body. A single exposure of 100 Hz f-PEMF for 30s to LPS stimulated cells resulted in reverse regulation of gene expression profile. The expression of genes that were increased by a known immune-stimulatory stimuli LPS were in most cases, reversed either by complete restoration or by bringing closure to the level of that was seen in the unstimulated (baseline control) cells. In some cases, focused PEMF downregulated the gene expression even below the constitutive level. For example, the PTGS2 gene was suppressed by 40% (from a RPKM value of. 143 in LPS treated cells to RPKM value of 3.579 in LPS + f-PEMF treated cells) by a low-frequency f-PEMF for the minuscule amount of time (30s) opens a new possibility for filling the safety and efficacy gaps in current therapeutic approaches for the treatment of chronic diseases.
In conclusion, alterations in epigenetic homeostasis that occur in various acute and chronic health conditions' resulting symptoms, syndromes, pain, and suffering might be associated with a dysregulated EM energy field within the cells of the affected tissues, organs, or organ systems. A reversal of such EM energy dysbalance might be possible through the creation of an optimum EM field within the damaged body parts. The main limitation of this study that it is of exploratory nature with a limited number of experiments. Therefore, our data need to be confirmed in larger studies with additional controls and more detailed comparative analysis. Further studies of EM therapy mechanisms and dosimetry for specific health conditions with devices such as the one used in our study may contribute significantly in the reduction of both health-and economic burdens caused by many chronic health conditions for which modern medicine offers no solution.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Allen RJ, Porte J, Braybrooke R, Flores C, Fingerlin TE, Oldham JM, et al
. Genetic variants associated with susceptibility to idiopathic pulmonary fibrosis in people of European ancestry: A genome-wide association study. Lancet Respir Med 2017;5:869-80.
David T, Ling SF, Barton A. Genetics of immune-mediated inflammatory diseases. Clin Exp Immunol 2018;193:3-12.
Kerstein A, Schüler S, Cabral-Marques O, Fazio J, Häsler R, Müller A, et al
. Environmental factor and inflammation-driven alteration of the total peripheral T-cell compartment in granulomatosis with polyangiitis. J Autoimmun 2017;78:79-91.
Kunnumakkara AB, Sailo BL, Banik K, Harsha C, Prasad S, Gupta SC, et al
. Chronic diseases, inflammation, and spices: How are they linked? J Transl Med 2018;16:14.
Smith JA, Zhao W, Wang X, Ratliff SM, Mukherjee B, Kardia SL, et al
. Neighborhood characteristics influence DNA methylation of genes involved in stress response and inflammation: The multi-ethnic study of atherosclerosis. Epigenetics 2017;12:662-73.
Langgartner D, Palmer A, Rittlinger A, Reber SO, Huber-Lang M. Effects of prior psychosocial trauma on subsequent immune response after experimental thorax trauma. Shock 2018;49:690-7.
Ritzel RM, Doran SJ, Barrett JP, Henry RJ, Ma EL, Faden AI, et al
. Chronic alterations in systemic immune function after traumatic brain injury. J Neurotrauma 2018;35:1419-36.
Kezić A, Stajic N, Thaiss F. Innate immune response in kidney ischemia/reperfusion injury: Potential target for therapy. J Immunol Res 2017;2017:6305439.
Ravichandran S, Michelucci A, Del Sol A. Integrative computational network analysis reveals site-specific mediators of inflammation in Alzheimer's disease. Front Physiol 2018;9:154.
Egawa M, Mitamura Y, Akaiwa K, Semba K, Kinoshita T, Uchino E, et al
. Changes of choroidal structure after corticosteroid treatment in eyes with Vogt-Koyanagi-Harada disease. Br J Ophthalmol 2016;100:1646-50.
Winkelman C, Higgins PA, Chen YJ, Levine AD. Cytokines in chronically critically ill patients after activity and rest. Biol Res Nurs 2007;8:261-71.
Nagahisa A, Asai R, Kanai Y, Murase A, Tsuchiya-Nakagaki M, Nakagaki T, et al
. Non-specific activity of (+/-)-CP-96,345 in models of pain and inflammation. Br J Pharmacol 1992;107:273-5.
Joshi P, Dhaneshwar SS. An update on disease modifying antirheumatic drugs. Inflamm Allergy Drug Targets 2014;13:249-61.
Cudaback E, Jorstad NL, Yang Y, Montine TJ, Keene CD. Therapeutic implications of the prostaglandin pathway in Alzheimer's disease. Biochem Pharmacol 2014;88:565-72.
Bolay H, Durham P. Pharmacology. Handb Clin Neurol 2010;97:47-71.
Aghazadeh-Habashi A, Asghar W, Jamali F. Drug-disease interaction: Effect of inflammation and nonsteroidal anti-inflammatory drugs on cytochrome P450 metabolites of arachidonic acid. J Pharm Sci 2018;107:756-63.
Chung EY, Tat ST. Nonsteroidal anti-inflammatory drug toxicity in children: A clinical review. Pediatr Emerg Care 2016;32:250-3.
de Pouvourville G, Tasch RF. The economic consequences of NSAID-induced gastrointestinal damage. Eur J Rheumatol Inflamm 1993;13:33-40.
Trippel SB. The unmet anti-inflammatory needs in orthopedics. Am J Orthop (Belle Mead NJ) 1999;28 Suppl 3:3-7.
Pardutz A, Schoenen J. NSAIDs in the acute treatment of migraine: A review of clinical and experimental data. Pharmaceuticals (Basel) 2010;3:1966-87.
Hunsche E, Chancellor JV, Bruce N. The burden of arthritis and nonsteroidal anti-inflammatory treatment. A European literature review. Pharmacoeconomics 2001;19 Suppl 1:1-15.
Iwasa K, Reddi AH. Pulsed electromagnetic fields and tissue engineering of the joints. Tissue Eng Part B Rev 2018;24:144-54.
Aragona SE, Mereghetti G, Lotti J, Vosa A, Lotti T, Canavesi E. Electromagnetic field in control tissue regeneration, pelvic pain, neuro-inflammation and modulation of non-neuronal cells. J Biol Regul Homeost Agents 2017;31 2 Suppl 2:219-25.
Bilgin HM, Celik F, Gem M, Akpolat V, Yildiz I, Ekinci A, et al
. Effects of local vibration and pulsed electromagnetic field on bone fracture: A comparative study. Bioelectromagnetics 2017;38:339-48.
Ongaro A, Pellati A, Bagheri L, Fortini C, Setti S, De Mattei M. Pulsed electromagnetic fields stimulate osteogenic differentiation in human bone marrow and adipose tissue derived mesenchymal stem cells. Bioelectromagnetics 2014;35:426-36.
Ferroni L, Tocco I, De Pieri A, Menarin M, Fermi E, Piattelli A, et al
. Pulsed magnetic therapy increases osteogenic differentiation of mesenchymal stem cells only if they are pre-committed. Life Sci 2016;152:44-51.
Jiao M, Lou L, Jiao L, Hu J, Zhang P, Wang Z, et al
. Effects of low-frequency pulsed electromagnetic fields on plateau frostbite healing in rats. Wound Repair Regen 2016;24:1015-22.
Panja A. A novel method for the establishment of a pure population of nontransformed human intestinal primary epithelial cell (HIPEC) lines in long term culture. Lab Invest 2000;80:1473-5.
Martin CA, Panja A. Cytokine regulation of human intestinal primary epithelial cell susceptibility to Fas-mediated apoptosis. Am J Physiol Gastrointest Liver Physiol 2002;282:G92-104.
Kim YO, Han SB, Lee HW, Ahn HJ, Yoon YD, Jung JK, et al
. Immuno-stimulating effect of the endo-polysaccharide produced by submerged culture of Inonotus obliquus. Life Sci 2005;77:2438-56.
Haller D, Bode C, Hammes WP. Cytokine secretion by stimulated monocytes depends on the growth phase and heat treatment of bacteria: A comparative study between lactic acid bacteria and invasive pathogens. Microbiol Immunol 1999;43:925-35.
Pardon MC. Lipopolysaccharide hyporesponsiveness: Protective or damaging response to the brain? Rom J Morphol Embryol 2015;56:903-13.
Zielen S, Trischler J, Schubert R. Lipopolysaccharide challenge: Immunological effects and safety in humans. Expert Rev Clin Immunol 2015;11:409-18.
Voss OH, Murakami Y, Pena MY, Lee HN, Tian L, Margulies DH, et al
. Lipopolysaccharide-induced CD300b receptor binding to toll-like receptor 4 alters signaling to drive cytokine responses that enhance septic shock. Immunity 2016;44:1365-78.
Umeno J, Esaki M, Hirano A, Fuyuno Y, Ohmiya N, Yasukawa S, et al
. Clinical features of chronic enteropathy associated with SLCO2A1 gene: A new entity clinically distinct from Crohn's disease. J Gastroenterol 2018;53:907-15.
Kallwellis-Opara A, Dörner A, Poller WC, Noutsias M, Kühl U, Schultheiss HP, et al
. Autoimmunological features in inflammatory cardiomyopathy. Clin Res Cardiol 2007;96:469-80.
Stephenson J, Nutma E, van der Valk P, Amor S. Inflammation in CNS neurodegenerative diseases. Immunology 2018;154:204-19.
Dalle S, Rossmeislova L, Koppo K. The role of inflammation in age-related sarcopenia. Front Physiol 2017;8:1045.
Lucas SM, Rothwell NJ, Gibson RM. The role of inflammation in CNS injury and disease. Br J Pharmacol 2006;147 Suppl 1:S232-40.
Gea J, Orozco-Levi M, Barreiro E, Ferrer A, Broquetas J. Structural and functional changes in the skeletal muscles of COPD patients: The “compartments” theory. Monaldi Arch Chest Dis 2001;56:214-24.
Chung KF, Adcock IM. Pathophysiological mechanisms of asthma. Application of cell and molecular biology techniques. Mol Biotechnol 2001;18:213-32.
Bohadana A, Teculescu D, Martinet Y. Mechanisms of chronic airway obstruction in smokers. Respir Med 2004;98:139-51.
Ogata T, Shibamura H, Tromp G, Sinha M, Goddard KA, Sakalihasan N, et al
. Genetic analysis of polymorphisms in biologically relevant candidate genes in patients with abdominal aortic aneurysms. J Vasc Surg 2005;41:1036-42.
Masser DR, Otalora L, Clark NW, Kinter MT, Elliott MH, Freeman WM. Functional changes in the neural retina occur in the absence of mitochondrial dysfunction in a rodent model of diabetic retinopathy. J Neurochem 2017;143:595-608.
Nishida K, Otsu K. Inflammation and metabolic cardiomyopathy. Cardiovasc Res 2017;113:389-98.
Rainard P, Cunha P, Gilbert FB. Innate and adaptive immunity synergize to trigger inflammation in the mammary gland. PLoS One 2016;11:e0154172.
Biswas G, Bilen S, Kono T, Sakai M, Hikima J. Inflammatory immune response by lipopolysaccharide-responsive nucleotide binding oligomerization domain (NOD)-like receptors in the Japanese pufferfish (Takifugu rubripes). Dev Comp Immunol 2016;55:21-31.
Salomão R, Martins PS, Brunialti MK, Fernandes Mda L, Martos LS, Mendes ME, et al
. TLR signaling pathway in patients with sepsis. Shock 2008;30 (Suppl 1):73-7.
Stroeher UH, Jedani KE, Manning PA. Genetic organization of the regions associated with surface polysaccharide synthesis in Vibrio cholerae O1, O139 and Vibrio anguillarum O1 and O2: A review. Gene 1998;223:269-82.
[Table 1], [Table 2], [Table 3], [Table 4], [Table 5], [Table 6]