High-entropy alloys (HEAs) are attracting attention because of their excellent properties and functions; however, they are relatively expensive compared with commercial alloys. Therefore, various efforts have been made to reduce the cost of raw materials. In this study, MIM is attempted using coarse equiatomic CoCrFeMnNi HEA powders. The mixing ratio (powder:binder) for HEA feedstock preparation is explored using torque rheometer. The block-shaped green parts are fabricated through a metal injection molding process using feedstock. The thermal debinding conditions are explored by thermogravimetric analysis, and solvent and thermal debinding are performed. It is densified under various sintering conditions considering the melting point of the HEA. The final product, which contains a small amount of non-FCC phase, is manufactured at a sintering temperature of 1250oC.

The 17th APEC Future Education Forum (AFEF) and the 19th International ALCoB Conference were held from 28 September (Tuesday) to 1 October (Friday) in a virtual manner. The annual forum and conference serve as an invaluable opportunity to discuss future directions of education and human resources development in the Asia-Pacific region. Since 2005, the forum and conference have served their role as the largest and longest thematic meeting in the APEC Human Resources Development Working Group (HRDWG). The 2021 forum and conference have gathered 780 participants from 20 APEC member economies, guest economies and international institutions. Under the theme “Strengthening lifelong competencies and skills development for individuals’ career, education, training, and life cycles”, the forum and conference reflected the current educational agenda from 21 APEC member economies and other international organizations such as G20, UNESCO. As a way forward, the forum and conference speakers recommend APEC to 1) consider individual and industrial needs for curriculum development, 2) expand lifelong learning policies and practices at both domestic and international level and 3) develop a public-private partnership to prepare its future talents with adequate competencies. This review explains the 17th AFEF and the 19th International ALCoB Conference background, summary and outcomes. The review also briefs on collective actions member economies can take as future steps to continue the forum and conference discussions.

Oocyte is the central factor in the bi-directional communication axis in the ovarian follicles. It controls the cumulus or granulosa cells to perform functions which are beneficial for its own development via secreting paracrine growth factors, including GDF9 and BMP15. The aim of this study was to investigate whether the recombinant GDF9 and BMP15 are able to promote meiotic resumption and cumulus expansion of canine COCs during IVM, as well as to demonstrate the actions of GDF9 and BMP15 in regulating the expression of connexin transcripts in the ovarian granulosa cells. As results, GDF9 and BMP15 significantly improved the meiotic resumption rate and cumulus expansion by activating ERK1/2 signaling. Treatments with GDF9 significantly improved the expression of CyclinB1 but inhibited the expression of Cx43 transcripts. In addition, cumulus expansion genes (MAPK1, Ptgs2, Tnfaip6 and Ptx3) were differentially improved by GDF9 and BMP15. In the ovarian granulosa cells, GDF9 suppressed the expression of Cx43 transcripts by binding ALK4/5/7 receptors and activation Smad2/3 signaling, whereas, BMP15 stimulated the expression of Cx43 transcripts by binding ALK2/3/6 receptors and activating Smad1/5/8 signaling. In conclusion, by regulating functions of granulosa/cumulus cells, oocyte has the potential to enhance the growth and maturation of itself.

Somatic cell nuclear transfer (SCNT) is the technique which generates embryos by transferring diploid nucleus into an enucleated oocyte, it has produced specific animals successfully in a variety of species. However, the developmental capacity of SCNT embryos is still relatively lower than that of embryos produced in vivo. Oocyte is a kind of lipid rich cells, its quality limits the efficiency of embryo production. L-carnitine is a co-enzyme facilitating the transportation of long chain fatty acids across the inner mitochondria membrane where fatty acids are used for generating adenosine triphosphate (ATP) via beta-oxidation. It also has antioxidant actions which may protect mitochondrial membranes and DNA against damage induced by reactive oxygen species (ROS). Whether L-carnitine is functional in bovine SCNT embryos are unknown. Therefore, the objective of this study was to examine the effects of L-carnitine on oocyte maturation and developmental competence of subsequent SCNT embryos. L-carnitine was supplemented during IVM, then intracellular ROS and GSH levels, mitochondrial activity, gene expression of COCs were analyzed at the end of IVM. SCNT embryos were produced subsequently, apoptosis detection and gene expression evaluation were performed in blastocysts. In the results, treatments with 1.5 mM and 3 mM L-carnitine significantly improved maturation rates (P<0.05). Treatments with 3 mM L-carnitine effectively induced improvement in nuclear maturation, intracellular GSH levels and mitochondrial activity, as well as a reduction in intracellular ROS levels (P<0.05). mRNA levels of CPT1A, ACAA1, ACAA2, AREG, EREG, SOD1, GPX4, GLUT1 and CDC2 transcripts were effectively up-regulated by 3 mM L-carnitine treatments (P < 0.05). Similarly, 3mM L-carnitine induced an increase in blastocyst developmental rates and an improvement in blastocyst quality (P<0.05). Our study indicates that L-carnitine treatment during IVM improves oocyte nuclear maturation and subsequent SCNT embryo development.

The mesenchymal stem cells (MSC) has been investigated as a source of stem cell therapy to replace and treat damaged cells. Human endometrial epithelial and stromal cells was isolated from hysterectomy tissue and the direct evidence of stem/progenitor cells in the human endometrium was identified. Endometrium derived stem cells (EnMSCs) are known to have a high proliferative ability, genetic stability, lack of tumorigenicity and low immnunogenicity during long-term cultivation. Here, we aimed to identify MSC in canine endometrium and characterize its potential to differentiate into decidua cells. EnMSCs were isolated from thrown-away spayed uterus of adult canine depending on their estrus cycle, and identified by flow cytometry, immunocytochemistry and flow cytometry with MSC specific markers. We then characterized the ability of EnMSCs by the doubling-time analysis, colony-forming units and MSC differentiation assays. Isolated EnMSCs expressed stem cell specific genes (Sox2, Oct4, Nanog, MCAM, Endoglin, Susd2 and IGTB) and MSC surface markers (CD90, CD44 and CD117). EnMSCs are also differentiated into adipogenic, osteogenic and chondrogenic cells morphologically under modified conditions with the expression of lineage specific genetic markers. EnMSCs showed higher proliferation ability than canine amniotic fluid derived MSCs which were used as a positive control. EnMSCs were cultured at low density (10, 20, cells/cm2) and initiated to form small colonies of loosely-arranged cells and gradually formed large colonies of densely-packed cells which underwent self-renewal with high proliferative potential which is similar to the clonogenicity feature of human endometrium-derived stem cells. EnMSCs were then induced to differentiate into decidua cells with 0.5 mM dbcAMP. After 14 days, EnMSCs changed their morphology into the elongated and rounded shape. The induced decidual cells expressed PRL and IGFBP1 which are typically expressed in decidua cells. In conclusion, we successfully isolated and characterized MSC in the canine endometrium which differentiated into decidua cells. These results showed that endometrium may be a promising source of stem cells, and furthermore raise the possibility of canine EnMSCs as a novel hypothetical decidualisation model of infertility associated with decidualisation insufficiency and implantation failure.

Oocyte is the central factor in the bi-directional communication axis in the ovarian follicles. It controls the cumulus or granulosa cells to perform functions which are beneficial for its own development via secreting paracrine growth factors, including GDF9 and BMP15. The aim of this study was to investigate whether the recombinant GDF9 and BMP15 are able to promote meiotic resumption and cumulus expansion of canine COCs during IVM, as well as to demonstrate the actions of GDF9 and BMP15 in regulating the expression of connexin transcripts in the ovarian granulosa cells. As results, GDF9 and BMP15 significantly improved the meiotic resumption rate and cumulus expansion by activating ERK1/2 signaling. Treatments with GDF9 significantly improved the expression of CyclinB1 but inhibited the expression of Cx43 transcripts. In addition, cumulus expansion genes (MAPK1, Ptgs2, Tnfaip6 and Ptx3) were differentially improved by GDF9 and BMP15. In the ovarian granulosa cells, GDF9 suppressed the expression of Cx43 transcripts by binding ALK4/5/7 receptors and activation Smad2/3 signaling, whereas, BMP15 stimulated the expression of Cx43 transcripts by binding ALK2/3/6 receptors and activating Smad1/5/8 signaling. In conclusion, by regulating functions of granulosa/cumulus cells, oocyte has the potential to enhance the growth and maturation of itself.

Human interferon alpha 2b (hIFNα-2b) is an important immune regulator widely used in clinic, for the treatment of chronic hepatitis, hairy cell leukemia, chronic myelogenous leukemia and multiple myeloma, etc. The clinically used hIFNα-2b is generally produced by E. Coli, which lacks the post-translational O-glycosylation of naturally synthesized protein, and has a short serum half-life. In this study, we report the successful generation of transgenic chickens that produce hIFNα-2b in the egg white using a feline immunodeficiency virus (FIV)-based lentiviral vector. In preliminary in vitro study, the hIFNα-2b gene under the control of CMV promoter expressed as much as 2,650 ng/㎖ in CEF-LNC-hIFNα-2bW cell. A FIV vector packaged with vesicular stomatitis virus G glycoprotein (VSV-G) was injected underneath the blastoderm of freshly laid chicken eggs (stage X) to produce a hIFNα -2b transgenic chicken. Out of 187 injected eggs, 55 chicks were hatched after 21 days of incubation, and 27 of the G0 hatched chicks expressed the vector-encoded hIFNα-2b gene. The expression of recombinant hIFNα-2b in transgenic chickens constitutes an important step towards low-cost and full biological activity production of this protein drug in bioreactor. This work was supported by the Bio-industry Technology Development Program, Ministry of Agriculture, Food and Rural Affairs, Republic of Korea, and by a grant from the Next-Generation BioGreen 21 Program (No. PJ011178), Rural Development Administration, Republic of Korea.

In the present study, using a MoMLV-based retrovirus vector, we successfully generated a new transgenic chicken line expressing high levels of hEPO. A replication-defective Moloney murine leukemia virus (MoMLV)-based vectors packaged with vesicular stomatitis virus G glycoprotein (VSV-G) was injected beneath the blastoderm of non-incubated chicken embryos (stage X). One rooster was mated to wild-type hens to produce 748 G1 progeny. PCR analysis of blood samples from these progeny revealed that there were seven G1 transgenic offspring, corresponding to a 0.9% germline transmission rate. Subsequently, Southern blot analysis of the genomic DNA from three G1 transgenic chickens was carried out to verify the stable genomic integration and copy number of the transgene in the genome. Quantitative analyses of the blood samples taken from G1 transgenic chickens resulted in 4,150 ～ 10,823 IU/㎖ (34.6 ～ 90.2 ㎍/㎖) of hEPO in the blood. The biological activity of the recombinant hEPO in transgenic chicken serum was comparable to its commercially available counterpart. Red blood cell numbers were more than three-fold higher in the transgenic chickens compared to the non-transgenic chickens. Successful germline transmission of the transgene was also confirmed in G2 transgenic chicks produced from crossing G1 transgenic roosters with non-transgenic hens. We confirmed that 13 transgenic chicks of 45 G2 progeny, corresponding to a 28.9% germline transmission rate. These results will help establish a useful transgenic chicken model system for studies of embryonic development and for efficient production of transgenic chickens as bioreactors. This work was supported by the Bio-industry Technology Development Program, Ministry of Agriculture, Food and Rural Affairs, Republic of Korea, and by a grant from the Next-Generation BioGreen 21 Program (No. PJ011178), Rural Development Administration, Republic of Korea.

Somatic cell nuclear transfer (SCNT) has been considered for preserving genetically valuable or endangered animals. Sapsaree is a Natianal Monument in Korea to maintian a pure pedigree. The aim of this study was to produce azoospermia Sapsaree using SCNT and identify normal reproductive ability of cloned azoospermia Sapsaree. Ear skin biopsy was performed on a thirteen-year-old azoospermia Sapsaree and ear skin fibroblasts were isolated for SCNT as donor cells. The fibroblasts were injected into enucleated in vivo matured oocytes, the couplets were electrical fused by two pulse of direct current (55 V for 15 μs) using titanium and platinum fusion needle and activated by calcium ionophore. Cloned embryos were surgically transferred into oviducts of natuarally estrus cycle synchronized recipient dogs. The fusion rate of platinum needle was 70%, which was higher than those of titanium needle (64.1%). Developmental rate to the 8 cells and 10 cell stages was higher in platinum needle group (24% and 16%, respectively) than those of platinum needle group (14.8% and 3.1%, respectively). Total 35 SCNT embryos were transferred into oviducts of 3 recipient dogs and one recipient finally delivered a puppy by caesarean section. As results, this study demonstrated that platinum fusion needle could be successfully make the reconstructed embryos and improve the efficiency of canine SCNT. Cloning azoospermia Sapsaree may contribute to conserve genetically valuable and unique pedigree. And further study should be confirm whether cloned live dog is azoospermia.

Prolonged communication between oocytes and the surrounding somatic cells is one of the unique reproductive physiology in canine. Paracrine Kit ligand (KITL) signaling is a well-known communication between granulosa cells and the oocyte. KITL is a cytokine growth factor secreted by granulosa cells that signals via the c-kit receptor expressed by oocytes. Paracrine factors, including growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15), exert their effects by binding with the kinase receptors expressed on the granulosa cells. However, the regulations of GDF9 and BMP15 in the canine KITL expression are currently poorly understood. Therefore, we investigated the effects of GDF9 and BMP15 on the expression of KITL in canine ovarian granulosa cells in vitro. In Annexin V assay recombinant GDF9 and BMP15 did not induce apoptosis in the cultured ovarian granulosa cells. When treated, FSH significantly increased KITL expression, and hCG suppressed its expression. When both FSH and hCG were treated, the expression of KITL was affected by GDF9 and BMP15 in dose and time dependent manner in the luteal granulosa cells. GDF9 (10 ng/mL) significantly decreased KITL expression after12 h. BMP15 (10 ng/mL) significantly also decreased KITL expression after 24 h. Western blot and immunochemistry results indicate that GDF9 activated Smad2/3. After blocking ALK 4/5/7 receptors by SB, GDF9 failed to activate Smad2/3, also BMP15 did not activate Smad1/5/8 after blocking ALK 2/3/6 receptors by DM. So GDF9 exerts its effects via using ALK 4/5/7 receptors to activate SMAD2/3 signaling, and BMP15 binds ALK 2/3/6 receptors to activate SMAD1/5/8 signaling. The expression of KITL was not changed by SB or DM treatment. However, the effect of GDF9 and BMP15, which decreased the expression of KITL, was suppressed by SB or DM treatment. In conclusion, this study provides the first evidence that recombinant GDF9 and BMP15 decrease KITL expression in canine ovarian granulosa cells.