1.Role of calcineurin-nuclear factor of activated T cells signaling pathway in myoblast apoptosis induced by cyclic tensile strain.
Xian DING ; Chenlei XIA ; Miao HE ; Wenna SUN ; Fang WANG ; Wenxin JIANG ; Caixia ZHANG ; Shuangyu WANG ; Qiang ZHANG ; Ruyong YAO ; Xiao YUAN
West China Journal of Stomatology 2015;33(5):456-461
OBJECTIVEThis study investigated the role and mechanism of calcineurin (CaN)-nuclear factor of activated T cells (NFAT) pathway in the myoblast apoptosis induced by cyclic tensile strain.
METHODSMyoblasts were cultured using an in vitro-mechanical stimulation model and imposed with tension for different hours with a multi-channel cell stress loading system. Cyclosporine (CsA) was used as CaN inhibitor to clarify the role of CaN in the apoptosis induced by cyclic stress. Hochest 33258 staining and flow cytometry detection were performed to detect the apoptotic cells. Real-time polymerase chain reaction was conducted to detect the mRNA expression of CaN and NFAT. Protein levels of NFAT3 were evaluated by Western blot.
RESULTSThe apoptosis rate increased with the extension of loading time. The mRNA expression of the CaN subunits, CnA and CnB, and the protein levels of NFAT3 also increased. When the myoblasts were incubated with CsA, the apoptosis rate decreased, the mRNA expression of CnA and NFAT3 significantly decreased, and the NFAT3 protein expression levels became significantly lower than those of the groups without CsA.
CONCLUSIONContinuous cyclic tensile stress can induce myoblast apoptosis. The CaN-NFAT signaling pathway may be involved in the cyclic stretch-induced apoptosis of myoblasts.
Apoptosis ; Calcineurin ; genetics ; Cyclosporine ; Flow Cytometry ; Myoblasts ; physiology ; NFATC Transcription Factors ; metabolism ; Real-Time Polymerase Chain Reaction ; Signal Transduction ; T-Lymphocytes
2.The effect of ultra high molecular weight polyethylene particles on macrophage.
Yuan-qing MAO ; Zhen-an ZHU ; Ting-ting TANG ; Hong-bin LI
Chinese Journal of Surgery 2006;44(12):852-855
OBJECTIVETo study the effect of ultra high molecular weight polyethylene (UHMWPE) particles on macrophages and evaluate the expression of NFAT2, a key transcriptional factor for osteoclast differentiation.
METHODSFrom November 2004 to February 2005, macrophages were co-cultured with UHMWPE particles. When observed at different times, the proliferation activity of macrophages was analyzed by MTT and the expression of calcineurin (CaN) and NFAT2 by immunohistochemical and RT-PCR method respectively.
RESULTSThe macrophages phagocytosed UHMWPE particles in an early time, the expression of CaN and NFAT2 was increased, while the proliferation activity was not enhanced.
CONCLUSIONSUHMWPE particles can stimulate macrophages to phagocytose significantly, and enhance the expression of the transcriptional factor NFAT2.
Animals ; In Vitro Techniques ; Joint Prosthesis ; Macrophages ; drug effects ; physiology ; Mice ; NFATC Transcription Factors ; biosynthesis ; genetics ; Phagocytosis ; drug effects ; Polyethylenes ; pharmacology ; RNA, Messenger ; genetics
3.Hypertrophy of the heart: calcineurin-NFAT pathway may be a new therapeutic target.
Ying-Jun ZHANG ; Lan-Feng DONG ; Yong-Li WANG
Acta Pharmaceutica Sinica 2006;41(10):913-916
Animals
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Calcineurin
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genetics
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physiology
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Cardiomegaly
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genetics
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physiopathology
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prevention & control
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Cyclosporine
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pharmacology
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therapeutic use
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Gene Expression
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Humans
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Immunosuppressive Agents
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pharmacology
;
therapeutic use
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NFATC Transcription Factors
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genetics
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physiology
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Signal Transduction
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drug effects
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genetics
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physiology
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Tacrolimus
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pharmacology
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therapeutic use
4.Nuclear factor of activated T cells negatively regulates expression of the tumor necrosis factor receptor-related 2 gene in T cells.
Woon Ki KIM ; Ok Ju SUL ; Jung Sook KWAK ; Hye Young HUR ; Anne M LATOUR ; Beverly H KOLLER ; Byoung S KWON ; Choon Soo JEONG
Experimental & Molecular Medicine 2010;42(12):805-810
Tumor necrosis factor receptor-related 2 (TR2, HVEM or TNFRSF-14) plays an important role in immune responses, however, the mechanisms regulating its expression are unclear. To understand the control of TR2 gene expression, we studied the upstream region of the gene. Gel supershift assays revealed inducible binding of nuclear factor of activated T cells (NFAT) to a putative NFAT site within the TR2 promoter. Furthermore, cotransfection of a dominant negative NFAT construct, or siRNA for NFAT, resulted in increased expression of a TR2 reporter gene. Our findings demonstrate that NFAT negatively regulates TR2 expression in activated T cells.
Animals
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Base Sequence
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CD4-Positive T-Lymphocytes/metabolism
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Cells, Cultured
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Down-Regulation
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Mice
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Mice, Inbred C57BL
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Molecular Sequence Data
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NFATC Transcription Factors/*physiology
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Receptors, Tumor Necrosis Factor, Member 14/*biosynthesis
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T-Lymphocytes/*metabolism
5.Platinum nanoparticles reduce ovariectomy-induced bone loss by decreasing osteoclastogenesis.
Woon Ki KIM ; Jin Chun KIM ; Hyun Jung PARK ; Ok Joo SUL ; Mi Hyun LEE ; Ji Soon KIM ; Hye Seon CHOI
Experimental & Molecular Medicine 2012;44(7):432-439
Platinum nanoparticles (PtNP) exhibit remarkable antioxidant activity. There is growing evidence concerning a positive relationship between oxidative stress and bone loss, suggesting that PtNP could protect against bone loss by modulating oxidative stress. Intragastric administration of PtNP reduced ovariectomy (OVX)-induced bone loss with a decreased level of activity and number of osteoclast (OC) in vivo. PtNP inhibited OC formation by impairing the receptor activator of nuclear factor-kappaB ligand (RANKL) signaling. This impairment was due to a decreased activation of nuclear factor-kappaB and a reduced level of nuclear factor in activated T-cells, cytoplasmic 1 (NFAT2). PtNP lowered RANKL-induced long lasting reactive oxygen species as well as intracellular concentrations of Ca2+ oscillation. Our data clearly highlight the potential of PtNP for the amelioration of bone loss after estrogen deficiency by attenuated OC formation.
Animals
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Metal Nanoparticles/*administration & dosage
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Mice
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Mice, Inbred C57BL
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NFATC Transcription Factors/metabolism
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*Osteoclasts/drug effects/physiology
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Osteoporosis/drug therapy
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Ovariectomy/adverse effects
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Oxidative Stress/drug effects
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Platinum/*administration & dosage
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*RANK Ligand/genetics/metabolism
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Reactive Oxygen Species/metabolism
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Signal Transduction
6.NFAT2 is implicated in corticosterone-induced rat Leydig cell apoptosis.
Wei-Ran CHAI ; Qian WANG ; Hui-Bao GAO
Asian Journal of Andrology 2007;9(5):623-633
AIMTo investigate the activation of the nuclear factor of activated T cells (NFAT) and its function in the corticosterone (CORT)-induced apoptosis of rat Leydig cells.
METHODSNFAT in rat Leydig cells was detected by Western blotting and immunohistochemical staining. Cyclosporin A (CsA) was used to evaluate potential involvement of NFAT in the CORT-induced apoptosis of Leydig cells. Intracellular Ca(2+) was monitored in CORT-treated Leydig cells using Fluo-3/AM. After the Leydig cells were incubated with either CORT or CORT plus CsA for 12 h, the levels of NFAT2 in the nuclei and in the cytoplasm were measured by semi-quantitative Western blotting. The role of NFAT2 in CORT-induced Leydig cell apoptosis was further evaluated by observing the effects of NFAT2 overexpression and the inhibition of NFAT2 activation by CsA on FasL expression and apoptosis.
RESULTSWe found that NFAT2 was the predominant isoform in Leydig cells. CsA blocked the CORT-induced apoptosis of the Leydig cells. The intracellular Ca(2+) level in the Leydig cells was significantly increased after the CORT treatment. The CORT increased the level of NFAT2 in the nuclei and decreased its level in the cytoplasm. CsA blocked the CORT-induced nuclear translocation of NFAT2 in the Leydig cells. Both CORT-induced apoptosis and FasL expression in the rat Leydig cells were enhanced by the overexpression of NFAT2 and antagonized by CsA.
CONCLUSIONNFAT2 was activated in CORT-induced Leydig cell apoptosis. The effects of NFAT2 overexpression and the inhibition of NFAT2 activation suggest that NFAT2 may potentially play a pro-apoptotic role in CORT-induced Leydig cell apoptosis through the up-regulation of FasL.
Animals ; Apoptosis ; drug effects ; Calcium ; metabolism ; Cell Nucleus ; metabolism ; Corticosterone ; pharmacology ; Cytoplasm ; metabolism ; Immunohistochemistry ; Kinetics ; Leydig Cells ; cytology ; drug effects ; physiology ; Male ; NFATC Transcription Factors ; metabolism ; Rats ; Rats, Sprague-Dawley
7.Three transcription factors and the way immune cells affected by different plasma change in opposite ways in the development of the syndrome of pre-eclampsia.
Zhou LIANG ; Jing ZHU ; Yunfei WANG ; You WANG ; Yu ZHANG ; Jianhua LIN ; Wen DI ;
Chinese Medical Journal 2014;127(12):2252-2258
BACKGROUNDHow the transcriptional factors regulated the innate and adaptive immune system in pregnancy and pre-eclampsia are less understood. Nevertheless, what the plasma work in the development of this disease was not sure. The present study was design to evaluate what the transcriptional factors change in innate and adaptive immune system and what the plasma do in this filed.
METHODSPeripheral blood mononuclear cells (PBMC) from non-pregnant women (n = 18), women with clinically normal pregnancies (n = 23) and women with pre-eclampsia (n = 20) were separated from peripheral blood to isolate monocytes and T cells. The purity of monocytes and T cells were analysed by flow cytometry. Monocytes and T cells were stimulated in either lipopolysaccharides (LPS) or phorbol-myristate-acetate (PMA), respectively. Transcription Factor Arrays were used to screen the transcription factors of interest in comparing of different groups. PBMC were isolated from another 8 non-pregnant samples were co-incubated with different groups of plasma. Polymerase chain reaction (PCR) was performed using whole cell extractions of the samples.
RESULTSNuclear factor of activated T-cells-1 (NFAT-1), signal transducers and activators of transcription-1 (STAT-1) and activator protein-1 (AP-1) are up-regulated in monocytes in pregnancy and more so in pre-eclampsia. On the the contrary, NFAT-1, STAT-1 and AP-1 are down-regulated in T cells in pregnancy and more so in pre-eclampsia. A reduction was observed in interferon (IFN)-γ, interleukin (IL)-12 and IL-4 expression in T cells incubated with pre-eclamptic plasma. An elevation was observed in tumor necrosis factor (TNF)-α, IL-1 and IL-12 expression in monocytes incubated with pre-eclamptic plasma.
CONCLUSIONSInnate immunity is over activated and adaptive immunity is over suppressed in the development of pre-eclampsia. NFAT-1, STAT-1 and AP-1 might be the central transcription factors in the pathogenesis of pre-eclampsia. They induced some changes in plasma and "educate" the monocytes and T cells for relevant cytokine production. Successful completion of this study will enhance our understanding of pre-eclampsia and will discover new knowledge beyond pregnancy. The work will inform future therapies for the treatment of a wide range of condition such as transplantation immunology and a wide range of immune and inflammatory conditions.
Adult ; Female ; Humans ; Immunity, Innate ; physiology ; Interferon-gamma ; metabolism ; Interleukin-12 ; metabolism ; Interleukin-4 ; metabolism ; Leukocytes, Mononuclear ; metabolism ; Male ; NFATC Transcription Factors ; genetics ; metabolism ; Pre-Eclampsia ; immunology ; metabolism ; Pregnancy ; STAT1 Transcription Factor ; genetics ; metabolism ; Transcription Factor AP-1 ; genetics ; metabolism ; Transcription Factors ; genetics ; metabolism ; Young Adult
8.Relations of transcription expression of IL-2 with nuclear factor of activated T cells as well as changes of C-Fos and C-Jun after trauma.
Yan LUO ; Huaping LIANG ; Chengxiang HU ; Xiang XU ; Zhengguo WANG
Chinese Journal of Traumatology 2002;5(5):275-278
OBJECTIVETo observe the relations among expression of interleukin-2 (IL-2) in spleen lymphocytes, DNA binding activity of nuclear factor of activated T cells (NFAT) and expression of the partly family members C-Fos, C-Jun after trauma.
METHODSA murine closed trauma model was used, animals were sacrificed 6, 12 hours and 1, 4, 7, 10, 14 days, respectively after injury. Spleen lymphocytes were isolated from injured mice and stimulated with concanavalin-A. The culture supernatants were harvested and assayed for IL-2 activity. Total RNA was extracted from spleen lymphocytes and assayed for IL-2 mRNA. Nuclear protein was extracted, and the DNA binding activity of NFAT was measured using an electrophoretic mobility shift assay (EMSA), the expressions of C-Fos, C-Jun protein determined by Western blot analysis.
RESULTSThe expressions of IL-2 activity and IL-2 mRNA in spleen lymphocytes were decreased in injured mice compared with those in control mice, and the most obvious decrease appeared on the 4th day after injury. The DNA binding activity of NFAT decreased gradually and reached the minimum that was only 41% of the control on the 4th day after injury, which was closely associated with the decline of IL-2 activity and IL-2 mRNA. An decrease in the expression of C-Fos on the 1st and 4th day after injury, trauma had no significant effect on the C-Jun expression.
CONCLUSIONSThese results suggest that the inhibition of IL-2 expression is partly due to the impairment in the activation of NFAT in injured mice; and the decline in the DNA binding activity of NFAT is partly due to trauma block in the C-Fos expression.
Animals ; Blotting, Western ; DNA-Binding Proteins ; metabolism ; Electrophoretic Mobility Shift Assay ; Female ; Interleukin-2 ; metabolism ; Lymphocyte Activation ; physiology ; Male ; Mice ; NFATC Transcription Factors ; Nuclear Proteins ; Proto-Oncogene Proteins c-fos ; metabolism ; Proto-Oncogene Proteins c-jun ; metabolism ; Random Allocation ; T-Lymphocytes ; physiology ; Transcription Factors ; metabolism
9.Protein kinase C micron plays an essential role in hypertonicity-induced heat shock protein 70 expression.
Yun Sook LIM ; Jae Seon LEE ; Tai Qin HUANG ; Jeong Sun SEO
Experimental & Molecular Medicine 2008;40(6):596-606
Heat shock protein 70 (HSP70), which evidences important functions as a molecular chaperone and anti-apoptotic molecule, is substantially induced in cells exposed to a variety of stresses, including hypertonic stress, heavy metals, heat shock, and oxidative stress, and prevents cellular damage under these conditions. However, the molecular mechanism underlying the induction of HSP70 in response to hypertonicity has been characterized to a far lesser extent. In this study, we have investigated the cellular signaling pathway of HSP70 induction under hypertonic conditions. Initially, we applied a variety of kinase inhibitors to NIH3T3 cells that had been exposed to hypertonicity. The induction of HSP70 was suppressed specifically by treatment with protein kinase C (PKC) inhibitors (Go6976 and GF109203X). As hypertonicity dramatically increased the phosphorylation of PKC micron, we then evaluated the role of PKC micron in hypertonicity-induced HSP70 expression and cell viability. The depletion of PKC micron with siRNA or the inhibition of PKC micron activity with inhibitors resulted in a reduction in HSP70 induction and cell viability. Tonicity-responsive enhancer binding protein (TonEBP), a transcription factor for hypertonicity-induced HSP70 expression, was translocated rapidly into the nucleus and was modified gradually in the nucleus under hypertonic conditions. When we administered treatment with PKC inhibitors, the mobility shift of TonEBP was affected in the nucleus. However, PKC micron evidenced no subcellular co-localization with TonEBP during hypertonic exposure. From our results, we have concluded that PKC micron performs a critical function in hypertonicity-induced HSP70 induction, and finally cellular protection, via the indirect regulation of TonEBP modification.
Animals
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Carbazoles/pharmacology
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Cell Line
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Flavonoids/pharmacology
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HSP70 Heat-Shock Proteins/*biosynthesis
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Humans
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Indoles/pharmacology
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Isoquinolines/pharmacology
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MAP Kinase Signaling System/physiology
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Maleimides/pharmacology
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Mice
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NFATC Transcription Factors/metabolism
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Phosphorylation
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Promoter Regions, Genetic
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Protein Kinase C/antagonists & inhibitors/*physiology
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Protein Transport
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Saline Solution, Hypertonic/*pharmacology
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Signal Transduction
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Sulfonamides/pharmacology
10.Novel natural product therapeutics targeting both inflammation and cancer.
Jiangjiang QIN ; Wei WANG ; Ruiwen ZHANG
Chinese Journal of Natural Medicines (English Ed.) 2017;15(6):401-416
Inflammation is recently recognized as one of the hallmarks of human cancer. Chronic inflammatory response plays a critical role in cancer development, progression, metastasis, and resistance to chemotherapy. Conversely, the oncogenic aberrations also generate an inflammatory microenvironment, enabling the development and progression of cancer. The molecular mechanisms of action that are responsible for inflammatory cancer and cancer-associated inflammation are not fully understood due to the complex crosstalk between oncogenic and pro-inflammatory genes. However, molecular mediators that regulate both inflammation and cancer, such as NF-κB and STAT have been considered as promising targets for preventing and treating these diseases. Recent works have further demonstrated an important role of oncogenes (e.g., NFAT1, MDM2) and tumor suppressor genes (e.g., p53) in cancer-related inflammation. Natural products that target these molecular mediators have shown anticancer and anti-inflammatory activities in preclinical and clinical studies. Sesquiterpenoids (STs), a class of novel plant-derived secondary metabolites have attracted great interest in recent years because of their diversity in chemical structures and pharmacological activities. At present, we and other investigators have found that dimeric sesquiterpenoids (DSTs) may exert enhanced activity and binding affinity to molecular targets due to the increased number of alkylating centers and improved conformational flexibility and lipophilicity. Here, we focus our discussion on the activities and mechanisms of action of STs and DSTs in treating inflammation and cancer as well as their structure-activity relationships.
Animals
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Anti-Inflammatory Agents
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pharmacology
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Antineoplastic Agents, Phytogenic
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pharmacology
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Humans
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Inflammation
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drug therapy
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etiology
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NF-kappa B
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antagonists & inhibitors
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NFATC Transcription Factors
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antagonists & inhibitors
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Neoplasms
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drug therapy
;
etiology
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Proto-Oncogene Proteins c-mdm2
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antagonists & inhibitors
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physiology
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Sesquiterpenes
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chemistry
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pharmacology
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Structure-Activity Relationship