• 약학회지
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• 제54권6호
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• pp.461-465
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• 2010

We investigated the role of L-type $Ca^{2+}$ channel in receptor activator of nuclear factor-kappaB ligand (RANKL)-induced osteoclast formation. BayK 8644, a L-type $Ca^{2+}$ channel agonist, was shown to increase the RANKLinduced osteoclastogenesis and actin ring formation in mouse bone marrow-dereived macrophage (BMM) culture system. BayK 8644 stimulated RANKL-induced extracellular signal-regulated kinase (ERK) and p38 MAP kinase (MAPK) activation, which leads to increased nuclear factor of activated T cells (NFAT)c1 expression. Taken together, these data indicate that L-type $Ca^{2+}$ channel regulates osteoclast formation possibly through ERK- and p38-mediated NFATc1 expression.

• Asian Pacific Journal of Cancer Prevention
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• 제13권11호
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• pp.5709-5714
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• 2012

Objective: To evaluate the effects of curcumin on matrixmetalloproteinase-9 (MMP-9) and invasion ability induced by transforming growth factor-${\beta}1$ (TGF-${\beta}1$) in MDA-MB-231 cells and potential mechanisms. Methods: Human breast cancer MDA-MB-231 cells were used with the CCK-8 assay to measure the cytotoxicity of curcumin. After treatment with 10 ng/ml TGF-${\beta}1$, with or without curcumin (${\leq}10{\mu}M$), cell invasion was checked by transwell chamber. The effects of curcumin on TGF-${\beta}1$-stimulated MMP-9 and phosphorylation of Smad2, extracellular-regulated kinase (ERK), and p38 mitogen activated protein kinases (p38MAPK) were examined by Western blotting. Supernatant liquid were collected to analyze the activity of MMP-9 via zymography. Following treatment with PD98059, a specific inhibitor of ERK, and SB203580, a specific inhibitor of p38MAPK, Western blotting and zymography were employed to examine MMP-9 expression and activity, respectively. Results: Low dose curcumin (${\leq}10{\mu}M$) did not show any obvious toxicity to the cells, while $0{\sim}10{\mu}mol/L$ caused a concentration-dependent reduction in cell invasion provoked by TGF-${\beta}1$. Curcumin also markedly inhibited TGF-${\beta}1$-regulated MMP-9 and activation of Smad2, ERK1/2 and p38 in a dose- and time-dependent manner. Additionally, PD98059, but not SB203580, showed a similar pattern of inhibition of MMP-9 expression. Conclusion: Curcumin inhibited TGF-${\beta}1$-stimulated MMP-9 and the invasive phenotype in MDA-MB-231 cells, possibly associated with TGF-${\beta}$/Smad and TGF-${\beta}$/ERK signaling.

• Biomolecules & Therapeutics
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• 제25권6호
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• pp.593-598
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• 2017

The $Na^+/H^+$ exchanger-1 (NHE-1) is a ubiquitously expressed pH-regulatory membrane protein that functions in the brain, heart, and other organs. It is increased by intracellular acidosis through the interaction of intracellular $H^+$ with an allosteric modifier site in the transport domain. In the previous study, we reported that glutamate-induced NHE-1 phosphorylation mediated by activation of protein kinase C-${\beta}$ (PKC-${\beta}$) in cultured neuron cells via extracellular signal-regulated kinases (ERK)/p90 ribosomal s6 kinases (p90RSK) pathway results in NHE-1 activation. However, whether glutamate stimulates NHE-1 activity solely by the allosteric mechanism remains elusive. Cultured primary cortical neuronal cells were subjected to intracellular acidosis by exposure to $100{\mu}M$ glutamate or 20 mM $NH_4Cl$. After the desired duration of intracellular acidosis, the phosphorylation and activation of PKC-${\beta}$, ERK1/2 and p90RSK were determined by Western blotting. We investigated whether the duration of intracellular acidosis is controlled by glutamate exposure time. The NHE-1 activation increased while intracellular acidosis sustained for >3 min. To determine if sustained intracellular acidosis induced NHE-1 phosphorylation, we examined phosphorylation of NHE-1 induced by intracellular acidosis by transient exposure to $NH_4Cl$. Sustained intracellular acidosis led to activation and phosphorylation of NHE-1. In addition, sustained intracellular acidosis also activated the PKC-${\beta}$, ERK1/2, and p90RSK in neuronal cells. We conclude that glutamate stimulates NHE-1 activity through sustained intracellular acidosis, which mediates NHE-1 phosphorylation regulated by PKC-${\beta}$/ERK1/2/p90RSK pathway in neuronal cells.

• Journal of Microbiology and Biotechnology
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• 제15권4호
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• pp.756-766
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• 2005

The signaling mechanism underlying aburatubolactam C-induced FasL upregulation was investigated in human Jurkat T cells. After treatment with aburatubolactam C, the src-family PTKs $p56^{lck}\;and\;p59^{fyn}$, and MAP kinases ERK2 and JNK1, were activated prior to FasL upregulation; Both $p56^{lck}\;and\;p59^{fyn}$ were directly activated 2.4- and 2.2-fold, respectively, in vitro by aburatubolactam C. The aburatubolactam C-induced cellular changes, including the activation of ERK2 and INK1, and FasL upregulation, were completely prevented by the PTK inhibitor genistein. The activation of protein kinase C (PKC$\delta,\;\epsilon\;and\;\mu$ was also induced following aburatubolactam C treatment. Although the activation of $p56^{lck}$ and tyrosine phosphorylation of the cellular proteins were not blocked by the PKC inhibitor GFl09203X, the activation of ERK2 was completely abrogated, along with a detectably enhanced JNK1 activation; FasL upregulation, and apoptosis. However, the FasL upregulation and apoptosis were significantly inhibited by the PKC activator PMA, with a remarkable increase in the ERK2 activation. The cytotoxic effect of aburatubolactam C was reduced in the presence of the anti-Fas neutralizing antibody ZB-4. Although ectopic expression of Bcl-2 failed to completely block the cytotoxicity of aburatubolactam C, it was clearly suppressed. The c-Fos mRNA expression was upregulated in a biphasic manner, where the second phasic expression overlapped with the FasL upregulation. Accordingly, these results demonstrate that aburatubolactam C-induced apoptosis is exerted, at least in part, by FasL upregulation dictated by activation of the PTK ($p56^{lck}\;and\;p59^{fyn}$) /JNKI pathway, which is negatively affected by the concurrent activation of the PKC/ERK2 pathway proximal to PTK activation.

• 동의생리병리학회지
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• 제21권6호
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• pp.1555-1563
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• 2007

To idenifty effect of Bojungbangam-tang kakambang on Cisplatin-Induced G2/M Phase Arrest in Human Renal Proximal Tubular HK-2 Cells. Cytotoxicity of cisplatin was detected in HK-2 cells and the value of IC50 is about $25\;{\mu}M$. The treatment of cisplatin to HK-2 showed the G2/M phase cell cycle arrest. The ethanol extract of Bojungbangam-tang kakambang (EBTKB), a new herbal prescription composed of ten crude herbs, inhibited cisplatin-induced G2/M phase arrest in HK-2 cells. EBTKB increased G0/G1 peak in cisplatin-treated HK-2 cells. p53, p21 and p27 expression were increased in cisplatin-treated HK-2 cells. Inhibitory effect of EBTKB on cisplatin-induced G2/M phase arrest was accomplished through inhibition of p53, p21 and p27 expression. Also, reduced CDK2 and cyclin A expression by cisplatin were increased by EBTKB, but cyclin E was not changed. Reduction of ERK activation and increment of p38 activation by cisplatin were increased ERK activation and decreased p38 activation by EBTKB. Cisplatin had no effect on JNK activation, but EBTKB increased JNK activation. These results can suggest that EBTKB inhibits cisplatin-induced G2/M phase arrest in HK-2 cell through reduction of p53-dependent p21 and p27 protein, ERK activation and p38 inactivation.

• 생명과학회지
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• 제31권9호
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• pp.806-817
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• 2021

우울증 발생 기전에 신경가소성의 손상이 관여한다. Sirtuin 1은 신경가소성의 조절에 중요한 역할을 담당하고 있다. 또한 mTORC1 신호전달의 활성화가 신경가소성을 향상시키는 것으로 알려져 있다. 본 연구는 sirtuin 1이 mTORC1 신호전달을 통해 수상돌기 성장과 가시형성에 미치는지 영향을 조사하였다. 덱사메타손이 처치된 신경세포와 정상 배양 신경세포에 resveratrol (sirtuin 1 활성제)과 sirtinol (sirtuin 1 억제제)을 각각 처치하였다. Western blot 분석법을 사용하여, sirtuin 1 발현 및 ERK1/2, mTORC1, p70S6K의 인산화 양을 분석하였고, 면역형광측정법으로 수상돌기의 길이와 가시밀도를 분석하였다. Resveratrol은 덱사메타손 환경에서 농도 의존적으로 sirtuin 1의 발현을 증가시켰으며, ERK1/2 (sirtuin 1의 하위 타겟), mTORC1, 그리고 p70S6K (mTORC1의 하위 타겟)의 인산화를 유의하게 증가시켰다. 또한 resveratrol은 수상돌기 성장과 가시밀도를 증가시켰다. 반면, sirtinol은 정상 배양액에서 sirtuin 1의 발현을 유의하게 감소시켰으며, ERK1/2, mTORC1, p70S6K의 인산화 양을 농도 의존적으로 유의하게 감소시켰다. 또한 sitinol은 수상돌기 성장과 가시밀도를 유의하게 감소시켰다. 신경세포에 sirtuin 1의 siRNA를 transfection시켜 sirtuin 1을 knockdown 시켰을 때, ERK1/2 및 mTORC1의 인산화 양이 감소하였을 뿐만 아니라, 수상돌기 성장과 가시밀도도 감소하였다. 본 연구는 sirtuin 1이 ERK1/2-mTORC1 신호전달을 통해서 수상돌기 성장과 가시밀도를 변화시켜 신경가소성을 조절한다는 것을 보여주었다.

• Journal of Oral Medicine and Pain
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• 제32권4호
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• pp.357-364
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• 2007

고농도 포도당이 뼈모세포와 치주인대세포의 세포자멸사에 미치는 영향과 그 경로를 알아보기 위하여 뼈모세포주인 MC3T3-E1 (E1) 세포와 사람 치주인대로부터 일차배양을 통해 얻은 치주인대세포를 1,000 mg/L 농도의 포도당이 포함된 배양액 (대조군)과 4,500 mg/L 농도의 포도당이 포함된 배양액 (실험군)으로 나누어 24시간과 48시간 배양하였다. 그 후, ELISA assay를 통해 p38 MAPK와 caspase-3의 발현을 평가하고 Western blot을 통해 JNK-1과 ERK-1의 발현을 평가하여 다음과 같은 결론을 얻었다. 1. 뼈모세포와 치주인대세포 모두 대조군에 비해 실험군에서 caspase-3와 p38 MAPK 발현이 증가하였다. 2. 실험군에서의 caspase-3와 p38 MAPK 발현은 뼈모세포에 비해 치주인대세포에서 더욱 크게 증가하였다. 3. 뼈모세포와 치주인대세포 모두 대조군에 비해 실험군에서 JNK-1 발현이 증가하였다. 4. 뼈모세포와 치주인대세포 모두 ERK-1 발현에는 변화가 없었다. 이상의 결과로 보아, 고혈당 조건에 의해 뼈모세포와 치주인대세포의 세포자멸사가 증가하며, 치주인대세포가 고혈당 조건에 더욱 민감하게 반응하여 세포자멸사가 크게 증가하는 것으로 생각된다. 또한 이들 세포의 세포자멸사 과정은 p38 MAPK와 JNK-1 경로가 관여하며 ERK-1 경로는 관여하지 않는 것으로 추정된다.

• The Korean Journal of Physiology and Pharmacology
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• 제20권1호
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• pp.69-74
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• 2016

The present study was undertaken to investigate the influence of cardamonin on vascular smooth muscle contractility and to determine the mechanism(s) involved. Denuded aortic rings from male rats were used and isometric contractions were recorded and combined with molecular experiments. Cardamonin significantly relaxed fluoride-, phenylephrine-, and phorbol ester-induced vascular contractions, suggesting that it has an anti-hypertensive effect on agonist-induced vascular contraction regardless of endothelial nitric oxide synthesis. Furthermore, cardamonin significantly inhibited the fluoride-induced increase in pMYPT1 level and phenylephrine-induced increase in pERK1/2 level, suggesting inhibition of Rho-kinase and MEK activity and subsequent phosphorylation of MYPT1 and ERK1/2. This study provides evidence that the relaxing effect of cardamonin on agonist-induced vascular contraction regardless of endothelial function involves inhibition of Rho-kinase and MEK activity.

• Clinical and Experimental Pediatrics
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• 제60권6호
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• pp.181-188
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• 2017

Purpose: Hypoxic-ischemic encephalopathy is a significant cause of neonatal morbidity and mortality. Erythropoietin (EPO) is emerging as a therapeutic candidate for neuroprotection. Therefore, this study was designed to determine the neuroprotective role of recombinant human EPO (rHuEPO) and the possible mechanisms by which mitogen-activated protein kinase (MAPK) signaling pathway including extracellular signal-regulated kinase (ERK1/2), JNK, and p38 MAPK is modulated in cultured cortical neuronal cells and astrocytes. Methods: Primary neuronal cells and astrocytes were prepared from cortices of ICR mouse embryos and divided into the normoxic, hypoxia (H), and hypoxia-pretreated with EPO (H+EPO) groups. The phosphorylation of MAPK pathway was quantified using western blot, and the apoptosis was assessed by caspase-3 measurement and terminal deoxynucleotidyl transferase dUTP nick end labeling assay. Results: All MAPK pathway signals were activated by hypoxia in the neuronal cells and astrocytes (P<0.05). In the neuronal cells, phosphorylation of ERK-1/-2 and apoptosis were significantly decreased in the H+EPO group at 15 hours after hypoxia (P<0.05). In the astrocytes, phosphorylation of ERK-1/-2, p38 MAPK, and apoptosis was reduced in the H+EPO group at 15 hours after hypoxia (P<0.05). Conclusion: Pretreatment with rHuEPO exerts neuroprotective effects against hypoxic injury reducing apoptosis by caspase-dependent mechanisms. Pathologic, persistent ERK activation after hypoxic injury may be attenuateed by pretreatment with EPO supporting that EPO may regulate apoptosis by affecting ERK pathways.

• 생약학회지
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• 제44권4호
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• pp.379-383
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• 2013

Portulaca oleracea L. is known to have many biological benefits such as anti-oxidant, anti-inflammatory, anti-allergic and anti-tumor. The objective of this study is to explore the neuroprotective effect of P. oleracea L. against glutamate-induced oxidative stress in mouse hippocampal HT22 cells. P. oleracea L. 70% ethanol extract and solvent fractions have the potent neroprotective effects on glutamate-induced nerotoxicity by induced the expression of heme oxygenase (HO)-1 in HT22 cells. Especially, ethyl acetate fraction showed higher protective effect. In HT22 cell, P. oleracea L. treatment with ERK inhibitor (PD98059) and c-JUN N-terminal kinase (JNK) inhibitor (SP600125) reduced P. oleracea L. ethyl acetate fraction induced HO-1 expression and P. oleracea L. ethyl acetate fraction also increased ERK and JNK phosphorylation. Furthermore, we found that treatment of P. oleracea L. caused the nuclear accumulation of Nrf2. In conclusion, the ethyl acetate fraction of 70% ethanol extract of P. oleracea L. significantly protect glutamate-induced oxidative damage by induction of HO-1 via Nrf2, ERK and JNK pathway in mouse hippocampal HT22. Taken together these finding suggest that P. oleracea L. ethyl acetate fraction is good source for taking active compounds and may be a potential therapeutic agent for brain disorder that induced by oxidative stress and neuronal damage.