GH₃ 뇌하수체 종양세포의 분화에 미치는 laminin, Matrigel 및 EGF의 영향

정경아(Gyung-Ah Jung),남선영(Seon Young Nam),이병란(Byung-Lan Lee) 대한해부학회 2001 Anatomy & Cell Biology Vol.34 No.3

뇌하수체 앞엽에는 여러 가지 종류의 호르몬 분비세포가 존재하므로 한가지 종류의 호르몬 세포만을 상대로 연구를 진행하기에 어려움이 있다. GH3 세포는 뇌하수체종양세포로서 젖샘자극호르몬 (PRL)과 성장호르몬을 분비하는 뇌하수체 세포의 특성을 유지하지만 배양기질에 부착력이 낮으며 작은 비율의 세포만이 PRL을 발현하고 장기간 배양시에는 세포의 과증식이 일어나므로 장기배양이 어렵다. 세포사이물질(ECM)은 단세포배양시에 일어나는 세포의 특성소실을 방지하기 위하여 배양기질로 사용되고 있다. ECM과 성장조절인자는 서로 상호작용을 통하여 세포에 영향을 미치나 이때 성장조절인자가 세포에 미치는 영향은 배양기질로 사용된 물질의 종류에 따라 다르다고 알려져 있다. 따라서 본 연구에서는 성장조절인자인 EGF와 세포사이물질을 이용하여 GH3 세포의 PRL 세포로서의 분화를 최대화하고자 하였다. 이를 위하여 세포들의 성장, 증식 및 PRL 발현세포의 빈도를 관찰하기 위하여 세포수측정, bromodeoxyuridine (BrdU) 표지, BrdU 및 PRL에 대한 면역세포화학 염색을 시행하였다. 본 연구의 결과로부터 EGF는 본 연구에 사용된 배양기질인 배양용 플라스틱, laminin 및 Matrigel 위에서 GH3 세포수를 감소시키고 PRL 세포로의 분화를 증가시킴을 알 수 있었고 특히 Matrigel과 함께 사용시 가장 높은 분화도를 나타냄이 관찰되었다. 따라서 GH3 세포의 장기배양시에 과증식을 막고 특성소실을 최소화 하기 위해서는 Matrigel 위의 배양에서 EGF를 투여하는 것이 가장 효과적인 방법으로 사료되었다. This study was performed in order to establish the culture system optimal for the study on pituitary prolactin cells using growth factor and extra cellular matrix components as the culture substrate. The effect of epidermal growth factor (EGF) alone or along with extracellular marix components on GH3 cell growth and PRL expression was assessed using cell count, BrdU-immunocytochemistry and PRL-immunocytochemistry in in vitro cultures on plastic, laminin and Matrigel. EGF decreased the cell growth, BrdU-labeling and increased the PRL-immunoreactive cells regardless of the culture substrate by day 3 of the culture. Matrigel was the best culture substrate to decrease the cell growth and to increase the PRL expression. EGF treatment in the Matrigel culture showed about 80.5% of PRL-immunoreactive cells by day 6 of the culture. These results indicated that Matrigel is the better culture substrate than plastic or laminin to inhibit the overgrowth and to increase the prolactin expression of the GH3 cell and that EGF and Matrigel causes very effective culture environment for the long-term culture of the GH3 cell by synergistic mechanism.

톨루엔이 흰쥐 태반의 PRL-GH군 유전자발현과 발생에 미치는 영향

최병수(Byong-Soo Choe),이채관(Chae-Kwan Lee),문덕환(Deog-Hwan Moon),손병철(Byung-Chul Son),김대환(Dae-Hwan Kim),이창희(Chang-Hee Lee),김휘동(Hwi-Dong Kim),김정원(Jung-Won Kim),김종은(Jong-Eun Kim),이채언(Chae-Un Lee) 대한직업환경의학회 2004 대한직업환경의학회지 Vol.16 No.3

Objectives: The purpose of this experimental study was to investigate the toxic effects of toluene on the placental functions and reproductionin the rat. In this study, the expression of placental prolactin-growth hormone (PRL-GH) and Pit-1 genes, the frequency of placental trophoblast cells, and the reproductive data were analyzed. Methods: The pregnancy of the Sprague-Dawley rats (250±25 g) was determined by verifying the presence of the copulatory plug or sperm in the vaginal smear and the day on which this was observed was defined as pregnancy day 0. The pregnant rats were divided into three groups. The control group was intraperitoneally (ip) injected with sesame oil, and the other two groups were given either 150 or 750 ㎎/㎏ BW/day of toluene resuspended in sesame oil during pregnancy days 7-11 and 16-20. The rats from the three experimental groups were sacrificed on pregnancy days 11 and 20, respectively. The mRNA levels of the PRL-GH, Pit-1a and b isotype genes were analyzed by Northern blot hybridization and Reverse transcription-polymerase chain reaction (RT-PCR), respectively. The hormonal concentration was analyzed by Radioimmunoassay. The frequency of the placental trophoblast cells was determined by means of a histochemical study. Reproductive data, such as the placenta and infnat weight, pregnancy period and litter size were surveyed at pregnancy day 20 and after birth. Statistical analysis was carried out by means of the SAS program (version 8.1). Results: The mRNA levels of the PRL-GH family genes were reduced in a linear fashion by exposure to toluene. The mRNA levels of the Pit-1a and b isotype genes, which induce the expression of the PRL-GH family genes, were also reduced by exposure to toluene. The placental lactogen Iv and Ⅱ concentrations in the rat placenta, fetus and maternal blood were also decreased by exposure to toluene. During the last stage of gestation, exposure to a high dose of toluene reduced the frequency of the spongiotrophoblast cells that secrete the PRL-GH hormones. Reproductive data such as the placenta and infant weight, and litter size were reduced, and the pregnancy period was extended in the toluene exposed group as compared with the control group. Conclusions: Toluene disrupts the PRL-GH hormone metabolism in the rat placenta and this leads to reproductive disorder.

Combined Expression of Metastasis Related Markers Naa10p, SNCG and PRL-3 and its Prognostic Value in Breast Cancer Patients

Min, Li,Ma, Ruo-Lan,Yuan, Hua,Liu, Cai-Yun,Dong, Bing,Zhang, Cheng,Zeng, Yan,Wang, Li,Guo, Jian-Ping,Qu, Li-Ke,Shou, Cheng-Chao Asian Pacific Journal of Cancer Prevention 2015 Asian Pacific journal of cancer prevention Vol.16 No.7

Combinations of multiple biomarkers representing distinct aspects of metastasis may have better prognostic value for breast cancer patients, especially those in late stages. In this study, we evaluated the protein levels of N-${\alpha}$-acetyltransferase 10 protein (Naa10p), synuclein-${\gamma}$ (SNCG), and phosphatase of regenerating liver-3 (PRL-3) in 365 patients with breast cancer by immunohistochemistry. Distinct prognostic subgroups of breast cancer were identified by combination of the three biomarkers. The Naa10p+SNCG-PRL-3-subgroup showed best prognosis with a median distant metastasis-free survival (DMFS) of 140 months, while the Naa10p-SNCG+PRL-3+subgroup had the worst prognosis with a median DMFS of 60.5 months. Multivariate analysis indicated Naa10p, SNCG, PRL-3, and the TNM classification were all independent prognostic factors for both DMFS and overall survival (OS). The three biomarker combination of Naa10p, SNCG and PRL-3 performed better in patients with lymph node metastasis, especially those with more advanced tumors than other subgroups. In conclusion, the combined expression profile of Naa10p, SNCG and PRL-3, alone or in combination with the TNM classification system, may provide a precise estimate of prognosis of breast cancer patients.

Matrigel 배양상태에서 정상 흰쥐로부터 얻은 뇌하수체 전엽세포의 호르몬 발현에 미치는 영향

이병란,김현준,황덕호,이한경,유영복,황일태 대한해부학회 2000 Anatomy & Cell Biology Vol.33 No.3

6가 크롬 폭로가 랫트의 태반 기능과 출산에 미치는 영향

이헌,문덕환,이채언,강성구,손병철,김대환,이창희,김정원,이채관,전진호,Lee, Heun,Moon, Deog-Hwan,Lee, Chae-Un,Kang, Sung-Goo,Son, Byung-Chul,Kim, Dae-Hwan,Lee, Chang-Hee,Kim, Jung-Won,Lee, Chae-Kwan,Chun, Jin-Ho 대한예방의학회 2004 예방의학회지 Vol.37 No.2

Objectives : This study aimed to investigate the toxic effects of chromium (VI) on the placental function and reproduction in rats. For the study, the placental prolactin-growth hormone (PRL-GH) gene expression, placental trophoblast cell differentiation and reproductive data were analyzed. Methods : The pregnancies of F344 Fisher rats were checked by the presence of a copulatory plug or sperm in the vaginal smear, which was defined as day 0 of the pregnancy. Pregnant rats were divided into the three groups. The control group was given tap water (chromium level < 0.001 ppm) and the remaining groups were given 250 or 750 ppm of chromium (VI) [as potassium dichromate], from day 7 to 19 of the pregnancy. Rats were sacrificed at days 11 and 20 of pregnancy. The mRNA levels of PRL-GH and Pit-1a and b isotype genes were analyzed by Northern blot hybridization and reverse transcription-polymerase chain reaction (RT-PCR). The hormonal concentration was analyzed by radioimmunoassay, and the differentiation of placental trophoblast cells were observed by histochemical studies. Reproductive data, such as placental and fetal weights, pregnancy period, and litter size, were surveyed at day 20 of pregnancy and after birth. A statistical analysis was carried out using the SAS program (version 8.1). Results : The mRNA levels of the prolactin-growth hormone (PRL-GH) family of genes were dose dependently reduced by chromium exposure. The mRNA levels of Pit-1a and b isotype genes that induce the expression of the PRL-GH family of genes were also reduced by chromium exposure. The PRL-GH hormonal concentration in the rat placenta, fetus and maternal blood were decreased by chromium exposure. In the middle stage of pregnancy (day 11), a high dose of chromium suppressed the differentiation of spongiotrophoblast cells that secret the PRLGH hormones. In the last stage of pregnancy (day 20), a high dose of chromium induced apoptosis of placental cells. Reproductive data, such as placental and fetal weights, litter size, were reduced, but the pregnancy period was extended in the group exposed to chromium compared with the controls. Conclusion : Chromium (VI) disrupts the ordered functions of the placenta, which leads to reproductive disorders in rats.

Bisphenol A가 흰쥐의 태반 기능과 출산에 미치는 영향

이채관,김석현,문덕환,김정호,손병철,김대환,이창희,김휘동,김정원,김종은,이채언,Lee, Chae-Kwan,Kim, Seog-Hyun,Moon, Deog-Hwan,Kim, Jeong-Ho,Son, Byung-Chul,Kim, Dae-Hwan,Lee, Chang-Hee,Kim, Hwi-Dong,Kim, Jung-Won,Kim, Jong-Eun,Lee, Chae 대한예방의학회 2005 예방의학회지 Vol.38 No.3

Objectives : The aim of this study was to investigate the effects of bisphenol A (BPA), an estrogen-like environmental endocrine disrupter, on the placental function and reproduction in rats. The mRNA levels of the placental prolactin-growth hormone(PRL-GH) gene family, placental trophoblast cell frequency and reproductive data were analyzed. Methods : The pregnancies of F344 Fisher rats ($160g{\pm}20g$) were detected by the presence of the copulatory plug or sperm in the vaginal smear, which marked Day 0 of pregnancy. Pregnant rats were divided into three groups. The control group was intraperitoneally injected with a sesame oil vehicle. The two remaining groups were injected with 50 or 500 mg/kg B.W/day of BPA, resuspended in sesame oil, on either days 7 to 11 or 16 to 20 of pregnancy, with the rats sacrificed on either day 11 or 20, respectively. The mRNA levels of PRL-GH and Pit-1a and b isotype genes were analyzed by Northern blot hybridization and reverse transcription-polymerase chain reaction. The hormone concentrations were analyzed by radioimmunoassay, and the frequency of the placental trophoblast cells observed by a histochemical study. Reproductive data, such as the placental weight and litter size, were surveyed on day 20. The fetal weight was surveyed for 4 weeks after birth. A statistical analysis was carried out using the SAS program (version 8.1). Results : The mRNA levels of the PRL-GH gene family, such as placental lactogen I, Iv and II, prolactin like protein A, C and Cv, and decidual prolactin-related protein were significantly reduced due to BPA exposure. The mRNA levels of the Pit-1a and b isotype genes, which induce the expression of the PRL-GH gene family in the rat placenta, were also reduced due to BPA exposure. The PL-Iv and PL-II concentrations were reduced in the BPA exposed group. During the middle to last stage of pregnancy (Days 11-20), a high dose of BPA exposure reduced the frequency of spongiotrophoblast cells, which are responsible for the secretion of the PRL-GH hormones. Reproductive data, such as the placental and fetal weights and the litter size, were reduced, but that of the pregnancy period was extended in the BPA exposed compared to the control group. Conclusions : BPA disrupts the placental functions in rats, which leads to reproductive disorders.

Matrigel이 배양상태에서 정상 흰쥐로부터 얻은 뇌하수체 전엽세포의 호르몬 발현에 미치는 영향

이한경(Han-Kyung Lee),김현준(Hyun-Joon Kim),유영복(Young-Bok Yoo),황일태(Il-Tae Hwang),황덕호(Douk-Ho Hwang),이병란(Byung-Lan Lee) 대한해부학회 2000 Anatomy & Cell Biology Vol.33 No.3

This study was performed to investigate the effect of Matrigel, a reconstituted basement membrane, on the expression of the anterior pituitary hormones in culture. Rat pituitary cells cultured for 6 days on Matrigel showed 3-dimensional, lobular structures with connecting cells while those on plastic showed flat, polygonal cells forming a monolayer. Western blot analysis showed that prolactin (PRL) content in the anterior pituitary cells was higher compared to those cultured on plastic. In comparison, TSH expression was not increased in cultures on Matrigel. The total cell number and the proportion of fibroblasts was decreased. These results suggested that Matrigel is a useful culture substrate for the enhanced expression of PRL but not for TSH. Further studies are needed in order to find a useful culture substrate for TSH cells. 본 연구는 세포배양에서 기저막 성분인 Matrigel이 뇌하수체 세포의 호르몬 발현에 미치는 영향을 파악하기 위하여 시 행되었다. 위상차 현미경과 주사전자현미경을 사용한 관찰에서 Matrigel 위에서 배양 된 쥐의 정상 뇌하수체 세포는 3차원적 구조를 형성하였는데, 이 때 세포덩어리들은 마치 생체내 분비샘들의 소엽과 유사한 형태를 나타내었다. Western blot을 사용한 관찰에서 세포 내에 포함된 젖샘자극호르몬 (PRL)의 양이 유의하게 증가됨이 관찰되었으나, reverse transcriptionpolymerase chain reaction (RT-PCR) 분석의 결과로 볼 때 갑상샘자극호르몬 (TSH)의 발현에는 유의한 차이가 없었다. 이로부터 Matrigel은 PRL의 발현에는 영향을 미치나 모든 종류의 뇌하수체 호르몬 발현에 영향을 미치지는 않는 것으로 관찰 되었으므로 각 종류의 뇌하수체 호르몬세포에 적합한 배양기질을 개발해야 할 것으로 사료되었다.

트리클로로에틸렌이 흰쥐의 태반 기능과 발생에 미치는 영향

강정학 ( Jeong Hak Kang ),이채관 ( Chae Kwan Lee ),문덕환 ( Deog Hwan Moon ),손병철 ( Byung Chul Son ),김대환 ( Dae Hwan Kim ),이창희 ( Chang Hee Lee ),김휘동 ( Hwi Dong Kim ),김정원 ( Jung Won Kim ),김종은 ( Jong Eun Kim ),이채언 한국산업위생학회 2004 한국산업보건학회지 Vol.14 No.3

This study aimed at investigating the toxic effects of trichloroethylene (TCE) on the placental function and reproduction in rat. For these study placental prolactingrowth hormone (PRL-GH) family gene expression, placental trophoblast cell frequency and reproductive data were analyzed. Pregnancy of the Sprague-Dawley rats were checked by the presence of the copulatory plug or sperm in the vaginal smear and defined as the pregnant day (PD) 0. The pregnant rats were divided into the three groups. The control group was intraperitoneally injected with sesame oil. The remaining groups were injected with 100 or 500mg/kg B.W/day of TCE resuspended with sesame oil from PD 7-11 or 16-20. Rats were sacrificed at PD 11 and 20. mRNA levels of PRL-GH family and Pit-1a, b isotype genes were analyzed by Northern blot hybridization and Reverse transcription-polymerase chain reaction. Hormone concentration was analyzed by radioimmunoassay. Frequency of placental trophoblast cells were observed by histochemical study. Reproductive data such as placental and fetal weights pregnancy period, and litter size were surveyed at PD 20 and after birth. Statistical analysis was carried out by the SAS program (version 8.1). mRNA levels of PRL-GH family genes such as placental lactogens and placental prolactin like proteins were reduced by TCE administration. mRNA level of Pit-1a, b isotype genes that induce the expression of PRL-GH family genes were also reduced by TCE administration. Placental lactogen II concentration in the placenta, fetus and maternal blood were decreased by TCE administration. Exposure to a high dose of TCE reduced the frequency of the spongiotrophoblast cell in junction zone. Reproductive data such as placental and fetal weights, litter size were reduced, and pregnancy period was extended in the TCE exposed group than control. These data suggested that TCE disrupts the ordered functions of placenta and these effects lead to the reproductive disorder in rat.

Biflavonoids inhibited phosphatase of regenerating liver-3 (PRL-3)

Choi, Sung-Kyu,Oh, Hyun-Mi,Lee, Su-Kyung,Jeong, Dae Gwin,Ryu, Seong Eon,Son, Kwang-Hee,Han, Dong Cho,Sung, Nack-Do,Baek, Nam-In,Kwon, Byoung-Mog Taylor & Francis Health Sciences 2006 NATURAL PRODUCT RESEARCH Vol.20 No.4

<P> Two biflavonoids, ginkgetin (1) and sciadopitysin (2), were isolated from the MeOH extract of the young branches of Taxus cuspidata, which inhibited phosphatase of regenerating liver-3 (PRL-3) with IC50 values of 25.8 and 46.2 μM, respectively. This is the first report on PRL-3 inhibitors, isolated from natural sources.</P>

카드뮴이 흰쥐의 태반 기능과 발생에 미치는 영향

안진홍(Jin-Hong Ahn),고광수(Kwang-Soo Ko),이채관(Chae-Kwan Lee),문덕환(Deog-Hwan Moon),손병철(Byung-Chul Son),김대환(Dae-Hwan Kim),이창희(Chang-Hee Lee),김휘동(Hwi-Dong Kim),김정원(Jung-Won Kim),김종은(Jong-Eun Kim),전진호(Jin-Ho Chu 대한직업환경의학회 2004 대한직업환경의학회지 Vol.16 No.3

Objectives: This study investigated the toxic effects of cadmium on placental function and reproduction in rats. For this study, the mRNA levels of the placental prolactin-growth hormone (PRL-GH) gene family, placental trophoblast cell frequemcy and reproductive data were analyzed. Methods: Pregnant F344 Fisher rats (200 g±23 g) were intraperitoneally injected with 0, 0.5, and 5.0 ㎎/㎏ B.W/day of cadmium (CdCl2) dissolved in saline from days 7-11 or 16-20 of pregnancy, and were sacrificed at days 11 or 20, respectively. The mRNA levels were analyzed by Northern blot hybridization and reverse transcription-polymerase chain reaction. The hormone concentration was analyzed by radioimmunoassay and the frequemcy of the placental trophoblast cells was observed by histochemical study. Reproductive data were surveyed at day 20 of the pregnancy and after the births. Statistical analysis was carried out using the SAS program (version 8.1). Results: The mRNA levels of the PRL-GH gene family were reduced dose dependently by cadmium. The mRNA levels of Pit-1a and -b isotype genes were also reduced by cadmium. The hormone concentration of PL-Iv and -Ⅱ was decreased by cadmium. During the second half of pregnancy (days 11-21), a high dose of cadmium exposure significantly reduced the frequency of spongiotrophoblast and trophoblast giant cells that secrete the PRL-GH hormones. In the last stage of pregnancy (day 20), a high dose of cadmium exposure induced the apoptosis of spongiotrophoblast cells in the junctional zone of the placenta. Reproductive data such as placental and infant weight, number of live fetuses were decreased, and number of resorptions and dead fetuses, post-implantation loss were increased significantly in the cadmium exposed group compared with the control. Conclusions: Cadmium disrupts the functions of the placenta and these effects leads to reproductive disorders in rats.