Sung-Il Yang

The critical period of valproate exposure to induce autistic symptoms in Sprague–Dawley rats

Prenatal exposure to valproic acid (VPA) induces neural tube defects and impairment in social behaviors related to autistic spectrum disorder in newborns, which make it a useful animal model of autism. In this study, we compared the effects of different time window of prenatal valproic acid exposure for inducing the altered social behaviors relevant to autism from embryonic day 7 to embryonic day 15 in Sprague-Dawley rats to determine the critical periods for the impairment. Compared to E7, E9.5 and E15 exposure, VPA exposure at E12 showed most significant changes in behaviors over control animals with reduced sociability and social preference. E9.5 exposure to valproic acid showed strong reproductive toxicity including decrease in the number of live birth. In general, exposure at E15 showed only marginal effects on reproduction and social behaviors. Finally, VPA-exposed rats at E12 were more sensitive to electric shock than VPA-exposed rats at any other periods. These results suggested that E12 is the critical period in rats when valproate exposure has prominent effects for inducing the altered social behavior similar to human autistic behavior.

Psammaplin A, a natural bromotyrosine derivative from a sponge, possesses the antibacterial activity against methicillin-resistantStaphylococcus aureus and the DNA gyrase-inhibitory activity

Psammaplin A, a natural bromotyrosine derivative from an associated form of two sponges (Poecillastra sp. andJaspis sp.) was found to possess the antimicrobial effect on the Grampositive bacteria, especially on methicillin-resistantStaphylococcus aureus (MRSA). The minimal inhibitory concentration of psammaplin A against twenty one MRSAs ranged from 0.781 to 6.25 μg/ml, while that of ciprofloxacin was 0.391∼3.125 μg/ml. Psammaplin A could not bind to penicillin binding protein, but inhibited the DNA synthesis and the DNA gyrase activity with the respective 50% (DNA synthesis) and 100% (DNA gyrase) inhibitory concentration 2.83 and 100 μ/ml. These results indicate that psammaplin A has a considerable antibacterial activity, although restricted to a somewhat narrow range of bacteria, probably by inhibiting DNA gyrase.

Inhibition of GSK-3β mediates expression of MMP-9 through ERK1/2 activation and translocation of NF-κB in rat primary astrocyte

Abstract Glycogen synthase kinase (GSK)-3β and extracellular signal-regulated kinase (ERK) regulate several cellular signaling pathways in common, including embryonic development, cell differentiation and apoptosis. In this study, we investigated whether GSK-3β inhibition is involved in ERK activation, which affects the activation of NF-κB and induction of MMP-9 in cultured rat primary astrocytes. Here, we found that GSK-3β inhibition using GSK-3β inhibitor TDZD-8 increased the phosphorylation of GSK-3β at Ser9 site as well as the phosphorylation of ERK1/2 and Akt at Ser473 site. In this condition, GSK-3β inhibition increased MMP-9 but not MMP-2 activity in a concentration-dependent manner. In RT-PCR analysis, MMP-9 mRNA level was increased by GSK-3β inhibition in a concentration-dependent manner. MMP-9 promoter reporter activity was similarly increased by GSK-3β inhibition. Pretreatment of U-0126 (MEK1/2 inhibitor) completely abolished the GSK-3β inhibition-induced phosphorylation of ERK1/2. U-0126 prevented GSK-3β inhibition-mediated induction of MMP-9 reporter activity as well as the MMP-9 gene expression. The transcriptional activity of NF-κB was significantly increased by GSK-3β inhibition, which was determined by nuclear translocation of NF-κB. Inhibition of ERK1/2 activity by U-0126 also completely blocked the nuclear translocation of NF-κB. Transfection of dominant negative plasmid (S9A) of GSK-3β significantly decreased phosphorylation of ERK, MMP-9 expression and nuclear translocation of NF-κB by GSK-3β inhibition as compared to wild type GSK-3β. These data suggest that GSK-3β inhibition mediates ERK1/2 activation followed by NF-κB activation, which directly regulates the induction of MMP-9 in rat primary astrocytes.

Activation of adenosine A2A receptor up-regulates BDNF expression in rat primary cortical neurons.

As a member of neurotrophin family, brain derived neurotrophic factor (BDNF) plays critical roles in neuronal development, differentiation, synaptogenesis, and neural protection from the harmful stimuli. There have been reported that adenosine A2A receptor subtype is widely distributed in the brain regions, such as hippocampus, striatum, and cortex. Adenosine A2A receptor is colocalized with BDNF in brain regions and the functional interaction between A2A receptor stimulation and BDNF action has been suggested. In this study, we investigated the possibility that the activation of A2A receptor modulates BDNF production in rat primary cortical neuron. CGS21680, an adenosine A2A receptor agonist, induced BDNF expression and release. An antagonist against A2A receptor, ZM241385, prevented CGS21680-induced increase in BDNF production. A2A receptor stimulation induced the activation of Akt-GSK-3β signaling pathway and the blockade of the signaling pathway with specific inhibitors abolished the increase in BDNF production, possibly via modulation of ERK1/2-CREB pathway. The physiological roles of A2A receptor-induced BDNF production was demonstrated by the protection of neurons from the excitotoxicity and increased neurite extension as well as synapse formation from immature and mature neurons. Taken together, activation of A2A receptor regulates BDNF production in rat cortical neuron, which provides neuro-protective action.

Cellular stress-induced up-regulation of FMRP promotes cell survival by modulating PI3K-Akt phosphorylation cascades

BackgroundFragile X syndrome (FXS), the most commonly inherited mental retardation and single gene cause of autistic spectrum disorder, occurs when the Fmr1 gene is mutated. The product of Fmr1, fragile X linked mental retardation protein (FMRP) is widely expressed in HeLa cells, however the roles of FMRP within HeLa cells were not elucidated, yet. Interacting with a diverse range of mRNAs related to cellular survival regulatory signals, understanding the functions of FMRP in cellular context would provide better insights into the role of this interesting protein in FXS. Using HeLa cells treated with etoposide as a model, we tried to determine whether FMRP could play a role in cell survival.MethodsApoptotic cell death was induced by etoposide treatment on Hela cells. After we transiently modulated FMRP expression (silencing or enhancing) by using molecular biotechnological methods such as small hairpin RNA virus-induced knock down and overexpression using transfection with FMRP expression vectors, cellular viability was measured using propidium iodide staining, TUNEL staining, and FACS analysis along with the level of activation of PI3K-Akt pathway by Western blot. Expression level of FMRP and apoptotic regulator BcL-xL was analyzed by Western blot, RT-PCR and immunocytochemistry.ResultsAn increased FMRP expression was measured in etoposide-treated HeLa cells, which was induced by PI3K-Akt activation. Without FMRP expression, cellular defence mechanism via PI3K-Akt-Bcl-xL was weakened and resulted in an augmented cell death by etoposide. In addition, FMRP over-expression lead to the activation of PI3K-Akt signalling pathway as well as increased FMRP and BcL-xL expression, which culminates with the increased cell survival in etoposide-treated HeLa cells.ConclusionsTaken together, these results suggest that FMRP expression is an essential part of cellular survival mechanisms through the modulation of PI3K, Akt, and Bcl-xL signal pathways.

RPS3a Over-Expressed in HBV-Associated Hepatocellular Carcinoma Enhances the HBx-Induced NF-κB Signaling via Its Novel Chaperoning Function

Hepatitis B virus (HBV) infection is one of the major causes of hepatocellular carcinoma (HCC) development. Hepatitis B virus X protein (HBx) is known to play a key role in the development of hepatocellular carcinoma (HCC). Several cellular proteins have been reported to be over-expressed in HBV-associated HCC tissues, but their role in the HBV-mediated oncogenesis remains largely unknown. Here, we explored the effect of the over-expressed cellular protein, a ribosomal protein S3a (RPS3a), on the HBx-induced NF-κB signaling as a critical step for HCC development. The enhancement of HBx-induced NF-κB signaling by RPS3a was investigated by its ability to translocate NF-κB (p65) into the nucleus and the knock-down analysis of RPS3a. Notably, further study revealed that the enhancement of NF-κB by RPS3a is mediated by its novel chaperoning activity toward physiological HBx. The over-expression of RPS3a significantly increased the solubility of highly aggregation-prone HBx. This chaperoning function of RPS3a for HBx is closely correlated with the enhanced NF-κB activity by RPS3a. In addition, the mutational study of RPS3a showed that its N-terminal domain (1–50 amino acids) is important for the chaperoning function and interaction with HBx. The results suggest that RPS3a, via extra-ribosomal chaperoning function for HBx, contributes to virally induced oncogenesis by enhancing HBx-induced NF-κB signaling pathway.

Essential role of mitogen-activated protein kinase pathways in protease activated receptor 2-mediated nitric-oxide production from rat primary astrocytes

Protease-activated receptors (PARs) play important roles in the regulation of brain function such as neuroinflammation by transmitting the signal from proteolytic enzymes such as thrombin and trypsin. We and others have reported that a member of the family, PAR-2 is activated by trypsin, whose involvement in the neurophysiological process is increasingly evident, and is involved in the neuroinflammatory processes including morphological changes of astrocytes. In this study, we investigated the role of PAR-2 in the production of nitric oxide (NO) in rat primary astrocytes. Treatment of PAR-2 agonist trypsin increased NO production in a dose-dependent manner, which was mediated by the induction of inducible nitric-oxide synthase. The trypsin-mediated production of NO was mimicked by PAR-2 agonist peptide and reduced by either pharmacological PAR-2 antagonist peptide or by siRNA-mediated inhibition of PAR-2 expression, which suggests the critical role of PAR-2 in this process. NO production by PAR-2 was mimicked by PMA, a PKC activator, and was attenuated by Go6976, a protein kinase C (PKC) inhibitor. PAR-2 stimulation activated three subtypes of mitogen-activated protein kinases (MAPKs): extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 MAPK. NO production by PAR-2 was blocked by inhibition of ERK, p38, and JNK pathways. PAR-2 stimulation also activated nuclear factor-kappaB (NF-kappaB) DNA binding and transcriptional activity as well as IkappaBalpha phosphorylation. Inhibitors of NF-kappaB pathway inhibited PAR-2-mediated NO production. In addition, inhibitors of MAPK pathways prevented transcriptional activation of NF-kappaB reporter constructs. These results suggest that PAR-2 activation-mediated NO production in astrocytes is transduced by the activation of MAPKs followed by NF-kappaB pathways.

Synthesis and in vitro evaluation of 1,8-diazaanthraquinones bearing 3-dialkylaminomethyl or 3-(N-alkyl- or N-aryl)carbamoyloxymethyl substituent

A series of 1,8-diazaanthraquinone derivatives carrying a 3-dialkylaminomethyl or a 3-(N-alkyl or aryl)carbamoyloxymethyl substituent was synthesised and their in vitro cytotoxic activities were evaluated against eight human cancer cell lines (HOP62, SK-OV-3, HCT-15, SF295, MCF7, SNU-354, KB-3-1 and KB-V-1). A number of compounds including 8c, 8d and 11c showed cytotoxic activity comparable to that of doxorubicin against all human cancer cell lines tested. The compounds 8c and 8d were 2-100 times more potent than doxorubicin against HCT-15, MCF7 and SNU-354 cancer cell lines. Furthermore, these compounds retained considerable cytotoxic activity against the doxorubicin-resistant cell line KB-V-1, implying their therapeutic potential to treat doxorubicin-resistant tumours. These compounds inhibited topoisomerase II-mediated DNA relaxation in vitro, suggesting that this inhibitory effect be attributable to their cytotoxicity.

Positive feedback regulation of Akt-FMRP pathway protects neurons from cell death.

J. Neurochem. (2012) 123, 226–238.

Oroxylin A Induces BDNF Expression on Cortical Neurons through Adenosine A2A Receptor Stimulation: A Possible Role in Neuroprotection

Oroxylin A is a flavone isolated from a medicinal herb reported to be effective in reducing the inflammatory and oxidative stresses. It also modulates the production of brain derived neurotrophic factor (BDNF) in cortical neurons by the transactivation of cAMP response element-binding protein (CREB). As a neurotrophin, BDNF plays roles in neuronal development, differentiation, synaptogenesis, and neural protection from the harmful stimuli. Adenosine A2A receptor colocalized with BDNF in brain and the functional interaction between A2A receptor stimulation and BDNF action has been suggested. In this study, we investigated the possibility that oroxylin A modulates BDNF production in cortical neuron through the regulation of A2A receptor system. As ex-pected, CGS21680 (A2A receptor agonist) induced BDNF expression and release, however, an antagonist, ZM241385, prevented oroxylin A-induced increase in BDNF production. Oroxylin A activated the PI3K-Akt-GSK-3β signaling pathway, which is inhibited by ZM241385 and the blockade of the signaling pathway abolished the increase in BDNF production. The physiological roles of oroxylin A-induced BDNF production were demonstrated by the increased neurite extension as well as synapse formation from neurons. Overall, oroxylin A might regulate BDNF production in cortical neuron through A2A receptor stimulation, which promotes cellular survival, synapse formation and neurite extension.