11:10-12:00, Rm 103

Cell: differentiation, division and deathC-17-01Expressions of apoptosis-related genes during the mouse ovariandevelopments*Chang-Eun Park and Suk-Ryul JungDepartment of Biomedical Laboratory Science, Namseoul University, Cheonan, Chungnam 331-707, KoreaDespite of the importance of the primordial follicle recruitment, its factors andmechanisms are poorly understood. This study was to find out the differentially expressedfactors between primordial and primary follicles and to evaluate the expression and roleof the apoptosis-related genes in the follicular transition from primordial follicles (PMF) toprimary follicles (PRI). The apoptosis-related genes were analyzed by reversetranscription-PCR in ovary of postnatal stages (1st day, 5th day, 14th day, 21th day and28th). The genes were Cideb, Aatk, Egln3, Pdcd7, Casp3, Pdcd5 and Faf1.Cideb andAatk were detected in ovary of all developmental stages. Pdcd7, Casp3 and Faf1 werevery highly expressed in 1st day but were lowly expressed in 5th day, 14th day, 21th dayand 28th. Egln3 and Pdcd5 were highly expressed in 1st day and gradually decreased inother development stages (5th day, 14th day, 21th day and 28th). We detected severalapoptosis-related genes in ovary by using RT-PCR and thus suggest that the apoptosisrelatedgenes may play role(s) in the mouse ovary (oocyte) of early folliculardevelopment.C-17-04TREK-2-induced apoptotic cell death was attenuated by antidepressantfluoxetineGyujin Sim and Kee Ryeon KangDepartment of Biochemistry, MRC for Neural Dysfunction and BK 21 for Medical Science, Schoolof Medicine and Institute of Health Sciences, Gyeongsang National University, Jinju 660-751,KoreaIt has been known that TREK-2, one of K2P channel family, is inhibited its activity byselective serotonin reuptake inhibitors (SSRIs) such as paroxetine and fluoxetine.However, the underlying mechanisms between TREK-2 and psychiatric disorders suchas depression is still unclear. It is well known that antidepressants have anti-apoptoticeffects in various cell types. To understand the role of TREK-2 involved in thepathophysiology of depression, over-expressed rat TREK-2 in 293A cells (293A-rTREK-2) were treated with fluoxetine and observed its protective effects against apoptotic celldeath. We found that the expression of TREK-2 could elicit the activation of JNK andERK activities, and induce PARP cleavage in 293A-rTREK-2 cells up to 48 h after thedrug treatment. Furthermore, we also observed that the effect of fluoxetine in 293ArTREK-2cells correlated with the increased sub-G1 phase of the cell cycle by FACSanalysis. Thus, it is demonstrated that TREK-2 expression could induce apoptotic celldeath by abolishing anti-apoptotic effects of fluoxetine through JNK and ERK. [Fundingsource: National Research Foundation of Korea (NRF) grant by the Korea government(MEST): R13-2005-012-01003-0]C-17-02Identification of DNA methylation markers for lineage commitment ofin vitro hepatogenesisMirang Kim¹, Tae-Wook Kang¹, Han-Chul Lee¹, Yong-Mahn Han², Hyemin Kim²,Hyoung Doo Shin³ , ⁴, Hyun Sub Cheong³, Daeyoup Lee², Seon-Young Kim¹, and YongSung Kim¹¹Medical Genomics Research Center, Korea Research Institute of Bioscience and Biotechnology(KRIBB), Daejeon 305-806, ²Department of Biological Sciences, Korea Advanced Institute of Scienceand Technology (KAIST), Daejeon 305-701, ³Department of Genetic Epidemiology, SNP GeneticsInc., Seoul 153-803, ⁴Department of Life Science, Sogang University, Seoul 121-742, KoreaHepatocytes that have differentiated from human embryonic stem cells have great potentialfor the treatment of liver disease as well as for drug testing. Moreover, in vitro hepatogenesisis a powerful model system for studying the molecular mechanisms underlying liverdevelopment. DNA methylation is an important epigenetic mechanism that influencesdifferential gene expression during embryonic development. We profiled gene expressionand DNA methylation of three cell states of in vitro hepatogenesis-human embryonic stemcell, definitive endoderm, and hepatocyte-using microarray analysis. Among 525 statespecificexpressed genes, 67 showed significant negative correlation between geneexpression and DNA methylation. State-specific expression and methylation of target geneswere validated by quantitative reverse transcription-PCR and pyrosequencing, respectively.To elucidate genome-scale methylation changes beyond the promoter, we also performedhigh-throughput sequencing of methylated DNA captured by MBD2 protein. We founddynamic methylation changes in intergenic regions of the human genome duringdifferentiation. This study provides valuable methylation markers for the lineage commitmentof in vitro hepatogenesis and should help elucidate the molecular mechanisms underlyingstem cell differentiation and liver development.C-17-03The pro-apoptotic effect of hydroquinone in human neutrophils andeosinophilsEun Ju Yang, Ji-Sook Lee¹, Chi-Young Yun²and In Sik Kim³Department of Clinical Laboratory Science, Daegu Haany University, Gyeongsan, Korea,¹Department of Clinical Laboratory Science, Wonkwang Health Science University, Iksan 570-750,Korea, ²Department of Biology, College of Natural Sciences, Daejeon University, Daejeon 300-716,Korea, ³Department of Biomedical Laboratory Science, School of Medicine, Eulji University andEulji University Medical Sciences Research Center, Daejeon 301-746, KoreaHydroquinone (HQ) is a benzene metabolite that is involved in hematopoiesis via itsaccumulation into bone marrow. HQ also acts as a toxic agent that influences variousimmune responses. Both neutrophils and eosinophils function as important leukocytes inimmunological regulation and immune diseases. In this study, we examined the toxiceffects of HQ on the apoptosis of human neutrophils and eosinophils isolated from theblood of healthy donors. HQ markedly increased the apoptosis of neutrophils andeosinophils in a concentration- and a time-dependent manner. The pro-apoptotic effect isinvolved in activation of caspase 9 and caspase 3. Reactive oxygen species (ROS)production was enhanced after HQ treatment in a dose-dependent manner. In addition,HQ up-regulated the release of IL-8 and MCP-1 from neutrophils and eosinophils,respectively. Taken together, the results of this study demonstrated that HQ stronglyinduces the apoptosis of neutrophils and eosinophils through the caspase 9/3-dependentpathway and the increased ROS production. HQ exerts a cytotoxic effect in humanneutrophils and eosinophils and may impair the regulation of immune responses.C-17-05Myostatin inhibits brown adipocyte differentiation via Smad3-mediatedβ-catenin stabilizationWon Kon Kim, Hye-Ryung Choi, Sung Goo Park, Byoung Chul Park, Kwang-HeeBae* and Sang Chul Lee*Medical Proteomics Research Center, KRIBB, Daejeon 305-806, KoreaBrown adipocytes play important roles in regulating energy balance, and alteration ofthese cells has been associated with metabolic diseases such as obesity and diabetes.Although molecular mechanism of white adipocyte differentiation has been widelyrecognized, brown adipogenesis has not been extensively studied. Here, we investigatedthe mechanism of the regulatory action of myostatin in brown adipogenic differentiationusing primary brown preadipocytes. The results clearly showed that differentiation ofbrown adipocyte was significantly inhibited by myostatin treatment. In addition, weobserved that myostatin-induced suppression of brown adipogenesis was appeared atinitiation phase of differentiation. Myostatin induced the phosphorylation of Smad3, whichleading to increased β-catenin stabilization. This effect was blocked by treatment withSmad3 inhibitor. Expression of brown adipocyte-related genes, such as PPAR-γ, UCP-1,PGC-1αand PRDM16, was dramatically down-regulated by treatment with myostatin,and these phenomena were further decreased by co-treatment with β-catenin activator.Taken together, the present study demonstrated that myostatin is a potent negativeregulator of brown adipogenic differentiation by modulation of Smad3-induced β-cateninstabilization.C-17-06A 4-bp deletion mutation in DLX3 enhances anti-adipogenic activity ofDLX3Hye-Lim Lee, Kyung Hwa Baek, Kyung-Mi Woo, Hyun-Mo Ryoo, Gwan-Shik Kimand Jeong-Hwa BaekDepartment of Molecular Genetics, School of Dentistry, Seoul National University, Seoul 110-749,KoreaDeletion of 4 bp in DLX3 is causally related to tricho-dento-osseous syndrome. In thisstudy, we examined whether there are any differences in regulatory effect onadipogenesis between wild type DLX3 (wtDLX3) and 4-bp deletion mutant DLX3(mtDLX3). Over-expression of wtDLX3 and mtDLX3 suppressed adipogenicdifferentiation of 3T3-L1 cells. However, anti-adipogenic effect of mtDLX3 was muchstronger than that of wtDLX3. The expression of PPARγ, a master transcription factor foradipogenesis, was suppressed by wtDLX3 and mtDLX3. In silico analysis showed thatthere was no putative DLX-binding element within 2 kb of the PPARγpromoter sequence.C/EBPα-induced PPARγpromoter activity was suppressed by both wtDLX3 and mtDLX3whereas CREB-induced PPARγpromoter activity was suppressed only by mtDLX3.Results from co-IP and GST-pulldown assays demonstrated that mtDLX3 binds to CREBand C/EBPαwhereas wtDLX3 binds only to C/EBPα. The results from ChIP showed thatbinding of C/EBPαto the PPARγpromoter was decreased by both wtDLX3 and mtDLX3whereas that of CREB was suppressed only by mtDLX3. These results suggest thatDLX3 is a negative regulator for adipocyte differentiation and that a frameshift by 4-bpdeletion provides enhanced anti-adipogenic activity to mtDLX3.200 Korean Society for Biochemistry and Molecular Biology