Due to modern trends with postponing child-bearing and getting worse living environment in women, an ovarian aging increased pregnancy failure and other complications with menopause or premature ovarian failure. Although several theories have been suggested such as mitochondrial malfunction, DNA damage/repair/methylation, caloric restriction, studies regarding ovarian aging-related molecular mechanisms for development of therapeutic methods are insufficient so far. Our objective is to determine molecular pathways of ovarian aging that result in pregnancy failure and other complications in women health to develop treatment strategies. This study is consisted of two parts: in Phase I stage, we analyzed distinct gene expression profile between young and aged mouse ovaries, and in Phase II stage several preferentially expressed genes in both ovaries were selected and analyzed their physiological functions and involved molecular networks related to ovarian aging for development of diagnostic markers and therapeutic methods. Ovaries from 10 week and 11 month-old FVB/NJ female mice with synchronized estrus cycle were collected for this study. A half of each ovary was used for RNA preparation and the other half for histological analysis. Using the Illumina HiSeq 2000 System, preferentially expressed genes were identified. Functional annotation database-based gene-set enrichment analyses and Pathway Studio® were employed to evaluate aging-related molecular networks. These findings were confirmed through qRT-PCR and immunohistochemistry. To validate RNA-Seq data, we examined expression patterns of marker genes (Amh, Bmp15 and Nobox) that were wellknown to be decreased in ovarian aging process. In young or aged ovary, preferentially expressed 876 genes were identified and extracellular matrix (ECM; p<0.001) and chromatin/nucleosome-related (p<0.001) protein-coded genes have the majority in these genes by GOTERM analysis. Amongthem, we selected several candidate genes and confirmed their expression profiles by qRT-PCR and immunohistochemistry followed by molecular network analysis. Regarding molecular interactions in these genes, PathwayStudio® was employed to predict aging-involved molecular networks in mouse ovary. Here we report a couple of candidate molecular networks and medicines (chemicals) for targeting these preferentially expressed genes/proteins. Further analyses are scheduled to produce transgenic animal models and with human ovarian tissues/cell lines.

내분비계 교란물질은 환경으로 방출되어 다양한 경로를 통해 조직에 축적되며, 각종 형태의 교란을 일으킴으로써 생식 이상과 성장억제 등을 초래한다. 본 연구에서는 내분비계 교란물질로 분류되는 Phthalate 및 대체물질 안전성 평가를 위해 TG407 실험을 수행하였다. TG407에 따라 (8주령) 수컷 Mouse에 Sonde를 이용하여 Phthalate (DEHP, 양성대조군; 4, 400 mg/kg/day) 및 대체후보물질 2종 (ATEC; 4, 40, 400 mg/kg/day), (ATHC; 400 mg/kg/day)를 28일 동안 경구투여 하였으며, 경구투여 종료 후 Sampling, 혈당 측정, CBC 측정, 호르몬 측정을 수행하였다. CBC와 혈당, Insulin 측정 및 뇌, 뇌하수체, 심장, 부정소, 정낭의 무게 측정 결과, 대조군과의 차이를 보이지 않았으며, T4와 T3의 농도는 ATHC를 제외하고는 대조군과의 차이를 보이지 않았다. ATHC(400 mg/kg)를 처리한 경우, 대조군에 비해 T4가 높았으며, T3는 낮았다. 간의 경우 DEHP(400 mg/kg)을 처리했을 때 대조군에 비해 무게가 증가하였다. 신장의 경우, DEHP(400 mg/kg), ATHC(400 mg/kg)을 처리했을 때 대조군에 비해 무게가 증가하였으며, 부신의 경우, DEHP(400 mg/kg), ATEC(40, 400 mg/kg), ATHC(400 mg/kg)을 처리했을 때 대조군에 비해 무게가 증가하였다. 흉선의 경우 DEHP(400 mg/kg), ATHC(400 mg/kg)을 처리했을 때 대조군에 비해 무게가 증가하였으며, 비장의 경우, DEHP(400 mg/kg)을 처리했을 때 대조군에 비해 무게가 증가하였다. 정소의 경우, DEHP(400 mg/kg), ATEC(40, 400 mg/kg), ATHC(400 mg/kg)을 처리했을 때 대조군에 비해 무게가 증가하였으며, 전립선의 경우, DEHP(400 mg/kg), ATHC(400 mg/kg)을 처리했을 때 대조군에 비해 무게가 증가하였다. 간, 신장, 부신, 비장, 정소, 부정소, 전립선을 대상으로 조직학적 분석을 진행하였으며, 특이적인 소견을 보이지 않았다. 본 연구를 통해 ATEC가 가장 독성이 낮은 것으로 사료되며, 따라서 ATEC와 같은 대체소재가 향후 플라스틱 시장에서 Phthalate를 대체할 수 있을 것으로 생각된다.

Interleukin(IL)은 cytokines의 group으로 주로 macrophages에 의해 생산되어지며, 그 중 IL-1 system 은 염증반응과 면역 및 항상성을 조절한다. Interleukin-1은 Interleukin-1α(IL-1α)와 Interleukin-1β (IL-1β) 2개의 isoform이 존재하며, 이는 IL-1Receptor1(IL-1R1)과 IL-1Receptor2(IL-1R2) 두 가지 타입의 receptor에 결합하여 작용한다. 또한 IL-1R1의 신호전달에는 IL-1 receptor accessory protein (IL-1RAcP)가 필요로 한다. 또한 IL-1RAcP와 IL-1R1은 Toll interacting protein(TOLLIP)과도 작용한 다고 알려져 있다. Interleukin-1 cytokine family member 중 Interleukin-1 receptor antagonist(IL-1Ra)는 IL-1α와 IL-1β의 작용을 저해한다. 선행된 다른 연구에서는 IL-1의 과발현이 정자형성에 대한 부정 적인 효과를 주는 것이 밝혀진바 있다. 따라서 본 연구에서는 정자에서 IL-1α, IL-1β, IL-1R1, IL-1RAcP와 TOLLIP의 발현 여부와 IL-1α, IL-1β와 IL-1Ra가 정자에 미치는 영향을 알아보고자 하였으며, 정자에게 미치는 영향 중에서도 정자의 운동성에 관여한다고 알려져 있는 GSK3의 억제성 인산화 형태인 p-GSK3에 대해서 알아보고자 하였다. 실험 결과, 생쥐의 정자에서 IL-1α, IL-1β, IL-1R1, IL-1RAcP와 TOLLIP이 모두 정자에서 발현되는 것을 확인하였다. Western blot 실험 결과, IL-1α, IL-1β가 단독 처리된 생쥐 정자에서 p-GSK3의 발현이 증가한 것을 확인하였다. 그리고 IL-1Ra가 처리된 경우 p-GSK3의 발현이 생쥐 정자에서 대조군에 비해 감소한 것을 확인하였다. IL-1 α, IL-1β를 IL-1Ra와 함께 처리 시에는 IL-1α, IL-1β를 단독처리 했을때 보다 생쥐의 정자에서 p-GSK3의 발현이 감소한 것을 확인하였다. 이러한 결과를 토대로 IL-1과 IL-1R1이 정자에서 발현되며, IL-1 및 IL-1Ra 처리시 p-GSK3의 발현에 영향을 미치는 것으로 보아 IL-1이 정자운동성에도 관여하는 것으로 사료된다.

Spatiotemporal expressions of microRNAs (miRNAs) are altered by the physiological states of cells which could be influenced by microenvironment. Function of miRNAs has been focused as a new regulator of gene expressions and cell differentiation in human health and diseases. We found and identified the several miRNAs, which were related to developmental competence of preimplantation and implantation process of mouse blastocysts and outgrowth embryos by microarray-based bioinformatical studies. In this study, we evaluated three miRNAs expressions related to third cleavage event in conditioned media (CM) and blastocysts. Mouse 2-cell stage embryos were collected and monitored for 9 hours. The embryos were divided two groups as early third cleavage before 9 hours of collection and late third cleavage after 9 hours of collection. They were cultured to blastocyst stage up to day-5 after hCG injection. The total number of cells and the number of cells with fragmented DNA were assessed in blastocysts by terminal dUTP nick-end labelling (TUNEL) staining and DAPI staining. Mean cell number of early third cleavage group was significantly higher than that of late third cleavage group (105.3±8.0 vs 81.8±7.0, p<0.05), but apoptotic index was not different. The miRNAs of CM and blastocysts from early and late group were prepared, and quantified by qRT-PCR with TaqMan probes. The expression levels of three miRNAs (mmu-let-7b, mmu-miR-183, and mmu-miR-429) in CM and blastocysts were slightly upregulated in late third cleavage group. Our study suggested that the expression level of miRNAs could be altered with embryo quality, and miRNAs in CM may be used to predict miRNAs expression of embryos and developmental competence.

The Hippo signaling pathway is essential for regulating proliferation, differentiation, and apoptosis in mammalian cells. Hippo signaling pathway exists in most body tissues and organs, where it controls the size of organs and tissues by keeping cell growth in check and promoting cell death as needed. It has been reported that the members of Hippo signaling pathway are highly expressed in mammalian ovaries and uteri. However, the regulatory mechanism of this pathway in the uterus during estrous cycle regulation remains unclear. Serine/Threonine Protein Kinase 4 (STK4, also known as MST1, a homolog of Hippo in Drosophila) is a major factor of Hippo signaling pathway. However STK4 in the mouse uterus has not yet been examined. The purpose of our study was to determine the expression of STK4 during the estrous cycle and regulation by estrogen in the mouse uterus. We found that STK4 was dynamically expressed in uterine endometrium during the estrous cycle. STK4 highly expressed at the estrus, diestrus, and were found to dramatically decrease as it progressed to the proestrus, metestrus stage of uterus during the estrous cycle. Expression of STK4 was dominant in glandular epithelial and luminal epithelial of proestrus, estrus, and diestrus stage, whereas in metestrus stage, expression of gene intensity was faint. Estrogen or estrogen receptor antagonist ICI 182,780 treatment, in ovariectomized mouse uterus, Expression of STK4 and its downstream genes were increased by estrogen. Our results show that the Hippo signaling pathway is estrogen-dependent in the mouse uterus. These informations will give us on sights to understand uterine dynamics during the estrous cycle.

Background & Objectives : Nonylphenol (NP), a member of alkylphenols, has been widely used in manufacturing antioxidants, The present study was undertaken to examine whether short-term exposure to NP can alter the onset of puberty and associated reproductive parameters such as hormone receptor expressions in prepubertal female rats. Methods : NP (0.1, 1, 10 mg/kg BW) was administered orally at 10:00 from postnatal day 25 (PND 25) through the PND 35. The control group was administered orally each day 200 uL of the sesame oil. At PND 36, all groups were sacrificed at 18:00. The first vaginal opening (V.O.) day was monitored, and the weights of reproductive tissues were measured. Ovaries and uteri were used for analysis of mRNA levels, protein levels and histology. The transcriptional activities of the ovaries (StAR, P405scc, 3β-HSD, 17β-HSD, Aromatase and LH-R) and uteri(ER) were measured using quantitative RT-PCRs. The uterine proteins were used for the ER western blotting. Histological analysis of the reproductive organs was carried out. Results: Administration of NP induced early V.O. when compared to control. The ovarian weights of 1 mg group (0.164±0.010 mg/g BW) was significantly higher than those of control group (0.116±0.007 mg/g BW). Among the steroidogenesis-related genes, mRNA levels of the ovarian P450scc, 3β-HSD and Aromatase are increased significantly in all NP-treated groups. The mRNA levels of StAR and 17β- HSD were unchanged. NP administration (1 mg group) significantly increased the LH-R transcript level. Among the steroid hormone receptor genes, the mRNA level of ER-alpha was significantly elevated in 1mg NP group, while the mRNA levels of ER-beta and PR were unchanged in all NP-treated groups. Interestingly, wetern blotting data for uterine steroid hormone receptors revealed the complete disagreement with the transcriptional activities. Histological status of ovaries and uteri also exerted the occureence of advanced maturation in NP-treated groups. Conclusion: The present study demonstartes that the short-term administration of NP accelerates V.O. and maturation of reproductive organs such as ovary and uterus.

Glycogen synthase kinase 3 α/β(GSK3α/β)는 serine/threonine kinase로 Wnt와 insulin은 GSK3α/ β의 활성을 감소시켜 하위에 있는 많은 표적단백질의 활성과 분해를 조절한다. 본 연구에서는 정자 의 성숙과 수정능력획득 과정에서 GSK3α/β의 기능을 규명하고자 하였다. 생쥐 부정소 상피에서 GSK3α/β와 p-GSK3α/β가 발현하였고, 정자의 첨체, 첨체후부(postacrosomal region), 중편(midpiece) 에서 발현하였다. p-GSK3α (Ser21)의 발현은 두부 부정소보다는 미부 부정소 정자에서 높았 다. 부정소에서 Wnt2b, 3a, 10a이 발현하였고, 특히 부정소 두부 상피의 principal cell에 다량으로 발현하였다. 정자성숙에 기여하는 epididymal exosome에서 Wnt2b, 3a, 10a를 확인하였다. 두부 부정 소 정자에 Wnt3a를 처리하면 GSK3α/β의 억제성 인산화의 변화 없이 운동성이 유의하게 증가하였다. 따라서 부정소 두부에서 exosome을 통해 분비된 Wnt가 정자 성숙과정에서 운동성의 증가에 관여 하는 것으로 사료된다. 생쥐와 사람 정자의 수정능획득 과정에서 total phosphotyrosine proteome의 증가와 함께 p-GSK3α(Ser21)가 증가하였다. 특히 두부 부정소보다 미부 부정소 정자에서 p-GSK3α (Ser21)가 높았다. 이는 GSK3α의 억제성인산화가 정자의 운동성에 중요함을 의미한다. 생쥐와 사람 의 정자에서 insulin like growth factors-1(IGF-1)은 GSK3α의 억제성 인산화 및 운동성을 유의적으로 증가시켰다. Protein phosphatase의 억제제인 Calyculin A는 GSK3α의 억제성 인산화 및 운동성을 유의적으로 증가시켰다. 이와는 반대로, AKT inhibitor는 GSK3α의 억제성 인산화 및 운동성을 감소 시켰다. 이는 insulin 수용체 하위에서 PI3/AKT kinase 회로를 경유하여 GSK3α/β의 억제성 인산화가 진행되어 정자의 운동성이 증가하는 것으로 사료된다. 결론적으로 정자의 성숙과정에서 부정소 기원의 Wnt는 정자의 운동성을 촉진하며, 성숙한 정자의 수정능력획득 과정에서 IGF-1등의 성장인자에 의해 GSK3α/β의 억제성 인산화가 촉진되어 정자의 운동성이 증가하는 것으로 사료된다.

Background & Objectives: Methoxychlor(MET), an organochlorine insecticide, has been thought a potent endocrine disrupting chemical. The present study was undertaken to examine whether short-term exposure to MET can alter the onset of puberty and the associated reproductive parameters such as hormone receptor expressions in prepubertal female rats. Method: MET (1, 10 and 100 mg/kg/day) was administered daily from postnatal day 25 (PND 25) through the PND 34, and the animals were sacrificed on the PND 35. The first V.O. day was monitored, and the weights of reproductive tissues were measured. To assess the structural alterations in the ovary and uterus, the tissues were embedded in paraffin and stained for histological analysis. The transcriptional activities of hypothalamic and pituitary genes were measured using quantitative RT-PCRs. The uterine and hypothalamic proteins were extracted and used for the ER western blotting. Results: As a result, 100 mg group showed advanced V.O. than control, 1 mg group and 10 mg group. The wet weights of ovaries from MET-treated animal dose-dependently increased. The uterine weights were increased in 1 mg group and 10 mg group, while the 100 mg group samples were not significantly different from control tissues. The adrenal, kidney, spleen and thymus weights were not shown any significant change. Corpora lutea and fully grown follicles were observed in the ovaries from the 100 mg group, while numerous primary and secondary follicles were observed in the ovaries from control group. Myometrial thickness of MET-treated group was dose-dependently increased. Epithelial hypertrophy and well-developed glands were observed in the uterus from the 10 mg groups. Conclusions: The present study demonstrated that the short-term exposure to MET during the critical period of prepubertal stage could activate a reproductive endocrine system, resulting the early onset of puberty in immature female rats. Our study suggests that MET’s disrupting effect might be derived from premature activation of key reproduction-related genes in hypothalamus-pituitary neuroendocrine circuit.

Nonylphenol (NP) is known to be an endocrine disruptor with estrogen-like activity. In this study, we tested low dose of NP and its alternative candidate Octyl-β-D-glucopyranoside (OG) on F1 female mice (n=10) from parents (P) generation to postnatal days(PND) 56. Animals [Control (tap water), NP-50 (50ug/L, drinking water), OG-50(50ug/L, drinking water)] were fed with normal chow and drinking water. ad libitum. Body weights and tissue weights were measured after sacrifice, and sera were measured using creatinine assay (n=8) and blood urea nitrogen (BUN) ELISA kit (n=8). Histopathology of kidney was used Hematoxylin & Eosin staining method and Periodic Acid-Schiff(PAS) staining method. NP treatment significantly increased body weight and kidney weight of F1 female(p<0.05). The serum levels of creatinine in NP-50 and OG-50 significantly decreased(p<0.001 and p<0.01, respectively) compared to the control group. In histopathological study, renal tubules spaces were increased and severe glomerular anomalies were found in NP-50 and OG-50. In NP-50, the thickened basal membrane was observed. The present study demonstrated that NP and OG has renal toxicity, and this toxicity due to long-term low dose treatment seems to be occurred in the next generation and female-specific.

Nonylphenols (NP) are used in manufacturing antioxidants and lubricating oil additives, which are considered to be as potent endocrine disruptor. The aim of this study was to examine the effects of NP on reproductive system in adult male mice. Mice were divided into 4 groups; (1) tap water (CON group), (2) 50 μg/L (NP50), (3) 500 μg/L (NP500) and (4) 5000 μg/L (NP5000) of NP via the drinking water for 4 weeks. Mice were sacrificed and the reproductive organ weights were measured. The caudal epididymal sperms were count to assess the toxicity on germ cells. Histopathological changes of tissues were observed by using hematoxylin/eosin staining. The weights of testis in NP5000 group were significantly lighter than those in CON group (104.9±2.9 mg vs 90.7±5.1 mg, p<0.05). And weights of epididymis significantly increased in NP500 group (44.2±2.6 mg vs 54.42±3.44 mg, p<0.05). As concentration of NP increased, the number of sperms significantly decreased (NP50 and NP500, p<0.01; NP5000, p<0.001). In histopathological analyses, the sperms in seminiferous tubules showed a concentration-dependent decrease in mice treated with NP. In epididymis, treatment with NP resulted in empty space and the reduced sperm numbers in dose-dependent manner. Our results confirmed the dose-dependent decrease in the number of sperm and histopathological abnormality of testis and epididymis in mice exposed from 50 μg/L to 5000 μg/L of NP for 4 weeks. The present study suggests that oral exposure of NP might have negative effects on reproductive function, particularly on germ cells, in adult male mice.

S100 protein family is small calcium-binding proteins with two EF-hand motifs and comprises more than 20 proteins in human. Although S100A proteins are known to play important roles in proinflammatory responses including damage-associated molecular pattern (DAMP) signaling and in the establishment of pregnancy, the expression of S100As have not been determined in the uterine endometrium during the estrous cycle in pigs. Thus, this study was performed to investigate expression and localization of S100A8, S100A9, and S100A12 in the uterine endometrial tissues during the estrous cycle in pigs. Real-time RT-PCR analysis showed that S100A8, S100A9, and S100A12 mRNAs were expressed in the uterine endometrium during the estrous cycle with higher levels on days 15 and 18 of the estrous cycle than the other days of cycle. Immunohistochemistry analysis showed that S100A9 and S100A12 proteins were mainly localized to the immune cells in the uterine endometrium. Especially, S100A9- and S100A12-positive immune cells were detected in the uterine blood vessels on day 15 of the estrous cycle, and also localized to stroma near to luminal epithelium on days 0 and 18 of the estrous cycle. These results showed that S100As were expressed in the uterine endometrium during the estrous cycle in a cyclic stage-specific manner, and these proteins were localized to the immune cells in the endometrium. These suggest that immune cells expressing S100A proteins may be recruited into the endometrium during the estrous cycle and play an important role in regulating endometrial function in pigs.

Bovine mammalian gland has biopotential in therapeutic protein production and could be used as a model in further lactation researches. In this study, we have isolated bovine mammary gland-derived epithelial cells (BMECs) from Korean Holstein dairy cattle and themselves show differential dynamic ability in in-vitro culture. BMECs enables to form lobulo-alveolar structure, express milk production related gene. Functional studies indicated that BMECs secret exogeneous antibacterial fragment-Bovine Lactoferricin B (bLfcin-B), which is inserted in PiggyBac system, and this bioactive fragment inhibits the growths of Escherichia coli and Staphylococcus aureus. These data demonstrated that BMECs open new scope in either bioactive fragment, heading to prevent the spread of mastitis or post-mastitis damage in dairy graze and could be an ideal bioreactor for antibacterial proteins.

Interleukin-12 (IL12) and IL23 are members of the IL12 family and secreted from antigen presenting cells (APCs) such as dendritic cells and macrophages. IL12 and IL23 are composed of two subunits of a sharing subunit, IL12B, and a unique subunit, IL12A for IL12 and IL23A for IL23. IL12 is involved in induction of T helper (Th) type 1 response, whereas IL23 is associated with the differentiation of naive T cells into Th17 cells. It has shown that IL12, a proinflammatory cytokine, is down-regulated during pregnancy for successful establishment and maintenance pregnancy and increases in plasma levels in women with preeclampsia. IL23 decreases IL12 expression to change the immune microenvironment at the maternal fetal interface in humans and mice. In the present study we determined the expression of IL12 and IL23 and their receptors in the endometrium and placenta during pregnancy. Real-time RT-PCR analysis showed levels of IL12A and IL12B mRNAs in the endometrium were high during early pregnancy, but maintained low during mid- to term pregnancy. During pregnancy, levels of IL12RB1 mRNA in the endometrium showed a biphasic pattern with the highest levels on Days 15 and 60 of pregnancy, while levels of IL12RB2 mRNA did not change. Levels of IL23A and IL23R mRNAs in the endometrium decreased toward term pregnancy. Immunohistochemical analysis showed that IL12A protein was localized specifically to scattered cells in endometrial stroma, but barely detected during mid- to term pregnancy. Conceptuses from early pregnancy expressed IL12, IL23, and their receptors, except IL12RB2, and corioallantoic tissues during mid- to late pregnancy expressed IL12, IL23, and their receptors, but not IL23R. These results showed that IL12 and IL23 and their receptors were expressed at the maternal-conceptus interface, suggesting that IL12 and IL23 may play a key role in regulating maternal immune environment for the establishment and maintenance of pregnancy in pigs.

Asymmetric cell divisions play crucial roles during ascidian embryogenesis. In these processes, an FGF signaling is an essential inductive signal for establishing cell fate polarization, such as mesenchyme and notochord. It was well reported that the FGF signaling cascade is composed of FGF, FGF receptor, Ras, MEK, Erk and Ets. However, mechanisms of communication between the FGF and other signaling pathways and of integrated regulation of signaling pathways have remained largely unknown. In this study, we isolated HrS6K, a homologue of the S6K gene that belongs to the S6-H1 kinase of the ribosomal S6 kinase family, and HrNck1, a homologue of Nck1 gene that encodes an adaptor protein containing Src homology 2 and 3 domains, from the ascidian Halocynthia roretzi, to elucidate the mechanisms. Zygotic expression of HrS6K was initiated as early as the 16-cell stage. In the 64-cell stage embryos, expression of HrS6K was seen in mesenchyme precursor cells. The signal was detected in dorsal midline cells and mesenchyme clusters of the early tailbud embryos, and then down-regulated by the late tailbud stage. In adults, HrS6K mRNA was highly detected in muscle and stomach by QPCR method. On the other hand, HrNck1 transcripts are detected maternally. Zygotic HrNck1 mRNA was strongly expressed in mesenchyme clusters of the neurula, and in tail tip cells of the early tailbud embryos. These results suggest that HrS6K and HrNck1 are involved in formation of mesenchyme cells, which are specified by the FGF signaling.

Gonadotropin-releasing hormone (GnRH) is a key neuropeptide regulating reproduction in humans and other vertebrates. Here, we evaluated the reproductive control system mediated by GnRH in the Pacific abalone Haliotis discus hannai. We cloned a prepro-GnRH cDNA (Hdh-GnRH) from the pleural-pedal ganglion (PPG) in H. discus hannai, and analyzed its spatiotemporal gene expression pattern. The open reading frame of Hdh-GnRH encodes a protein of 101 amino acids, consisting of a signal peptide, a GnRH dodecapeptide, a cleavage site, and a GnRH-associated peptide. This structure and sequence are highly similar to GnRH-like peptides reported for mollusks and other invertebrates. Quantitative polymerase chain reaction demonstrated that Hdh-GnRH mRNA was more strongly expressed in the ganglions (PPG and cerebral ganglion [CG]) than in other tissues (gonads, gills, intestine, hemocytes, muscle, and mantle) in both sexes. In females, the expression levels of Hdh-GnRH mRNA in the PPG and branchial ganglion (BG) were significantly higher at the ripe and partial spent stages than at the early and late active stages. In males, Hdh-GnRH mRNA levels in the BG showed a significant increase in the partial spent stage. Unexpectedly, Hdh-GnRH levels in the CG were not significantly different among the examined stages in both sexes. These results suggest that Hdh-GnRH mRNA expression profiles in the BG and possibly the PPG are tightly correlated with abalone reproductive activities.

In fish, light (photoperiod, intensity and spectra) is main regulator in many physiological actions including body growth. The present study was to investigate the effect of light spectra on the body growth in red spotted grouper Epinephelus akaara. Experiment performed from December, 2016 to March, 2017 and fish (n=200, body weight 6.9±0.1, total length 7.5±0.1) was reared under different light spectra (blue, green, red and white) for 15 week under 12 hour light and 12 hour dark photoperiod and natural water temperature (13.6~16.7℃). We examined the mRNA expression levels of growth hormone (GH) in the brain with pituitary and photoreceptors mRNA expression in the retina. Red spotted grouper of BW was no different in each for the four groups (light spectra of blue, green, red and white) to five week of rearing period, but BW of rearing blue light spectra group fish was significantly increased in ten week of rearing period. After that, in 15week of rearing period, BW of four group fish was no showed difference respectively. The total length (TL) of fish was no differed in four experimental groups under the experimental duration respectively. The gh mRNA was more highly expressed in rearing blue light spectra group fish than other experimental groups in 15 weeks. The short wavelength sensitive opsin (sws) mRNA was more increased in rearing blue light spectra group fish than other groups in 15 weeks. The middle wavelength sensitive opsin (mws) and rod opsin mRNA expression was no differed in four experimental groups under all experimental durations respectively. The long wavelength sensitive opsin (lws) mRNA expression was more increased in rearing red light spectra group fish than other groups in 15 weeks. Our present results suggest that somatic growth of red spotted grouper is induced under blue light condition under low water temperature duration. Also, these results are thought to be affected of the gh and sws mRNA expression blue wavelength. However, in this experiment we did not confirm the correlation between GH and photoreceptor. Therefore, further studies are needed to clarify the correlation of the light spectra and photoreceptor with body growth in red spotted grouper.

동물의 성은 유전학적 성과 형태학적 성으로 구분할 수 있으며, 성에 관한 표현은 일반적으로 형태 학적 성을 기준으로 한다(Lee, 2015). 둥근전복속 전복의 형태학적 성분화 및 성성숙에 관한 연구는 이들의 생식생물학적 기초자료 제공에 필요하다. 또한, 연체동물의 성분화 및 성성숙은 유전적 요인과 환경적 요인에 영향을 받을 수 있다(Yusa, 2007; Chávez-Villalba et al., 2011). 본 연구는 둥근전복 속(Haliotis spp.) 교잡종의 성성숙을 생식생물학적 지표를 이용하여 확인하고자 하였다. 연구에는 162개체의 평균 각장 49.12(±1.05) mm인 둥근전복속 교잡종을 사용하였다(북방전복, H. discus hannai♀* 둥근전복, H.discus discus♂, H. discus hannai♀* 왕전복, H. madaka♂, H. discus discus♀*H. discus hannai♂, H. discus discus♀*H. madaka♂, and H. discus discus♀* 말전복, H. gigantea♂). 교잡종의 성성숙을 확인하기 위해 생식소 부위가 포함된 간췌장을 적출하여 10% 중성 포르말린에 고정 후 조직표본을 제작하였다. 조직표본은 Mayer’s hematoxylin-0.5% eosin 비교염색을 실시하여 광학현미경으로 관찰하였다. 성성숙을 확인하기 위한 생식생물학적 지표로는 성비, 성성숙율 (%) 및 생식소지수(gonad index)를 이용하였다. 둥근전복속 교잡종의 성성숙과 관련한 생식생물학적 지표는 Table 1에 나타내었다.

Ascidian embryos have become an important model for embryological studies, offering a simple example for mechanisms of cytoplasmic components segregation. It is a well-known example that segregation of mitochondria into muscle lineage cells occurs during ascidian embryogenesis. It is, however, still unclear what signaling and molecular event control polarized distribution of mitochondria in the early ascidian embryonic development. To obtain molecular markers for studying mechanisms involved in polarized distribution of mitochondria, we have produced monoclonal antibodies, Mito-1, Mito-2 and Mito-3, that specifically recognize mitochondria-rich cytoplasm in all cells of the ascidian Halocynthia roretzi embryos. These antibodies stained cytoplasm like a mesh structure in epidermis cells, except for nuclei, at the early tailbud stage. Similar pattern was observed in vital staining of mitochondria with DiOC2, a fluorescent probe of mitochondria. These antibodies showed that mitochondria were distributed evenly in the animal hemisphere blastomeres at cleavage stages, whereas did not in the vegetal hemisphere blastomeres. Mitochondria were transferred more into cells of the marginal zone, such as muscle and nerve cord lineage cells, than into cells of the central zone, such as mesenchyme, notochord and endoderm lineage, in the vegetal hemisphere. Therefore, it is suggested that these antibodies may be useful as markers for analysing mechanisms involved in polarized distribution of mitochondria during ascidian embryogenesis.

The author has investigated four Manila clam populations of the family Veneridae, belonging to the order Veneroida. The clam is also indigenous to some parts of the sandy regions of the West Sea in the Korean Peninsula, as well as in several areas in China. Clams are the most popular marine products in Korea because of their taste and nutritional value, and Koreans consume them in large quantities. However, in spite of their economic and scientific consequences, a little information currently exist regarding the physiological and ecological levels only of clam species in Korea. This study attempt is to elucidate the genetic distances within and between clam populations from the West Sea. Four populations of Manila clam (R. philippinarum) were obtained in adjacent district to the West Sea in Korea. Four populations of clam muscle was collected in sterile tubes, placed on ice immediately, and stored under refrigeration until needed. Genomic DNA was extracted and purified under the conditions described previously (Yoon and Kim, 2004). The degree of variability was calculated by use of the Dice coefficient (F), which is given by the formula: F=2 nab / (na+nb), where nab is the number of bands shared between the samples a and b, na is the total number of bands for sample a and nb is the total number of bands for sample b (Jeffreys and Morton, 1987; Yoke-Kqueen and Radu, 2006). Euclidean genetic distances within- and between-species were also calculated by complete linkage method with the support of the hierarchical dendrogram program Systat version 10. The genomic DNA isolated from four Manila clams populations in the West Sea, were amplified several times by PCR reaction. The dendrogram obtained by the six oligonucleotides primers indicates three genetic clusters. The hierarchical dendrogram indicates four main branches: cluster 1 (GOCHANG 01, 02, 04 and 05), cluster 2 (SEOCHEON 06, 07, 08, 09 and 10), cluster 3 (TAEAN 11, 12, 13, 14 and 15) and cluster 4 (ANMYEON 16, 17, 18, 19, 20 and GOCHANG 03). Multiple comparisons of average bandsharing values among Manila clam populations from four sections were generated according to the bandsharing values and similarity matrix. Ultimately, individuals from SEOCHEON clam population (0.637±0.227) exhibited higher bandsharing values than did individuals from GOCHANG clam population (0.402±0.115) (P<0.05).