The aim of this study was performed to evaluate the effects of ice-binding protein from the arctic yeast Leucosporidium (LeIBP) supplementation on cryopreservation of boar sperm. The collected semen was diluted (1.5×108/ml) in lactose egg yolk (LEY) and cooled at 5°C for 3 h. The cooled semen was then diluted (1×108/ml) in LeIBP containing LEY with 9% glycerol and maintained at 5°C for 30 min. The semen was divided into six experimental groups (control, 0.001, 0.005, 0.01, 0.05 and 0.1 mg/ml of LeIBP). The straws were kept on above the liquid nitrogen (LN2) vapors for 20 minutes and then plunged into LN2. After thawing, computer-assisted sperm analysis was used for sperm motility and flow cytometry was performed to assess the viability, acrosome integrity (FITC-PSA/PI), ROS (DCF/PI), lipid peroxidation (BODIPY C11/PI) and apoptosis (Annexin V/PI), respectively. No significant responses were observed for sperm motility. However, sperm viability was significantly increased on 0.05 and 0.1 mg/ml of LeIBP groups compared to control (P < 0.05). In addition, acrosome integrity was significantly increases LeIBP groups (P < 0.05) and both ROS and lipid peroxidation level were lower in all LeIBP groups than those of control (P < 0.05). On the other hand, a significant higher apoptosis rate was observed in 0.05 and 0.1 mg/ml of LeIBP groups compared to control (P < 0.05). It can be assumed that a supplementation of LeIBP in boar sperm freezing extender is an effective method to increase the sperm qualities after cryopreservation.

This study is to investigate the effects of binding agents which contain sodium alginate, corn starch, carrageenan, and guar gum and on the physicochemical and sensory characteristics of pork patties. The pork patty formulation was prepared with 0.5 percent of the each binding agents. Some selected properties of binding agents, such as, pH, cooking yield, shrinkage, water holding capacity, shear force, color (L*, a* and b*), and TPA were determined. The pH of pork patties containing corn starch and control was significantly same but the others were significantly difference. The cooking yield of pork patties containing sodium alginate were significantly higher than the control (p<0.05). The shrinkage of pork patties containing corn starch, carrageenan, and was significantly lower than the control (p<0.05). The water holding capacity of pork patties containing corn starch was significantly lower than the others. The shear force of pork patties containing corn starch, carrageenan, and guar gum, and were significantly lower than control. The color of pork patties containing corn starch, carrageenan, guar gum was significantly low L* and high a*. The hardness and springiness of pork patties containing sodium alginate were significantly higher than the control. The cohesiveness of pork patties containing each binding agents was significantly lower than the control. The gumminess and chewiness of pork patties containing corn starch, carrageenan, and guar gum were significantly lower than the control. Therefore, pork patties containing sodium alginate are useful for making pork patties with desirable quality characteristics.

As the ridges become larger and larger, a structural type that enables effective utilization of the long span and space is required. In the construction stage, the steel column supports the installation load. However, in order to secure the stability against the out - of - plane deformation of the steel column due to the lateral pressure when the concrete is laid, a binding frame is installed inside the steel pipe at constant intervals to resist the concrete installation pressure. When the concrete is cured and its performance as a composite section is exerted, a stress is generated which pushes the steel pipe out of the plane by the column compressive force. In this case, since the binding frame controls the deformation, the local buckling is delayed and the constraining effect on the concrete is increased. In order to evaluate the structural performance and behavior of the composite mega column according to the eccentricity effect and the effect of the binding frame, we carried out a structural test by fabricating eight monopole specimens with the binding frame reinforcement, reinforcing gap, reinforced cross section and eccentricity , And the experimental results are compared with the KBC2016 design formula.

Composite columns are increasingly used due to the construction of super-tall buildings and large-scale buildings. Studies on the shapes of and construction technologies for structural members using steel tubes are being conducted actively. Welded built-up CFT columns previously developed and commercialized by the authors of this study (ACT-1 columns) are structurally stable and economically efficient. However, the 1m limit in the width of the columns and their small interior spaces impose a difficulty in installing reinforcing materials and thus deteriorate the ease and efficiency with which they are constructed. This study suggests placing thick plates at the centers of the surfaces of the existing ACT-1 column and installing a binding frame (binding frames) at the central thick plates to enhance the integrity and resist lateral pressure caused by concrete casting. Finite element analysis was conducted with the variables of the number and cross-sectional size of the binding frame and the cross-sectional size of the steel tube to estimate the structural behavior of the steel tubes. Hydraulic tests were conducted to analyze load-displacement relations and identify the influence of the binding frames on the relations. The variables in the tests were the number and cross-sectional size of the binding frame, welding details, column joint and the cross-sectional size of the steel tube

Actin nucleation factors, which promote the formation of new actin filaments, have emerged in the last decade as key regulatory factors controlling asymmetric division in mammalian oocytes. Actin nucleators such as formin-2, spire, and the ARP2/3 complex have been found to be important regulators of actin remodeling during oocyte maturation. We found that actin nucleation promoting factor called WASP homolog-associated protein with actin, membranes and microtubules (WHAMM) play crucial roles in mouse oocyte maturation by generation of ER-associated actin filaments during meiotic spindle migrations. We also investigate regulatory mechanism of actin nucleator spire and discovered the novel roles of Zinc in regulating spire localization and cytoplasmic actin mesh formation. Another class of actin-binding proteins including cofilin, tropomyosin, capping proteins and tropomodulin, are thought to control actin cytoskeleton dynamics at various steps of oocyte maturation. The heterodimeric actin-capping protein (CP) binds to the fast-growing (barbed) ends of actin filaments and plays essential roles in various actin-mediated cellular processes. When CP is knockdowned or inhibitory component was overexpressed, asymmetric divisions of oocytes have been compromised. It turns out that knockdown or inhibition of CP deplete cytoplasmic actin mesh level, which have been known to be essential for maintain cytoplasmic actin mesh. Another actin binding proteins, tropomodulin 3 (Tmod3), binds to the slow-growing end of actin filaments and knockdown or expression deletion mutant of Tmod3 also decrease actin mesh level in maturing oocyte and it severely ablated asymmetric division of oocyte. Finally, tropomyosin 3, actin filament binding proteins protect actin filament from depolymerization, is also important to maintain cortex integrity in maturing oocyte, therefore showed the importance maintenance of actin filaments during oocyte maturation. Taken together, our study on various actin nucleator and actin binding study showed the importance of actin dynamics in mammalian oocyte maturation and early embryonic developments.

This study was carried out to investigate the effects of tissue inhibitor of matalloproteinase-1 (TIMP-1), Activin A and Heparin binding epidermal growth factor (HB-EGF) on in vitro production of bovine embryos. In experiment 1, presumptive zygotes were cultured in the medium supplemented with TIMP-1 (0.5 μg/ml), Activin A (100 ng/ml), or HB-EGF (100 ng/ml) at 39 ℃ in a humidified atmosphere of 5% (v/v) CO2, 5% (v/v) O2 and 90% (v/v) N2. In experiment 2, TIMP-1 + HB-EGF or Activin A + HB-EGF combinations were supplemented in the culture medium. The developmental rate to blastocysts, hatching rate and total cell numbers of the blastocysts were evaluated in both experiments. The embryos cultured in medium without growth factor supplementation was used as control group. In experiment 1, the embryos cultured in medium supplemented with TIMP-1 and Activin A showed significantly higher developmental rate to blastocysts than those cultured with HB-EGF and control (36.9%, 34.1%, 21.2% and 23.1%, respectively) (P<0.0001). However, the hatching rate of blastocyst was significantly higher in embryos with HB-EGF than those with TIMP-1, Actvin A and Control groups (84.4%, 58.8%, 51.4% and 49.3%, respectively) (P<0.001). Total cell number per blastocyst was also significantly higher in embryos with HB-EGF group (174.3±2.5) than those with TIMP-1, Activin A (149.7 and 150.0, respectively) (P<0.05) and Control (119.0) (P<0.001). In experiment 2, embryos cultured with combined treatment of Activin A and HB-EGF resulted in significantly higher rates of blastocysts formation (48.0%), hatching rate (89.7%) and total cell number in blastocyst (182.3±2.1) than those with TIMP-1 and HB-EGF combination group (32.0%, P<0.001; 76.6%, P<0.05; 165.7±4.2, P<0.001, respectively). Our data demonstrate that in vitro production of bovine embryos could be improved by combined supplementation of Activin A and HB-EGF in culture medium.

Toll–interleukin 1 receptor (TIR) superfamily는 intracellular TIR domain에 존 재하며, proinflammatory cytokines을 생산하는 transcription factor NF-κB의 활성 화에 의해 선천성 면역을 개시한다. 본 연구에서는 갈색거저리 유충을 이용하여 Mycoplasma genus와 유사한 acholeplasma lysate를 접종하여 비교 유전체학적 방 법을 통하여 갈색거저리에서 면역반응에 관여하는 t1/st2 receptor binding protein 을 동정하였으며, 그 구조 분석을 위한 기초 데이터를 확보하고자 하였다. IL1RL1 (Interleukin 1 receptor-like 1) gene는 Toll-like receptor superfamily로써 사람에서 발견되며 ST2는 Toll-interleukin 1 receptor family의 member이자 endotoxin tolerance 유지에 중요한 역할을 하는 것으로 알려졌다. 갈색거저리 유충에 acholeplasma lysate를 처리하기 전과 후의 각 샘플들로부터 cDNA library를 구축 한 후 random sequencing 을 통해 분석되어진 서열들 중 증감하는 유전자들을 동정 하였고, 그 중 acholeplasma 처리 후 약 4배 정도 발현이 증가한 t1/st2 receptor binding protein 의 서열을 추출한 후 단백질의 2차 구조를 예측한 결과 alpha helix 구조는 서열상에서 8영역으로 예측되었으며 beta sheet 구조는 서열상에서 1영역 에서 존재하고 있어 후속연구를 진행하고 있다.

Methoxychlor (MXC) was developed to be a replacement for the banned pesticide DDT. HPTE [2,2-bis (p-hydroxyphenyl) -1,1,1-trichloroethane], which is an in vivo metabolite of MXC, has strong oestrogenic and anti-androgenic effects. MXC and HPTE are thought to produce potentially adverse effects by acting through oestrogen and androgen receptors. Of the two, HPTE binds to sex-steroid receptors with greater affinity, and it inhibits testosterone biosynthesis in Leydig cells by inhibiting cholesterol side-chain cleavage enzyme activity and cholesterol utilisation. In a previous study, MXC was shown to induce Leydig cell apoptosis by decreasing testosterone concentrations. I focused on the effects of MXC on male mice that resulted from interactions with sex-steroid hormone receptors. Sexsteroid hormones affect other organs including the kidney and liver. Accordingly, I hypothesised that MXC can act through sex-steroid receptors to produce adverse effects on the testis, kidney and liver, and I designed our experiments to confirm the different effects of MXC exposure on the male reproductive system, kidney and liver. In these experiments, I used pre-pubescent ICR mice; the puberty period in ICR mice is from postnatal day (PND) 45 to PND60. I treated the experimental group with 0, 100, 200, 400 mg MXC/kg b.w. delivered by an intra-peritoneal injection with sesame oil used as vehicle for 4 weeks. At the end of the experiment, the mice were sacrificed under anaesthesia. The testes and accessory reproductive organs were collected, weighed and prepared for histological investigation. I performed a chemiluminescence immune assay to observe the serum levels of testosterone, LH and FSH. Blood biochemical determination was also performed to check for other effects. There were no significant differences in our histological observations or relative organ weights. Serum testosterone levels were decreased in a dose-dependent manner; a greater dose resulted in the production of less testosterone. Compared to the control group, testosterone concentrations differed in the 200 and 400 mg/kg dosage groups. In conclusion, I observed markedly negative effects of MXC exposure on testosterone concentrations in pre-pubescent male mice. From our biochemical determinations, I observed some changes that indicate renal and hepatic failure. Together, these data suggest that MXC produces adverse effects on the reproductive system, kidney and liver.

Cry3 toxins from Bacillus thuringiensis are used as biopesticides and the transgenic crops to control of leaf-feeding beetles. Cadherin in insect midgut epithelium is identified as receptor for Cry toxins in several insects and some domains of it synergizes Cry toxicity. Cadherin (DvCad1-CR8-10) fragment of Diabrotica virgifera virgifera enhanced Cry3Bb toxicity to Colorado potato beetle (CPB), Leptinotarsa decemlineata. Single cadherin repeat (CR) fragment of DvCad1-CR8-10, have a strong binding affinity to the active Cry3Bb toxin. The dissociation constant Kd value of CR8, CR9, and CR10 were 4.9 nM, 28.2 nM, and 4.6 nM, respectively. Interestingly, CR8 and CR10 enhanced Cry3Bb toxicity against CPB and Lesser mealworm (LMW), Alphitobius diaperinus, neonates in up to 2-folds. The DvCad1-CR10 peptide is further analyzed by in-frame deletion to determine the active site for Cry3Bb toxin. The active site is narrowed down to a 26 amino acid locating in the N-terminal region of DvCad1-CR10 that either synergized Cry3Bb toxicity on the CPB and LMW neonates in 3-folds or bound to the toxin with high affinity. The extent of Cry3Bb toxin enhancement by the activie site in DvCad1-CR10 may have practical application for control of CPB and LMW.

Immune defense is indispensible for insect survival. However, uncontrolled and excessive immune responses would be highly detrimental and energy-consuming processes. An insect cytokine, plasmatocyte-spreading peptide (PSP), induces hemocyte-spreading behavior as well as activating phenoloxidase (PO) in the beet armyworm, Spodoptera exigua. A hemocyte transcriptome of S. exigua contains a partial sequence of a putative PSP-binding protein (SePSP-BP). SePSP-BP was expressed in all developmental stages especially in hemocytes and fat body. A quantitative RT-PCR showed that the bacterial infection significantly up-regulated the expression level of SePSP-BP. A double-stranded RNA specific to SePSP-BP (dsRNASePSP-BP) was injected and suppressed SePSP-BP expression even in response to bacterial challenge. The larvae treated with dsRNASePSP-BPsuffered high mortality to infection of nonpathogenic bacteria and prolonged high PO activity after the immune challenge. These results suggest that SePSP-BP may play a role in suppressing immune responses as a negative controller

To clarify the role of stem cells in hepatocarcinogenesis, octamer-binding transcription factor 4 (Oct4) expression was investigated in mouse liver and embryonic cell lineages. In vivo, at 14 days of age, ten ICR mice were divided into two groups and treated with saline or diethylnitrosamine (DEN), and were sacrificed at 6 h after treatment. Livers were fixed in 10% neutral phosphate buffered formalin, embedded in paraffin, sectioned to a thickness of 5 μm, and immunohistochemical analysis of Oct4 was performed. In vitro, mouse embryonic stem cells, hepatic progenitor cells and hepatocytes, representing 0, 22, and 40 days of differentiation, respectively, were treated with DEN at four doses (0, 1, 5 and 15 mM; G1, G2, G3 and G4, respectively) for 24 h and RNA was isolated; Oct4 and Gadd45a mRNA were investigated. In vivo, Oct4 expression was not detected in saline-treated livers. However, its expression was observed in hepatocytes of mice treated with DEN, showing cytoplasmic staining. In vitro, Oct4 expression differed significantly for G4 on day 0 (P<0.05) and for G2 on day 22 (P<0.01) and G3 and G4 on day 40 (P<0.05 and P<0.01, respectively) compared with G1 at each time point. Gadd45a expression differed significantly in G4 (P<0.01) on day 0 and G4 on day 40 (P<0.01), compared with that of G1 at each time point. Taken together, Oct4 expression was increased by treatment with DEN in hepatocytes, however, not in embroyonic stem cells and hepatic progenitor cells. This finding suggests that Oct4 expression may be modulated in hepatocarcinogenesis induced by DEN.

Insulin/insulin-like peptide-binding protein (IBP) is abundantly found in venom of the solitary hunting wasp, Eumenes pomiformis (Hymenoptera: Eumenidae). E. pomiformis IBP (EpIBP) is most similar to insect IBP-like proteins that are known to inhibit insect growth and insulin signaling. To investigate the toxicity and target protein, EpIBP was in vivo expressed by Escherichia coli. Spodoptera exigua (Lepidoptera: Noctuidae) larvae injected with EpIBP showed a 20% lower pupation rate than the control larvae, although their body weight was not significantly different from the control when the larvae were provided artificial diet after the injection. EpIBP extended the larval stage without inducing paralysis of S. exigua larvae. To investigate the effects of EpIBP on caterpillar under a starvation condition, survivorship and body weight of the EpIBP-injected were evaluated without providing artificial diet until all the larvae died. The survivorship of the EpIBP-injected larvae was 24-36% higher than the control larvae at 4-5 d post-injection. The body weight of the control larvae reduced to 59% that is approximately 10% lower than the body weight of the EpIBP-injected larvae. These results suggest that EpIBP might inhibit the metabolism of the caterpillars, which is likely related with insulin-like peptide signaling pathway, suppress the loss of body weight and eventually extend the larval stage. An EpIBP-binding protein (EpIBPBP) isolated by immunoprecipitation was matched with a coiled-coil domain-containing protein of the fruit fly. The full-length sequence analysis of EpIBPBP is in progress.

The Oct-4 (octamer-4), a member of the POU family transcription factor, is expressed in early mouse embryogenesis and in pluripotent embryonic stem (ES) lines. Oct-4 expression is thought to remain confined to the germline after gastrulation in the embryo. Therefore, the study was designed to, study the location of Oct-4 protein in the ovaries, placenta and testis of Korean native cattle (Hanwoo). Expression of Oct-4 mRNA in the ovaries and placenta of bovine was confirmed by RT-PCR and immunohistochemical analysis. Oct-4 was expressed in granulosa, thecal cells irrespective of the shape and size of follicles and endometerium of Korean native cattle (Hanwoo). Expression of Oct-4 was profound in all the tissues of Korean native cattle (Hanwoo) suggestung their role in them. Oct-4 localization and expression could contribute to further developmental studies in Korean native cattle (Hanwoo).

alcineurin (CN) is a calcium and calmodulin-depedent serine/threonine phosphatase. CN plays an important role in various biological processes including cell proliferation, cardiovascular, skeletal muscle development and apoptosis. In rheumatoid arthritis (RA), CN plays a role synoviocyte activation and arthritis progression. The selective inhibition of CN by the over-expression of CN-binding protein 1 (Cabin1). In the present study, joint restricted transgenic mice expressing the human Cabin1(hCabin1) were generated, driven by type II collagen promoter and efficiency of these mice was investigated by experimental arthritis. These transgenic mice successfully expressed hCabin1 in joint tissue as well as other organs like the liver, the heart, and the brain. The joint specific over-expression of hCabin1 reduced the disease severity during collagen-induced arthritis. In fibroblast-like synoviocytes (FLSs) from hCabin1 transgenic mice, the productions of these cytokines including, TNF-α, IL-1β and IL-6 were decreased and MMPs was also depressed in transgenic mice FLS. In addition, the expression of proapoptotic p53, p21, caspase-3, caspase-9 and Bax increased in transgenic mice, indicating that hCabin1 may induce FLS death by regulating the expression of Bcl-2, p53, p21, caspase-3, casepase-9 and Bax. It is expected that these findings will provide a more knowledge about the pathogenic mechanisms of rheumatoid arthritis and a potential animal model of the choronic inflammatory conditions, including atherosclerosis and transplantation.