과배란 미성숙 흰쥐에서 prolactin이 난소 내 tissue type plasminogen activator와 prostaglandin E2에 미치는 영향

김경원 ( Kyoung Won Kim ),심재철 ( Jae Chul Sim ),양회생 ( Hoe Saeng Yang ) 대한산부인과학회 2007 Obstetrics & Gynecology Science Vol.50 No.2

목적: 고PRL혈증이 불임, 무월경, 배란장애 등 여성 생식기능 장애를 유발시키는 난소 내 원인을 알아보고자 한다. 연구방법: 고PRL혈증이 배란 및 난소 형태적 변화에 영향을 미치는지 확인하기 위해 미성숙 암컷 흰쥐를 과배란 시킨 후 control, PRL 150 ug, PRL 300 ug, PRL 300 ug과 naloxone 300 ug을 복강 주사한 group으로 각각 분류 후 시간경과에 따를 배란수 및 조직학적 형태 관찰한다. 고PRL혈증으로 인한 난소내 t-PA 및 PGE2 변화를 확인하기 위해 미성숙 암컷 흰쥐를 동일한 방법으로 배란 유도 후 hCG투여 4시간 경과하여 control과 culture medium내 PRL 300 ng을 혼합한 group으로 분류하고, gonadotropin 투여로 인한 PGE2 농도변화를 확인하기 위해 PRL과 hMG 혼합군, PRL 300 ng과 FSH 혼합군으로 임의 분류 후 도살한 뒤 상층액을 채취해 EIA방법으로 t-PA 및 PGE2농도를 측정한다. 각 변수는 이원배치분석 후 각 군별 또는 각 시간별 종속변수의 변화치를 Mann-Whitney U 검사 또는 Kruskal-Wallis 검사를 시행하여, p<0.05일 때 유의하다고 판정하였다. 결과: PRL농도와 상관없이 시간이 경과될수록 배란된 난자수는 증가되었고 (p<0.05), 각 시간별 배란수는 PRL농도가 증가될수록 감소되었으나 시간이 경과됨에 따라 배란 억제율은 감소되어 PRL의 배란억제 효과는 일시적임을 알 수 있다. 또한 opioid antagonist인 naloxone 투여로 억제된 배란이 회복되는 것으로 보아 PRL은 배란과정중 opioid pathway에 영향을 미침이 간접적으로 확인되었다. PRL투여 후 t-PA 농도를 변화는 PRL투여군에서 control보다 낮은 농도를 나타내는 것으로 보아 PRL은 t-PA를 억제하는 것으로 사료된다. 그러나 PRL투여로 인한 PGE2 억제소견은 없었으며 hMG나 FSH와 같은 gonadotrophin을 병합 투여한 경우 난소내 PGE2농도는 상승하였다. 결론: 고PRL혈증은 대단히 복잡한 난소내 작용에 의해 배란을 억제시키며 앞으로 배란에 관여하는 더 많은 인자들에 대한 고PRL혈증의 영향에 대한 연구는 고PRL혈증 환자들의 임신능력 회복 및 여성 생식기능 유지에 많은 기여를 하리라 생각된다. Objective: This study is to analyze the direct effects of hyperprolactinemia, cause of anovulation and infertility, on ovarian function. Methods: The prepubertal female Sprague-Dawley (SD) rats were obtained and ovulation was induced with PMSG and hCG s.c.. The rats were divided into four groups, which received the following treatments IP : saline 0.2 ml, 150 ug PRL, 300 ug PRL, 300 ug PRL plus 300 ug naloxone. The animals were killed and the oviducts were evaluated for the presence of ova. The ovary were then removed and evaluated under light microscopy. For changes of follicular t-PA and PGE2 concentration after PRL, immature female SD rats were stimulated as described above. At four hours after the hCG injection the rats were killed and the ovaried were removed. Each isolated ovaries were incubated in culture plate containing incubation medium or 300 ng PRL to be tested. And PRL plus gonadotropin in incubation medium was tested because of change of PGE2 concentration. After incubation period, t-PA and PGE2 were measured by EIA. Differences between groups were assessed by two-way ANOVA of variance followed Mann-Whitney U test or Kruskal-Wallis test for multiple comparisons. p<0.05 was considered statistically significant. Results: As result, prolactin transiently suppresses ovulation, especially with its increased concentration not by altering the ovarian morphology. But ovulation inhibition was reversed by naloxone injection. The level of t-PA in control and prolactin-treated group increased steadily in response to human chorionic gonadotropin administration, yet lower in prolactin-treated group. But PGE2 concentration was increased in gonadotropin mixed groups but not affected in prolactin-treated group despite a significant blockade of ovulation. Conclusion: Thus, further studies on the effect of high level prolactin on ovulatory function would significantly contribute toward the patient with hyperprolactinemia for managing infertility and maintaining appropriate female reproductive function.

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.

Increased Phosphatase of Regenerating-1 by Placental Stem Cells Promote Hepatic Regeneration in a Rat Model with Bile Duct Ligation

( Jong Ho Choi ),( Gi Dae Kim ),( Jin Seok ),( Si Hyun Bae ),( Soon Koo Baik ),( Hoon Oh ),( Gi Jin Kim ) 대한간학회 2016 춘·추계 학술대회 (KASL) Vol.2016 No.1

Aims: Phosphatase of regenerating liver-1 (PRL-1) controls diverse cellular processes including liver regeneration. However, it is still unknown whether MSCs influence PRL-1 expression during regeneration of a damaged liver. We therefore investigated PRL-1 expression and its functions in bile duct ligation (BDL) following transplantation (Tx) with CP-MSCs. Methods: CP-MSCs and WI-38 cells labeled with fluorescent dye were engrafted into BDL model via intravenous Tx. Expression markers related to engraftment and proliferation in hepatic cells were analyzed by quantitative real time-PCR, Western blot and immunofluorescence. Furthermore, BrdU incorporation and FACS analysis were performed to confirm the CP-MSC effect on the regeneration of injured hepatic cells in in vitro. Results: CP-MSC Tx decreased the level of cirrhosis in a BDL rat model compared with others. The expression of PRL-1 and Rho family-related genes in the liver tissue of a BDL rat model was increased by CP-MSC Tx. Interestingly, CP-MSC migration was also decreased by PRL-1 siRNA treatment. Furthermore, CP-MSC Tx increased the expression of albumin and PRL-1 in liver tissue compared with others as well as the proliferation of hepatocytes in in vitro. However, proliferation and albumin production in primary hepatocytes were decreased by the PRL-1 siRNA treatment. Conclusions: Taken together, the increase in PRL-1 expression induced by CP-MSC Tx enhanced liver regeneration in a rat hepatic failure model via the dual function of PRL-1 controlling CP-MSC migration and hepatic proliferation. Therefore, these findings reveal a fundamental mechanism of the therapeutic effects of PRL-1 on hepatic diseases resulting from CP-MSC Tx.

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>