This study was conducted to examine the effect of IRES controlled reporter gene on screening and production of recombinant human erythropoietin (EPO) proteins from cultured CHO cells. The cDNA was cloned for EPO from human liver cDNA. Using site-directed mutagenesis, we generated recombinant human EPO (rhEPO) with two additional N-glycosylations (Novel erythropoiesis-stimulating protein: NESP). Wild type hEPO and NESP were cloned into expression vectors with GFP reporter gene under regulatory control of CMV promoter and IRES so that the vectors could express both rhEPO and GFP. The expression vectors were transfected to cultured CHO-K1 cells. Under microscopy, expression of GFP was visible. Using supernatant of the culture, ELISA assay, immunocytochemistry and in vitro assay using EPO dependant cell line were performed to estimate biological activity to compare the production characteristics (secretion levels, etc.) between rhEPO and NESP. The activity of NESP protein, obtained by mutagenesis, was described and compared with its rhEPO counterpart produced under same conditions. Although NESP had less secretion level in CHO cell line, the biological activity of NESP was greater than that of rhEPO. These results are consistent with previous researches. We also demonstrated that rhEPO and GFP proteins expressed simultaneously from transfected CHO cell line. Therefore we conclude that use of GFP reporter gene under IRES control could be used to screen and produce rhEPO in cultured CHO cells.

The purpose of this study was to investigate how comparative menu price, human service, amenity, and menu quality affected menu value, and how menu value influenced revisit intention. The model was tested in a family restaurant setting using a sample of customers visiting and enjoying menu in Daegu metropolitan city. Empirical results confirmed that not only do human service, amenity and menu quality increase menu value but that comparative menu price reduces menu value. Menu value was also found to be a significant antecedent of revisit intention. The results obtained have major implications for family restaurant marketers as well as for future research. First, family restaurant marketers should pay attention to menu pricing, as menu price decreased menu value. Second, family restaurant marketers should try to increase menu value through training of human service. Third, family restaurant marketers should try to add menu value by way of recruiting high-skilled cook. Fourth, family restaurant marketers should make efforts to attract customers through interior design.

This experiment was performed to study the biocompatibility of xenograft materials (ABBM. coralline HA). Both autogenous bone grafts and allogenic banked bone were frequently and successfully used to promote regeneration of parts of skeleton. The use of these types of grafts were limited by the cost of donor site operation for autogenous boneor by fear of the risk of infection of allogenic materials. Another type of graft is xenograft which include ABBM and coralline HA. For investigating the biocompatibility, generally many investigators used cancer cell lines or animal cell lines. But cancer cell lines and animal cell lines had functioned different metabolism from normal human cell. So the experiment used normal human osteoblast for compare the biocompatibility of ABBM with coralline HA which were fixed in 24 well base contained culture medium. After 1st, 3rd, 7th, 14th, 28th days, the culture medium were taken out and checked the concentrations ofcalcium( Ca), inorganic phosphate(IP) and alkaline phosphatase(ALP). In another method, histologic samples were investigated after 8weeks of xenograft materials implantated on rabbit's tibia, the bone was cut and made undecalcified ground samples and checked with fluorecent microscope, polarizing microscope, reflection electron microscope and electron probe microanalysis. The statistical results of concentrations (Ca, IP, ALP) of materials in the culture medium have decreasedby day's, which meant that xenograft materials were effective for bone remodelling. The concentrations in the culture medium of ABBM were lower than that of coralline HA, that meant that biocompatibility of ABBM were superior than that of coralline HA. Histologic samples showed that ABBM had better bone remodelling effect than coralline HA. ABBM showed good alizarin red marking lines, more deposition of Ca, IP, and dense color of bone around newly formed osteon and bone trabeculae. it was concluded that ABBM was more biocompatible than corallineHA in vivo and in vitro test

It has been reported that light-emitting diodes(LED) can be used in the treatment of oral diseases. Although bio-stimulatory effects of LED irradiation such as promotion of wound healing have been well known, there are few reports about molecular mechanisms associated with wound healing by LED irradiation. The purpose of the present study was to investigate the expression pattern of various extracellular matrix(ECM) molecules in relation to wound healing after LED irradiation on primary human gingival fibroblasts(hGFs) in vitro. The source of light for irradiation was a continuous-wave LED emitting at a wavelength of 635 nm, and manufactured that energy density was 5 mW/cm2 on sample surfaces. The hGFs were irradiated for 1 hour at 37℃ in 5% CO2 humidified chamber. Experimental samples were acquired at 0 (right after irradiation), 24 and 48 hour after irradiation. To investigate the molecular mechanisms associated with wound healing, we examined the mRNA expression of 6 types of collagens, 7 types of matrix metalloproteinases(MMPs) and 4 types of tissue inhibition of metalloproteinases(TIMPs) after LED irradiation by RT-PCR. The mRNA expression of collagen 4, MMP-3, 9, and 16, and TIMP-3 was influenced by LED irradiation. Generally, the collagen expression of the irradiation group was slightly increased, particularly collagen 4 was significantly increased at 0 hour. The expression of MMP-3 was increased at 0 and 24 hours and MMP-16 was increased at 24 hours, respectively. The expression of MMP-9 was decreased at 0 hour and increased at 24 and 48 hours. The mRNA expression of TIMP-3 was significantly decreased at 24 and 48 hours after irradiation. These results suggest that the altered expression of ECM molecules after LED irradiation may contribute to the accelerated wound healing.

Pro-inflammatory cytokines are important mediators of cutaneous cellular activities during many oral mucosal diseases. IHOK culture model transfected by E6/E7 genes provide further evidence for the role of HPV in tumorogenesis. It is interesting to investigate cytokine expression of immortalized human oral keratinocyte(IHOK). The purpose of this study were to analysis cytokine mRNA expression levels of NHOK and IHOK by RT-PCR. IHOK showed about 5 fold increases of IL-6 compared with NHOK, while TNF-α was the lowest. It suggested that immortalization of NHOK with E6/E7 could result in elevated expression of IL-6, and IHOK be in the intermediate stage of oral carcinogenesis.

본 논문에서는 세계적으로 널리 사용되어지고 있는 초고층 건축물의 진동사용성 평가기준이 검토되었다. 초고층 건축물의 진동에 대한 사용성 평가기준은 바람에 의한 초고층 건축물의 가속도 응답의 크기로 나타내는 것이 일반적이다. 건축물의 가속도의 응답의 크기를 산정함에 있어서 두 가지의 서로 다 른 척도, 즉 최대가속도 또는 RMS가속도를 각 기준에서 채택하고 있는데 이에 대한 상이점을 토의하고 각 국의 진동사용성 평가기준을 고찰함으로써 우리나라에는 어떤 진동사용성 평가기준이 적합한가에 대한 논의를 시작하고자 하였다. 그리고 건축물의 각기 다른 응답을 조사하여 최대가속도와 RMS가속도와 관계를 면밀히 검토하고 각각에 대해 기술적인 논거를 기술하였다.

Since ancient Eygypt, various dental materials were used for lost teeth including gold. The key point of this materials were nontoxic to human body. Since early of 1990's, dental implant was done for recovery of maxillofacial defects. From middle of 1970's, osseointergation concept of implant was introduced and performed in dental field. Biocompatibility of titanium showed good effect for osseointergration but had some problems (Galvance current and toxic corrosion) with suprastructures such as gold crowns. This study was performed to make safe dental implants which have reduced Galvanic currents and corrosion. 3 kind of dental casting gold alloys (different Gold contents, 1㎝×1㎝×0.1㎝ plates.) were used as experimental group, while Titanium were used as control group. Normal human osteoblasts(NHOsts)were cultured during 1-4weeks for histologic study. For analysing the calcium(Ca), Phosphorus(P) and alkaline phosphatase(ALP), NHosts were cultred during 2-23days. After experiments, histologic finding were observed by LSM and SEM. Ca, P, ALP concentration by automatic biochemical analyzer were analyzed by ANOVA test and linear regression method. The results were as follows. Biocompatibility of dental casting gold alloys were similar to titianium alloys histolgically. Biochemical analysis of dental casting gold alloys had no significant difference to titianium alloy except AIGIS-Fine. We could conclude that biocompatibility of dental casting gold alloys with high contents of gold were superior to that of low contents and alloys with high contents of gold had no significant difference from titanium on NHost culture. Gold dental implant might be better than titanium implant due to similar biocompatibility and no galvanic currency.

It has been reported that light-emitting diodes(LED) can be used in the treatment of oral diseases. Although bio-stimulatory effects of LED irradiation such as promotes stimulation of wound healing have been well known, there are few reports about molecular mechanism associated with cell cycle by LED irradiation. The purpose of present study was to examine the molecular event in cell cycle of LED irradiation on primary human gingival fibroblast(hGF) in vitro. The source of light for irradiation was a continuous-wave LED emitting at a wavelength of 635nm, and manufactured that energy density was 5mW/cm2 on sample surface. The hGF were irradiated for 1 hour at 37℃ in 5% CO2 humidified chamber. Experimental samples were acquired at 0 (right after irradiation), 8 and 24 hour after irradiation. To investigate the molecular mechanisms associated with cell cycle, growth phase was determined by flow cytometry and mRNA expression of cyclin A, cyclin B, cyclin D1, cyclin E, cdc2, PCNA, p18, p27, p21, and p53 were determined by real time RT-PCR. Flow cytometric analysis demonstrated the percentage of cells in the G1 and S phase were decreased, but the G2 phase increased, which showed cells irradiated by LED were transitioned from S to G2 phase. For mRNA expression, cyclin B, cdc2, PCNA and p53 were increased at 0 hour after irradiation, and most of cell cycle molecules were increased at 8 hour after irradiation. At 24 hour after irradiation, cyclin A, cyclin E, PCNA and p18 were increased. Taken together, LED irradiation induced proliferation of hGF cells through transition from S to G2 phase.

Basis frame-work's base in a semiconductor industry have gas, chemical, electricity and various facilities in bring to it. That it is a foundation by fire, power failure, blast, spill of toxicant huge by large size accident human and physical loss and damage because it can bring this efficient, connect with each kind mechanical, physical thing to prevent usefully need that control finding achievement factor of human factor of human action. Large size accident in a semiconductor industry to machine and human and it is involved that present, in system by safety interlock defect of machine is conclusion for error of behaviour. What is not construing in this study, do safety in a semiconductor industry to do improvement. Control human error analyzes in human control with and considers mechanical element and several elements. Also, apply achievement factor using O'conner Model by control method of human error. In analyze by failure mode effect using actuality example.

Oral squamous cell carcinoma is the 1st most common malignancy in oral and maxillofacial area. HPV 16 has been strongly linked to progression of cervical carcinoma. E6 and E7 as a small DNA virus encoding two major oncoproteins of HPV 16 can act together to produce efficient immortalization of primary human epithelial cells. Thus it is important to pursue the development of Immortalized human oral keratinocyte(IHOK) culture model which could be related to the pathogenesis of oral squamous cell carcinoma. If we establish IHOK transfected by E6/E7 genes, IHOK will be accepted as a model system for HPV-linked oral carcinogenesis. The purpose of this study were to culture primary normal human oral keratinocyte(NHOK), and to establish IHOK for studying oral carcinogenesis in the future. NHOK was primarily cultured under normal culture condition, and transformed into IHOK by transfection of E6/E7 genes. After 100 passages depend on Ca++ condition, cultured IHOK was confirmed by growth curve, cornified cell envelope measurement, TGase 1activity, mRNA detection, tumorogenecity and anchorage independence assay. After 100 passages, cultured IHOK showed most basal cell and monolayer of polyhedral cells under 0.15mM Ca++, and small area of stratification and flattened epithelial cells with irregular border under 1.2mM Ca++. The cultured IHOK showed relatively resistant growth under high calcium condition. The E6/E7 mRNA was detected in cultured IHOK by RT-PCR. During the terminal differentiation in cultured IHOK, increased insoluble cornified cell envelope formation was accompanied with induction of TGase 1 activity. But the cultured IHOK showed less CEM and TGase 1 activity than those of cultured NHOK. Cultured IHOK showed non-tumorogenecity, but slight anchorage independence. We had developed a technique to transform NHOK into IHOK by transfection of E6/E7 genes. Cultured IHOK was established as intermediate stage cell to study the pathogenesis of human oral squamous cell carcinoma.

Amino acid transporters are essential for the growth and proliferation in all living cells. Among the amino acid transporters, the system L amino acid transporters are the major nutrient transport system responsible for the Na+-independent transport of neutral amino acids including several essential amino acids. The L-type amino acid transporter 1 (LAT1) is over-expressed to support cell growth in malignant tumors. The double stranded RNA-mediated RNA interference (RNAi) analysis can be in a wide variety of eukaryotes to induce the sequence-specific inhibition of gene expression. In this study, we examined the effect of LAT1 short interfering RNA (siRNA) on cell growth using siRNA of LAT1 in the KB human oral squamous cell carcinoma. In the RT-PCR analysis and western blot analysis, the siRNA of LAT1 inhibited expressions of LAT1 mRNA and protein. The uptake of [14C]L-leucine was inhibited by siRNA of LAT1. In the MTT assay, the siRNA of LAT1 inhibited the growth of the KB cells in the time-dependent manner, indicating that the growth inhibition of KB cell by the siRNA of LAT1 is induced by the blocking of neutral amino acid transport mediated by LAT1. These results suggest that the transport of neutral amino acids including several essential amino acids into the KB human oral squamous cell carcinoma is mediated mainly by LAT1. Further, the LAT1 would be a new target for the inhibition of cancer cell growth.

Resveratrol (3,4',5-trihydroxy-trans-stilbene), a naturally occuring polyphenol compound which present in the skin of grapes and red wine has been considered to posses chemopreventive and antioxidant properties. However, little is known about the cellular actions by which resveratrol mediates its therapeutic effects. In this study, the effect of resveratrol on cell proliferation and induction of apoptosis in human osteogenic sarcoma (HOS) cells was investigated. IC50 value was determined to be approximately 60μg/mℓ. Chromosomal DNA framgmentation analysis showed the appearance degraded DNA in time-and dose-dependent manner upon treatment of resveratrol. In order to observe the molecular mechanism involved in resveratrol-induced apoptosis, Western blot analysis was performed. We observed the decrease in the level of procaspase-3, the zymogen form of active caspase-3 in resveratrol-treated cells. This result implies that caspase-3 is activated upon treatment of resveratrol. The activation of caspase-3 was confirmed by the cleavage of poly(ADP-ribose) polymerase. Taken together, our data demonstrate that resveratrol has anti-proliferative effect on HOS cells and induced apoptosis through activation of caspase-3 and PARP cleavage.

Cultured normal human oral kera tinocyte(NHOK) & inunortalized human oral keratinocyte(IHOK) provide a valuable model in ce llular proliferation and differentiation after proper stimulation , And it is interesting to study these estab lished cell lines esca ping normal control on their growth and differentiation, SPRR1 is induced during t erminal differ entiation 0 1' human epiderma l kerat inocytes but is rarely in anaplastic cells of keratinocyte origin, But SPR1 expression has not yet been explained during differ entiation uf NHOK and Lransformed oral keraLinocyLes , The purpose of this study were to examine mHNA and protein expression of SPR1 in response to a known differentiation signal, calcium conc in NHOK, lHOK a nd oral SCC ce ll line(HN 4) , and to apply these results for investigating the molecular mecha nisms of tra nsformed cellular differentiation , Primary cultured NHOK, established IHOK and HN 4 cell line were cul tured in KBM bullet kit Preconfluency of NHOK as control group was used Under O, 15mM Ca++ conc(Precon, Postcon) , and 1, 2mM Ca++ conc(Pos tcon)‘ the insoluble final pellets were measured fo1' cornified cell envelope measurements, and RT- PCR for SPRR1 mRNA meas urement, and immunoblotting for SPRR1 protein measurements in tripli cate , resp ectively , The terminal different ia tion of cu ltured NHOK and IHOK was depend on calcium concentration, while HN4 cell line was not SPRR1 mRNA and protein expression of cultured NHOK showed the highest among cultured IHOK & HN 4 cell line in hi gher ca lcium condition , SPRR1 mRNA and protein expression of cultured IHOK showed higher‘ than tha t of HN 4 cell line in hjgher cacium condi tion , SPRH1 was expressed in differentiation of NHOK and IHOK t ransfected by E6/E7 genes but ra rely expressed in malignant oral keratinocytes , It suggested that SPRR1 ex pression as kera tinocyte terminal diff‘erent ia tion marker involved in cellular cornification would be differentially effected by immorta li zation and ca rcinogenic transforma tion