Cryopreservation of boar semen is continually researched in reproductive technologies and genetic resource banking in breed conservation. For evaluating the boar semen quality, sperm motility (MOT) is an important parameter because the movement of spermatozoa indicates active metabolism, membrane integrity and fertilizing capacity. Various researches have been trying to improve the quality of semen post-thawed in boar. Recently, polymorphism (g.358A>T) of cluster-of-differentiation antigen 9 (CD9) gene reported to be significant association with MOT. Also, CD9 gene was expressed in the male germ line stem cells is crucial for sperm-egg fusion, and was therefore selected as candidate gene for boar semen. This study was conducted to evaluate the pig SNP (g.358A>T) of CD9 gene as a positional controlling for semen parameters of post-thawed boar semen. To results, the g.358A>T SNP of the CD9 gene was significantly associated with the traits such as MOT, curve linear velocity, straight line velocity, average path velocity and amplitude of lateral head displacement. Particularly, the g.358A>T SNP significantly has the highest association with MOT and animals with AA genotype (p<0.001). Therefore, we suggest that the g.358A>T in the intron 6 region of the porcine CD9 may be used as a molecular marker for Duroc boar Post-thawed semen quality, although its functional effect was not defined yet.

Obesity, most often defined as a body mass index (BMI) of ≥30 kg/m2 caused by an imbalance between energyintake and expenditure. It is widely recognized as the largest and fastest growing public health problem in the developed anddeveloping world. Obesity results by an increase in adipose tissue mass, resulting from the multiplication of fat cells fol-lowed by adipogenesis and increased deposition of intracellular cytoplasmic triglycerides. In this study, we investigated theanti-adipogenic properties of the extracts (KEs) obtained from Kimchi (Korean traditional fermented vegetable) fermentedusing Kimchi refrigerator under different storage conditions. The intracellular lipid droplets in the differentiated adipocyteswere also measured. We observed that the KEs significantly lower the contents of cellular triglyceride and formation ofintracellular lipid droplets. These results suggested that KEs have anti-adipogenic effects and may contribute to food indus-tries in developing functional.

A Duplex polymerase chain reaction (PCR) was developed for the simultaneous detection and differentiation among Nosema spp. and Vairimorpha spp. from Lepidoptera insects. Two sets of primers were selected from different genomic sequences to specifically amplify an 831 bp amplicon within the SSU rRNA gene, specific for both Nosema spp. and Vairimorpha spp. (MSSR primer); a 542 bp amplicon within the SSU rRNA gene, specific for Vairimorpha spp. (VSSU primer). Using the primers in conjunction (duplex PCR) it was possible to detect Nosema spp. and Vairimorpha spp. to differentiate between them. The sensitivity of this PCR assay was approximately 10 spores per milliliter. It is proposed that the duplex PCR is a sensitive, specific and rapid tool that can serve as a useful differential diagnostic tool for detecting Nosema spp. and Vairimorpha spp. in Lepidoptera insect.

본 연구에서는 식육원료 4종(소, 돼지, 말 및 닭)을 각 각 판별하기 위하여 real-time PCR을 이용한 판별법을 개발하였다. 종판별을 위한 유전자로는 미토콘드리아 유전자 중 12S rRNA와 16S rRNA 부분을 대상으로 하였으며, 특이 프로브의 Reporter dye는 FAM, Quencher dye는 TAMRA로 설계하였다. 개발된 프라이머 및 프로브 세트 로 유사종에 대한 비 특이적 signal의 생성 유무를 관찰하기 위하여 총 10종을 대상으로 특이성을 확인한 결과, 비 특이적 signal은 확인되지 않았다. 식육원료에 대하여 realtime PCR을 통한 식육 판별 시 식육 혼합 방법에 따른 CT값의 유의적인 차이는 없었다. 의도적 혼합 및 비의도적 혼입을 판별하기 위한 정량법 개발에서는 의도적 혼합의 경우 100% 식육과의 CT 값 차이가 4 cycle 이내이고, 비의도적 혼입일 경우 100% 식육과의 CT 값 차이가 6 cycle 이상이었다. 따라서 본 연구를 통하여 개발한 식육 원료의 의도적, 비의도적 혼입 판별법은 불량식품 유통 근절 및 소비자 인권보호에 크게 기여할 것으로 기대된다.

Phasianus colchicus is not only a beautiful bird but also a great value in science and under the threat of endanger. Hence, the aim of this study was to isolate the pheasant male germ cells (mGCs) and then induce them into elongated sperm-like cells in vitro. The mGCs were purified and enriched by a two-step plating method based on the different adherence velocities of mGCs and somatic cells. The percentage of the c-kit positive cells and c-kit negative cells examined by flow cytometry analysis (FCA) was 92.87% and 2.57%, respectively. Subsequently, the mGCs were induced for 48h in DMEM/F12 medium supplemented factors such as retinol acid, testosterone and bovine FSH, followed by 5 weeks in culture. We found that some elongated sperm-like cells appeared initially in vitro under inducement of stimulated factors. The elongated sperm-like cells showed in the expression of changed morphology and post-transcriptional marker such as spermatid associated (SPERT), spermatid perinuclear RNA binding protein (STRBP), round spermatid basic protein 1 (RSBN1) and SPER1L. Moreover, in DNA content identified assay, induced cells showed that the 1C DNA population markedly increased in differentiated group but it was not change in undifferentiated group. Successful in vitro differentiation of pheasant testicular germline cells into spermatids appears to offer extremely attractive potential for the conservation of endangered birds and treatment of male infertility.

Skin is the outermost organ and acts as a barrier between the organism and environment. Skin protects the organism from environmental insults, such as chemicals, pathogens, and UV light. Much of the protective function of skin is dependent on the epidermis, a multi-layered epithelium that is composed of various cell types such as keratinocytes and melanocytes. Keratinocytes produce protective components through a sophisticated differentiation process. Disturbance of keratinocyte differentiation is related to several skin diseases such as psoriasis and atopic dermatitis. In this study, we prepared extract of combined medicinal plants (ECMP) consisting of Taraxacum platycarpum H. Dahlstedt, Heartleaf Houttuynia, Glycyrrhiza uralensis Fischer, and root bark of Ulmus davidiana. We demonstrated that ECMP enhanced keratinocyte differentiation and barrier functionality using an in vitro cell culture system and in vivo animal test. Treatment of cultured keratinocytes with ECMP resulted in induction of keratinocyte differentiation, as evidenced by increased differentiation markers such as involucrin, loricrin, and filaggrin. In line with these results, ECMP decreased proliferation of keratinocytes cultured in vitro. ECMP applied topically to tape-stripped mouse skins accelerated reduction of transepidermal water loss (TEWL), indicating fast recovery of barrier function. Immunohistochemistry showed that ECMP increased the filaggrin level in tape-stripped mouse skins. These results suggest that ECMP may be applicable for keratinocyte differentiation-related skin diseases.

A tremendous increase in the human population has put poultry industry under an increased pressure to meet steep increase in the demand. Poultry is contributing 25% of the total world’s meat production and lesser cost of investment per bird makes it more suitable for the further breeding programmes. Major poultry diseases frequently lead to cardiac damage and cause huge economic losses to poultry industry due to mortality. The in vitro embryonic stem cell (ESC) technology has a futuristic approach for homogeneous populace of differentiated cells, for their further transplantations. During in vitro conditions the differentiated cell populace can be used in grafting and transplantation processes to regenerate damaged tissues. Therefore, the current study targeted the use of spermatogonial stem cells (SSCs) in the poultry production system through cardiac regeneration. The current study will also open new boulevard for the similar kind of research in other livestock species for the management of heart diseases.

A poor prognosis of oral squamous cell carcinoma (SCC) is partly due to the invasiveness and metastasis of the tumor. One of key elements in tumor invasion and metastasis in the degradation of extracellular matrix is tissue inhibitors of metalloproteinases (TIMPs). This study was performed to determine the expression of TIMP-1 and TIMP-2 of oral SCCs with regard to the histologic invasiveness and differentiation in 5 normal oral mucosa and 36 oral SCCs. The histologic invasiveness of oral SCCs were classified into 4 grades. The differentiation of oral SCCs was divided into 3 grades. The StreptAvidin-Biotin immunohistochemical process, using TIMP-1 and TIMP-2 monoclonal antibodies, was performed to determine the expression of TIMP-1 and TIMP-2. The expression of TIMP-1 was positive in 5 of 17 oral SCCs with weak invasiveness and was positive in 8 of 19 oral SCCs with strong invasiveness. The TIMP-1 expression did not increase significantly with respect to the invasiveness of oral SCCs (P>0.05). The expression of TIMP-2 was strongly positive in 5 out of 17 SCCs with weak invasiveness and was strongly positive in 15 of 19 SCCs with strong invasiveness. The TIMP-2 expression increased significantly with respect to the invasiveness of oral SCCs; the stronger the expression, the stronger the invasiveness (P<0.05). The expression of TIMP-1 and TIMP-2 did not increase significantly with respect to the histologic differentiation. We concluded that with respect to the invasiveness, the TIMP-2 expression increases significantly in oral SCCs but the TIMP-1 expression does not; and that with respect to the histologic differentiation, their expressions do not increase significantly. These results suggested that TIMP-2 can be used as a tool to evaluate the invasiveness of oral SCCs.

During bone remodeling, there is requirement of differentiation of osteoblastic cells. Previously, we identified proteins differentially expressed in soft tissue during bone healing. Of these proteins, we focused the effect of LTF on differentiation of osteoblast. In order to analyze the osteogenic ability of LTF, we treated conditioned media collected from human LTF-stably transfected HEK293T cells into osteoblastic MC3T3-E1. The results showed that the activity and expression of alkaline phosphatase were increased in MC3T3-E1 cells treated with conditioned media containing LTF in dose- and time-dependent manner. At the same time, we observed the significant increase of the expression of osteoblastic genes, such as ALP, BSP, COL1A1, and OCN, and along with matrix mineralization genes, such as DMP1 and DMP2, in LTF conditioned media-treated groups. Moreover, the result of treating recombinant human LTF directly into osteoblastic MC3T3-E1 showed the same pattern of treating conditioned media containing LTF. Our study demonstrated that LTF constitutively enhances osteoblastic differentiation via induction of osteoblastic genes and activation of matrix mineralization in MC3T3-E1 cells.

Fucoidan has been extensively studied as medicinal materials due to its biological activities including osteoblastic differentiation effect. However, osteoblastic effect by fucoidan is unknown in alveolar bone marrow derived mesenchymal stem cells (ABM-MSCs). The present study was undertaken to evaluate the effect of fucoidan on Osteoblastic differentiation in ABM-MSCs and explore its mechanism. Cell proliferation was analyzed by crystal violet staining. Osteoblast differentiation was determined by alkaline phosphatase activity, calcium accumulation assay and gene expression of osteoblast markers. We found that fucoidan induced cell proliferation of ABM-MSCs. Furthermore, fucoidan increased the ALP activity, calcium accumulation, and osteoblast specific genes such as Runx2, type I collagen alpha 1. Moreover, fucoidan induces the expression of asporin and bone morphogenic protein (BMP)-2 and asporin. Based on these results, these finding indicate that fucoidan induces osteoblast differentiation in ABM-MSCs and partially enhanced the mRNA expression of BMP-2 and asporin.

Acetylcholinesterase (AChE) plays a pivotal role in the synaptic transmission in the cholinergic nervous system of most animals, including insects. Most insects possess two AChEs (i.e., AChE1 vs. AChE2), which are encoded by two paralogous loci originated from the duplication that occurred before the radiation of insects. The phylogenetic analysis suggested that the last common ancestor of ace1 and ace2 shared its origin with those of Platyhelminthes. In addition, ace1 lineage showed a lower evolutionary rate (d and dN/dS ratio) compared to ace2 lineage, suggesting that the ace1 lineage has maintained relatively more essential functions following duplication. Furthermore, structural modeling of AChEs revealed that consistent structural alteration in their active-site gorge topology was caused by amino acid substitution, likely leads to functional differentiation between two AChEs. The functional transition of ace in some hymenopteran insects appears to have occurred by only a few mutations resulting in dramatic alteration of AChE activity. Taken together, our findings provide basic information on when the ace duplication occurred and what structural features have been associated with the differentiation of two AChEs during evolution.