• Journal of Veterinary Science
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• v.24 no.5
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• pp.49.1-49.15
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• 2023

Background: Hystricomorpha rodents display a similar placentation model to humans. The present study was carried out considering the scarcity of information concerning the placental development in agouti. Objective: Describe the microscopy of the placenta, subplacenta and yolk sac of agoutis in early pregnancy and report on the inversion of the yolk sac. Methods: Fifteen females between the 14^th-32^nd day of gestation were used following euthanasia. Gestational buttons were collected, fixed, processed, stained to optical microscopy or immunohistochemistry. Results: Chorioallantoic placenta (CP) ranged from conical to a half-sphere, as follows: from the 14^th to 17^th day, the CP displays an inverted "V" shape, predominantly formed by cytotrophoblasts; from 20 to 22 days, formed almost entirely by cytotrophoblasts; at 28 days, a half sphere, with distinct lobes and interlobular area, numerous maternal gaps delimited by syncytiotrophoblasts and trophoblast giant cells; at 32 days, globose and undergoing the maturation process. Subplacenta, located between decidua and CP, initially presents septa consisting of simple columnar epithelium and after 17 days, comprising stratified epithelium. Visceral yolk sac (VYS) is attached to two CP projections between 14 and 17 days, formed by a simple cubic epithelium and inverted. Between 20 and 22 days, the epithelium displays apical villous projections with cytoplasmic vacuoles and a vascularized mesoderm. After the 24^th day, the VYS near the placenta is pleated, very vascularized and villous, with decreased villi sizes further away from the placenta. Conclusion: The agouti CP displays similar characteristics to other hystricomorpha, including placenta lobulation, a subplacenta and an inverted vitelline placenta.

• Development and Reproduction
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• v.6 no.2
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• pp.77-82
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• 2002

In ascidians, a primitive chordate, maternal cytoplasmic factors and inductive interactions are involved in the specification of cell fates in early embryos. The larval structure of ascidians is relatively simple, and the major mesodermal tissues of the tadpole larva are notochord, muscle, and mesenchyme. Formation of muscle cells is a cell-autonomous process, and localized maternal macho-l mRNA specify muscle fate in the posterior marginal zone of the early embryo. In contrast, inductive influence from endoderm precursors plays important roles in the specification of notochord and mesenchyme fates. FGF-Ras-MAPK signaling is involved In the induction of both tissues. The difference in responsiveness of the posterior mesenchyme and anterior notochord precursors to FGF signaling is caused by the presence or absence of intrinsic factors that inherited from the posterior-vegetal egg cytoplasm, respectively. In these inductions, directed signal polarizes the induced cells and promotes asymmetric cell divisions to produce two daughter cells with distinct fates.

• Development and Reproduction
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• v.15 no.4
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• pp.339-347
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• 2011

Human eyelid adipose-derived stem cells (hEAs) and amniotic mesenchymal stem cells (hAMs) are very valuable sources for the cell therapeutics. Both types of cells have a great proliferating ability in vitro and a multipotency to differentiate into adipocytes, osteoblasts and chondrocytes. In the present study, we evaluated their stem cell characteristics after long-time cryopreservation for 6, 12 and 24 months. When frozen-thawed cells were cultivated in vitro, their cumulative cell number and doubling time were similar to freshly prepared cells. Also they expressed stem cell-related genes of SCF, NANOG, OCT4, and TERT, ectoderm-related genes of NCAM and FGF5, mesoderm/endoderm-related genes of CK18 and VIM, and immune-related genes of HLA-ABC and ${\beta}$2M. Following differentiation culture in appropriate culture media for 2-3 weeks, both types of cells exhibited well differentiation into adipocyte, osteoblast, and chondrocyte, as revealed by adipogenic, osteogenic or chondrogenic-specific staining and related genes, respectively. In conclusion, even after long-term storage hEAs and hAMs could maintain their stem cell characteristics, suggesting that they might be suitable for clinical application based on stem cell therapy.

• Reproductive and Developmental Biology
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• v.35 no.1
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• pp.1-8
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• 2011

Techniques to evaluate gene expression profiling, such as sufficiently sensitive cDNA microarrays or real-time quantitative PCR, are efficient methods for monitoring human pluripotent stem cell (hESC/iPSC) cultures. However, most of these high-throughput tests have a limited use due to high cost, extended turn-around time, and the involvement of highly specialized technical expertise. Hence, there is an urgency of rapid, cost-effective, robust, yet sensitive method development for routine screening of hESCs/hiPSCs. A critical requirement in hESC/hiPSC cultures is to maintain a uniform undifferentiated state and to determine their differentiation capacity by showing the expression of gene markers representing all three germ layers, including ectoderm, mesoderm, and endoderm. To quantify the modulation of gene expression in hESCs/hiPSC during their propagation, expansion, and differentiation via embryoid body (EB) formation, we developed a simple, rapid, inexpensive, and definitive multimarker, semiquantitative multiplex RT-PCR platform technology. Among the 9 gene primers tested, 5 were pluripotent markers comprising set 1, and 3 lineage-specific markers were combined as set 2, respectively. We found that these 2 sets were not only effective in determining the relative differentiation in hESCs/hiPSCs, but were easily reproducible. In this study, we used the hES/hiPS cell lines to standardize the technique. This multiplex RT-PCR assay is flexible and, by selecting appropriate reporter genes, can be designed for characterization of different hESC/hiPSC lines during routine maintenance and directed differentiation.

• Reproductive and Developmental Biology
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• v.33 no.4
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• pp.263-271
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• 2009

Cardiovascular diseases (CVDs) are one of the most cause of death around the world and fields of interest for cardiac stem cells. Also, current use of terminally differentiated adult cardiomyocytes for CVDs has limited regenerative capacity therefore any significant cell loss may result in the development of progressive heart failure. Human embryonic stem cells (hESCs) derived from blastocyst-stage embryos spontaneously have ability to differentiate via embryo-like aggregates (endoderm, ectoderm and mesoderm) in vitro into various cell types including cardiomyocyte. However, most effective molecule or optimized condition which can induce cardiac differentiation of hESCs is rarely studied. In this study, we developed both spontaneous and inductive cardiomyocyte-like cells differentiation from hESCs by treatment of induced-factors, 5-azacytidine, BMP-4 and cardiogenol C. On the one hand, spontaneous and inductive cardiomyocyte-like cells showed that cardiac markers are expressed for further analysis by RT-PCR and immunocytochemistry. Interestingly, BMP-4 greatly improved homogeneous population of the cardiomyocyte-like cells from hESCs CHA15 and H09. In conclusion, we verified that spontaneously differentiated cells showed cardiac specific markers which characterize cardiac cells, treated extrinsic factors can manage cellular signals and found that hESCs can undergo differentiation into cardiomyocytes better than spontaneous group. This finding offers an insight into the inductive factor of differentiated cardiomyocytes and provides some helpful information that may offer the potential of cardiomyocytes derived from hESCs using extrinsic factors.

• BMB Reports
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• v.47 no.12
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• pp.673-678
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• 2014

During Xenopus early development, FGF signaling is involved in mesoderm formation and neurogenesis by modulating various signaling cascades. FGF-MAPK signaling induces Xbra expression, which maintains mesodermal fate through an autocatalytic-loop. Interestingly, previous reports have demonstrated that basic FGF (bFGF) treatment alone does not induce neurogenesis in ectodermal explants, even though FGF signaling inhibits BMP signaling via phosphorylation in Smad1 linker region. In addition, the overexpression of dominantnegative Xbra induces neurogenesis in ectodermal explants. However, the detailed mechanism underlying these phenomena has not yet been clarified. In this work, we showed that bFGF-Xbra signaling increased the PV.1 expression. DN-Xbra was found to decrease PV.1 expression, and the co-injection of PV.1 with DN-Xbra reduced neurogenesis in ectodermal explants. Furthermore, the knockdown of PV.1 induced neurogenesis in bFGF-treated ectodermal explants. Taken together, our results demonstrate that FGF-Xbra signaling induces PV.1 expression and that PV.1 functions as a neural repressor in the FGF-treated ectoderm.

• Journal of Veterinary Science
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• v.21 no.1
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• pp.9.1-9.15
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• 2020

Regenerative therapy holds great promise in the development of cures of some untreatable diseases such as cardiovascular diseases, and pluripotent stem cells (PSCs) including induced PSCs (iPSCs) are the most important regenerative seed cells. Recently, differentiation of human PSCs into functional tissues and cells in vitro has been widely reported. However, although porcine reports are rare they are quite essential, as the pig is an important animal model for the in vitro generation of human organs. In this study, we reprogramed porcine embryonic fibroblasts into porcine iPSCs (piPSCs), and differentiated them into cluster of differentiation 31 (CD31)-positive endothelial cells (ECs) (piPSC-derived ECs, piPS-ECs) using an optimized single-layer culture method. During differentiation, we observed that a combination of GSK3β inhibitor (CHIR99021) and bone morphogenetic protein 4 (BMP4) promoted mesodermal differentiation, resulting in higher proportions of CD31-positive cells than those from separate CHIR99021 or BMP4 treatment. Importantly, the piPS-ECs showed comparable morphological and functional properties to immortalized porcine aortic ECs, which are capable of taking up low-density lipoprotein and forming network structures on Matrigel. Our study, which is the first trial on a species other than human and mouse, has provided an optimized single-layer culture method for obtaining ECs from porcine PSCs. Our approach can be beneficial when evaluating autologous EC transplantation in pig models.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.10
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• pp.5705-5711
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• 2013

Background: Embryonic stem cells (ESCs) have the potential to form teratomas when implanted into immunodeficient mice, but data in immunocompetent mice are limited. We therefore investigated teratoma formation after implantation of three different mouse ESC (mESC) lines into immunocompetent mice. Materials and Methods: BALB/c mice were injected with three highly germline competent mESCs (129Sv, BALB/c and C57BL/6) subcutaneously or under the kidney capsule. After 4 weeks, mice were euthanized and examined histologically for teratoma development. The incidence, size and composition of teratomas were compared using Pearson Chi-square, t-test for dependent variables, one-way analysis of variance and the nonparametric Kruskal-Wallis analysis of variance and median test. Results: Teratomas developed from all three cell lines. The incidence of formation was significantly higher under the kidney capsule compared to subcutaneous site and occurred in both allogeneic and syngeneic mice. Overall, the size of teratoma was largest with the 129Sv cell line and under the kidney capsule. Diverse embryonic stem cell-derived tissues, belonging to the three embryonic germ layers, were encountered, reflecting the pluripotency of embryonic stem cells. Most commonly represented tissues were nervous tissue, keratinizing stratified squamous epithelium (ectoderm), smooth muscle, striated muscle, cartilage, bone (mesoderm), and glandular tissue in the form of gut- and respiratory-like epithelia (endoderm). Conclusions: ESCs can form teratomas in immunocompetent mice and, therefore, removal of undifferentiated ESC is a pre-requisite for a safe use of ESC in cell-based therapies. In addition the genetic relationship of the origin of the cell lines to the ability to transplant plays a major role.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.10 no.2
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• pp.71-96
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• 1977

Early development Linuparus trigonus(von Siebold) has been studied based on the samples collected monthly in Je-ju Island, Korea from February, 1975 to January, 1977. Gametogenesis, reproductive cycle, embryonic development were investigated by histological mettled, and morphological description was made on the first phyllosoma larva which reared in the laboratory. Testis is composed of two tubular duct which are symmetrical with H-shaped appearance. Outer layer of testis is of fibrous connective tissue capsule. In the lumen there is a convoluted seminiferous tubule with interstitial tissue. Ovary is a pair of symmetrical blind tubular lobes, and the midportions are connected each other. The ovary consists of a couple of ovarian sacs partitioned by two-layered connective tissue fibers. Proliferation of spermatogonia are observed all the year around on the germinal epithelium of seminiferous tubule. Partial spermatogenesis is always in progress, and the spermatozoa appear all the year around in the tubules. Nutrition of early oogonia is supplied by fibrous mesenchyme which is abundantly distributed in ovarian sacs. Oocytes grow and couplete maturation divisions in the follicle layers. They finally develop into mature ova before spawning. Reproductive cycle is classified into four successive stages; multiplication stage from September to December, growing stage from January to March, maturation division stage from April to May and mature stage from June to August. Spawning takes place from May to August with peak spawning from Into July to early August. Cleavage type is superficial. Blastopore is formed in blasto-disc region which is proliferation of blastoderm cells. Germinal layers are also derived from tile region. Mesoderm formation is originated from endodermal cells which are formed front the blasto-disc region. The endodermal cells are separated by the process of delamination from yolk sac and take part in the formation of the mid-gut. Morphological characteristics of first phyllosoma larva are different from the larvae of other Palinurid and Scyllarid species.

• Journal of the korean academy of Pediatric Dentistry
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• v.28 no.4
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• pp.709-727
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• 2001

The pattern of programmed cell death(PCD) has been examined during the early developmental period of development in mouse embryos, from embryonic day 4.5(E4.5) to E11.5 Embryos from Balb/c breedings were harvested at various embryonic stages between E4.5 and El1.5. Cell death was analysed by in situ terminal deoxynucleotidyl transferase mediated dUTP nick end labeling(TUNEL) staining in tissue sections and whole embryos. At the blastocyst stage(E4.5), a very few apoptotic cells were found in the inner cell mass of the blastocyst. In the early egg cylinder stage(35.0-5.5), a few apoptotic cells were detected in the embryonic ectoderm, the embryonic endoerm and the proamniotic cavity. In the advanced egg cylinder stage(E5.5-6.5), TUNEL-posifive cells were observed in the extra-embryonic ectoderm and extra-embryonic endoderm as well as in the embryonic ectoderm, embryonic visceral endoderm and proamniotic cavity. In the streak stage(E6.75-7.75), many TUNEL-positive cells were found in the ectoplacental cone. In contrast, only very few apoptotic cells were found in the chorion and extra-embryonic endoderm in extra-embryonic regions. In intra-embryonic region, a few apoptotic cells were randomly found in the embryonic ectoderm, mesoderm and visceral endoderm. At the early somitogenesis stage(E8.0-8.5), most apoptotic cells were observed in the most cranial portion of neural fold (neural ectoderm and adjacent ectoderm). At the mid somitogenesis stage(39.0-9.5), the otic placode first showed TUNEL-positive at this stage. Small number of TUNEL-positive cells were also first seen around optic placode and branchial arches. Three streams of TUNEL-positive cells were clearly seen in the cranial region at 59.5-9.75. At E10.5, apoptotic cells were localized in the developing eye, the junctional portion of medial nasal, lateral nasal and maxillary processes, the lateral portion of branchial arches, the junction of bilateral mandibular processes, and apical ectodermal ridges of limb buds. At E11.5, apoptotic cells were noticeably decreased in most area, except the developing limbs and several somites in the tail region. In this study, the global temporospatial pattern of PCD throughout early development of mouse embryos was discussed. It may provide the basis for further studies on its role in the morphogenesis of the embryo.