• Biomolecules & Therapeutics
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• v.19 no.1
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• pp.45-51
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• 2011

These experiments were designed to use typical makers from behaviors and molecular basis in addicted animals of morphine and cocaine. Morphine has been widely abused with a high physical dependence liability. Morphine withdrawal activates the intracellular cAMP signaling pathway and further leads to changes in the expression of the cAMP response element binding protein (CREB), which may be important to the development and expression of morphine dependence. From these experiments, repeated morphine (10 mg/kg, twice per day for 7 days) developed physical dependence. Withdrawal signs were precipitated by naloxone and also increased the expression of the CREB. In addition, repeated exposure of cocaine (15 mg/kg) to mice develops locomotor sensitization and produced lasting behavioral sensitivity. Cocaine- and amphetamine-regulated transcript peptide (CART) peptide was up-regulated by repeated administration of cocaine in the striatum. Therefore, repeated morphine induced the development of physical dependence and increased pCREB. In addition, repeated cocaine induced locomotor sensitization and over-expressed CART peptide. In conclusion, the development of physical dependence and pCREB for morphine, and locomotor sensitization and CART peptide over-expression for cocaine would be useful markers to predict the abuse potential of opioid analgesics and pychostimulant drugs in animals, respectively.

• Journal of Plant Biotechnology
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• v.43 no.1
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• pp.30-36
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• 2016

Brassinosteroids (BRs) are essential plant steroid hormones required for cell elongation, plant growth, development and abiotic and biotic stress tolerance. BRs are recognized by BRI1 receptor kinase that is localized in the plasma membrane, and the BRI1 protein will eventually autophosphorylate in the intracellular domain and transphosphorylate BAK1, which is a co-receptor in Arabidopsis thaliana. However, little is known of the role OsBRI1 receptor kinase plays in Oryza sativa, monocotyledonous plants, compared to that in Arabidopsis thaliana, dicotyledonous plants. As such, we have studied OsBRI1 receptor kinase in vitro and in vivo with recombinant protein and transgenic plants, whose phenotypes were also investigated. A OsBRI1 cytoplasmic domain (CD) recombinant protein was induced in BL21 (DE3) E.coli cells with IPTG, and purified to obtain OsBRI1 recombinant protein. Based on Western blot analysis with phospho-specific pTyr and pThr antibodies, OsBRI1 recombinant protein and OsBRI1-Flag protein were phosphorylated on Threonine residue(s), however, not on Tyrosine residue(s), both in vitro and in vivo. This is particularly intriguing as AtBRI1 protein was phosphorylated on both Ser/Thr and Tyr residues. Also, the OsBRI1 full-length gene was expressed in, and rescued, bri1-5 mutants, such as is seen in normal wild-type plants where AtBRI1-Flag rescues bri1-5 mutant plants. Root growth in seedlings decreased in Ws2, AtBRI1, and 3 independent OsBRI1 transgenic seedlings and had an almost complete lack of response to brassinolide in the bri1-5 mutant. In conclusion, OsBRI1, an orthologous gene of AtBRI1, can mediate normal BR signaling for plant growth and development in Arabidopsis thaliana.

• Biomolecules & Therapeutics
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• v.24 no.1
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• pp.99-107
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• 2016

Triclosan is an antimicrobial or sanitizing agent used in personal care and household products such as toothpaste, soaps, mouthwashes and kitchen utensils. There are increasing evidence of the potentially harmful effects of triclosan in many systemic and cellular processes of the body. In this study, we investigated the effects of triclosan in the survivability of cultured rat neural stem cells (NSCs). Cortical cells from embryonic day 14 rat embryos were isolated and cultured in vitro. After stabilizing the culture, triclosan was introduced to the cells with concentrations ranging from $1{\mu}M$ to $50{\mu}M$ and in varied time periods. Thereafter, cell viability parameters were measured using MTT assay and PI staining. TCS decreased the cell viability of treated NSC in a concentration-dependent manner along with increased expressions of apoptotic markers, cleaved caspase-3 and Bax, while reduced expression of Bcl2. To explore the mechanisms underlying the effects of TCS in NSC, we measured the activation of MAPKs and intracellular ROS. TCS at $50{\mu}M$ induced the activations of both p38 and JNK, which may adversely affect cell survival. In contrast, the activities of ERK, Akt and PI3K, which are positively correlated with cell survival, were inhibited. Moreover, TCS at this concentration augmented the ROS generation in treated NSC and depleted the glutathione activity. Taken together, these results suggest that TCS can induce neurodegenerative effects in developing rat brains through mechanisms involving ROS activation and apoptosis initiation.

• Molecules and Cells
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• v.26 no.3
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• pp.285-290
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• 2008

Estrogen-induced proliferation in estrogen receptor (ER)-positive breast cancer cells is primarily mediated through two distinct intracellular receptors, $ER{\alpha}$ and $ER{\beta}$. Although tumor necrosis factor alpha ($TNF{\alpha}$) and $E2/ER{\alpha}$ are known to exert opposing effects on cell proliferation in MCF-7 cells, the mechanism by which $TNF{\alpha}$ antagonizes $E2/ER{\alpha}$-mediated cell proliferation is not well understood. The present study suggests that reduced cell survival in response to $TNF{\alpha}$ treatment in MCF-7 cells may be associated with the down-regulation of $ER{\alpha}$ protein. The decrease in $ER{\alpha}$ protein level was accompanied by an inhibition of $ER{\alpha}$ gene transcription. Cell viability was decreased synergistically by the combined treatment with $ER{\alpha}$-siRNA and $TNF{\alpha}$. Furthermore, pretreatment of cells with the PI3-kinase (PI3K)/ Akt inhibitor, LY294002, markedly enhanced $TNF{\alpha}$-induced down-regulation of the $ER{\alpha}$ protein, suggesting that the PI3K/Akt pathway might be involved in control of the $ER{\alpha}$ level. Moreover, down-regulation of $ER{\alpha}$ by $TNF{\alpha}$ was not inhibited in cells that were pretreated with the proteasome inhibitors, MG132 and MG152, which suggests that proteasome-dependent proteolysis does not significantly influence $TNF{\alpha}$-induced down-regulation of $ER{\alpha}$ protein. In contrast, the effect of the PI3K/Akt inhibitor on $ER{\alpha}$ was blocked in cells that were treated with LY294002 in the presence of the proteasome inhibitors. Collectively, our findings show that the $TNF{\alpha}$ may partly regulate the growth of MCF-7 breast cancer cells through the down-regulation of $ER{\alpha}$ expression, which is primarily mediated by a PI3K/Akt signaling.

• Korean Journal of Acupuncture
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• v.36 no.4
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• pp.200-209
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• 2019

Objectives : The liver is rich in mitochondria and it plays a key role in whole-body energy homeostasis. Mitochondria is double membrane-bound organelle that supplies energy for intracellular metabolism including Krebs cycle and beta-oxidation. Acupuncture is known to stimulate and regulate the flow of energy. To explore the effect of acupuncture on the mitochondrial respiratory chain activity in the rats' livers, the activity of mitochondrial respiratory chain complexes I to IV was observed. Methods : The rats were divided into 4 groups; Normal 1 (no acupuncture treatment and anesthesia for 5 min), Normal 2 (no acupuncture treatment and anesthesia for 10 min), MA1 (acupuncture treatment at bilateral LR3 under anesthesia for 5 min), and MA2 (acupuncture treatment at bilateral LR3 under anesthesia for 10 min). All rats were sacrificed and the livers were examined for respiratory chain change. Results : There was no difference in ubiquinon oxidoreductase, succinate dehydrogenase, and ubiquinol cytochrome C oxidoreductase after acupuncture at LR3. Acupuncture at LR3 for 10 min increased the activity of cytochrome C oxidase compared with no acupuncture groups. Conclusions : Acupuncture at LR3 mediated mitochondrial respiratory chain activity via the cytochrome C oxidase signaling pathway in the livers of rats.

• KSBB Journal
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• v.18 no.3
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• pp.207-210
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• 2003

The locus coeruleus (LC) contains about half of the total number of noradrenergic neurons in the brain and those noradrenergic neurons from the LC innervate entire brain regions. The LC is a major common target region in several neurodegenerative disorders such as Alzheimer's, Pakinson's and Huntington's diseases. The brain-derived neurotrophic factor (BDNF) regulate neuronal cell survival and differentiation of central nervous system neurons, including LC noradrenergic neurons. In this study, various small molecules and growth factors were tested as candidates to promote the production of BDNF in LC noradrenergic neuronal cells. The molecules tested include neuropeptides, cytokines, growth factors, neurotransmitters, and intracellular signaling agents. Four small molecules or growth factors, FGF8b, BMP-4, forskolin, and dibutyryl cGMP, were found to increase the release of BDNF in LC noradrenergic neurons. Especially, BMP-4 significantly enhanced BDNF production over 2.5-fold in LC noradrenergic neurons.

• The Korea Journal of Herbology
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• v.31 no.5
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• pp.47-53
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• 2016

Objectives : Mori Follium (Morus alba L. leaf) has been cultivated in many Asian countries. Especially, mulberry leaf has been used as an anti-diabetic remedy in oriental medicine. However, anti-obesity effect of mulberry has not been unknown. In this study, our objectives of study is to investigate the anti-adipogenic effect of mulberry water extract (MLE) and to reveal potential molecular anti-obesity mechanism in 3T3-L1 adipocytes differentiation model.Methods : The cytotoxicity of MLE in 3T3-L1 was examined by MTT assay. Anti-adipogenic effect of MLE was evaluated by Oil Red O (ORO) staining. To elucidate the molecular mechanism, inhibitor assay was employed. The mRNA expression levels of adipogenic transcriptional factors such as PPARγ and fatty acid synthase (FAS) were analyzed by reverse transcription-polymer chain reaction (RT-PCR) analysis.Results : The MLE treatment for 24 h did not affect to the 3T3-L1 cells at concentrations of 1, 10, 100, 200, 400, 800 and 1,000 ㎍/㎖. Thus, non-toxic concentration rages of MLE were used during adipogenesis period (day -2 to 7). Intracellular lipid accumulation in MLE-treated 3T3-L1 adipocytes (day 6) were quantitatively evaluated by ORO staining. The MLE treatment significantly and dose-dependently suppressed 3T3-L1 adipogenesis by 60.42%, 38.24%, and 5.97% at 10, 100, and 200 ㎍/㎖, respectively. In addition, our inhibitor assay and RT-PCR analysis revealed that the MLE-inhibited 3T3-L1 adipogenesis through inhibition of PPARγ mediated by Wnt/β-catenin signaling pathway.Conclusions : In conclusion, these findings indicate that the MLE could be used in prevent and/or treatment of obesity-related diseases.

• Animal cells and systems
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• v.1 no.4
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• pp.641-646
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• 1997

Lysophosphatidic acid (LPA; 1-acyl-glycerol-3-phosphate) has been known as an intercellular phospholipid messenger with a wide range of biological activities. In this study, the effect of LPA on both the proliferation and differentiation of rat E63 myoblasts has been investigated. In the serum-free Insulin-Transferrin-Selenium (ITS) media, the proliferation of E63 cells was largely restricted. Addition of LPA into the ITS media strongly promoted the cell proliferation and resulted in two to four fold increase of cell number. Furthermore, it appeared to increase the percent fusion in a dose-dependent manner up to 15 ug/ml. The synthesis of myosin heavy chain (MHC) was increased by LPA as well. These results indicate that LPA is able to promote both cell proliferation and differentiation in rat E63 myoblasts. Suramin, known to have uncoupling activity on growth factor-receptor interaction, was tested for antagonistic activity in myoblast proliferation and differentiation. Myoblasts grown in the ITS medium containing LPA were able to proliferate well even in the presence high concentration of suramin whereas myoblast differentiation was completely blocked by 30 ug/ml of suramin. The inhibitory effect of suramin on the myoblast differentiation was completely reversible by removing the suramin. This result indicates that the intracellular signaling pathway of LPA leading to cell proliferation might be distinct from that leading to cell differentiation on E63 myoblasts. Also, the antagonistic effect of suramin suggests that the differentiation activity elicited by LPA might be mediated by a specific G protein-coupled receptor.

• Journal of Life Science
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• v.17 no.1 s.81
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• pp.18-23
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• 2007

Fe65, a neuron-specific adaptor protein, has two phosphotyrosine binding (PTB) domains. The second PTB (PTB2) domain interacts with intracellular domain fragment (AICD) of amyloid beta precursor protein (APP). Recent studies suggested that tile complex is composed of AICD and Fe65 transactivates genes that are responsible for neuronal cell death in Alzheimer's disease (AD). Therefore, a compound inhibiting the interaction between Fe65 and AICD can be a drug candidate to treat AD. However, it remains unclear how Fe65 recognizes AICD at a molecular level. Here, we report high-level production of the PTB2 domain of Fe65 in the baculovirus system. We found that the baculovirus system is an efficient method to obtain the Fe65 PTB2 domain, compared with the bacterial and mammalian expression systems. The purified recombinant protein was used for crystallization to determine its crystal structure helping to understand the molecular mechanism of Fe65-dependent signaling and to design its inhibitors.

• Molecules and Cells
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• v.19 no.1
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• pp.60-66
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• 2005

Low density lipoproteins (LDL) play important roles in the pathogenesis of atherosclerosis. Diabetes is associated with accelerated atherosclerosis leading to cardiovascular disease in diabetic patients. Although LDL stimulates the proliferation of arterial smooth muscle cells (SMC), the mechanisms are not fully understood. We examined the effects of native LDL and glycated LDL on the extracellular signal-regulated kinase (ERK) pathway. Addition of native and glycated LDL to rat aorta SMCs (RASMCs) stimulated ERK phosphorylation. ERK phosphorylation was not affected by exposure to the $Ca^{2+}$ chelator BAPTA-AM but inhibition of protein kinase C (PKC) with GF109203X, inhibition of Src kinase with PP1 ($5{\mu}M$) and inhibition of phospholipase C (PLC) with U73122/U73343 ($5{\mu}M$) all reduced ERK phosphorylation in response to glycated LDL. In addition, pretreatment of the RASMCs with a cell-permeable mitogen-activated protein kinase kinase (MEK) inhibitor (PD98059, $5{\mu}M$) markedly decreased ERK phosphorylation in response to native and glycated LDL. These findings indicate that ERK phosphorylation in response to glycated LDL involves the activation of PKC, PLC, and MEK, but is independent of intracellular $Ca^{2+}$.