• Journal of Life Science
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• v.10 no.5
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• pp.456-466
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• 2000

Aquaperin 4 (AQP4) is the mercurial water channel expressed abundantly in brain, especially the region related with cerebrospinal fluid reabsorption and osmoregulation. The primary structure of AQP4 water channel was elucidated but the molecular mechanism of AQP4 channel regulation is still unknown. To investigate the possible regulation of AQP4 water channel by phosphorylation via various protein kinases, osmotic water permeability of AQP4 expressed in Xenopus oocytes was measured by videomicroscopy technique. Forskolin (10 $\mu$M) did not affect osmotic water permeability of oocytes injected with AQP4 cRNA, excluding the regulation of AQP4 water cnannel by protein kinase A. Osmotic water permeability (P아래첨자) of AQP4-expressed oocytes was ingibited by the pretreatmeat of BAPTA/AM (up to 500$\mu$M), an intracellular Ca윗첨자 chelator, and calmidazolium (100$\mu$M), a specific Ca윗첨자/calmodulin antagonist, in a dose-dependent manner. The inhibition of osmotic water permeability (P아래첨자) by the calmidazolium treatment was completely reversed by the addition of calyculin A (0.1$\mu$M), a nonspecific phosphatase inhibitor. Phorbol 12-myristate 13-acetate (PMA), a protein kinase C activator, had biphasic effects on osmotic water permeability in AQP4 cRNA injected oocytes depending on its concentration; 21% increase by 100 nM PMA, 35% decrease by 1$\mu$M PMA. These effects were reversed with 2$\mu$M staurosporine, a nonspecific PKC inhibitor. These results suggest that phosphorylation of AQP4 water channel by Ca윗첨자/calmodulin kinase and protein kinase C might regulate the osmotic water permeability.

• Natural Product Sciences
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• v.21 no.1
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• pp.59-65
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• 2015

In this study, we investigated whether cynaroside, cynarin and linarin derived from Chrysanthemum indicum L. affect the secretion, production and gene expression of MUC5AC mucin in airway epithelial cells. Confluent NCI-H292 cells were pretreated with cynaroside, cynarin or linarin for 30 min and then stimulated with PMA (phorbol 12-myristate 13-acetate) for 24 h. The MUC5AC mucin gene expression, mucin protein production and secretion were measured by RT-PCR and ELISA, respectively. Effect of linarin on EGF (epidermal growth factor) - or TNF-${\alpha}$ (tumor necrosis factor-${\alpha}$)-induced MUC5AC mucin gene expression and mucin protein production was also examined. The results were as follows: (1) Cynaroside and cynarin did not significantly affect PMA-induced MUC5AC mucin secretion from NCI-H292 cells. However, linarin decreased MUC5AC mucin secretion; (2) Cynaroside did not affect PMA-induced MUC5AC mucin production and gene expresion from NCI-H292 cells. However, cynarin and linarin inhibited the production and gene expression of MUC5AC mucin; (3) Linarin also inhibited the production and gene expression of MUC5AC mucin induced by EGF- or TNF-${\alpha}$ from NCI-H292 cells. These results suggest that linarin can regulate the gene expression, production and secretion of mucin, by directly acting on airway epithelial cells.

• The Journal of Korean Medicine
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• v.21 no.4
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• pp.193-203
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• 2000

Objectives : Hindered barrier function of vascular endothelium has been implicated in the initiation and progression of degenerative vascular diseases such as atherosclerosis. In this study, the effect of Sunghyangchungisan(SHCS) as a protectant against oxidant-induced destruction of endothelial barrier function was assessed. Methods : Toward this end, endothelial cells derived from the human umbilical vein were cultured as monolayers on permeable membrane filters. Endothelial permeability was monitored by measuring transendothelial electrical resistance and movement of low density lipoprotein (LDL) across the endothelial monolayer. Results : Along with increased movement of LDL, $H_2O_2$-induced increase in endothelial permeability was paralleled by a decrease in transendotheliaI electrical resistance. The effect of $H_2O_2$ was mimicked by phorbol 12-myristate 13-acetate (PMA), a potent activator of proteinkinase C. Calphostin-C, a protein kinase C inhibitor, effectively blocked the increase in endothelial permeability induced by $H_2O_2$ or PMA, indicating that activation of protein kinase C is associated with the $H_2O_2-induced$ permeability change. SHCS effectively protected the endothelial monolayer against $H_2O_2-induced$ increase in permeability, whereas, it did not affect PMA-induced change. Forskolin, a potent activator of adenylyl cyclase, antagonized $H_2O_2$ to increase endothelial permeability. In addition, in ${H_2O_2}-treated$ cens, intracenular cAMP concentration was significantly decreased, indicating that impaired cAMP production as well as activation of proteinkinase C is a mechanism underlying ${H_2O_2}>-induced$$H_2O_2$ with regard to its effect on intracellular cAMP content. However, SHCS itself did not affect resting cAMP concentration in endothelial cells. Conclusions : These results suggest that SHCS might operate as an effective protectant against oxidant-induced destruction of endothelial barrier function. The mechanism does not appear to involve direct interaction with protein kinase C- or cAMP-associated signaling mechanism.

• International Journal of Oral Biology
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• v.38 no.1
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• pp.37-42
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• 2013

Receptor activator of NF-${\kappa}B$ ligand (RANKL) is an essential cytokine for osteoclast differentiation, activation and survival. T lymphocytes such as $T_{17}$ cells, a subset of T helper cells that produce IL-17, play an important role in rheumatoid arthritic bone resorption by producing inflammatory cytokines and RANKL. It has not yet been clearly elucidated how T cell activation induces RANKL expression. T cell receptor activation induces the activation of nuclear factor of activated T cell (NFAT) and expression of its target genes. In this study, we examined the role of NFAT in T cell activation-induced RANKL expression. EL-4, a murine T lymphocytic cell line, was used. When T cell activation was induced by phorbol 12-myristate 13-acetate (PMA) and ionomycin, RANKL expression increased in a time-dependent manner. In the presence of cyclosporin, an inhibitor of NFAT activation, this PMA/ionomycin-induced RANKL expression was blocked. Overexpression of either NFATc1 or NFATc3 induced RANKL expression. Chromatin immunoprecipitation results demonstrated that PMA/ionomycin treatment induced the binding of NFATc1 and NFATc3 to the mouse RANKL gene promoter. These results suggest that NFATc1 and NFATc3 mediates T cell receptor activation-induced RANKL expression in T lymphocytes.

• Journal of Physiology & Pathology in Korean Medicine
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• v.28 no.1
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• pp.45-52
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• 2014

Hoesaeng-san is known to be effective for treating diarrhea and vomiting. However the therapeutic mechanism of Hoesaeng-san is still not well understood. The aim of the present study was to demonstrate the effects of Hoesaeng-san ethanol extract (HSSEE) on the expression of pro-inflammatory cytokines, as well as to elucidate its mechanism of action in the human mast cell line (HMC-1). Mast Cell were stimulated with phorbol 12-myristate 13-acetate (PMA) plus A23187 in the presence or absence of HSSEE. To study the possible effects of HSSEE, ELISA, RT-PCR, Western blot analysis were used in this study. HSSEE significantly inhibited the PMA plus A23187-induction of inflammatory cytokines such as tumor necrosis factor (TNF)-${\alpha}$, interleukin (IL)-6 and IL-8. In activated HMC-1 cells, phosphorylation of extra-signal response kinase (ERK) 1/2 and c-jun n-terminal kinase (JNK)1/2 decreased after treatment with HSSEE. Moreover HSSEE inhibited PMA plus A23187-induced nuclear factor (NF)-${\kappa}B$ activation and $I{\kappa}B$ degradation. HSSEE suppressed the expression of TNF-${\alpha}$, IL-6, IL-8 through a decrease in the ERK 1/2 and JNK 1/2, as well as activation of NF-${\kappa}B$. These results indicated that HSSEE exerted a regulatory effect on inflammatory reactions mediated by mast cells.

• Journal of Physiology & Pathology in Korean Medicine
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• v.22 no.2
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• pp.377-384
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• 2008

The root of Vitis labrusca, is used as a source of health promoting drug in Korean traditional medicine. It has been reported that root of Vitis labrusca has antioxidant, anti lipid peroxidation and anti-reactive nitrogen species (RNS) activities. The aim of this study was to elucidate the molecular changes of apoptotic signaling pathways in phorbol 12-myristate 13 acetate (PMA)-induced human hepatocellular carcinoma cell line (Hep G2). The root of Vitis labrusca, ethanol extract (RVLEE) was tested for cell viability on Hep G2 cell using the MTT assay. RVLEE exhibited weak cytotoxic activity. However, treatment of Hep G2 cells with RVLEE suppressed PMA-induced cell proliferation. Also, dramatic changes of cell death signals in cellular molecules such as Chk2/Cds1, CIDE-B, CLIMP-63, Bax, Bcl-xL, C-myc, Bcl-2, Bric-5, NIP-3, TRAF2 and BAR but not CIDE-B and DR4. Futhermore, our results showed that the treatment of Hep G2 cells with 25 and $50\; {\mu}g/ml$ of RVLEE suppressed PMA-induced COX-2 gene activity. These data suggest that RVLEE have inhibitory effect of cell proliferation, induction of apoptosis and, thus, may offer therapeutic potential in Hep G2.

• Journal of Microbiology and Biotechnology
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• v.32 no.1
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• pp.81-90
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• 2022

Peucedanum japonicum Thunberg (PJT) has been used in traditional medicine to treat colds, coughs, fevers, and other inflammatory diseases. The goal of this study was to investigate whether 3'-isovaleryl-4'-senecioylkhellactone (IVSK) from PJT has anti-inflammatory effects on lung epithelial cells. The anti-inflammatory effects of IVSK were evaluated using phorbol 12-myristate 13-acetate (PMA)-stimulated A549 cells and regular human lung epithelial cells as a reference. IVSK reduced the secretion of the inflammatory mediators interleukin (IL)-8 and monocyte chemoattractant protein-1 (MCP-1), and the mRNA expression of IL-6, IL-8, MCP-1, and IL-1β. Additionally, it inhibited the phosphorylation of IκB kinase (IKK), p65, Iκ-Bα, and mitogen-activated protein kinases (MAPKs) p38, JNK, and ERK in A549 cells stimulated with PMA. Moreover, the binding affinity of activator protein-1 (AP-1) and nuclear factor-κB (NF-κB) was significantly reduced in the luciferase assay, while nuclear translocation was markedly inhibited by IVSK in the immunocytochemistry. These findings indicate that IVSK can protect against inflammation through the AP-1 and NF-κB pathway and could possibly be used as a lead compound for the treatment of inflammatory lung diseases.

• Molecules and Cells
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• v.24 no.1
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• pp.9-15
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• 2007

To investigate the effects of chitosan on the redifferentiation of dedifferentiated chondrocytes, we used chondrocytes obtained from a micromass culture system. Micromass cultures of chick wing bud mesenchymal cells yielded differentiated chondrocytes, but these dedifferentiated during serial monolayer subculture. When the dedifferentiated chondrocytes were cultured on chitosan membranes they regained the phenotype of differentiated chondrocytes. Expression of protein kinase $C{\alpha}$ ($PKC{\alpha}$) increased during chondrogenesis, decreased during dedifferentiation, and increased again during redifferentiation. Treatment of the cultures with phorbol 12-myristate 13-acetate (PMA) inhibited redifferentiation and down-regulated $PKC{\alpha}$. In addition, the expression of p38 mitogen-activated protein (MAP) kinase increased during redifferentiation, and its inhibition suppressed redifferentiation. These findings establish a culture system for producing chondrocytes, point to a new role of chitosan in the redifferentiation of dedifferentiated chondrocytes, and show that $PKC{\alpha}$ and p38 MAP kinase activities are required for chondrocyte redifferentiation in this model system.

• Biomolecules & Therapeutics
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• v.17 no.1
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• pp.79-85
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• 2009

Polyphenolic compounds are reported to have various pharmacological activities such as anti-oxidative, anti-cancerous, anti-inflammatory and anti-aging effects. Although numerous papers explore their functional roles in many different cellular actions, not many studies handle their structural features in anti-inflammatory responses. In this study, therefore, we examined structural role of substituted transstilbenes in their NO inhibitory and NF-${\kappa}B$ suppressive activities. Of 10 compounds tested, 4 compounds (cinnamic acid, resveratrol, piceatannol and curcumin) displayed NO inhibitory activities in a dose-dependent manner. Similarly, these compounds blocked LPS-induced cytotoxicity of RAW264.7 cells. All NO inhibitory compounds also inhibited $I{\kappa}B{\alpha}$ phosphorylation, a hallmark for NF-${\kappa}B$ activation. However, these inhibitory compounds exhibited distinct suppressive pattern in tumor necrosis factor (TNF)-${\alpha}$- or phorbol-12-myristate-13-acetate (PMA)-induced NF-${\kappa}B$ and AP-1 activation. According to structure-activity relationship study, polarity and size of ring B seem to be important for diminishing NO production. Therefore, our data suggest that substituted trans-stilbenes can be developed as novel anti-inflammatory drug or further developed as lead compounds for another improvement.

• Archives of Pharmacal Research
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• v.18 no.4
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• pp.262-266
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• 1995

Bacterial lipopolysaccharide (LPS) is one of the most potent inducers of various cytokines nad other proinflammatory mediators in macrophages. Although pathophysiological consequences of LPS-induced responses are well established, the mechanisms through which LPS-generated singals are transduced remain unclear. In the present study, we attempted to determine early intracellular events after LPS binding which transduced the signal for the induction of arachidonic acid metabolism in rat alveolar macrophages. While H-7, a protein kinase C(PKC) inhibitor, did not affect LPS-stimulated prostaglandin synthesis, staurosporine enhanced archidonic acid etabolism in macropahages treated with LPS. Phorbol-12-myristate-13 acetate snesitive to LPS compare with control group. PMA and H-7 did not alter the effect of flucose. Pertussis toxin did not show nay effect, thus pertussis toxin snesitive G-protein pathway appears not to play a role in this experimental system. Genistein and tyrphostin 25, protein tyrosine kinase 9PTK) inhibitors, markedly inhibited prostaglandin synthesis in macrophages nal transduction events leading to icnreased macrophage arachidonic acid metabolism.