The successful establishment and maintenance of pregnancy is achieved by well-coordinated interactions between the maternal uterus and the implanting conceptus. In pigs, the conceptus undergoes dramatic morphological and functional changes, and secretes various biological products such as estrogens and cytokines, interleukin-1beta (IL1B), interferon-gamma (IFNG), and IFN-delta (IFND) during the implantation period. The uterine endometrium in response to the conceptus-derived molecules and ovarian progesterone becomes receptive to the conceptus by changing cell adhesion molecule expression, epithelial cell depolarization and secretory activity. Conceptus-derived estrogen acts as the maternal pregnancy recognition signal which changes the direction of endometrial prostaglandin (PG) F2 secretion from the uterine vasculature into the uterine lumen. Estrogen also induces the expression of a variety of endometrial genes, including AKR1B1, FGF7, LPAR3, and SPP1. The function of cytokines, IL1B, IFNG, and IFND, in the endometrium is not fully understood, but some recent work shows that IL1B is involved in the synthesis and transport of endometrial PGs by regulating endometrial expression of PG-synthetic enzymes, PTGS1, PTGS2, and AKR1B1, and PG transporters, ABCC4 and SLCO2A1. Estrogen and IL1B also stimulate endometrial expression of IFN signaling molecules, suggesting that estrogen and IL1B act cooperatively on priming the endometrial function of conceptus IFNG and IFND. In turn, IFNG derived from the elongating conceptuses, induces many endometrial genes, including CXCL9, CXCL10, CXCL12, and SLA-DQ. The role of IFND at the maternal-conceptus interface is not well understood yet. Further analysis of the molecules derived from the endometrium and conceptus will provide insights into the cellular and molecular basis of maternal-conceptus interactions for the establishment of successful pregnancy in pigs.

In this study, a energy recovery ventilation system was applied to the pig model, intake and exhaust duct were installed at a height of top(2.1 m), middle(1.25 m) and bottom(0.4 m). In each of the 9 test sites, one test was performed from 10:00 to 18:00, optimal temperature exchange efficiency and temperature distribution were analyzed. In order to analyze the effect of the energy recovery ventilation system, the temperature of 31 points(4 points of the ventilation system and 27 points in the pig house model) was measured from 10:00 to 18:00. The test that showed the highest heat exchange efficiency was the configuration of middle intake duct and top exhaust duct, which was 75.44%. The test that showed the most uniform distribution was the configuration of middle intake duct and bottom exhaust duct. These results will be utilized to optimally design feeding environment of actual pig houses and to reduce fuel cost.

The cryopreservation of sperm has become the subject of research for successful artificial insemination technologies. Antifreeze proteins (AFPs), one of the factors necessary for effective cryopreservation, are derived from certain Antarctic organisms. These proteins decrease the freezing point of water within these organisms to below the temperature of the surrounding seawater to protect the organism from cold shock. Accordingly, a recent study found that AFPs can increase the motility and viability of spermatozoa during cryopreservation.To evaluate this relationship, we performed cryopreservation of boar sperm with AFPs produced in the Arctic yeast Leucosporidium sp. AFP expression system at four concentrations (0, 0.01, 0.1, and 1 μg/ml) and evaluated motility using computer assisted sperm analysis. DNA damage to boar spermatozoa was measured by the comet assay, and sperm membrane integrity and acrosome integrity were evaluated by flow cytometry. The results showed that motility was positively affected by the addition of AFP at each concentration except 1 μg/ml (p<0.001).Although cryopreservation with AFP decreased the viability of the boar sperm using, the tail DNA analyses showed that there was no significant difference between the control and the addition of 0.1 or 0.01 μg/ml AFP. In addition, the percentage of live sperm with intact acrosomes showed the least significant difference between the control and 0.1 μg/ml AFP (p<0.05), but increased with 1 μg/ml AFP (p<0.001). Our results indicate that the addition of AFP during boar sperm cryopreservation can improve viability and acrosome integrity after thawing.

The objective of this study was to investigate the physicochemical characteristics of bear, pig, and cow biles to determine their authenticities for safe food distribution. The main bile acids of bear bile were tauroursodeoxycholic acid and taurochenodeoxycholic acid, while glycochenodeoxycholic acid and taurochenodeoxycholic acid for pig bile and taurocholic acid and glycocholic acid for cow bile were majorly detected by HPLC analysis. HPLC analysis was effective in monitoring of several samples imported as bear bile if employed to determine the authenticity of each bile. After the analysis of fatty acid composition by GC analysis, the ratio of the oleic acid of bear bile was relatively low compared to pig and cow biles. The ratio of the linoleic acid of bear bile was also similar to pig bile, whereas it had a tendency to be high compared to cow bear. The genetic analysis of the imported bile samples was mostly in agreement with the results of HPLC and GC analysis to identify the origin of imported biles. Finally, this study on the identification of bile origin by physicochemical analysis can give basic information to monitor the origin of biles and further to establish a reliable system for bear bile distribution.

This study was performed to compare the healing quality of the allogenic acellular dermal matix (ADM) and xenogenic ADM combined with autologous skin graft. Xenogenic ADM was obtained from two GalT knock-out pigs. Allogenic ADM was obtained from cynomolgus monkeys. ADM was stored with lyophilization. Full-thickness skin wounds were made on the back of two cynomolgus monkeys. In one monkey, wounds were covered by xenogenic ADM combined with autologous skin graft or autologous skin graft only. In another monkey, wounds were covered by allogenic ADM combined with autologous skin graft or autologous skin graft only. Skin healing process was observed during 2 weeks and skin biopsies were performed on 3 months after skin transplantation. We obtained IACUC approval (ORIENT-IACUC-16053) Skin on the xenogenic ADM was necrotized 1 week after skin transplantation. Possibly due to the thickness of ADM, which block the blood supply from the subcutaneous tissue to the autologous skin graft. Skin biopsy revealed that less fibrotic change of the skin on the ADM compared with the skin without ADM. Xenogenic ADM can be used in high degree burn patients who can suffered from contracture after healing since it can reduce fibrotic change.

Ribosomal protein L21 (RPL21) plays an important role in ribosome assembly. It is considered to be a major cause for the occurrence of the hypotrichosis simplex (HTS), a type of sustained hair loss from early childhood to adulthood. In this study, the full-length sequence of pig RPL21 gene (GenBank accession number: KU891824) was cloned and identified for the first time. We found it contains a 483-bp open reading frame (ORF) encoding 160 amino acids. It is located in the plus strand of chromosome 11, which spans 2,167 bp from 4,199,792 to 4,201,958. We found RPL21 expression level is closely related to cell proliferation and cell cycle arrest. In the knockdown group, the cell proliferation activity was significantly decreased (P<0.01) and an obvious accumulation of cells at the G2/M phase with a simultaneous up-regulation of p53 and p21 was observed. This likely due to knockdown of RPL21 triggered ribosomal stress, which affected the normal ribosome assembly and caused defective ribosome biogenesis. The unassembled RPs were released consequently from the nucleolus to the nucleoplasm where they can activate p53-dependent cell-cycle responsive factors and led to a G2/M arrest. We expect these results may provide valid information for further study on the pig RPL21 gene and the cause of hypo trichosis simplex.

Alzheimer's disease (AD) has caused by expression of amyloid precursor protein (APP), Tau and presenilin (PS) as known as plaques and tangle accumulation. AD transgenic porcine model is necessary for preclinical testing of therapeutic agent because of similar metabolic system between porcine and human. The objective of study was to generate AD transgenic pig by somatic cell nuclear transfer (SCNT) with multi-cistronic vector system. AD multi-cistronic vector was 6 well-known mutation on 3 AD related genes, hAPP (K670N/M671L, I716V, V717I), hTau (P301L) and hPS1 (M146V, L286P). Establishment of AD transgenic cell lines was used from Jeju black pig ear fibroblast cells (JB-PEFAD) with the AD multi-cistronic vector. The JB-PEFAD cell was confirmed on mRNA expression, protein synthesis of hAPP, hTau and hPS1 and identification of integration and karyotype. Although fusion rate was no difference in SCNT with JB-PEF AD (SCNTAD) embryos, cleavage and blastocyst formation rates were slightly lower than in SCNT with non-transgenic JB-PEF (SCNTnon-TG). Individual SCNTAD blastocysts were detected hAPP, hTau and hPS1 genomic integration which showed 93.2% (n=30) efficiency in genomic DNA (gDNA) level. It will give us a possibility to develop porcine animal model for AD study in the future.

Our objective was to evaluate the function of treahlose and erythritol in reducing ROS concentrations, which is associated with a general improvement in the quality of frozen-thawing miniature pig sperm. Semen was mixed in modified Modena B extender, added to cooling media and freezing media, followed by the supplement of 100 mM trehalose and/or 100 mM erythritol with spermatozoa (1000x 109cells/straw). The trehalose plus erythritol (TE) added group had less intracellular H2O2 than did control and trehalose (36.6±1.6 vs. 49.0±5.8 and 48.8±7.9; P<0.05). The percentage of viable acrosome-intact sperm (FITC-PNA-/PI-) was higher in erythritol and TE than controls (57.0±5.5% and 62.5±4.3% vs. 45.4±5.4%; P<0.05 and P<0.001). The percentage of sperm with high fragmented DNA was observed in control group when compared with erythritol and TE also trehalose (65.5±1.3% vs 59.3±0.7% and 59.0±0.3% vs 62.2± 0.8%; P<0.001). The percentage of sperm LPO was higher in control and trehalose than erythritol (4.4±0.5% and 5.0±0.5% vs. 3.5±0.2; P<0.01 and P<0.001), and was lowest in the TE (control and trehalose vs. TE: P<0.001, erythritol vs. TE: P<0.05). Also, we performed that surgical insemination based on above data to evaluate the function of new cryoprotectant such as trehalose plus erythritol in vivo. Finally, 1 pregnant gilt showed natural estrus was allowed to go to term and 8 live piglets were born. In conclusion, miniature pig sperm was successfully cryopreserved with trehalose plus erythritol provided the increasing the sperm quality and reducing the ROS.

Crocin is a carotenoid that may protect cells against oxidative stress by scavenging free radicals particularly superoxide anions. It has been reported that oocyte maturation is influenced by the free radicals generated during in vitro culture (IVC) process. The objective of study was to examine the effect of crocin in in vitro maturation (IVM) medium as an antioxidant on oocyte maturation and embryonic development after parthenogenesis (PA). Cumulus-oocyte complexes (COCs) were collected from ovaries of prepubertal gilts. The basic medium for IVM was medium-199 containing 10% pig follicular fluid, cysteine, pyruvate, epidermal growth factor, kanamycin, insulin, and hormones. Oocytes were treated for 44 hours with crocin at 0, 25, 50, and 100 μg/ml during IVM. Oocytes reached the metaphase II stage were induced for PA and cultured for 7 days in porcine zygote medium-3. Nuclear maturation of oocytes was not influenced by various concentrations of crocin (89.0, 87.3, 84.3, and 94.1% for control, 25, 50, and 100 μg/ml crocin, respectively). IVM oocytes treated with 50 μg/ml crocin showed a higher (P<0.05) intraoocyte glutathione (GSH) contents than untreated oocytes (1.00 vs. 1.29 pixels/oocyte). Blastocyst formation of PA embryos treated with 50 (42.9%) and 100 μg/ml crocin (43.8%) was significantly higher (P<0.05) than oocytes treated with 25 μg/ml crocin (30.5%) but not different from that (35.2%) of untreated oocytes. In summary, crocin increases cytoplasmic maturation in terms of intraoocyte GSH content which may be beneficial for later embryonic development by protecting from harmful effect of reactive oxygen species. Further studies are needed to determine whether the beneficial effect of crocin treatment during IVC would be shown in embryonic development after in vitro fertilization and somatic cell nuclear transfer.

Oocytes from small antral follicles (< 3 mm in diameter; SAFO) show lower developmental competence compared to those from medium antral follicles (3-8 mm in diameter; MAFO) in pigs. This study was designed to evaluate the effect of various macromolecules such as fetal bovine serum (FBS), porcine follicular fluid (PFF), bovine serum albumin (BSA) and polyvinyl alcohol (PVA) in in vitro growth (IVG) medium on oocyte growth, maturation, and embryonic development after parthenogenesis (PA). The base medium for IVG was α-MEM supplemented with dibutyryl cyclic AMP, pyruvate, kanamycin, hormone. This medium was further supplemented with 10% FBS, 10% PFF, 0.4% BSA, or 0.1% PVA. The in vitro maturation (IVM) medium was medium-199 supplemented with 10% PFF, cysteine, pyruvate, epidermal growth factor, kanamycin, insulin, and hormones. SAFO were cultured for 2 days for IVG and then cultured for 44 h for IVM. After IVG, the mean diameter of SAFO treated with FBS, PVA, and no IVG-MAFO (114.1, 113.0, and 114.8 μm, respectively) was significantly larger (P<0.01) than that of no IVG-SAF (111.8 μm). Oocyte diameter after IVM was greater (P<0.01) in SAFO treated with FBS, BSA and PVA (112.8, 112.9 and 112.6 μm, respectively) than other groups (110.4, 109.6, and 109.8 μm for no IVG-MAFO, no IVG-SAFO and PFF, respectively). Intraoocyte GSH content was not influenced by the macromolecules in IVG medium (0.92, 0.93, 1.05, and 1.12 pixels/oocyte for FBS, PFF, BSA and PVA, respectively). The proportion of oocytes reached the metaphase II stage was higher in PFF (73.6%) than in BSA (43.5%) and PVA (53.7%) but not different from that of FBS treatment (61.5%). The cumulus expansion score of oocytes after IVG was significantly influenced (P<0.01) by the macromolecules (2.94, 2.24, 1.84, and 1.38 for PFF, FBS, PVA, and BSA treatments, respectively). Blastocyst formation of PA oocytes that were treated with FBS (51.8%), PFF (50.4%), and PVA (45.2%) during IVG was higher (P<0.05) than that of BSA-treated oocytes (20.6%) but was not significantly different from that (54.8%) of no IVG-MAFO oocytes. Our results demonstrated that growth, maturation, and embryonic development of SAFO are greatly influenced by macromolecules in IVG medium and that PFF or FBS can be replaced with a chemically defined synthetic macromolecule PVA.

Crocin is a carotenoid that may protect cells against oxidative stress by scavenging free radicals particularly superoxide anions. It has been reported that oocyte maturation is influenced by the free radicals generated during in vitro culture (IVC) process. The objective of study was to examine the effect of crocin in in vitro maturation (IVM) medium as an antioxidant on oocyte maturation and embryonic development after parthenogenesis (PA). Cumulus-oocyte complexes (COCs) were collected from ovaries of prepubertal gilts. The basic medium for IVM was medium-199 containing 10% pig follicular fluid, cysteine, pyruvate, epidermal growth factor, kanamycin, insulin, and hormones. Oocytes were treated for 44 hours with crocin at 0, 25, 50, and 100 μg/ml during IVM. Oocytes reached the metaphase II stage were induced for PA and cultured for 7 days in porcine zygote medium-3. Nuclear maturation of oocytes was not influenced by various concentrations of crocin (89.0, 87.3, 84.3, and 94.1% for control, 25, 50, and 100 μg/ml crocin, respectively). IVM oocytes treated with 50 μg/ml crocin showed a higher (P<0.05) intraoocyte glutathione (GSH) contents than untreated oocytes (1.00 vs. 1.29 pixels/oocyte). Blastocyst formation of PA embryos treated with 50 (42.9%) and 100 μg/ml crocin (43.8%) was significantly higher (P<0.05) than oocytes treated with 25 μg/ml crocin (30.5%) but not different from that (35.2%) of untreated oocytes. In summary, crocin increases cytoplasmic maturation in terms of intraoocyte GSH content which may be beneficial for later embryonic development by protecting from harmful effect of reactive oxygen species. Further studies are needed to determine whether the beneficial effect of crocin treatment during IVC would be shown in embryonic development after in vitro fertilization and somatic cell nuclear transfer.

Mitochondrial dysfunction is found in oocytes and transmitted to the offspring due to maternal obesity. This is curable by endoplasmic reticulum (ER) stress inhibitors such as salubrinal (SAL). Recently pigs are considered as a model animal for biomedical research due to its physiological similarity with human. Pig oocytes have shown ER stress mostly in metaphase II stage. ER stress is hindering the in vitro embryo production (IVP). This study investigated the effect of ER stress inhibition by using SAL during 44 h of in vitro maturation (IVM) of oocytes at 1, 10, 50 and 100 nM concentrations. Firstly, we defined the concentration of SAL during IVM of pig oocytes. SAL at 10 nM showed higher (44.2 to 55.6%, P<P0.05) development competence to the blastocyst state than control and other concentrations after parthenogenetic activation (PA). Secondly, we sorted out the time-dependent treatment at 10 nM of SAL for IVM of oocytes. It revealed that treatment with SAL during 22 to 44 h and 0 to 44 h of IVM improved PA embryonic development significantly (40.5, 51.7 and 60.2% for control, 22 to 44 h and 0 to 44 h of IVM, respectively, P<0.05). Glutathione (GSH) level is an indicator of cytoplasmic maturation of oocytes. Reactive oxygen species (ROS) have a harmful effect on development competence of oocytes. For this, we determined the intraoocyte levels of GSH and ROS after 44 h of IVM. It was found that SAL increased intraoocyte GSH level and also decrease ROS level (P<0.05). Finally, we performed somatic cell nuclear transfer (SCNT) after treating oocytes with 10 nM SAL during IVM. SAL treatment significantly improved blastocyst formation of SCNT embryos compared to control (24.7 vs. 39.6%, P<0.05). Our results indicate that treatment of pig oocytes with ER stress inhibitor SAL during IVM improves preimplantation development cloned pig embryos by influencing cytoplasmic maturation in terms of increased GSH content and decreased ROS level in IVM pig oocytes.

For useful research animal to study human’s disease and for xenotransplantation donor, pig was studied to improve the quality of in vitro production (IVP). But, still the developmental ability of in vitro porcine embryos is still lower than in vivo embryos. Using a antioxidant is one of the strategy to overcome the drawback of in vitro producted embryos by protecting the oocyte from free radicals during in vitro maturation (IVM). Resveratrol, one of the plant-derived polyphenol antioxidants, have been used as effective antioxidants. Therefore, resveratrol treatment during IVM of porcine oocytes is expected to improve efficiency of the IVP by reducing free radical accumulation. In this study, we designed control (no treated) and resveratrol treatment groups (0, 2 and 4uM), evaluated maturation rate, cleavage rate, blastocyst formation rate and total cell number. Additionally GSH and ROS accumulation levels were measured via staining oocytes. In the results, maturation rate had not shown significant difference among the groups. However, in further development, not only the results of cleavage rate (0uM : 84.64±2.65 vs 4uM : 93.67±2.36, p<0.05) and blastocyst formation rate (0uM : 6.39± 0.90, vs 4uM : 13.67±2.32, P<0.05) were significantly increased in 4uM resveratrol treated group, and result of total cell number (0uM : 22.47±0.76 vs 2, 4uM : 30.35±1.76, 27.65±1.23, P<0.05) also shown significant difference in 2, 4uM resveratrol groups with control. GSH accumulated levels of matured oocytes in resvetrol treated groups were significantly higher than control. Meanwhile ROS levels of treated groups were significantly reduced [GSH (0uM : 142±10.49 vs 2, 4uM : 163.2±3.29, 169.7±0.94, P<0.05), ROS (0uM : 170.2±7.76 vs 2, 4uM : 118.6±7.90, 130±7.07, P<0.05)]. From these results, we conclude the treatment of resveratol improved further development of porcine embryos by regulating intracellular GSH, ROS levels during porcine IVM. Therefore, exogenous antioxidants such as a resveratol can be supportive substances for obtaining the improved quality of IVP.

α-solanine is toxic to human health by disturbing digestive and central nervous systems. However, little information has been focused on investigated with respect to α-solanine influence in mammal oocyte maturation and quality. In this study, we investigated the effects of α-solanine on oocyte maturation, quality and possible molecular mechanisms in a pig model. Porcine Cumulus-oocyte complexes (COCs) were treated with increasing concentration (0, 1, 10, 20, 50 μM) of α-solanine subjected to further in vitro maturation culture. The result showed that α-solanine significantly inhibited cumulus cells expansion and increased oocyte death rates when the concentration of α-solanine more than 10 μM. After cell cycle and cytoskeleton analysis, the results showed that α-solanine (10 μM) disturbed meiotic resumption, increased abnormal spindle formation and cortical granules (CGs) distribution rates when compared with the untreated group. α-solanine (10 μM) triggered autophagy by increasing the expression of autophagy-related genes (LC3, ATG7, LAMP2) and accumulation of LC3-specific puncta (an autophagy maker). TUNEL staining assay showed that α-solanine significantly increased apoptosis in porcine oocytes confirmed by up-regulated the levels of BAX and CAPS3 genes. Further study revealed that exposure α-solanine (10 μM) to porcine oocytes induced ROS generation, reduced mitochondrial membrane potential. In addition, our results suggested that α-solanine (10 μM) significantly increased the levels of H3K36me3 and H3K27me3 in porcine oocytes. Taken together, these data indicated that α-solanine toxic impaired oocyte maturation and quality by inhibited cumulus cells expansion, increased abnormal spindle and CGs distribution rates, triggered autophagy/apoptosis occur, accumulated ROS, decreased mitochondrial membrane potential, and changed epigenetic modifications.

Cluster-of-differentiation antigen 9 (CD9) gene expressed in the male germ line stem cells is crucial for sperm–egg fusion, and was therefore selected as candidate gene for boar semen motility and kinematic characteristics. This study was performed to investigatetheir association with semen motility and kinematic characteristics. DNA samples from 96 Duroc pigs with records of sperm motility and kinematic characteristics [Total motile spermatozoa (MOT), Curvilinear velocity(VCL), Straight-line velocity(VSL), the ratio between VSL and VCL(LIN), Amplitude of Lateral Head displacement(ALH)] were used in present study. A single nucleotide polymorphism (g.358A>T) in intron 6 was associated with MOT, VCL, VAP and ALH in Duroc population (p < 0.05). Therefore, we suggest that the porcine CD9 may be used as a molecular marker for Duroc boar semen quality, although its functional effect was not clear yet. These results will improve the understanding of the functions of the CD9 in spermatogenesis within the reproductive tracts, and will shed light on CD9 as a candidate gene in the selection of good sperm quality boars.

The structural diversity and localization of cell surface glycosphingolipids (GSLs), including gangliosides, in glycolipid-enriched microdomains (GEMs) render them ideally suited to play important roles in mediating cell recognition, adhesion, interactions, receptor function, and signaling. Gangliosides, sialic acid-containing GSLs, are most abundant in the nerve tissues. The quantity and expression pattern of gangliosides in brain change drastically throughout development and these changes are mainly regulated through stage-specific expression of glycosyltransferase genes. However, roles of gangliosides in neuronal differentiation of mesenchymal stem cells (MSCs) is unclear. We previously demonstrated for the first time that the glycosyltransferase genes during mouse embryogenesis. So, we investigated the effects of ganglioside gene in differentiation of adipose-derived MSCs (AD-MSCs). GM2 and GD3 ganglioside synthease were increased during neuronal differentiation of AD-MSCs. This study showed that the differentiation of neuronal marker was decreased on the first step of ganglioside synthase UDP-glucose ceramide glucosyltransferase(UGCG) and knock downed GM2 sythase (B4GALNT1). The result of suggested that GM2 and GD3 might be important roles in the neural differentiation of mini-pig AD-MSCs. This work was carried out with the funding of the cooperative research Program for Agriculture Science & Technology Development[Project No. PJ00999901], the Rural Development Adiministration, the KRIBB Research Initiative Program[KGM4251622].

Cryopreservation of miniature pig sperm is essential because of high demand of organ transplant in mass production. However, miniature pig sperm are vulnerable to oxidative stress more than other mammals. Erythritol is a naturally occurring sugar alcohol with powerful antioxidant property. Thus, the aim of our study is to verify if erythritol could reduce lipid peroxide and enhance viability of frozen thawed miniature pig sperm. Ejaculated semen samples were frozen with cryoprotectant subjected to erythritol treatment (0, 10, 100, 500 mM). After frozen thawed, spematozoa viability were examined using the computer assisted sperm analysis (CASA) system. The product of lipid peroxidation, malondialdehyde (MDA) were quantified using spectrophotometer with DPPH and ABTS assays as ROS scavenger markers. Our result showed that erythritol enhanced sperm viability (p<0.05), reduced lipid peroxides significantly (p<0.05), proving the concentration of 100 mM erythritol to be an effective for lowing oxidative damage. Data from our study suggest that erythritol exhibits significant lipid peroxidation scavenging characteristics which may prevent oxidative damage, enhance viability of frozen thawed sperm and thus could be a effective additive as cryoprotectant.

Due to their anatomical, physiological and genetic similarities, pig is attractive animal model in biomedical research. In the recent stem cell research era, porcine derived stem cells also gain attention due to its use for the preclinical application of human. Mesenchymal stem cells (MSCs) have been studied by many researchers over decade, and their prospect for clinical application is recognized. Although porcine derived MSCs (pMSCs) have confirmed to be differentiated into various types of cells, such as osteocyte, chondrocyte, neuronal cell, cardiomyocyte and pancreatic β cell, few report has been studied regarding hepatocyte differentiation in vitro. The present study was therefore aimed for bone marrow MSCs derived from pig femur to differentiate into hepatocyte. The cells were confirmed as MSCs by characterizing their morphology, lineage differentiation capacity and surface phenotype. They showed spindle like morphology and adipocytic, osteoblastic, and chondrocytic differentiation potentials and displayed positive expression of mesenchymal markers CD29, CD44 and CD90 while lacked the expression of hematopoietic marker CD45. Under appropriate differentiation conditions, MSCs displayed hepatocyte-like morphology depending on duration of differentiation. The differentiated MSCs into hepatocyte expressed hepatocyte-specific genes including hepatocyte nuclear factor 4 (HNF4), albumin (ALB), alpha fetoprotein (AFP), alpha-1-anti trypsin (A1AT). They also showed hepatocyte-like function, glycogen storage which is identified by PAS staining. Taken together, it concluded that the bone marrow MSCs have the potential to differentiate into hepatocyte. Further studies are needed on additional hepatocytic functional assays, such as low density lipoprotein (LDL) uptake and urea synthesis of differentiated MSC.

The migration, adhesion, and proliferation of conceptuses during pregnancy are tightly controlled processes that are mediated by various factors including cytokines, growth factors, and hormones. Among many factors, chemokines play key roles in lymphocyte trafficking, cellular proliferation, vascularization, and embryogenesis in many mammalian species. Especially, it has been shown that C-X-C chemokine ligand 12 (CXCL12) plays an important role in early pregnancy by promoting trophoblast invasion, proliferation, and differentiation through its receptor, C-X-C chemokine receptor 4 (CXCR4) in humans. However, expression and function of CXCL12 in the uterine endometrium during pregnancy have not been well studied in pigs. Thus, we determined expression of CXCL12 and its receptor, CXCR4, in the uterine endometrium during the estrous cycle and pregnancy in pigs. We obtained endometrial tissues from gilts on day (D) 12 and D15 of the estrous cycle and D12, D15, D30, D60, D90, and D114 of pregnancy, conceptus tissues from D12 and D15 of pregnancy, and chorioallantoic tissues from D30, D60, D90, and D114 of pregnancy. Real-time RT-PCR analysis showed that levels of CXCL12 and CXCR4 mRNAs changed in the uterine endometrium during pregnancy. Levels of CXCL12 and CXCR4 mRNAs on D15 of pregnancy were higher than those on D15 of the estrous cycle. After D15 of pregnancy levels of CXCL12 and CXCR4 mRNAs gradually decreased toward term of pregnancy, and CXCL12 and CXCR4 were expressed in the chorioallantoic tissues during the mid- to late pregnancy. CXCL12 and CXCR4 mRNAs were expressed in chorioallantoic tissues during mid- to late pregnancy, and RT-PCR analysis showed that CXCL12 and CXCR4 mRNAs were detectable in conceptus on D12 and D15 of pregnancy. Immunohistochemistry showed that CXCL12 proteins were localized to endometrial luminal and glandular epithelial cells during the estrous cycle and pregnancy, and to chorionic epithelial cells during mid- to late pregnancy. Abundance of CXCL12 mRNAs, but not CXCR4, in the uterine endometrium was increased by the treatment of IFNG. These results showed that CXCL12 and CXCR4 were expressed in the uterine endometrium, conceptus, and chorioallantoic tissues and IFNG increased endometrial CXCL12 expression in pigs, suggesting that CXCL12 and its receptor may play a key role in regulation of the establishment and maintenance of pregnancy by affecting the conceptus development in pigs. [supported by the Next Generation BioGreen 21 Program (#PJ01110301), Rural Development Administration]

S100As are calcium-binding proteins with two EF-hand calcium-binding motifs. In several studies, S100A proteins are described to play important roles in pro-inflammatory responses including damage-associated molecular pattern (DAMP) signaling and in the establishment of pregnancy. However, the role 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 regulation 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 other days of cycle. To investigate the effects of steroid hormones, estradiol (E2) and progesterone (P4), on expression of S100A8, S100A9, and S100A12 mRNAs, endometrial tissue explants from immature pigs were treated with steroid hormones. Levels of S100A8, S100A9, and S100A12 were increased by the treatment of P4, and the increased levels of S100A8, S100A9, and S100A12 by P4 were not inhibited by the treatment of progesterone receptor antagonist, RU486. However, levels of S100A8, S100A9, and S100A12 were decreased by treatment of MEK inhibitor, U0126. These results exhibited that S100As were expressed in the uterine endometrium during the estrous cycle in a cyclic stage-specific manner, and their expression was affected by P4. These suggest that S100As may play an important role in endometrial function during the proestrous period of the estrous cycle in pigs. [Supported by the Next Generation Biogreen 21 program (#PJ01119103), Rural Development Administration, and by Korea Research Foundation (#2015R1D1A1A01058356)]