Bisphenol A (BPA), known as a typical endocrine disruptor, has been used commercially and widely for plastics and epoxy resins. BPA-based plastic is used extensively for the production of water bottles, food containers, CDs, DVDs, and panels that can be applied in construction. Epoxy resins containing BPA are used for coatings on the insides of water pipes, food cans, and thermal papers that are used in sales receipts. As its estrogenic effects and other adverse health effects have published, BPA has been regulated in many countries, and there have been efforts made to replace BPA. Other bisphenols substitutes such as bisphenol S (BPS) and bisphenol F (BPF) have been used. Currently, BPS- and BPF-based products labeled BPA-free products have been widely consumed. Because of structural similarities with BPA, however, these alternatives also show endocrine disruption effects like BPA, and many studies on adverse health effects of these alternatives are being reported. In this review, we describe the adverse health effects of bisphenols and the current status of regulation.

Phthalates, known as typical endocrine disruptors, are plasticizers used to soften plastics such as polyvinyl chloride (PVC). Because of their material properties, phthalates are used extensively in the production of toys, flooring, wood processing, detergents, and even cosmetics as lubricants and perfume solvents. Due to their endocrine disrupting effect and other adverse health effects published, recently, phthalates have been regulated in many countries. Besides, in an effort to replace phthalates, several chemical plasticizers such as trioctyltrimellitate (TOTM) and dioctylterephthalate (DIOP) have been used instead of the existing harmful phthalates, and novel alternatives are continuously being developed. Nonetheless, phthalates are still being detected in several plastic products, and the safety of alternatives that are considered safe is being questioned. In this review, we describe the adverse health effects of phthalates, their regulation and the current status of their alternatives.

Pollution in the fresh water system in urban area has the adverse effect on the amphibians population. Restoration activity of amphibian in the urban stream has been growing in Korea as well as western country. For successful restoration water quality of urban stream should be sufficient for survival and normal development of amphibian. To monitor the biological safety of surface water in the Tancheon basin, the capital area of Korea, a 6-day exposure Bombina orientalis embryo developmental toxicity assay was examined. The toxicity of surface water of Tancheon mainstream were lower than those of tributaries of Tancheon. The survival rate of embryos negatively correlated with total dissolved solid, turbidity and electrical conductivity whereas the developmental abnormality and growth retardation of embryos was positively correlated with total dissolved solid, turbidity and electrical conductivity. An amphibian developmental toxicity assay would be helpful for the selection of point for construction of habitat and reintroduction of amphibian in interrupted urban stream.

Endocirne disruptors (EDs) can cause fertility decrease, developmental disorder, and even cancer in animals. Until 90’s, EDs were used in various synthetic products including paints, coatings, detergents, plastics, and plasticizers. Currently, in several countries, the production, trade and use of EDs or EDs-suspected chemicals have been regulated while activity to screen the alternatives for EDs including bisphenol-A, phthalate and nonylphenol is active. Although various toxicity test method was developed and applied for screening of alternatives, however, the safety of alternatives has been not fully demonstrated. Some alternatives have high structural similarity with existing EDs, raising the possible risk of endocrine disruption by alternatives. In an effort to develop the safe alternatives, we reviewed the effects of EDs such as bisphenol-A, phthalates, nonylphenol and their substituents. In addition, in-silico analysis for endocrine disrupting activities of some alternatives was presented.

도심하천에 양서류를 재입식하기 위한 연구의 일환으로 2013, 14년에 걸쳐 강원도, 경기도 일부 지역에서 옴개구리 (Rana rugosa)를 대상으로 먹이활동 특성을 분석하였다. 옴개구리 위 내용물을 적출하여 70% 에탄올에 고정한 후 해부현미경으로 관찰 및 촬영하였고, 옴개구리 서식지에서 포충망을 이용한 쓸어잡기 (sweeping), 트랩 (trap) 채집을 실시하여 포획 생물들을 동정한 결과를 위 내용물 분석 결과와 비교하였다. 결과로서, 옴개구리는 벌목 (Hymenoptera), 딱정벌레목 (Cleoptera)의 곤충을 주로 섭식하는 것으로 확인되었다. 특히 위 내용물에서 확인된 벌목의 98% 이상은 개미과 (Formicidae)로 확인되었다. 옴개구리 서식지 주변 배회성 서식곤충은 남양주의 경우 벌목 58% (개미과 99%), 톡토기목 17%, 메뚜기목 10%, 파리목 9% 순으로 출현했다. 양평의 경우 톡토기목 49%, 메뚜기목 14%, 거미목 9%, 파리목 9%, 딱정벌레목 7%, 벌목 3% 순으로 출현했다. 이러한 결과는 하천변에 노린재목, 파리목이 많이 분포하지만 옴개구리는 하천변 바위나 초지에서 발견되는 개미나 딱정벌레들을 비교적 쉽게 사냥한다는 것을 의미한다. 옴개구리는 특정 먹이를 선호하기보다는 하천변에서 쉽게 발견되는 소형 육상곤충을 섭식하는 것으로 사료된다. 도심하천에 먹이자원은 옴개구리 서식의 제한요인이 되지 않을 것으로 사료된다.

Aluminum flows into the river from the abandoned mine leachate, industrial wastewater, and sewage and is responsible for acute toxicity in aquatic organisms. Recently, the number of reports have indicated the increased toxicity in a variety of aquatic organisms’ due to the aluminum toxicity. In this study, we reviewed the toxicity of aluminum on aquatic invertebrates, fishes and amphibians and suggested the guideline for management of aluminum residues in aquatic environment and strategies for aluminum toxicity evaluation. In aquatic animals aluminum complexes evoke gill dysfunction primarily, the cytotoxicity, genotoxicity, oxidative stress, disruption of endocrine function, reproductive success, metabolism and homeostasis. Notably, at environmentally relevant concentration, aluminum complex can alter the hormone levels in fish in acidic condition. Further, since the solubility of aluminum is higher in the acidic and basic conditions, thus it is likely that the toxic effects of aluminum may not only occur in acidic water near the abandoned mines but also in lakes and rivers, where pH is raised by algal bloom.

양서류는 육상과 수상생태계를 연결하는 먹이사슬의 연결자로 진화적 생태적 지이를 갖는다. 양서류의 배아와 유생은 모체와 독립되어 수환경 내에서 초기발생 및 성장하기 때문에 수환경에 존재하는 다양한 화학물질에 직접적으로 노출될 수 있다. Azole계열 항곰팡이제는 농업 및 의료용으로 널리 사용되는 화학물질로서 농지, 하수처리장 등으로 부터 수계로 유입된다. 최근, 양서류에서 이러한 azole계 물질에 의한 기형유발, 내분비계장애 효과가 증가하고 있다. 본 소고에서는 azole계 물질의 양서류 독성을 파악하고 azole계 물질의 안전한 이용을 위한 가이드라인을 제공하고자 azole계열에 속하는 imidazole, triazole, thiazole, oxazole, pyrazole 항곰팡이 물질이 양서류의 발생, 분화, 생식 등에 미치는 영향에 대해 최근까지의 연구결과를 정리하였다.

지구적으로 양서류가 감소하고 있다. 오래전부터 중금속은 양서류 군집 감소의 원인 중 한가지로 지목받고 있다. 수정 후 변태에 이르는 생활사를 수중에서 진행하는 양서류는 수환경 내의 오염물질에 1차적으로 노출되며 독성효과에 대한 감수성이 높아 수환경의 오염에 특히 취약하다. 양서류는 수서생태계의 건강도 지표로서 유용할 뿐 아니라 분자 및 개체수준의 다양한 생체지표를 이용한 다양한 환경오염물질의 독성평가 모델로서도 유용하다. 양서류에서 얻어진 독성자료는 수환경 오염물질의 관리와 안전관리기준으로 사용될 수 있다. 본 논문에 서는 기존에 보고된 중금속이 양서류의 다양한 독성종말점에 미치는 영향들에 대해 검토하고 분석하였다.

For successful embryo implantation, the stromal cells of the endometrium are morphologically and functionally differentiated into decidual cells. In the endometrium, estrogen induces proliferation of epithelial cells, but progesterone regulates the differentiation of epithelial cells, leading to decidualization of stromal cells. Kruppel like factor (KLF) is a zinc finger DNA binding protein that regulates transcription and has a wide range of functions in the cell cycle, cell apoptosis and differentiation control. In the uterus, KLF9, 13 plays an important role in implantation and decidual cell differentiation. KLF4 and KLF15 regulate the proliferation and differentiation of endometrial epithelial cells, but their role in stromal cells is unknown. In this study, we investigated the role of KLF4 and KLF15 in endometrial stromal cells. In mouse uterus, KLF4 was expressed in proliferative phase of glandular and luminal epithelial cells. However in endometrial stromal cells, KLF4 was highly expressed in secretory phase and secondary decidual zone after implantation. The expression of KLF15 was little in cytoplasm of luminal and glandular epithelial cells and proliferated in nucleus of secretory phase stromal cell.herefore, KLF4 and 15 are thought to be important for decidualization. To investigate the effect of estrogen and progesterone on the expression of KLF4 and KLF15, uterus of ovariectomy (OVX) mice which were injcected 17β- estradiol (E2, 0.3 mg) and progesterone (P4, 1 mg) and both ERα-knock out and wild type (diestrus, estrus) mice were used. KLF4 in OVX+E2 group was significantly higher than OVX+E2 / P4 group was lower than OVX+E2 group. There was no significant difference between ERαKO and WT diestrus group and significantly lower than WT estrus group. Expression of KLF15 was higher in the OVX+ P4 group than in the OVX group and lower in the OVX+E2 group. The OVX+E2 / P4 group was higher than the OVX+E2 group. There was no difference between ERαKO and WT diestrus. The differences in expression of KLF4 and KLF15 by P4 in OVX mouse uterine tissues may be due to the tissue specific expression pattern of epithelial (KLF4) and stromal cells (KLF15). The expression of KLF4 and KLF15 was increased by treatment of cyclic adenosine monophostphate (cAMP, 0.5 mM) and medroxyprogesterone acetate (MPA, 1μM) in human endometrial stromal cells. KLF15 siRNA increased the expression of decidualization markers (BMP2, IGFBP-1 and prolactin) with increasing progesterone receptor A/B (PR A/B), while KLF4 siRNA treatment decreased expression of decidualization markers. There was no significant difference in cell proliferation and apoptosis markers in KLF4 / 15 siRNA treatment. Therefore, progesterone induces KLF4 to promote decidualization, while normally induced KLF15 inhibits progesterone receptor expression. Expression of KLF4 in endometrial epithelium is induced by estrogen but induced by progesterone to promote decidualization, and KLF15 is mainly induced by progesterone in stromal cells and inhibit excessive PR A / B activity.

Estrogen plays an important role both in male and female reproduction. Two estrogen receptor isoforms, Esr1 and Esr2, are expressed in male gonad. In the mouse, Esr1 is expressed in Leydig cells of testis and pituitary. Esr1-/- male mice show enhanced androgen synthesis, spermatogenic defect, and infertility. To evaluate the specific function of Esr1 in Leydig cells, we examined spermatogenesis and steroidogenesis in Esr1f/fCyp17iCre male mice in which Esr1 is deleted specifically in Leydig cells. These mice showed normal spermatogenesis and fertility when compared to wild type from young adulthood to old age. Testosterone synthesis in Esr1f/fCyp17iCre mice at 3-12 months old of age was not different from age-matched wild type mice, while, at 18 months old of age, circulating testosterone concentrations were significantly higher than wild type together with increased levels of Star, Cyp17a1, and Hsd17b3 mRNA and with a hypertropy of Leydig cells. In Esr1f/fCyp17iCre mouse pituitaries, Fshb and Lhb mRNA levels were not different from wild type from young adulthood to old age. Taken together, Esr1 in Leydig cells may be not essential for spermatogenesis and fertility under control of endogenous estrogens and may have a role in aged Leydig cell function.

Estrogen is an important regulator of reproduction in both male and female. The two forms of estrogen receptor (ER) are known, ERα and ERβ. To understand the role of ERα in the testis, we investigated the expression of ERα in the mouse Leydig cells during postnatal development and the effects of estrogen on steroidogenesis and proliferation in progenitor Leydig cells (PLCs). In the testis, ERα mRNA and protein levels were markedly increased from postnatal day (PND) 1 to 14 and decreased thereafter until PND 56. During postnatal development ERα immunoreactivity was strong in the nucleus of Leydig cells at PND 14 when PLCs were abundant in the interstitium and low in the mature adult Leydig cells (ALCs). In fetal Leydig cells (FLCs), ERα immunoreactivity was negligible at birth and became increased at PND 14. This suggests an important role of ERα in Leydig cells during neonatal period. In isolated PLCs, 17β-estradiol (E2) and ERα-selective agonist, PPT suppressed the hCG-induced progesterone production and steroidogenic pathway genes expression. The hCG-induced PLCs proliferation was significantly inhibited by E2 and PPT. In conclusion, estrogen - ERα signaling may negatively regulate functional differentiation and proliferation of PLCs.

Expression of claudin-11 (CLDN11), a tight junction (TJ) protein, was examined in the Korean soft-shelled turtle (Pelodiscus maackii) testes. Spermatogenesis began during the breeding season and peaked at the end of the breeding season. Spermiation started in summer and peaked in autumn. The deduced amino acid sequence of P. maackii CLDN11 was similar to those of avian and mammalian species. During the non-breeding season when spermatogenesis and testosterone production were active, testicular Cldn11 levels were high. In the seminiferous epithelium, strong wavy CLDN11 strands parallel to the basement membrane delaminate the spermatogonia, and early spermatocytes are in the open compartment. Otherwise, CLDN11 was found beneath the early spermatocytes and in the Sertoli cell cytoplasm. Punctate zonular occludens-1 (ZO-1) immunoreactivity was found within the CLDN11 strands parallel to the basement membrane or at the outermost periphery of the seminiferous epithelium close to the basal lamina. During the breeding season, when circulating testosterone levels and spermatogenic activity was low, testicular CLDN11 level was lower than those of non-breeding season. CLDN11 was found at apicolateral contact sites between adjacent Sertoli cells devoid of the postmeiotic germ cells. At this time, lanthanum tracer diffused to the adluminal compartment of seminiferous epithelium. In cultured testis tissues, testosterone propionate significantly increased the level of Cldn11 mRNA. In P. maackii testis, CLDN11 participates in the development of the BTB where the CLDN11 expression was coupled with spermatogenic activity and circulating androgen levels, indicating the conserved nature of TJ’s expressing CLDN11 at the BTB in amniotes.

Sirt1 belongs to class III histone/protein deacetylase. Sirt1 KO mice have spermatogenic defects. In this study, expression of Sirt1 and acetylated histone 3k9 (H3k9ac) was examined in developing mouse testes. Among two splicing variants of sirt1 mRNA long form was dominant in developing testis. Testicular sirt1 mRNA levels were low at birth, increased until 14 days post partum (pp) and remained constant thereafter. Sirt1 immunoreactivity was weak to negligible in gonocytes, moderate in spermatogonia, absent in preleptotene spermatocytes, moderate in zygotene spermatocytes, strong in pachytene spermatocytes, and weak in diplotene spermatocytes. Round and elongating spermatids were negative for Sirt1. Acetylated histone 3k9 (H3k9ac) immunoreactivity was moderate in spermatogonia and weak to negligible in preleptotene to pachytene spermatocytes but strong in round and elongating spermatids. In Sertoli cells, nuclear Sirt1 immunoreactivity was absent at birth, increased at 14 days pp and markedly increased at 28 days pp onwards. In immature Sertoli cells culture, FSH and testosterone increased sirt1 mRNA, suggesting that Sirt1 participates in protein deacetylation events during the differentiation of Sertoli cells by gonadotropin. In the Leydig cells, nuclear Sirt1 immunoreactivity was weak until 2 weeks pp and decreased in 4 weeks pp onward. suggesting the protein deacetylation by Sirt1 in Leydig cell precursor/progenitor cells. Mutually exclusive expression between Sirt1 and H3k9ac in pachytene spermatocytes in testis suggests that deacetylation of H3K9ac by Sirt1 participates in the gene silencing and/or chromosome behavior in pachytene sspermatocytes.

Water channel proteins, aquaporins (AQPs) contribute to transepithelial water movement in many tissues. To date, 13 mammalian AQPs have been identified. Of these, AQP5 plays an important role in the fluid homeostasis and cell volume control in epithelial cells. In an effort to understand the role of AQP5 in testis, we investigated the expression of AQP5 in developing mouse testis, its regulation by estrogen and LH, and the change of steroidogenesis by AQP5 knockdown. Testes and Leydig cells were isolated from male mice at postnatal day (PND) 1, 7, 14, 28, and 56 and estrogen receptor alpha knockout (ERαKO) mice. In mouse testis, AQP5 immunoreactivity was negligible by PND 14. From PND 28 onward, AQP5 immunoreactivity was found in Leydig cells. In ERαKO mouse Leydig cells, AQP5 mRNA level was significantly lower than wild type. In primary adult Leydig cell culture, the expression of AQP5 mRNA was increased by 17β-estradiol (E2) and human chorionic gonadotropin (hCG), but was not changed in ERαKO Leydig cells. Moreover, the expression of AQP5 mRNA was increased by E2 and ERα-selective agonist PPT, but was not changed by ERβ-selective agonist DPN in primary Leydig cells and mLTC-1. In silico analysis and chromatin immunoprecipitation (ChIP) assay revealed that there are putative estrogen response elements (EREs) and cAMP response elements (CRE) in AQP5 promoter region. Testosterone secretion and steroidogenic pathway genes (StAR, Cyp11a1, Cyp17a1, and 3β-HSD6) expression were decreased by AQP5 siRNA in primary Leydig cells. In conclusion, AQP5 expression was coupled with functional differentiation of adult Leydig cells. AQP5 may play an important role in the fluid homeostasis and cell volume control during development of adult Leydig cells. The expression of AQP5 in Leydig cells could be regulated by ERα and LH signaling and AQP5 may be involved in steroidogenesis.

Amnionless (AMN) is a plasma membrane protein that binds to cubilin and megalin in various epithelia and mediates endocytosis of extracellular ligands. This function has been studied in the kidney where it plays a key role in vitamin B12 and vitamin D homeostasis. Present study aimed to elucidate developmental pattern of expression of AMN during the peri-implantation period in mouse embryos. In an effort to understand functional role of AMN in the histiotropic nutrition in blastocyst, endocytotic function of AMN for apoplipoprotein was examined in blastocyst. Eight-week-old female mice were superovulated by intraperitoneal injection of 5 IU PMSG and 5 IU hCG 48h later. To obtain embryos, females were mated with males. Mouse embryos were collected at 12, 48, 56, 65, 72 and 96 h post-hCG by flushing oviducts and uterus, and we also obtained gestation day 6.5, 7.5 and 8.5 embryos in uterus. All samples were subjected to quantitative RT-PCR, whole-mount immunofluorescence and immunohistochemistry analysis. To analyze endocytotic function of AMN, we examined uptake experiment of FICT labeled apolipoprotein A-I (ApoA-I-FITC) following functional blocking of AMN in blastocysts. AMN and cubilin mRNA was expressed in all developmental stages of mouse embryos. Megalin was the first detected at morula stage. AMN protein was expressed in trophectoderm (TE) and inner cell mass (ICM). AMN and cubilin were expressed in visceral endoderm of GD 6.5 and 7.5 embryos and visceral yolk sec of GD 8.5 embryos. In normal IgG treated embryos, ApoA-I-FITC was detected in intracellular vesicles of TE and ICM. However, in the presence of anti-AMN antibody, ApoA-I-FITC was weakly detected in apical surface of plasma membrane of TE. To date, AMN has been believed to be expressed in visceral endoderm of post implantation embryos. Our results demonstrated that AMN is the important molecular partner of cubilin and megalin in the preimplantation embryos and that AMN mediates endocytosis of apoplipoprotein, which may play a crucial role in embryonic development and normal growth via supporting histiotropic nutrition during peri-implantation period.