• Korean Journal of Animal Reproduction
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• v.21 no.2
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• pp.177-183
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• 1997

The present study examined the effects of epidermal growth factor(EGF) and transforming growth factor-$\alpha$(TGF-$\alpha$) on in vitro maturation of porcine follicular oocytes. Basic medium used TCM-HEPES, and oocytes cultured for 42 hours in vitro. The results obtained are as follows; 1. The nuclear maturation rates of EGF-treated groups(10ng/ml, 75.9% ; 30ng/ml, 69.2% ; 50ng/ml, 67.2% ; 100ng/ml, 71.0%) on the porcine oocytes cultured in medium without pFF in vitro were significantly(P<0.01) higher than those of non-treated group(57.1%). When the oocytes were cultured in media with 10%(v/v) pFF, the nuclear maturation rates of 30ng EGF/ml(77.1%) treated group were significantly(P<0.01) higher than those of non-(59.2%) and EGF-treated groups(10ng/ml, 65.4% ; 50ng/ml, 65.5% ; 100ng/ml, 70.4%). 2. The nuclear maturation rates of 30ng TGF-$\alpha$/ml treated group(71.9%) in media without pFF in vitro were significatnly(P<0.01) higher than those of non-(57.1%) and TGF-$\alpha$ treated groups(10ng/ml, 60.4% ; 50ng/ml, 65.4% ; 100ng/ml, 60.0%). When the oocytes were cultured in media with 10%(v/v) pFF, the nuclear maturation rates of 30 and 50ng TGF-$\alpha$/ml(77.4% and 79.6%) treated groups(10ng/ml, 64.2% ; 100ng/ml, 61.6%). 3. On the effect of EGF(30ng/ml) and/or TGF-$\alpha$(30ng/ml) treated groups in medium without pFF in vitro, the nuclear maturation rates indicated 57.3, 60.4, 75.9 and 79.7% in media with no EGF & TFG-$\alpha$, TGF-$\alpha$ only, EGF only nad EGF＋TGF-$\alpha$ treated groups, respectively. The nuclear maturation rates in medium with EGF only or EGF＋TGF-$\alpha$ were significantly(P<0.01) higher than those non- and TGF-$\alpha$ treated groups. When the oocytes were cultured in media with 10%(v/v) pFF, the nuclear maturation ratesof EGF＋TGF-$\alpha$ treated group(75.9%) were significantly(P<0.01) higher than those of non-(59.2%), TGF-$\alpha$ only (64.2%) and EGF only(69.4%) treated groups.

• Microbiology and Biotechnology Letters
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• v.26 no.1
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• pp.61-67
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• 1998

Recombinant human epidermal growth factor (rhEGF) was produced by E. coli BL2l (DE3) harboring a plasmid pYHB101. The production of rhEGF was 44.5 mg/L when the E. coli BL2l (pYHB101) was cultured at 27$^{\circ}C$ for 48 hr in the modified MBL medium containing 10 $\mu\textrm{g}$/L glucose with 10 $\mu\textrm{m}$ IPTG/lactose induction at 2 hr after inoculation. It was shown that lactose is able to induce the rhEGF expression of E. coli BL2l (pYHB101) with the same efficiency as IPTG. In the batch culture system, when induced with 10 $\mu\textrm{m}$ lactose, E. coli BL2l (pYHB101) produced maximum 45 mg/L of the rhEGF at 28 hr culture in the modified MBL medium containing 10 g/L glucose. In the semi-fed batch culture system, the volumetric yield was 160 mg/L when the culture was added with 0.5% (w/v) lactose and 0.25% (w/v) yeast extract in the late logarithmic phase and 94.3% of rhEGF was secreted as soluble form. However, when the culture was added with them in the early logarithmic phase, the volumetric yield was 120 mg/L and 20.9% of rhEGF was found in cytoplasmic insoluble aggregates. It was found that the addition time of lactose was important for production of soluble rhEGF from E. coli BL21 (pYHB101).

• Microbiology and Biotechnology Letters
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• v.24 no.4
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• pp.478-484
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• 1996

Recombinant human epidermal growth factor (rhEGF) was produced by E. coli BL21 harboring a plasmid pYHB101. The maximum production was 68.7 mg/l when the E. coli strain was cultured at 25$\circ$C for 48 hours in the modified MBL medium containing 10 g/l glucose with 1 mM IPTG induction at 2 hours after inoculation. The rhEGF was purified upto 267 folds by Amberlite XAD- 7 chromatography, ultrafiltration, and DEAE Sepharose fast flow ion exchange chromatography with an overall yield of 66.6%. The purified rhEGF was further separated into two fractions by HPLC. The N-terminal amino acid sequence of the second fraction was Asn-Ser-Asp-Ser-Glu-Cys-Pro-Leu-Ser-His. The effect of rhEGF on the DNA synthesis was examined using in vitro biological assay based on the incorporation of 5'-bromo-2'- deoxy-uridine (BrdU). The purified rhEGF shows no difference with natural human epidermal growth factor (nhEGF) in N-terminal amino acids residues and biological activity. From the results, we concluded that rhEGF produced from E. coli harboring the plasmid pYHB101 was apparently the same as nhEGF.

• Proceedings of the KSAR Conference
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• 2001.03a
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• pp.29-29
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• 2001

This study was carried out to examine, by the reverse transcription chain reaction(RT-PCR)and Immunostain assays, epidermal growth factor mRNA expression in bovine ova during oocyte maturation in vitro(0-2lh)and after fertilization in vitro(6-144hr: zygotes to blastocysts). In this study, the transcripts of EGF was detected in oocytes using primers for EGF. Transcripts for EGF mRNA was not detected in oocytes through in vitro maturation. But EGF mRNA were present after fertilization up to the 2-cell stage and the blastocyst stage. The highest mRNA levels in 4-cell stage embryos were decreased at 8cell stage and then reincreased upto morulae and blastocysts. The results of this study showed EGF mRNA are present in embryo after fertilization and this factors are involved in the regulation of bovine embryo development.

• BMB Reports
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• v.32 no.1
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• pp.28-32
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• 1999

Annexin I (also called lipocortin 1), a 37-kDa member of the annexin family of proteins, has been implicated in the mitogenic signal transduction by epidermal growth factor (EGF). Annexin I is phosphorylated by the EGF signal, however, the role of annexin I in the EGF signal transduction is still unknown. To transduce extracellular signals into the intracellular targets, selective translocation of the signaling molecules to their targets would be necessary. In this study, we examined the subcellular locations of annexin I during EGF signal transduction. Treatment of A549 cells with EGF resulted in the translocation of cytoplasmic annexin I to the nucleus and perinuclear region as determined by Western blot and immunofluorescent staining. The nuclear translocation of annexin I was inhibited by tyrphostin AG 1478 and genistein, the inhibitors of EGF receptor kinase and downstream tyrosine kineses, respectively. Pretreatment of cells with cyclohexamide did not inhibit the nuclear translocation. The results suggest that nuclear translocation of annexin I is controlled by a series of kinase dependent events in the EGF receptor signaling pathway and may be important in tranducing the signals by EGF.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.20 no.4
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• pp.334-340
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• 1998

Stimulatory effects of epidermal growth factor (EGF) and transforming growth $factor-{\alpha}$($TGF-{\alpha}$) on the growth of squamous cancer cell lines established from human oral cancer tissue with moderate differentiation were studied in vitro. After culturing in serum-free media for 24 hours, growth factors-EGF only, $TGF-{\alpha}$ only and EGF, $TGF-{\alpha}$ together-were added to the media and numbers of cells were analyzed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and compared with the control at 96, 144 hours. Each of EGF and $TGF-{\alpha}$ showed statistically significant stimulatory effects on the growth of cells respectively. Dose-dependent relationship of the stimulatory effects were not clearly demonstrated. The effects of EGF were higher than those of $TGF-{\alpha}$ and combinative administration showed higher effects than those of single uses. In conclusion, EGF may play an important and major role in differentiation and growth of human oral squamous cancer cells. $TGF-{\alpha}$, produced from cells activated by EGF, also can stimulate the cell growth and could be an alternative ligand for EGF receptor.

• Korean Journal of Animal Reproduction
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• v.22 no.1
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• pp.51-59
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• 1998

This study was undertaken to investigate the effects of ITS and EGF on embryonic development of in vitro matured and fertilized bovine oocytes in culture medium su, pp.ementing with or without calf serum. When fertilized oocytes were cultured in TCM-199 containing 0, 0.1, 0.5 and 1.0\ulcorner/ml ITS with 5% calf serum, the rates of development to blastocyst stage and the cell number of blastocysts were not significantly different among all treatments. And also monolayer of cumulus/granulosa cells prepared in containing calf serum and ITS were no beneficial effects of embryonic development. On the other hand, when EGF was su, pp.imented to TCM-199 containing calf serum or calf serum free, embryonic development rates(24.0 2.8% to 29.2 1.7% or 8% to 9%) and cell number of blastocysts(p<0.05) were significantly increased compared with EGF-free(22.1 2.1 or 1.0%, p<0.05). But when fertilized oocytes were cultured with cumulus/granurosa cells in TCM-199 containing EGF and calf serum, the rate of embryos development to the blastocyst stage and cell number of blastocysts were not significantly different compared with EGF-free and any concentrations. These results showed that ITS and EGF was not improved the development of bovine embryo in vitro matured and in vitro fertilized with calf serum and/or monolayer of cumulus/granulosa cells.

• Korean Journal of Animal Reproduction
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• v.21 no.1
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• pp.39-46
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• 1997

본 연구는 체외생산된 소 배반포기배의 inner cell mass (ICM) 세포에서 epidermal growth factor-receptor (EGF-R)의 발현 유무를 immunosurgery와 indirect immunofluorescence (간접 면역 형광방법)을 이용하여 조사하고자 실시하였다. 본 실험에 사용된 ICM 세포는 체외수정 후 7∼8일째에 회수된 소 배반포기배로부터 immunosurgery 방법을 실시하여 얻어졌으며, 회수된 ICM세포는 live/dead 염색방법을 통한 생사 유무와 EGF-R 발현 유무 조사에 공시되었다. 특히, 배반포기배에 대한 immunosurgery를 위해 trophectoderm 세포에 대한 rabbit anti-bovine trophectoderm cell antibody (RABTE)를 제조하여 사용하였다. 결과를 요약하면 다음과 같다. ICM세포의 회수율은 RABTE와 guinea pig serum (complement)에 각각 15∼30분과 15∼60분동안 처리했을 경우 16.7∼74.2%였으며, 또한 처리시간이 각각 30분과 30분일 때 가장 높은 회수율(74.2%)을 얻었다. Immunosurgery 후 얻어진 ICM세포의 생존 유무를 조사하기 위해 live/dead 염색 방법을 이용하였던바, ICM세포의 생존율은 complement가 60분 처리된 군(69.3%)을 제외한 모든 처리군에서 84.0∼91.6%의 높은 생존율을 나타냈다. 또한, 회수된 ICM세포에 대한 EGF-R의 존재를 확인하였다. 따라서, ICM세포에서의 EGF-R의 발현은 인위적으로 첨가된 EGF의 이용 가능성을 높임으로서 체외에서의 착상전 배 발달을 증진시킬 수 있을 것으로 사료된다.

• YAKHAK HOEJI
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• v.41 no.6
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• pp.730-736
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• 1997

Distribution of radioactivity in the skin tissues, subcutaneous tissues, blood and body organs was examined following topical application of $^{125}I-rhEGF$(0.4 ${\mu}Ci$), in the form of a Pluronic F-127 gel, on the normal and damaged (burned and stripped) skins of SD male rats. The radioactivity in the skin tissues and subcutaneous tissues was 3-5 times higher for the damaged skins than for the normal skin. But pretreatment of the skin with rhEGF (1${\mu}g$)) twice at 24 hr dose intervals affected the distribution of the radioactivity yielding the order of burned skin> stripped skin=normal skin. The decrease for the stripped skin by the pretreatment might be related either to the pathophysiological change of the skin or to the down regulation of the EGF receptor. Liver showed the highest radioactivity in amount following single and multiple administration of the drug to the normal and damaged skins. But,in concentration, the kidney and stomach showed higher value than the liver which is consistent with that kidney is a major eliminating organ of EGF and that EGF exerts its pharmacological effect specifically for the stomach.

• The Korean Journal of Zoology
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• v.39 no.2
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• pp.215-222
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• 1996

In the previous studies, we have demonstrated that prorenin converting enzyme (PRECE) was identical to the epidermal grouch factor-binding protein (EGF-BP) type B, which was a member of the mouse glandular kallikrein family, To examine whether or not EGF-BP type A and C are involved in the processing of prorenin, we have cloned the CDNAS of the EGF-BP type h and C from a library of male ICR mouse submandibular gland (SMGI. And then CHO cells were transfected with the EGF-BP expression plasmids. and stable cell lines expressing a high level of the EGF-BPS precursor were obtained. The conditioned medium was then treated with trypsin, which has been knotvn to effectively convert the EGF-BP type A and C precursor to the active forms. 수ubsequentlv, the prorenin converting activity of the trypsin-treated or untreated medium was examined. PRECE converted exactly prorenin to renin, but the prorenin converting activities of EGF-BP type A and C were not detected. From these results, it seems that only type B of these EGF-BPs is involved in processing Ren 2 prorenin in mouse SMG.