• Molecules and Cells
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• v.23 no.1
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• pp.30-38
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• 2007

Dipyrithione (2, 2'-dithiobispyridine-1, 1'-dioxide, PTS2), a pyrithione derivate, is highly bactericidal and fungicidal. In this study we examined its apoptotic effect on HeLa cells. PTS2 induced HeLa cell death in a dose and time dependent manner. ERK1/2 and p38 were markedly activated, but little JNK1/2 activation was detected. Suppression of p38 activation by SB203580 reduced the extent of apoptosis of the HeLa cells and also prevented induction of p21, release of cytochrome c, and cleavage of caspase-3 and PARP. Inhibition of ERK1/2 with PD98059 increased apoptosis, indicating that ERK1/2 activation has an anti-apoptotic effect on PTS2-induced HeLa cell apoptosis. PTS2 also inhibited murine sarcoma 180 and hepatoma 22 tumor growth in an animal tumor model. Our findings indicate that PTS2 possesses anti-tumor activity, that caspase-3 and poly (ADP-ribose) polymerase (PARP) are involved in PTS2-induced HeLa cell apoptosis and that ERK1/2 and p38 have opposing effects on this apoptosis.

• Molecules and Cells
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• v.25 no.4
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• pp.479-486
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• 2008

Mitogen-activated protein kinases (MAPKs) play an indispensable role in activation of the myogenic program, which is responsive to mechanical stimulation. Although there is accumulating evidence of mechanical force-mediated cellular responses, the role of MAPK in regulating the myogenic process in myoblasts exposed to cyclic stretch is unclear. Cyclic stretch induced the proliferation of C2C12 myoblasts and inhibited their differentiation into myotubes. In particular, it induced persistent phosphorylation of p38 kinase, and decreased the level of phosphorylation of extracellular-signal regulated kinase (ERK). Partial inhibition of p38 phosphorylation increased cellular levels of MyoD and p-ERK in stretched C2C12 cells, along with increased myotube formation. Treatment with $10{\mu}M$ PD98059 prevented myogenin expression in response to a low dose of SB203580 ($3{\mu}M$) in the stretched cells, suggesting that adequate ERK activation is also needed to allow the cells to differentiate into myotubes. These results suggest that cyclic stretch inhibits the myogenic differentiation of C2C12 cells by activating p38-mediated signaling and inhibiting ERK phosphorylation. We conclude that p38 kinase, not ERK, is the upstream signal transducer regulating cellular responses to mechanical stretch in skeletal muscle cells.

• Molecules and Cells
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• v.39 no.10
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• pp.742-749
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• 2016

The cancer chemo-preventive effects of equol have been demonstrated for a wide variety of experimental tumours. In a previous study, we found that equol inhibited proliferation and induced apoptotic death of human gastric cancer MGC-803 cells. However, the mechanisms underlying equol-mediated apoptosis have not been well understood. In the present study, the dual AO (acridine orange)/EB (ethidium bromide) fluorescent assay, the comet assay, MTS, western blotting and flow cytometric assays were performed to further investigate the pro-apoptotic effect of equol and its associated mechanisms in MGC-803 cells. The results demonstrated that equol induced an apoptotic nuclear morphology revealed by AO/EB staining, the presence of a comet tail, the cleavage of caspase-3 and PARP and the depletion of cIAP1, indicating its pro-apoptotic effect. In addition, equol-induced apoptosis involves the mitochondria-dependent cell-death pathway, evidenced by the depolarization of the mitochondrial membrane potential, the cleavage of caspase-9 and the depletion of Bcl-xL and full-length Bid. Moreover, treating MGC-803 cells with equol induced the sustained activation of extracellular signal-regulated kinase (ERK), and inhibiting ERK by U0126, a MEK/ERK pathway inhibitor, significantly attenuated the equol-induced cell apoptosis. These results suggest that equol induces mitochondria-dependent apoptosis in human gastric cancer MGC-803 cells via the sustained activation of the ERK1/2 pathway. Therefore, equol may be a novel candidate for the chemoprevention and therapy of gastric cancer.

• Molecules and Cells
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• v.41 no.5
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• pp.436-443
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• 2018

The actin cytoskeleton plays a key role in the entry of mitosis as well as in cytokinesis. In a previous study, we showed that actin disruption delays mitotic entry at G2/M by sustained activation of extracellular signal-related kinase 1/2 (ERK1/2) in primary cells but not in transformed cancer cell lines. Here, we examined the mechanism of cell cycle delay at G2/M by actin dysfunction in IMR-90 normal human fibroblasts. We observed that de-polymerization of actin with cytochalasin D (CD) constitutively activated ribosomal S6 kinase (RSK) and induced inhibitory phosphorylation of Cdc2 (Tyr 15) in IMR-90 cells. In the presence of an actin defect in IMR-90 cells, activating phosphorylation of Wee1 kinase (Ser 642) and inhibitory phosphorylation of Cdc25C (Ser 216) was also maintained. However, when kinase-dead RSK (DN-RSK) was overexpressed, we observed sustained activation of ERK1/2, but no delay in the G2/M transition, demonstrating that RSK functions downstream of ERK in cell cycle delay by actin dysfunction. In DN-RSK overexpressing IMR-90 cells treated with CD, phosphorylation of Cdc25C (Ser 216) was blocked and phosphorylation of Cdc2 (Tyr 15) was decreased, but the phosphorylation of Wee1 (Ser 642) was maintained, demonstrating that RSK directly controls phosphorylation of Cdc25C (Ser 216), but not the activity of Wee1. These results strongly suggest that actin dysfunction in primary cells activates ERK1/2 to inhibit Cdc2, delaying the cell cycle at G2/M by activating downstream RSK, which phosphorylates and blocks Cdc25C, and by directly activating Wee1.

• The Korean Journal of Physiology and Pharmacology
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• v.25 no.6
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• pp.593-601
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• 2021

Primary cilia on kidney tubular cells play crucial roles in maintaining structure and physiological function. Emerging evidence indicates that the absence of primary cilia, and their length, are associated with kidney diseases. The length of primary cilia in kidney tubular epithelial cells depends, at least in part, on oxidative stress and extracellular signal-regulated kinase 1/2 (ERK) activation. Hydrogen sulfide (H₂S) is involved in antioxidant systems and the ERK signaling pathway. Therefore, in this study, we investigated the role of H₂S in primary cilia elongation and the downstream pathway. In cultured Madin-Darby Canine Kidney cells, the length of primary cilia gradually increased up to 4 days after the cells were grown to confluent monolayers. In addition, the expression of H₂S-producing enzyme increased concomitantly with primary cilia length. Treatment with NaHS, an exogenous H₂S donor, accelerated the elongation of primary cilia whereas DL-propargylglycine (a cystathionine γ-lyase inhibitor) and hydroxylamine (a cystathionine-β-synthase inhibitor) delayed their elongation. NaHS treatment increased ERK activation and Sec10 and Arl13b protein expression, both of which are involved in cilia formation and elongation. Treatment with U0126, an ERK inhibitor, delayed elongation of primary cilia and blocked the effect of NaHS-mediated primary cilia elongation and Sec10 and Arl13b upregulation. Finally, we also found that H₂S accelerated primary cilia elongation after ischemic kidney injury. These results indicate that H₂S lengthens primary cilia through ERK activation and a consequent increase in Sec10 and Arl13b expression, suggesting that H₂S and its downstream targets could be novel molecular targets for regulating primary cilia.

• The Korean Journal of Physiology and Pharmacology
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• v.8 no.4
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• pp.181-185
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• 2004

Extracellular regulated protein kinase1/2 (pERK1/2) is one of the major regulatory factors for transcription of the c-fos oncogene in neurons. The purpose of this study was to evaluate the expression of phosphorylated ERK1/2 within the vestibular nuclei (VN) of rats following acute arterial hypotension. Following the acute arterial hypotension induced by rapid hemorrhage, a significant number of pERK1/2-immunoreactive neurons appeared bilaterally in the caudal aspect of the medial and inferior VN. No labeling of pERK1/2 was observed in the lateral VN. The peak expression of pERK1/2 in these nuclei occurred within 5 min after hemorrhage. However, in bilaterally labyrinthectomized rats, the appearance of pERK1/2-immunoreactive neurons was eliminated in the VN. Western blot confirmed the effect of bilateral labyrinthectomy on pERK1/2 protein expression in the medial vestibular nucleus 5 min after hemorrhage. These results suggest that, following acute hypotension, afferent signals from the peripheral vestibular receptors are required for activation of ERK 1/2 in the VN.

• The Korean Journal of Physiology and Pharmacology
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• v.8 no.6
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• pp.339-344
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• 2004

Recent data have shown the importance of oxidative stresses in the pathogenesis of inflammatory bowel disease, crohn's disease and ulcerative colitis. $H_2O_2$, reactive oxygen species (ROS) donor, has been reported to act as a signaling molecule involved in a variety of cellular functions such as apo/ptosis and proliferation. In the present study, we investigated viability of cultured ileal smooth muscle cells (ISMC) after stimulation with $H_2O_2$. Trypan blue method revealed that the cell viability of ISMC treated with 1 mM $H_2O_2$ was not different from that of controls at up to 2 h time point, while treatment of ISMC with 1 mM $H_2O_2$ for 48 h finally induced significant decrease in the cell viability. Therefore, we evaluated whether $H_2O_2$ was capable of ERKs activation in ISMC for the short-term exposure and examined whether tyrosine kinase was involved in the process of ERK activation by $H_2O_2$ in ISMC. We also investigated the effects of $H_2O_2$ on activation of SAPK/JNK and p38 MAP kinase in ISMC. Thus, ISMC were cultured and exposed to $H_2O_2$, and western blot analysis was performed with phosphospecific MAP kinase antibodies. Robust activation of ERK occurred within 30 min of 1 mM $H_2O_2$ treatment. $H_2O_2-induced$ ERK activation was attenuated by a tyrosine kinase inhibitor, genistein, indicating that tyrosine kinase was probably involved in the ERK activation by $H_2O_2$. $H_2O_2$ was a moderate activator of SAPK/JNK, while p38 MAP kinase was not activated by $H_2O_2$. We suggest that ERK activation induced by short-term $H_2O_2$ treatment plays a critical role in cellular protection in the early stage of response to oxidative stress. The present study suggests the necessity of identification of MAPK signaling pathways affected by ROS, since it could ultimately elucidate cellular consequences involved in initiation and perpetuation of intestinal tissue damage in the diseases such as crohn's disease and ulcerative colitis, resulted from excessive ROS.

• Natural Product Sciences
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• v.10 no.5
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• pp.211-216
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• 2004

Citrus unshiu (Rutaceae) has long been known as an anti-inflammatory and anti-allergic agent. In the present study, the inhibitory effect of CUWE (Citus unshiu water extract) on the production of $TNF-{\alpha}$ and tryptase was examined. In addition, a possible mechanism for the inhibition of trypsin-stimulated human leukemic mast cell-1 (HMC- 1 ) activation was determined. To do so, $TNF-{\alpha}$ production from the HMC-1 cells that were stimulated by trypsin (100 nM) in the presence or absence of CUWE $(10,\;100,\;and\;100\;{\mu}g/ml)$ was measured by enzyme-linked immunosorbent assay (ELISA) and reverse transcription-PCR. The tryptase production was evaluated by reverse transcription-PCR. Extracellular signal-regulated kinase (ERK) activation was analyzed by Western blot. Trypsin activity was measured by using Bz-DL-Arg-p-nitroanilide (BAPNA) as substrate. Results showed that the CUWE inhibited production of both $TNF-{\alpha}$ and tryptase from the trypsin-stimulated HMC-1 in a dose-dependent manner. The CUWE a1so inhibited the ERK phosphorylation and trysin activity. These results indicate that the CUWE had an inhibitory effect on $TNF-{\alpha}$ and the tryptase productions by blocking the ERK phosphorylation and trypsin activity.

• Korean Journal of Plant Resources
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• v.27 no.3
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• pp.209-214
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• 2014

In this study, we investigated whether A. distichum decreases the production of inflammatory mediators through downregulation of the NF-${\kappa}B$ and ERK pathway. Our data indicated that A. distichum leaf inhibits the overexpression of iNOS in protein and mRNA levels, and subsequently blocked LPS-mediated NO overproduction in RAW264.7 cells. A. distichum leaf inhibited $I{\kappa}B-{\alpha}$ degradation and p65 nuclear translocation, and subsequently suppressed transcriptional activity of NF-${\kappa}B$ in LPS-stimulated RAW264.7 cells. In addition, A. distichum leaf suppressed LPS-induced ERK1/2 activation by decreasing phosphorylation of ERK1/2. These findings suggest that A. distichum leaf shows anti-inflammatory activities through suppressing ERK-mediated NF-${\kappa}B$ activation in mouse macrophage.

• Journal of Physiology & Pathology in Korean Medicine
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• v.33 no.1
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• pp.63-67
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• 2019

Interaction between epidermis and dermis plays an important role in wound healing and hair follicle formation. This study focused on investigating the potency of ethanol extract of Puerariae Radix (EPR) as cosmetic ingredient using human dermal fibroblasts (hDFn). Our results revealed that EPR suppressed collagenase activity dose-dependently. EPR inhibited activity of $5{\alpha}$-reductase I and II at the final concentration of $25{\mu}g/ml$ in hDFn cells. Also, EPR promoted the proliferation and the ERK activation of cells. ERK phosphorylation by EPR was blocked by specific inhibitor of ERK, PD98059. EPR-induced cell proliferation was blocked by PD98059. This means that EPR could promote the proliferation of hDFn cells via the activation ERK. Collectively, these results suggest that EPR may be used as a new cosmetic ingredient.