• IMMUNE NETWORK
• /
• v.6 no.4
• /
• pp.204-210
• /
• 2006

Background: Nitric oxide (NO) in articular chondrocytes regulates dedifferentiation and inflammatory responses by modulating MAP kinases. In this study, we investigated whether the Src kinase in chondrocytes regulates NO-induced dedifferentiation and cyclooxygenase-2 (COX-2) expression. Methods: Primary chondrocytes were treated with various concentrations of SNP for 24 h. The COX-2 and type II collagen expression levels were determined by immunoblot analysis, and prostaglandin $E_2\;(PGE_2)$ was determined by using a $PGE_2$ assay kit. Expression and distribution of p-Caveolin and COX-2 in rabbit articular chondrocytes and cartilage explants were determined by immunohistochemical staining and immunocytochemical staining, respectively. Results: SNP treatment stimulated Src kinase activation in a dose-dependent manner in articular chondrocytes. The Src kinase inhibitors PP2 [4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo(3,4-d)pyrimidine], a significantly blocked SNP-induced p38 kinase and caveolin-1 activation in a dose-dependent manner. Therefore, to determine whether Src kinase activation is associated with dedifferentiation and/or COX-2 expression and $PGE_2$ production. As expected, PP2 potentiated SNP-stimulated dedifferentiation, but completely blocked both COX-2 expression and $PGE_2$ production. And also, levels of p-Caveolin and COX-2 protein expression were increased in SNP-treated primary chondrocytes and osteoarthritic and rheumatoid arthritic cartilage, suggesting that p-Caveolin may playa role in the inflammatory responses of arthritic cartilage. Conclusion: Our previously studies indicated that NO caused dedifferentiation and COX-2 expression is regulated by p38 kinase through caveolin-1 (1). Therefore, our results collectively suggest that Src kinase regulates NO-induced dedifferentiation and COX-2 expression in chondrocytes via p38 kinase in association with caveolin-1.

• Biomedical Science Letters
• /
• v.15 no.1
• /
• pp.67-72
• /
• 2009

Microtubule-interfering agents (MIAs), including paclitaxel, have been attributed in part to interference with microtubule assembly, impairment of mitosis, and changes in cytoskeleton. But the signaling mechanisms that link microtubule disarray to destructive or protective cellular responses are poorly understood. This study investigated the effect of paclitaxel on differentiation such as type II collagen expression and sulfated proteoglycan accumulation in rabbit articular chondrocytes. Paclitaxel caused differentiated chondrocyte phenotype as demonstrated by increment of type II collagen expression and proteoglycan synthesis Paclitaxel treatment stimulated activation of ERK-1/2 and p38 kinase. Inhibition of ERK-1/2 with PD98059 enhanced paclitaxel-induced differentiation, whereas inhibition of p38 kinase with SB203580 suppressed paclitaxel-induced differentiation. Our findings suggest that ERK-1/2 and p38 kinase oppositely regulate paclitaxel-induced differentiation in chondrocytes.

• Proceedings of the Korean Society of Toxicology Conference
• /
• 2001.10a
• /
• pp.29-30
• /
• 2001

MAP kinase/ AP-1 signal transduction components are rapidly initiated by many extracellular stimuli, especially environmental carcinogens. We have investigated the role of MAP kinases (Erks, JNKs, and p38 kinases) and AP-1 signal transduction pathways in the process of cell transformation and carcinogenesis. Incubation of Cl 41 cells with tumor promoters such as TPA, EGF, arsenic, or TNF-$\alpha$ led to cell transformation and activation of MAP kinases.(omitted)

• Proceedings of the Korea Environmental Mutagen Society Conference
• /
• 2002.11a
• /
• pp.14-40
• /
• 2002

15-Deoxy-${\Delta}^{12, 14}$-prostaglandin $J_2$ (15-deoxy-$PGJ_2$), a naturally occurring ligand activates the peroxisome proliferator-activated $receptor-{\gamma}(PPAR-{\gamma}$). Activation of $PPAR-{\gamma}$ has been found to induce cell differentiation such as adipose cell and macrophage. Here it was investigated whether 15-deoxy-$PGJ_2$ has neuronal cell differentiation and possible underlying molecular mechanisms. Dopaminergic differentiating PC 12 cells treated with 15-deoxy-$PGJ_2$ (0.2 to 1.6 ${\mu}M$) alone showed measurable neurite extension and expression of neurofilament, markers of cell differentiation. However much greater extent of neurite extension and expression of neurofilament was observed in the presence of NGF (50 ng/ml). In parallel with its increasing effect on the neurite extension and expression of neurofilament, 15-deoxy-$PGJ_2$ enhanced NGF-induced p38 MAP kinase expression and its phosphorylation in addition to the activation of transcription factor AP-1 in a dose dependent manner. Moreover, pretreatment of SD 203580, a specific inhibitor of p38 MAP kinase inhibited the promoting effect of 15-deoxy-$PGJ_2$(0.8 ${\mu}M$) on NGF-induced neurite extension. This inhibition correlated well with the ability of SB203580 to inhibit the enhancing effect of 15-deoxy-$PGJ_2$ on the expression of p38 MAP kinase and activation of AP-1, The promoting ability of 15-deoxy-$PGJ_2$ did not occur through $PPAR-{\gamma}$, as synthetic PPAR-${\gamma}$ agonist andantagonist did not change the neurite promoting effect of 15-deoxy-PGJ$_2$. In addition, contrast to other cells (embryonic midbrain and SK-N-MC cells), $PPAR-{\gamma}$ was not expressed in PC-12 cells. Other structure related prostaglandins, PGD$_2$ and $PGE_2$ acting via a cell surface G-protein-coupled receptor (GPCR) did not increase basal or NGF-induced neurite extension. Moreover, GPCR (EP and DP receptor) antagonists did not alter the promoting effect of f 5-deoxy-$PGJ_2$ on neurite extension and activation of p38 MAP kinase, suggesting that the promoting effect of 15-deoxy-$PGJ_2$ may not be mediated GPCR. These data demonstrate that activation of p38 MAP kinase in conjunction with AP-1 single pathway may be important in the promoting activity of 15-deoxy-$PGJ_2$ cells.

• Proceedings of the Korean Society of Toxicology Conference
• /
• 2002.11b
• /
• pp.14-40
• /
• 2002

15-Deoxy- Δ$\^$12,14/-prostaglandin J$_2$ (15-deoxy-PGJ$_2$), a naturally occurring ligand activates the peroxisome proliferator-activated receptor-${\gamma}$ (PPAR-${\gamma}$). Activation of PPAR-y has been found to induce cell differentiation such as adipose cell and macrophage. Here it was investigated whether 15-deoxy-PGJ$_2$ has neuronal cell differentiation and possible underlying molecular mechanisms. Dopaminergic differentiating PC 12 cells treated with 15-deoxy-PGJ$_2$ (0.2 to 1.6 ${\mu}$M) alone showed measurable neurite extension and expression of neurofilament, markers of cell differentiation. However much greater extent of neurite extension and expression of neurofilament was observed in the presence of NGF (50 ng/$m\ell$). In parallel with its increasing effect on the neurite extension and expression of neurofilament, 15-deoxy-PGJ$_2$ enhanced NGF-induced p38 MAP kinase expression and its phosphorylation in addition to the activation of transcription factor AP-1 in a dose dependent manner. Moreover, pretreatment of SD 203580, a specific inhibitor of p38 MAP kinase inhibited the promoting effect of 15-deoxy-PGJ$_2$ (0.8 ${\mu}$M) on NGF-induced neurite extension. This inhibition correlated well with the ability of SB203580 to inhibit the enhancing effect of 15-deoxy-PGJ$_2$ on the expression of p38 MAP kinase and activation of AP-1. The promoting ability of 15-deoxy-PGJ$_2$ did not occur through PPAR-${\gamma}$, as synthetic PPAR-${\gamma}$ agonist and antagonist did not change the neurite promoting effect of 15-deoxy-PGJ$_2$. In addition, contrast to other cells (embryonic midbrain and SK-N-MC cells), PPAR-${\gamma}$ was not expressed in PC-12 cells. Other structure related prostaglandins, PGD$_2$ and PGE$_2$ acting via a cell surface G-protein-coupled receptor (GPCR) did not increase basal or NGF-induced neurite extension. Moreover, GPCR (EP and DP receptor) antagonists did not alter the promoting effect of 15-deoxy-PGJ$_2$ on neurite extension and activation of p38 MAP kinase, suggesting that the promoting effect of 15-deoxy-PGJ$_2$ may not be mediated GPCR. These data demonstrate that activation of p38 MAP kinase in conjunction with AP-1 signal pathway may be important in the promoting activity of 15-deoxy-PGJ$_2$ on the differentiation of PC12 cells.

• Journal of Physiology & Pathology in Korean Medicine
• /
• v.21 no.4
• /
• pp.1017-1024
• /
• 2007

Thymus and activation-regulated chemokine (TARC/CCL-17), produced by keratinocytes, is a CC chemokine known to selectively Th2 type T cells via $CCR4^+$ and is implicated in the development of atopic dermatitis (AD). TARC/CCL17 expression was induced by cytokines such as tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$) and interferon-${\gamma}$ (IFN-${\gamma}$). We recently found that the wogonin, a flavone isolated from Scutellaria baicalensis, suppressed TARC expression via heme oxygenase 1 (HO1) in human keratinocytes induced with mite antigen. However, little is known about the inhibitory mechanism of wogonin on TARC/CCL-17 expression stimulated with cytokines. To investigate the inhibitory mechanism, I determined the inhibitory effects of wogonin on the activation of nuclear factor-${\kappa}B$ (NF-${\kappa}B$) and $I{\kappa}B{\alpha}$ phosphorylation, and also examined the activation of p38 MAP kainase in HaCaT keratinocytes stimulated with TNF-${\alpha}$ and IFN-${\gamma}$. Wogonin inhibited NF-${\kappa}B$-DNA complex, NF-${\kappa}B$ binding activity, and the phosphorylation of $I{\kappa}B{\alpha}$ in a dose dependent manner. Wogonin also inhibited the translocation of NF-${\kappa}B$ from cytosol to nucleus. Moreover, the phosphorylation of of p38 MAP kinase in the TNF-${\alpha}$ and IFN-${\gamma}$-stimulated HaCaT keratinocytes were suppressed by wogonin in a dose dependent manner. These results suggest that wogonin may inhibit cytokine-induced NF-${\kappa}B$ activation by $I{\kappa}B{\alpha}$ degradation via suppression of p38 MAP kinase signaling pathway in keratinocytes and modulation of wogonin signaling pathway may be beneficial for the treatment of AD.

• Journal of Photoscience
• /
• v.9 no.2
• /
• pp.485-487
• /
• 2002

Cells receive signals for survival as well as death, and the balance between the two ultimately determines the fate of the cells. UV-triggered apoptotic signaling has been well documented, whereas UV-induced survival effects have received little attention. We have reported previously that UVB irradiation prevented apoptosis, which was partly dependent on activation of the phosphatidylinositol 3-kinase (PI3-kinase)/ Akt pathway. In this study, anti-apoptotic effects of UV with different wavelength ranges, UVA, UVB and UVC, were examined. NIH3T3 cells showed apoptotic cell death by detachment from the extracellular matrix under serum-free conditions, which was prevented by all wavelengths. However, the effect of UVA was less than those of UVB and UVC. Reduction of mitochondrial transmembrane potential and activation of caspase-9 and -3 were suppressed by all three wavelengths of UV, showing wavelength-dependent effects as mentioned above. The PI3-kinase inhibitor wortmannin partially inhibittrl the UVB and UVC-induced suppression of apoptosis, but not the inhibitoty effect of UVA. The Akt phosphotylation by UVB and UVC was completely inhibittrl by addition of wortmannin, but that by UVA was not P38 MAP kinase inhibitor SB203580 partially inhibited the UVB and UVC-induced suppression of apoptosis and Akt phosphotylation, and completely inhibited UVA-induced those. These results suggested the existence of two different survival pathways leading to suppression of apoptosis, one for UVA that is independent of the PI3-kinase/Akt pathway and dependent on p38 MAP kinase, and the other for UVB and UVC that is dependent on both pathways.

• The Korean Journal of Physiology and Pharmacology
• /
• v.6 no.6
• /
• pp.319-325
• /
• 2002

The induction of interleukin-6 (IL-6) using combined proinflammatory agents $(LPS/IFN-{\gamma}\;or\;TNF-{\alpha}/IFN-{\gamma})$ was studied in relation to p38 mitogen-activated protein kinase (MAPK) and $NF-{\kappa}B$ transcriptional factor in primary neonatal cardiomyocytes. When added to cultures of cardiomyocytes, the combined agents $(LPS/IFN-[\gamma}\;or\;TNF-{\alpha}/IFN-{\gamma})$ had stimulatory effect on the production of IL-6 and the elevation was significantly reduced by SB203580, a specific p38 MAPK inhibitor. SB203580 inhibited protein production and gene expression of IL-6 in a concentration-dependent manner. In this study, $IFN-{\gamma}$ enhancement of $TNF-{\alpha}-induced\;NF-{\kappa}B$ binding affinity as well as p38 MAP kinase activation was observed. However, a specific inhibitor of p38 MAPK, SB203580, had no effect on $TNF-{\alpha}/IFN-{\gamma}\;or\;LPS/IFN-{\gamma}-induced\;NF-{\kappa}B$ activation. This study strongly suggests that these pathways about $TNF-{\alpha}/IFN-{\gamma}$ or $LPS/IFN-{\gamma}-activated$ IL-6 release can be primarily dissociated in primary neonatal cardiomyocytes.

• BMB Reports
• /
• v.35 no.4
• /
• pp.377-383
• /
• 2002

Arsenic trioxide ($As_O_3$) was recently demonstrated to be an effective inducer of apoptosis in patients with relapsed acute promyelocytic leukemia (APL) as well as patients with APL in whom all-trans-retinoic acid and conventional chemotherapy failed. Chronic myelogenous leukemia cells are highly resistant to chemotherapeutic drugs. To determine if $As_O_3$ might be useful for the treatment of chronic myelogenous leukemia, we examined the ability of $As_O_3$ to induce apoptosis in K562 cells. In vitro cytotoxicity of $As_O_3$ was evaluated in K562 cells by a MTT assay: the $IC_50$ value for $As_O_3$ was determined to be $10\;{\mu}m$. When analyzed by agarose gel electorphoresis, the DNA fragments became evident after incubation of the cells with $20\;{\mu}m$ $As_O_3$ for 24 h. We also found morphological changes and chromatin condensation of the cells undergoing apoptosis. Activation of caspase-3 was observed 6 h after treatment with $20\;{\mu}m$ $As_O_3$ by a Western blot analysis. Next, we examined the MAP kinase-signaling pathway of $As_O_3$-induced apoptosis in K562 cells. $As_O_3$ at $10\;{\mu}m$ strongly induced the activation of p38, inhibited $As_O_3$ induced apoptotic cell death. These results suggest that $As_O_3$ is able to induce the apoptotic activity in K562 cells, and its apoptotic mechanism may be associated with the activation of p38.

• Proceedings of the Korean Society of Toxicology Conference
• /
• 2005.05a
• /
• pp.160-160
• /
• 2005