Epithelium maintains homeostasis by the signaling balance of growth stimulation and inhibition. Recently, loss of growth inhibitory effects of transforming growth factor-β(TGF-β) on epithelial cells is regarded as a possible mechanism of cancer. Although the genomic mutation in type I and type Ⅱ receptors of TGF-β is considered one of important mechanism of these inactivation, there might be another inactivation mechanism because the mutation rate is relatively low and inhibitory effect is not associated with the mutation. The purpose of this study is evaluating controlling mechanism type Ⅱ receptor of TGF-β by detecting effects of TGF-β on growth inhibition and on expression of cell cycle regulatory protein p21CIP1. Eight cancer cell lines derived from oral squamous cell carcinoma(OSCC) were examined. There was no growth inhibition effects by TGF-β except YD-8 cells. YD-8 cells which showed growth inhibition expresses p21CIP1 by TGF-β whether refractory cell lines, YD-9, did not. All of the tumor cells express mRNA of type Ⅱ receptor by RT-PCR and northern blot analysis, especially on YD-8 and YD-17M. From these results, most of oral cancer cell lines might loose the growth inhibitory effects by TGF-β, and the growth inhibition on YD-8 cells was mediated by expression of p21CIP1.

Cellular imrnortali zation is thought to be an ear ly event during tumorigenesis. Telomerase reacLi vaLion by ecLopic hTERT expression is widely used for cellular imrnortali zation. This study was a imed Lo a na lyze establi sh immortalized human ora l epithelial cells(IOEC) and to reconstruct oral precancerous lesion by Lhree dimens iona l cultures. Telomerase activity was analyzed by Telomerase assays and Telomere longLh W:lS dcLccLcd by Termina l restri ction I"ragment analysis. bTERT gene was assayecl by tbe RT-PCR. p16lNK4 a‘ pHb. CDK2. P21CIP1. p27 and p53 were examined by western blotting. Three dimensiona l cu1ture using air - liquicl inLe rl"ace was pe rl"ormed. As results. IOEC was establi shed by ectopic ex pression of catalytic subunit• of telomerase‘ h1'EH1'. which is con tinuously maintained for more tban 120 population doublings(PDs) . IOEC showecl the expression 01" h1'ER1' and h1'H mHNA‘ elongated telomere length and higher telomerase activity. These cel ls showed no ex pression of p16lNK4a with retention 01" pRb and CDK2. Expression of p21CIP1. p27 a nd p53 may have no relation to immorta li ze oraJ epithelial cell s. Three dimensional culture of IOEC showed dysplastic strat ilïed epithelia l cell s. These results may serve as a useful moclel system for the study of oral carcinogenesis.

The imbalance between epithelial cell growth and inhibitory factors may cause human epithelial cancer. The dysregulation of growth inhibitory effect of TGF-β1 has been recognized in a variety of carcinomas. This study aimed to investigate the expression of TGF-β1 type II receptors(TβR-II) in the carcinogenesis of oral squamous cell carcinoma(OSCC). Six cell lines established from OSCC in the department of Oral Pathology, Yonsei University College of Dentistry were used. DNA was extracted from harvested cells by phenol-chloroform method. Polymerase chain reaction (PCR) was done with each primer of exon 3, 4, 5, 6 of TβR-II gene. PCR products were inserted to cloning vector (pGEM-T easy vector) and then analyzed to automatic DNA sequencing analyzer. Reverse transcriptase-PCR (RT-PCR) was performed to confirm the mRNA expression of TβR-II gene. Western blotting was performed to detect the expression of the TβR-II protein. As results, a frameshift within a polyadenine region of exon 3 was found in YD-8 cell line. In YD-17 cell line, a missense mutation at codon 238 of exon 4 was found, suggesting the alteration of amino acid from asparagine to aspartic acid. TβR-II mRNA was detected in all cancer cell lines, but it was slightly decreased as compared to that of normal oral mucosal cells. In Western blotting, no TβR-Ⅱ protein was detected in all OSCC cell lines. These results suggested that the altered regulation of TGF-β1 function might play a role in the development of OSCC.

In order to investigate the species, serobiological characteristics and antibiotic sensitivity of Pseudomonas spp, we isolated Pseudomonas spp from 57 spring waters around Seoul area for spring, summer and autumn and identified Pseudomonas spp by biochemical characteristics and serological method. And also we tested the antibiotic sensitivity test by discdiffusion method. Of 57 spring waters tested, Pseudomonas spp were isolated from 33 spring waters(57.9%). Isolation rate of Pseudomonas spp in spring season was 28.1%, summer 21.1% and autumn 28.1%. Only 1 spring water was detected Pseudomonas spp in all seasons and 9 (15. 8%) were detected for 2 seasons and 13 (22.8%) were for only 1 season. Isolation rate of Pseudomonas spp at Mt. Cheonggye was 50% and followed by Mt. Bookhan 35.7%, Mt. Daemo 33.3%, Mt. Dobong 29.6%, Mt. Surak 25.9%, Mt. Woomyun 22.2% and Mt. Bulam 7.4%. Of 44 Pseudomonas spp, 22 strains (50%) were identified by Ps. putida, Ps. aeruginosa, Ps. tluorescens and Ps. mendocina were identified 6 strains (13.6%), respectively. 4 strains (9.1%) were identified by Ps. aureofaciens. Of 6 Ps. aieruginosa, serotype A was 2 strains, B, E, G, and K was 1 strain, respectively. Of 44 Pseudomonas spp, resistance rate to amoxicillin was 90.9% and followed by chloramphenicol 84.1%, tetracycline 84.1%, carbenicillin 81.8%, nalidixic acid 68.2%, neomycin 38.6%, streptomycin 31.8%, gentamicin 4.6%, kanamycin 4.6% and colistin 2.3%. Ps. aseruginosa was more sensitive to carbenicillin than other Pseudomonas spp isolated from spring waters in Seoul area but more resistant to kanamycin, and Ps. aureofaciens was no resistant to streptomycin. Among multiple drug resistance, resistance to 5 drugs was 31.8%, 4 drugs 15.9%, 7 drugs 13.6%, 1 drug and 2 drugs 4.6%, and 8 drugs 2.3%, respectively. The multiple resistance patterns detected highestly were NA-CB-C-TE-AMC (18.2%), NA-CB-N-C-TE-AMC (13.6%), CB-C-TE-AMC (11.4%) and NA-CB-N-C-TE-AMC-S (9.1%).

Background : This study was performed to investigate the morphological and growth characteristics and metabolomics approach for the diploid and tetraploid Polygonum multiflorum Thunb (PMT). Methods and Results : The present study was performed to investigate the effective induction method of tetraploid plants using colchicine treatment. To know the morphological and growth characteristics of PMT overground growing was investigated about the stem diameter, leaf length and width, a scape, inflorescence length, chlorophyll, seed length and seed width. Metabolite profiles from P. multiflorum were analyzed by an ultra performance liquid chromatography–quadrupole - time of flight (UPLC-Q-TOF) mass spectrometry (MS). The number of stomata and chlorophyll content was also higher in tetraploid. However, the physiological and morphological characteristics of the newly developed tetraploid showed noticeable results compared to the diploid plant. Also, the inherent characteristics of a newly acquired tetraploid also exhibited better results compared to diploid. Twenty one compounds including catechins from leaf and thirty compounds including stilbenes from roots were identified as major metabolites of PMT. The levels of most root metabolites except dethiobiotin, epigallocatechin gallate, procyanidin B1, epiafzelechin 3-O-gallate, and moupinamide from PMT were higher than those of diploid. The levels of 2,3-digalloylglucose, fallacinol, tricin, physcion-8-O-β-D-glucoside, torachrysone-8-O-β-D-glucopyranoside, questin, aloe-emodin 8-O-(6′-O-acetyl)-glucoside, polydatin, 2,5-dimethyl-7-hydroxychromone, 3,5,4'-trihydroxystilbene, aloe-emodin, myrciacitrin II and ω-hydroxyemodin from teraploid PMT were 342, 32, 31, 21, 11, 11, 9, 8, 6, 5, 4, and 2 times, respectively, higher. Conclusion : From above results, overground growing of tetraploid P. multiflorum is fairly better than that of native P. multiflorum.

Ionizing radiation directly and indirectly affects gene expression within the plant genome. To access the physiological response of rice to different types of ionizing radiation, rice seeds were exposed to gamma-ray and ion beam radiation. Exposure to ionizing radiation dramatically decreased the shoot length compared with non-irradiated plants. Fluorescence-activated-cell-sorting (FACs) was used to measure DNA contents. There were significant correlations of dose-dependent between irradiated plant and non-irradiated plant. The radicals induced by the ionizing radiation in the plant could be observed by electron spin resonance (ESR). It was confirmed that the number of free radicals in cell was greatly increased all irradiated plants than non-irradiated plant. A significant positive correlation was shown between ionizing radiation dose and signal intensity. In order to determine the Genetic diversity, AFLP analysis was conducted with the irradiated plant and non-irradiated plant. Based on band patterns, the cluster analysis was conducted to evaluate the genetic variation by using the UPGMA (Unweighted Pair Grouping Method of Averages). Genetic diversity of irradiated plants by low dose ion beam was the closest non-irradiated plant and irradiated by high dose gamma-ray was the furthest from non-irradiated. We describe the detailed methods of ionizing irradiation and discuss its applications in genetic research as well as plant breeding.

Ionizing radiation affects gene expression from plant genomes. To monitor the genome-wide transcriptional changes induced by three types of ionizing radiation, we used the rice RNA sequencing to identify genes that are up- or down-regulated by gamma rays (GAs), proton (PRs) and ion beams (IBs). The Oryza sativa jacalin-like lectin domain containing proteins (OsJAC1) gene was highly induced by GAs, PRs and IBs. OsJAC1 was selected based on the expression patterns of a genome-wide dataset of RNA sequencing. Many jacalin-related lectin genes have been shown to be associated with disease resistance, biotic and abiotic stress signaling. Therefore, we studied its expression pattern in response to different abiotic stress and phytohormone treatments. The expression patterns of OsJAC1 under two different abiotic stress conditions (salt and heat stress) and phytohormones (salicylic acid and methyl jasmonate) were examined. The transcripts of OsJAC1 were significantly induced in response to abiotic stress conditions, including salt and heat treatments. In addition, it was induced in response to the salicylic acid and methyl jasmonate treatments, respectively. To investigate the sub-cellular localization of OsJAC1, the gene was expressed as a fusion protein tagged with GFP, in tobacco leaf epidermis and examined under confocal microscope. The OsJAC1 was clearly localized at the nucleus. These results provide critical insights into the molecular functions of the rice jacalin-like lectin domain containing proteins as receptors of external signals.

‘Tocomi-1’, a new japonica rice cultivar derived from a 200 Gy gamma ray irradiation with high tocopherol content and red pericarp. The local adaptability test of MRXII-1001-1 was carried out from 2012 to 2014 and it was named as ‘Tocomi-1’ in 2014. This variety is medium matured with heading date of August 12 in honam plain area of Korea. This variety is about 80 cm tall culm length and 106 spikelets per panicle. Its 1,000 grain-weight of rice seeds is 25.4 g. The yield potential of this variety is about 5.15 MT/ha in local adaptability test for three years. This variety exhibited greater seed longevity than the Donganbyeo, indicating a crucial role for tocopherols in maintaining viability during quiescence, and displayed faster seedling growth during the early growth stage. Tocopherol contents was 50% higher than the Donganbyeo. To study the molecular mechanism underlying vitamin E biosynthesis, we examined the expression patterns of seven rice genes encoding vitamin E biosynthetic enzymes. Accumulation levels of the OsVTE2 transcript and OsVTE2 protein in the ‘Tocomi-1’ were significantly higher than in the Donganbyeo. Sequence analysis revealed that the ‘Tocomi-1’ harbored a point mutation in the OsVTE2 promoter region, which resulted in the generation of MYB transcription factor—binding cis-element. These results help identify the promoter regions that regulate OsVTE2 transcription, and offer insights into the regulation of tocopherol content in ‘Tocomi-1’.

To successful molecular breeding, identification and functional characterization of breeding related genes and development of molecular breeding techniques using DNA markers are essential. Although the development of a useful marker is difficult in the aspect of time, cost and effort, many markers are being developed to be used in molecular breeding and developed markers have been used in many fields. Single nucleotide polymorphisms (SNPs) markers were widely used for genomic research and breeding, but has hardly been validated for screening functional genes in olive flounder. We identified single nucleotide polymorphisms (SNPs) from expressed sequence tag (EST) database in olive flounder; out of a total 4,327 ESTs, 693 contigs and 514 SNPs were detected in total EST, and these substitutions include 297 transitions and 217 transversions. As a result, 144 SNP markers were developed on the basis of 514 SNP to selection of useful gene region, and then applied to each of eight wild and culture olive flounder (total 16 samples). In our experimental result, only 32 markers had detected polymorphism in sample, also identified 21 transitions and 11 transversions, whereas indel was not detected in polymorphic SNPs. Heterozygosity of wild and cultured olive flounder using the 32 SNP markers is 0.34 and 0.29, respectively. In conclusion, we identified SNP and polymorphism in olive flounder using newly designed marker, it supports that developed markers are suitable for SNP detection and diversity analysis in olive flounder. The outcome of this study can be basic data for researches for immunity gene and characteristic with SNP.

Polyethyleneglycol-adsorbed–superoxide dismutase (PEG-SOD), has been proposed as an effective agent for reducing free radical-mediated injury. The objective of this study was to investigate a protective effect of PEG-SOD supplementation on ovarian tissue during transplantation. Ovaries from F-1 mice were collected and vitrified. After warming, ovaries were autotransplanted under kidney capsule. Mice were randomly divided into four groups according to dose of PEG-SOD, (0 U/ml, 100 U/ml, 1,000 U/ml and 10,000 U/ml respectively). Grafted ovaries were retrieved 2, 7 and 21 days later. PEG-SOD was treated by intraperitoneal injection once every 48 hours and especially for 21 days group, after first week treatment, PEG-SOD was treated once every 4 days. Morphology of ovaries was assessed histological analysis and ELISA for FSH was performed to evaluate restoration of ovarian function. In 2 days groups, morphologically intact follicle ratio of 10,000 U/ml group was significantly higher than other groups. In 7 days groups, morphologically intact follicle ratio was significantly higher in all treatment groups. In 21 days groups, there was no significant difference of intact follicle ratio in total follicles in all groups but intact primordial, primary and secondary follicles ratio was higher in 10,000 U/ml group. FSH levels in blood serum were decrease as time goes on, but there is no statistical difference in each groups. In conclusion, the data of the present study show that PEG-SOD has a beneficial effect on preservation of the morphologically intact follicle.

Ovarian tissue cryopreservation and transplantation causes follicle depletion. To overcome this problem, we investigate the effect of Anti-Müllerian hormone (AMH), a follicle recruitment control hormone, supplementation before and/or after mouse ovarian transplantation. A total of 120 5-week-aged BD F-1 female mice were used. The mice were randomly divided into four groups according to AMH doses (0, 5, 25, 125 μg/mL, respectively). AMH was injected intraperitoneally on every other day for a week before, after, or before and after transplantation of ovaries under kidney capsules was performed. One week after transplantation, follicular normality was evaluated by histological analysis and TUNEL assay. In Group A and C, morphologically intact follicle (G1) ratios of AMH treated groups showed no statistically significant difference. In Group B, G1 ratios of 25 and 125 μg/mL of AMH treated groups were higher than those of 5 μg/mL treated group, but there was no improvement in G1 ratio after AMH treatment. In every group, apoptotic follicle ratios did not show any trend according to AMH treatment. Proportions of primordial follicle were not significantly different according to AMH treatment in all groups. The result of the present study demonstrated that AMH treatment during on transplantation of cryopreserved ovaries has no significant effect on follicle survival and prevention of follicle depletion.

Objective : To investigate the effects of Simvastatin and Methylprednisolone on ovarian tissue cryopreservation and transplantation using mouse models. Methods : The mice were randomly distributed into 1 control and 3 experimental groups. The B6D2F1 mice were given oral Simvastatin (5 mg/kg), intravenous Methylprednisolone (15 mg/kg), or a combination of both at 2 hours before ovariectomy. Same volume of normal saline was given perorally in the control group at 2 hours before ovariectomy. The ovarian tissues were vitrified accrording to our protocols. The vitrified ovaries were warmed 1 week later and auto-transplanted under bilateral kidney capsules. The ovaries and blood sera were collected at 2, 7 or 21 days after transplantation. Histological analysis, TUNEL assay, immuno-histochemistry for CD31, serum AMH level and embryonic development after in vitro fertilization were assessed for evaluation. Results : With regard to the total grade 1 follicle rate, both Simvastatin or Methylprednisolone treated groups were significantly increased at 2, 7 or 21 days after transplantation (except Simvastatin treated group at 7 days). A combination of Simvastatin and Methylprednisolone group was significantly improved in terms of the total G1 follicle rate, apoptotic follicle rate, CD31 positive area and serum AMH after ovarian tissue transplantation. However, there were no statistically difference with respect to the oocyte maturation rate, blastulation rate, and the other embryonic development parameters after in vitro fertilization procedure among the four groups. Conclusion : Our results suggest that combined donor Simvastatin and Methylprednisolone have beneficial effects on the quality and function of transplanted ovarian tissues.

Perilla frutescens (L.) is an edible plant, not only used as s food ingredient, but also in skin cream, soaps, and medicinal preparetions. ‘Atom-Ros’, a perilla (Perilla frutescenc (L.) Britt. cv. Chookyoupjaso was developed in 1995 by 200 Gy gamma irradiation-mutagenesis. This new cultivar has high rosmarinic acid content more than two fold compare with ‘Chookyoupjaso’ control. The observed phenotypical difference was changed leaf color of the ‘Atom-Ketone’ from violet to green. The yield potential of this cultivar (123.5 kg/10a) was 2.14 fold higher than that of ‘Chookyoupjaso’ (57.65 kg/10a). The methanol extracts of ‘Atom-Ros’ were tested for inhibition of nitric oxide (NO) production in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophase cells. Atom-Ros showed significant inhibition of NO production. This rosmarinic acid extracted from ‘Atom-Ros’ has a good potential to be developed as an antioxidant agent.

Perilla frutescens (L.) is an annual herbaceous and ornamental plant in the Lamiaceae family. Perilla frutescens (L.) Britt.cv.Chookyoupjaso were irradiated using a 200 Gy gamma ray in 1995. By HPLC analysis, this new cultivar significantly induced isoegomaketone content compared with ‘Chookyoupjaso’ control. The phenotypical difference was the changed leaf color of the ‘Atom-Ketone’ from violet to green. The yield potential of this cultivar (106 kg/10a) was 1.83 folds higher than that of ‘Chookyoupjaso’ (57.65 kg/10a). The methanol extracts of ‘Atom-Ketone’ inhibit nitric oxide (NO) production in LPS-stimulated RAW 264.7 cells. This extract was further partitioned using ethyl acetate (EtOAc), butanol (BuOH), and water. The EtOAc fraction (EF-Atom-Ketone) was evaluated for antiinflammatory activities. These results indicated that the EF-Atom-Ketone reduced NO production by inhibiting inducible nitric oxide synthase (iNOS) expression. The EF-Atom-Ketone treatment also significantly diminished expression of MCP-1 and IL-6. Therefore, ‘Atom-Ketone’ reveals the potential therapeutic use of bioactive

This study conducted a laboratory simulation using artificial and natural rainfall in order to investigate the runoff characteristics of livestock resources through the analysis of the surface runoff water and infiltration water by rainfall intensity and fertilization level. Cattle manure and pig liquid fertilizer was used as livestock resource. As a result of this study, it was observed that the surface runoff occurred over 32 mm/hr rainfall intensity, and flow rate of the surface runoff water and the runoff ratio of contaminant parameters from livestock resource was increased, as rainfall intensity was stronger. With doubled fertilization level, T-N increased in compost and the amount of CODMn runoff also considerably increased in liquefied fertilizer. In the case of natural rainfall, the runoff ratio of T-P clearly increased in compost and the T-N of final surface runoff ratio in compost and liquefied fertilizer was ranged from 0.13047 to 0.13623 with stronger rainfall intensity.

FGF9/16/20 signaling pathway specify the developmental fates of notochord, mesenchyme, and neural cells in ascidian embryos. Although a conserved Ras/MEK/Erk/Ets pathway is known to be involved in this signaling, the detailed mechanisms of regulation of FGF signaling pathway have remained largely elusive. In this study, we have isolated Hr-Erf, an ascidian orthologue of vertebrate Erf, to elucidate interactions of transcription factors involved in FGF signaling of the ascidian embryo. The Hr-Erf cDNA encompassed 3110 nucleotides including sequence encoded a predicted polypeptide of 760 amino acids. The polypeptide had the Ets DNA-binding domain in its N-terminal region. In adult animals, Hr-Erf mRNA was predominantly detected in muscle, and at lower levels in ganglion, gills, gonad, hepatopancreas, and stomach by quantitative real-time PCR (QPCR) method. During embryogenesis, Hr-Erf mRNA was detected from eggs to early developmental stage embryos, whereas the transcript levels were decreased after neurula stage. Similar to the QPCR results, maternal transcripts of Hr-Erf was detected in the fertilized eggs by whole-mount in situ hybridization. Maternal mRNA of Hr-Erf was gradually lost from the neurula stage. Zygotic expression of Hr-Erf started in most blastomeres at the 8-cell stage. At gastrula stage, Hr-Erf was specifically expressed in the precursor cells of brain and mesenchyme. When MEK inhibitor was treated, embryos resulted in loss of Hr-Erf expression in mesenchyme cells, and in excess of Hr-Erf in a-line neural cells. These results suggest that zygotic Hr-Erf products are involved in specification of mesenchyme and neural cells.

VitE (tocotrienols and tocopherols) are micronutrients with antioxidant properties synthesized by photosynthetic bacteria and plants that play important roles in animal and human nutrition. A new mutant line, T1001-1, was isolated from in vitro mutagenized population by ionizing radiation and shown to have increased VitE contents. The total VitE content was 26% increased in the T1001-1 mutant seeds compare with cv. Dongan (wild-type). In addition, we showed that the mutant confers retarded seedling growth during the early seedling growth stage in rice. To study the molecular mechanism of VitE biosynthesis, we used the rice microarray to identify genes that are upor down-regulated in T1001-1 mutant. In addition, we identified differentially regulated pathway using MapMan analysis, which provides deep insight into changes in transcript and metabolites. Our results enhanced the transcription of genes involved in starch and lipid metabolism in T1001-1 mutant. To identify the molecular mechanisms of the events involving transcription factors in tocopherol accumulation, we compared the expression patterns of transcription factors. The AP2-EREBP, WRKY, C2H2 transcription factor were up-regulated, whereas the MYB family was down-regulated in T1001-1 mutant. Our results demonstrate change of important transcript in high level of VitE accumulating rice mutant.

Manganese () is a trace element that is essential for normal physiology, and is predominantly obtained from food. Several lines of evidence, however, demonstrated that overexposure to exerts serious neurotoxicity, immunotoxicity and developmental toxicity, particularly in male. The present study aimed to evaluate the effect of 0, 1.0, 3.3, and 10 mg/kg/day doses of on the reproductive organs in the immature female rats. Rats (PND 22; S.D. strain) were exposed to () dissolved in drinking water for 2 weeks. The animals were sacrificed on PND 35, then the tissues were immediately removed and weighed. Histological studies were performed using the uteri tissue samples. Serum LH and FSH levels were measured with the specific ELISA kits. Body weights of the experimental group animals were not significantly different from those of control group animals. However, ovarian tissue weights in 1 mg and 3.3 mg dose groups were significantly lower than those of control animals (p<0.05 and p<0.01, respectively). Uterine tissue weights of 3.3 mg dose groups were significantly lower than those of control animals (p<0.01), while the 1 mg dose and 10 mg dose failed to induce any change in uterine weight. Similarly, only 3.3 mg dose could induce the significant decrease in the oviduct weight compared to the control group (p<0.05). Non-reproductive tissues such as adrenal and kidney failed to respond to all doses of exposure. The uterine histology revealed that the exposure could affect the myometrial cell proliferation particularly in 3.3 mg dose and 10mg dose group. Serum FSH levels were significantly decreased in 1mg dose and 10 mg groups (p<0.05 and p<0.01, respectively). In contrast, treatment with 1 mg dose induced a significant increment of serum LH level (p<0.05). The present study demonstrated that exposure is capable of inducing abnormal development of reproductive tissues, at least to some extent, and altered gonadotropin secretions in immature female rats. Combined with the well-defined actions of this metal on GnRH and prolactin secretion, one can suggest the might be a potential environmental mediator which is involved in the female pubertal process.