태반조직의 Nucleases 에 관한 연구 ( 1 ) 사람 태반조직의 Deoxyribonucleases

백문기,황병두,금기창 ( Moon Kee Paik,Byung Doo Hwang,Ki Chang Kum ) 생화학분자생물학회 1976 BMB Reports Vol.9 No.1

The properties of acid deoxyribonuclease purified from human placenta have been investigated, and serial estimation of acid and alkaline deoxyribonuclease in human placenta has been performed. The acid deoxyribonuclease is partially purified forty-fold with specific activity of 240 units/㎎ protein. The acid deoxyribonuclease having an optimal pH around 5 is heat-labile with complete loss of activity at 70 for 10 minutes. The acid deoxyribonuclease is inhibited by Mg^(2+), Mn^(2+), Car and Co^(2+) above 1 mM concentration, suggesting that the enzyme is not dependent on divalent cations. The acid deoxyribonuclease acts on both native and heat-denatured DNA with preference for the former. A gradual decrease in the acid deoxyribonuclease activity occurs with advancing gestation after 20 weeks, the activity at term being about one-fourth of that before 20 weeks. The alkaline deoxyribonuclease activity is maintained untill approximately the 16th week of gestation at which the functioning villi has been established, and thereafter decreases rapidly with no apparent activity about 28 weeks. From the activity changes with gestational age, it is discussed that the acid deoxyribonuclease is related to DNA synthesis of the placenta whereas the alkaline deoxyribonuclease plays an important role in the formation of the functioning villi, suggesting that alkaline deoxyribonuclease is a useful enzyme to evaluate the function of the early placenta.

사람 태반조직 DNA Topoisomerase Ⅰ에 관한 연구

이정복,권기량,임규,황병두 ( Jeong Bok Lee,Gi Ryang Kweon,Kyu Lim,Byung Doo Hwang ) 생화학분자생물학회 1992 BMB Reports Vol.25 No.4

DNA topoisomerase I has been purified from human term placenta approximately 520-folds with a 10% yield. The enzyme shows a broad pH optimum from pH 6 to 9 and is heatlabile, being completely inactivated by heat treatment at 50℃ for 5 min. The enzyme is activated by K^+ and Na^+ approximately 20 and 8-folds respectively. Magnesium ion (Mg^(2+)) is the most potent activator, the activity being 20 and 40-folds activated at 2 mM and 10 mM respectively. But copper ion (Cu^(2+)) is a potent inhibitor. In the presence of Mg^(2+) and K^+, the enzyme is inhibited by physiologic concentration of ATP and GTP. Inhibitory mechanism of ATP is considered to be a inhibition of readoptation of active enzyme conformation and that of GTP is to be a inhibition of the prior step of DNA rejoining. The molecular weight is around 68,000. Camptothecin and 10-hydroxycamptothecin inhibit this enzyme, and the inhibitory action of 10-hydroxycamptothecin is potentiated by Mg^(2+) and K^+. DNA fragmentation by 10-hydroxycamptothecin is more potent than that by camptothecin in the DNA cleavage assay. Heparin and Cu^(2+) inhibit the prior step of DNA rejoining by this enzyme. From the above results, the inhibitory action mechanism of ATP, GTP, heparin, Cu^(2+) and the possible role of Mg^(2+) and K^+ for potentiating the inhibitory action of 10-hydroxycamptothecin, and the development the anticancer drug against DNA topoisomerase I as a target enzyme are discussed.

일차배양 Sertoli Cell에서 Testosterone에 의한 세포성 핵암유전자 발현에 관한 연구

임규,유지헌,김계영,권기량,황병두,Lim, Kyu,Yoo, Ji-Hun,Kim, Kye-Young,Kweon, Gi-Ryang,Hwang, Byung-Doo 생화학분자생물학회 1993 한국생화학회지 Vol.26 No.1

일차배양 Sertoli cell에서 testosterone에 의한 c-myc 등 세포성 핵암유전자 발현 및 그 조절기전을 검색하여 다음과 같은 결과를 얻었다. Steady state에서 발현되는 Sertoli cell 세포성 핵암유전자는 c-fos, c-myc, c-jun이었으며 c-myb은 검출되지 않았고 이때 mRNA 크기는 각각 2.2 kb, 2.4 kb, 2.7 kb에 해당되었다. Testosterone에 의한 c-myc 및 c-jun mRNA의 발현은 시간경과에 따라 증가하였으며 유도되는 시간은 모두 16시간에 최고치에 달하여 유사하였다. c-Myc은 testosterone 농도에 비례해서 그리고 cycloheximide 처리로 각각 전사가 증가되었으나 actinomycin D 처리로 억제되었다. 연령에 따른 Sertoli cell에서의 c-myc mRNA 발현에 대한 testosterone의 영향은 steady state에서는 8일령이 가장 높았으며, testosterone에 의한 유도는 8일령, 14일령에서는 나타났으나 28일령에서는 유도되지 않았다. 이상의 결과로 미루어 보아 testosterone에 의해 유도된 c-myc mRNA가 testosterone에 의해 조절되는 Sertoli cell의 다른 유전자 발현의 조절에 관련되어 있을 것이라 시사된다. The expression of the nuclear protooncogenes have been associated with the cell proliferation and differentiation into changes in nuclear function. To evaluate the possibility that the nuclear protooncogenes play roles testosterone-dependent gene regulation, the effects of testosterone on the expression of the nuclear protooncogenes have been investigated in primary Sertoli cell cultures. c-Myc, c-jun and c-fos were expressed in steady state of primary Sertoli cells but c-myb was not detected. Testosterone increased c-myc and c-jun mRNA with maximal stimulation reached in 16 h. The induction of c-myc was dependent on the concentrations of testosterone. The testsoterone induced c-myc mRNA level was also increased in the cells treated with cycloheximide, but reduced by actinomycin D pretreatment. Even in the absence of testosterone, c-myc mRNA was clearly detectable in Sertoli cells from 8-days-old rats, but hardly detectable in cells from 14 and 28 days of age. Testosterone stimulated c-myc expression in the Sertoli cells from 8 and 14-days-old rats. These results suggest that transient expression of c-myc may play some roles to an intermediate step required for testosterone-dependent regulation of other genes expression of Sertoli cells.

세균내독소에 의한 포도막염에서 Pro-inflammatory Cytokine에 관한 연구

박종문(Jong Moon Park),황병두(Byung Doo Hwang) 대한안과학회 1999 대한안과학회지 Vol.40 No.1

일차배양 Sertoli cell 에서 Testosterone 에 의한 세포성 핵암유전자 발현에 관한 연구

임규,유지헌,김계영,권기량,황병두 ( Kyu Lim,Ji Hun Yoo,Kye Young Kim,Gi Ryang Kweon,Byung Doo Hwang ) 생화학분자생물학회 1993 BMB Reports Vol.26 No.1

The expression of the nuclear protooncogenes have been associated with the cell proliferation and differentiation into changes in nuclear function. To evaluate the possibility that the nuclear protooncogenes play roles testosterone-dependent gene regulation, the effects of testosterone on the expression of the nuclear protooncogenes have been investigated in primary Sertoli cell cultures. c-Myc, c-jun and c-fos were expressed in steady state of primary Sertoli cells but c-myb was not detected. Testosterone increased c-myc and c-jun mRNA with maximal stimulation reached in 16 h. The induction of c-myc was dependent on the concentrations of testosterone. The testsoterone induced c-myc mRNA level was also increased in the cells treated with cycloheximide, but reduced by actinomycin D pretreatment. Even in the absence of testosterone, c-myc mRNA was clearly detectable in Sertoli cells from 8-days-old rats, but hardly detectable in cells from 14 and 28 days of age. Testosterone stimulated c-myc expression in the Sertoli cells from 8 and 14-days-old rats. These results suggest that transient expression of c-myc may play some roles to an intermediate step required for testosterone-dependent regulation of other genes expression of Sertoli cells.

인체 혀의 편평세포암 세포에서 ω3-fatty acid desaturase 유전자 발현이 침윤 및 종양형성에 미치는 영향

홍태화(Tae-Hwa Hong),신소연(Soyeon Shin),한승현(Seung-Hyeon Han),황병두(Byung-Doo Hwang),임규(Kyu Lim) 한국생명과학회 2018 생명과학회지 Vol.28 No.8

오메가-3 지방산(오메가-3)은 수종의 암에 대해 종양형성 억제 및 침윤이 억제됨이 알려져 있다. 그러나 혀의 편평세포암 세포에서 내인성 오메가-3에 의한 침윤 및 종양형성 억제 대한 연구가 명확하게 보고된 바 없다. 이에 본 연구는 혀의 편평세포암 세포에서 ω3-fatty acid desaturase의 유전자 발현이 침윤 및 종양형성에 미치는 영향을 규명하였다. 먼저 SCC-4 및 SCC-9세포의 침윤능은 오메가-3인 DHA 처리에 의해 억제 됨을 확인 하였다. DHA 처리 후 MMP-9 및 MMP-2 활성이 감소 되었을 뿐만 아니라 그 promoter의 reporter 활성도 억제하였다. 또한 COX-2 및 VEGF promoter 활성 뿐만 아니라 NF-kB 활성도 DHA에 의해 억제 되었다. SCC-9의 ω3-desaturase 유전자 stable 세포(fSCC-9sc)의 세포증식 및 colony formation이 억제 되었으며, in vivo 동물실험에서 fSCC-9sc 세포의 종양형성능은 현저히 억제 되었고, 면역형광염색법을 이용한 fSCC-9sc 세포의 종양 조직에서의 TUNEL 양성세포는 대조군인 fSCC-9cc 세포에 비해 현저히 증가하였다. 이상의 결과로 오메가-3는 인체 혀의 편평세포암 세포의 침윤 뿐만 아니라 종양형성을 억제하여 항암작용을 나타낼 수 있으며 따라서 오메가-3는 인체 혀의 편평암의 예방 및 치료에 유용하게 사용될 수 있으리라 생각된다. Omega-3 polyunsaturated fatty acids (ω3-fatty acid) have been found to possess anticancer properties in a variety of cancer cell lines and animal models, but their effects in human tongue squamous cell carcinomas (SCCs) remain unclear. This study was designed to examine the effect of ω3-fatty acid desaturase (fat-1) gene expression on invasion and tumorigenicity in human tongue SCC cells and the molecular mechanism of its action. Docosahexaenoic acid (DHA) treatment inhibited in vitro invasion in a dose-dependent manner. In zymography, matrix metalloproteinase-9 (MMP-9) and Matrix metallopeptidase-2 (MMP-2) activities were reduced, and MMP-9 and MMP-2 promoter activities were inhibited by the DHA treatment. In addition, cyclooxygenase-2 (COX-2) and vascular endothelial growth factor (VEGF) promoter reporter activities were inhibited in SCC-4 and SCC-9 cells after the DHA treatment. To investigate the effect of a high level of endogenous ω3 fatty acids, a stable SCC-9 cell line expressing the ω3-desaturase gene (fSCC-9sc) was generated. The growth rate and colony-forming capacity of fSCC-9sc were remarkably decreased as compared with those of fSCC-9cc. Likewise, the tumor size and volume of fSCC-9sc implanted into nude mice were significantly inhibited, with increases in the cell death index. Furthermore, a transwell chamber invasion assay showed a reduction in cell invasion of the fSCC-9sc lines when compared with that of the fSCC-9cc line. These findings suggested that fat-1 gene expression inhibited tumorigenicity, as well as invasion in human tongue SCC cells. Thus, utilization of ω3 fatty acids may represent a promising therapeutic approach for chemoprevention and the treatment of human tongue SCCs.

백서의 일차배양 간세포에서 Plasminogen Activator와 Plasminogen Activator Inhibitor의 표피 성장인자 의존적 발현에 관한 연구

윤완희(Wan-Hee Yoon),최정훈(Jeong-Hun Choi),정연주(Yeon-Joo Jung),이병학(Byung-Hak Lee),김태동(Tae-Dong Kim),송인상(In-Sang Song),배진선(Jin-Sun Bae),임규(Kyu Lim),황병두(Byung-Doo Hwang) 대한외과학회 2001 Annals of Surgical Treatment and Research Vol.61 No.3

胎盤組織 Cytidine Deaminase에 關한 硏究 (Ⅰ) : Purification & Characterization

황병두,곽상태,임규,이재흔,백문기 충남대학교 의과대학 지역사회의학연구소 1984 충남의대잡지 Vol.11 No.2

Cytidine deaminase has been partially purified from human term placenta approximately 26-fold by combination of ammoninm sulfate saturation, sephacryl S-300, DEAE-cellulose and hydroxyapatite chromatography, and then its properties have been investigated. The enzyme shows a broad optimum pH from 5 to 9 and is heat-stable being, almost remained its activity by heat treatment at 70℃ for 8 minutes. The enzyme activity is inhibited to 23% and 77% by 2 mM of Cu^2+ and Co^2+, respectively, but, other cations do not affect the enzyme activity. The apparant Michaelis constant for cytidine and deoxycytidine are 74 uM and 69 uM, respectively. Kinetic analysis of cytidine deminase in presence of 0.25 mM Cu^2+ shows that the nature of the inhibition to cytidine deaminase is competitive, considering that Vmax is independent while km value is increased. The enzyme activity is completely inhibited by 0. 5 mM of p-hydroxymecuribenzoicacid(PHMB)but all of the activity is recovered in adding mercaptoethanol. The enzyme activity is decreased about 50% in prescence of 5 mM of polyamines. The enzyme activity is mainly located in the nuclei (80%) and cytosol(16%) fractions. he molecular weight of the enzyme is about 770, 000.

사람 胎盤組織 膜 結合 Adenosine Deaminase의 性狀에 關한 硏究

황병두,홍승원 충남대학교 의과대학 지역사회의학연구소 1982 충남의대잡지 Vol.9 No.1

The properties of plasma membrane bound adenosine deaminase(ADase) from human placenta have been investigated. The enzyme exhibits a broad pH optimum from pH 6 to 8 and is heat labile, being completely inactivated by heat treatment at 70℃ for 10 minutes. The enzyme is not effected at pH 7 by 2 mM. Li^+, Mg^2+, Ca^2+, Mn^2+, Co^2+, Fe^2+ and Zn^2+, while the enzyme is activated by 2 mM Ca^2+ and Co^2+ at pH 8 and inhibited by 2 mM Cu^2+ independent of pH. The enzyme activity is completely recovered by EDTA (4 mM) only in case of adding Cue+ after preincubation of enzyme with substrate. The apparent Michaelis constant of the enzyme for adenosine is 58 μM. From the above result, it is suggested that the enzyme is different from ADase of soluble fraction of human placenta.

사람 洋水의 Adenosine Diphosphoribose Pyrophosphohydrolase

黃炳斗,白汶基 충남대학교 의과대학 지역사회의학연구소 1975 충남의대잡지 Vol.2 No.2

Adenosine diphosphoribose pyrophosphohydrolase (ADPR-PPase) which catalyzes the hydrolysis at the pyrophosphate linkage of adenosine diphosphoribose (ADP-ribose) formed from nicotinamide adenine dinucleotide (NAD) by NAD nucleosidase has been partially purified from human amniotic fluid, and its properties have been investigated. This enzyme shows optimal pH around 9.5 and is heat-labile, being completely inactivated by heat treament at 65˚ for 10 minutes. In contrast to erythrocyte ADPR-PPase of rabbit, the enzyme does not require Mg^2+, but requires inoragnic phosphate for the activity, maximal activity being attained at 20mM phosphate. The apparent Michaelis constant for ADP-ribose is 1.4 mM. The enzyme is inhibited markedly by adenine nucleotides which act as a competitive inhibitor, although CMP and GMP are also inhibitory, yet lesser in magnitude, whereas oxidized and reduced NAD and pHMB are not inhibitory. These results indicate that the enzyme is controled by related compounds as well as reaction product.