• BMB Reports
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• v.41 no.6
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• pp.479-484
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• 2008

In the present study, we demonstrate that ephrin-A5 is able to induce a transient increase of MAP kinase activity in PC12 cells. However, the effects of ephrin-A5 on the MAP kinase signaling pathway are about three-fold less than that of EGF. In addition, we demonstrate that EphA4 is the only Eph member expressed in PC12 cells, and that tyrosine phosphorylation induced by ephrin-A5 treatment is consistent with the magnitude and longevity of MAP kinase activation. Experiments using the Ras dominant negative mutant N17Ras reveal that Ras plays a pivotal role in ephrin-A5-induced MAP kinase activation in PC12 cells. Importantly, we found that the EphA4 receptor is rapidly internalized by endocytosis upon engagement of ephrin-A5, leading to a subsequent reduction in the MAP kinase activation. Together, these data suggest a novel regulatory mechanism of differential Ras-MAP kinase signaling kineticsexhibited by the forward signaling of EphA4 in PC12 cells.

• Molecules and Cells
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• v.28 no.4
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• pp.365-368
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• 2009

Toll-like receptors (TLRs) play an important role in host defense by sensing invading microbial pathogens and initiating innate immune responses. The stimulation of TLRs by microbial components triggers the activation of myeloid differential factor 88 (MyD88)- and toll-interleukin-1 receptor domain-containing adapter inducing interferon-${\beta}$ (TRIF)-dependent downstream signaling pathways. Isoliquiritigenin (ILG), an active ingredient of Licorice, has been used for centuries to treat many chronic diseases. ILG inhibits the MyD88-dependent pathway by inhibiting the activity of inhibitor-${\kappa}B$ kinase. However, it is not known whether ILG inhibits the TRIF-dependent pathway. To evaluate the therapeutic potential of ILG, we examined its effect on signal transduction via the TRIF-dependent pathway of TLRs induced by several agonists. ILG inhibited nuclear factor-${\kappa}B$ and interferon regulatory factor 3 activation induced by lipopolysaccharide or polyinosinic-polycytidylic acid. ILG inhibited the lipopolysaccharide-induced phosphorylation of interferon regulatory factor 3 as well as interferon-inducible genes such as interferon inducible protein-10, and regulated activation of normal T-cell expressed and secreted (RANTES). These results suggest that ILG can modulate TRIF-dependent signaling pathways of TLRs, leading to decreased inflammatory gene expression.

• The Plant Pathology Journal
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• v.26 no.4
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• pp.386-393
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• 2010

In order to study disease resistance mechanisms in rice against the rice blast fungus Magnaporthe grisea, we screened fungal elicitor-responsive genes from rice suspension-cultured cells treated with fungal elicitors employing differential hybridization (DH). By DH screening, 31 distinct rice clones were isolated and a majority of them were full-length cDNAs encoding pathogenesisrelated (PR) genes. Sixteen of the 31 genes were upregulated at 4, 8, and 12 h following fungal elicitor treatment. To elucidate the effect of signal molecules and biotic elicitors on the regulation of rice defense genes, we further characterized the transcriptional expression patterns of representative isolated PR genes; OsGlu1, OsGlu2, OsTLP, OsRLK, and OsPR-10, following treatment with fungal elicitor, phytohormones, cycloheximide, and inhibitors of protein phosphorylation. Jasmonic acid (JA) induced transcriptional expression of OsGlu1, OsTLP, and OsRLK, but not of OsGlu2 and OsPR-10 at any of the tested time points. Salicylic acid (SA) and abscisic acid weakly induced the expression of OsTLP and OsRLK. SA showed an antagonistic effect with fungal elicitor and JA. Cycloheximide suppressed all these genes upon elicitor treatment, except for OsGlu2. Staurosporine only induced the expression of OsRLK. Application of calyculin A strongly induced OsRLK expression, but suppressed the expression of OsGlu2. Our study yielded a number of PR genes that play a role in defense mechanisms against the rice blast fungus, as well as contribute towards the elucidation of crosstalk between phytohormones and other modifications during defense signaling.

• Journal of Ginseng Research
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• v.46 no.2
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• pp.255-265
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• 2022

Background: Ginsenoside Rb1, a bioactive component isolated from the Panax ginseng, acts as a remedy to prevent myocardial injury. However, it is obscure whether the cardioprotective functions of Rb1 are related to the regulation of endogenous metabolites, and its potential molecular mechanism still needs further clarification, especially from a comprehensive metabolomics profiling perspective. Methods: The mice model of acute myocardial ischemia (AMI) and oxygen glucose deprivation (OGD)-induced cardiomyocytes injury were applied to explore the protective effect and mechanism of Rb1. Meanwhile, the comprehensive metabolomics profiling was conducted by high-performance liquid chromatography and quadrupole time-of-flight mass spectrometry (HPLC-Q/TOF-MS) and a tandem liquid chromatography and mass spectrometry (LC-MS). Results: Rb1 treatment profoundly reduced the infarct size and attenuated myocardial injury. The metabolic network map of 65 differential endogenous metabolites was constructed and provided a new inspiration for the treatment of AMI by Rb1, which was mainly associated with mitophagy. In vivo and in vitro experiments, Rb1 was found to improve mitochondrial morphology, mitochondrial function and promote mitophagy. Interestingly, the mitophagy inhibitor partly attenuated the cardioprotective effect of Rb1. Additionally, Rb1 markedly facilitated the phosphorylation of AMP-activated protein kinase α (AMPKα), and AMPK inhibition partially weakened the role of Rb1 in promoting mitophagy. Conclusions: Ginsenoside Rb1 protects acute myocardial ischemia injury through promoting mitophagy via AMPKα phosphorylation, which might lay the foundation for the further application of Rb1 in cardiovascular diseases.

• Journal of Ginseng Research
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• v.47 no.1
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• pp.97-105
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• 2023

Background: Hyperactivated airway mucosa cells overproduce mucin and cause severe breathing complications. Here, we aimed to identify the effects of saponins derived from Panax ginseng on inflammation and mucin overproduction. Methods: NCI-H292 cells were pre-incubated with 16 saponins derived from P. ginseng, and mucin overproduction was induced by treatment with phorbol 12-myristate 13-acetate (PMA). Mucin protein MUC5AC was quantified by enzyme-linked immunosorbent assay, and mRNA levels were analyzed using quantitative polymerase chain reaction (qPCR). Moreover, we performed a transcriptome analysis of PMA-treated NCI-H292 cells in the absence or presence of Rg5, and differential gene expression was confirmed using qPCR. Phosphorylation levels of signaling molecules, and the abundance of lipid droplets, were measured by western blotting, flow cytometry, and confocal microscopy. Results: Ginsenoside Rg5 effectively reduced MUC5AC secretion and decreased MUC5AC mRNA levels. A systematic functional network analysis revealed that Rg5 upregulated cholesterol and glycerolipid metabolism, resulting in the production of lipid droplets to clear reactive oxygen species (ROS), and modulated the mitogen-activated protein kinase and nuclear factor (NF)-kB signaling pathways to regulate inflammatory responses. Rg5 induced the accumulation of lipid droplets and decreased cellular ROS levels, and N-acetyl-ⳑ-cysteine, a ROS inhibitor, reduced MUC5AC secretion via Rg5. Furthermore, Rg5 hampered the phosphorylation of extracellular signal-regulated kinase and p38 proteins, affecting the NF-kB signaling pathway and pro-inflammatory responses. Conclusion: Rg5 alleviated inflammatory responses by reducing mucin secretion and promoting lipid droplet-mediated ROS clearance. Therefore, Rg5 may have potential as a therapeutic agent to alleviate respiratory disorders caused by hyperactivation of mucosa cells.

• Molecules and Cells
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• v.24 no.3
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• pp.431-436
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• 2007

Stem cell-based therapy is being considered as an alternative treatment for cardiomyopathy. Hence understanding the basic molecular mechanisms of cardiomyocyte differentiation is important. Besides BMP or Wnt family proteins, $TGF-{\beta}$ family members are thought to play a role in cardiac development and differentiation. Although $TGF-{\beta}$ has been reported to induce cardiac differentiation in embryonic stem cells, the differential role of $TGF-{\beta}$ isoforms has not been elucidated. In this study, employing the DMSO-induced cardiomyocyte differentiation system using P19CL6 mouse embryonic teratocarcinoma stem cells, we investigated the $TGF-{\beta}$-induced signaling pathway in cardiomyocyte differentiation. $TGF-{\beta}1$, but not the other two isoforms of $TGF-{\beta}$, was induced at the mRNA and protein level at an early stage of differentiation, and Smad2 phosphorylation increased in parallel with $TGF-{\beta}1$ induction. Inhibition of $TGF-{\beta}1$ activity with $TGF-{\beta}1$-specific neutralizing antibody reduced cell cycle arrest as well as expression of the CDK inhibitor $p21^{WAF1}$. The antibody also inhibited induction of the cardiac transcription factor Nkx2.5. Taken together, these results suggest that $TGF-{\beta}1$ is involved in cardiomyocyte differentiation by regulating cell cycle progression and cardiac gene expression in an autocrine or paracrine manner.

• Journal of Microbiology and Biotechnology
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• v.29 no.12
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• pp.2014-2021
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• 2019

Middle East respiratory syndrome coronavirus (MERS-CoV) belongs to the genus Betacoronavirus and causes severe morbidity and mortality in humans especially when infected patients have underlying diseases such as chronic obstructive pulmonary disease (COPD). Previously, we demonstrated that MERS-CoV-encoded ORF8b strongly inhibits MDA5- and RIG-I-mediated induction of the interferon beta (IFN-β) promoter activities. Here, we report that ORF8b seemed to regulate MDA5 or RIG-I differentially as protein levels of MDA5 were significantly down-regulated while those of RIG-I were largely unperturbed. In addition, ORF8b seemed to efficiently suppress phosphorylation of IRF3 at the residues of 386 and 396 in cells transfected with RIG-I while total endogenous levels of IRF3 remained largely unchanged. Furthermore, ORF8b was able to inhibit all forms of RIG-I; full-length, RIG-I-1-734, and RIG-I-1-228, the last of which contains only the CARD domains. Taken together, it is tempting to postulate that ORF8b may interfere with the CARD-CARD interactions between RIG-I and MAVS. Further detailed analysis is required to delineate the mechanisms of how ORF8b inhibits the MDA5/RIG-I receptor signaling pathway.

• Toxicological Research
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• v.20 no.3
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• pp.233-239
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• 2004

The present study was designed to determine the impact of mercury on endotoxin-induced inflammatory cytokine expression and corresponding signal transduction in mouse kidney. Male BALB/c mice were exposed continuously to 0, 0.3, 1.5, 7.5, or 37.5 ppm of mercury in drink-ing water for 14 days and at the end of the treatment period, lipopolysaccharide (LPS, 0.5 mg/kg) was injected intraperitoneally 2 h prior to euthanasia. The doses of mercury and LPS did not cause hepatotoxicity or renal toxicity as indicated by unaltered plasma alanine aminotransferase and aspartate aminotransferase levels, and terminal UTP nucleotide end-labeling assay from kidney, respectively. Mercury decreased kidney glutathione (GSH) and with LPS, it additively decreased GSH. Mercury activated p38 mitogen-activated protein kinase (MAPK) and additively increased LPS-induced p38 MAPK phosphorylation. In contrast, mercury inhibited LPS-induced activation of extra-cellular signal-regulated kinase (ERK) but had no effect alone. Mercury increased the gene expression of tumor necrosis factor $\alpha$ (TN F$\alpha$) and potentiated LPS-induced TNF$\alpha$ expression. Mercury did not affect LPS-induced interleukin-1$\beta$ (IL-1$\beta$) expression but decreased LPS-induced IL-6 expression. These results suggest that low levels of mercury might augment LPS-induced TNF$\alpha$ expression by altering GSH and p38 MAPK. Mercury modulates LPS-induced p38 and ERK activation, and downstream TNF$\alpha$ and IL-6 expression in kidney, respectively.

• Molecular & Cellular Toxicology
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• v.5 no.3
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• pp.187-192
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• 2009

Toll-like receptors (TLRs) play an important role in host defense by sensing invading microbial pathogens. The stimulation of TLRs by microbial components triggers the activation of the myeloid differential factor 88 (MyD88)- and toll-interleukin-1 receptor domain-containing adapter inducing interferon-$\beta$ (TRIF)-dependent downstream signaling pathways. TLR/MyD88 signaling pathway induces the activation of nuclear factor-kappa B (NF-${\kappa}B$) and the expression of inflammatory cytokine genes, including tumor necrosis factor-alpha, interleukin (IL)-6, IL-12, and IL-$1{\beta}$. On the other hand, TLR/TRIF signaling pathway induces the delayed-activation of NF-${\kappa}B$ and interferon regulatory factor 3 (IRF3), and the expression of type I interferons (IFNs) and IFN-inducible genes. The divalent heavy metal cadmium (Cd) is clearly toxic to most mammalian organ systems, especially the immune system. Yet, the underlying toxic mechanism(s) remain unclear. Cd inhibits the MyD88-dependent pathway by ceasing the activity of inhibitor-${\kappa}B$ kinase. However, it is not known whether Cd inhibits the TRIF-dependent pathway. Presently, Cd inhibited NF-${\kappa}B$ and IRF3 activation induced by lipopolysaccharide (LPS) and polyinosinic-polycytidylic acid. Cd inhibited LPS-induced IRF3 phosphorylation and IFN-inducible genes such as interferon inducible protein-10 and regulated on activation normal T-cell expressed and secreted (RANTES). These results suggest that Cd can modulate TRIF-dependent signaling pathways of TLRs.

• Molecules and Cells
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• v.21 no.2
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• pp.237-243
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• 2006

Brain-derived neurotrophic factor (BDNF) is a neuromodulator of nociceptive responses in the dorsal root ganglia (DRG) and spinal cord. BDNF synthesis increases in response to nerve growth factor (NGF) in trkA-expressing small and medium-sized DRG neurons after inflammation. Previously we demonstrated differential activation of multiple BDNF promoters in the DRG following peripheral nerve injury and inflammation. Using reporter constructs containing individual promoter regions, we investigated the effect of NGF on the multiple BDNF promoters, and the signaling pathway by which NGF activates these promoters in PC12 cells. Although all the promoters were activated 2.4-7.1-fold by NGF treatment, promoter IV gave the greatest induction. The p38 mitogen-activated protein kinase (MAPK) inhibitor, SB203580, phosphatidylinositol 3-kinase (PI-3K) inhibitor, LY294003, protein kinase A (PKA) inhibitor, H89, and protein kinase C (PKC) inhibitor, chelerythrine, had no effect on activation of promoter IV by NGF. However, activation was completely abolished by the MAPK kinase (MEK) inhibitors, U0126 and PD98059. In addition, these inhibitors blocked NGF-induced phosphorylation of extracellular signal-regulated protein kinase (ERK) 1/2. Taken together, these results suggest that the ERK1/2 pathway activates BDNF promoter IV in response to NGF independently of NGF-activated signaling pathways involving PKA and PKC.