Busulfan is the most commonly used drug for preconditioning during the transplantation of hematopoietic stem cells and male germ cells. Here, we describe side effects of high doses of busulfan in male mongrel dogs. Busulfan was intravenously administered to three groups of dogs at doses of 10, 15, and 17.5 mg/kg body weight. The total white blood cell, neutrophil, eosinophil, lymphocyte, monocyte, and platelet counts steadily reduced in a dose-dependent manner following busulfan treatment. The white blood cell, neutrophil, and monocyte counts recovered after 6 weeks of busulfan treatment, however, the eosinophil, lymphocyte, and platelet counts remained unaltered. Additionally, there was one fatality in the each of the groups that were administered 15 and 17.5 mg/kg busulfan. The gross lesions included severe hemorrhage in the stomach, intestinal tracts, mesentery and urinary bladder. Microscopic investigation revealed severe pulmonary edema and hemorrhage in the lungs, and severe multifocal to coalescing transmural hemorrhage in the intestines and urinary bladder. These results indicated that treatment with busulfan at doses higher than 15 mg/kg initiates severe bleeding in the internal organs and can have fatal results.

An experiment was carried out to assess the effect of feed selenium-lysine (Se-Lys) supplementation on milk compositions and serum biochemical indices in Saanen dairy goats in Korea. A total of twelve 36 months old Saanen lactating dairy goats (47±6.21 kg) fed the similar dry matter intake twice a day at 2% of BW (DMI) (10.9% moisture of concentrate and 19% moisture of roughage), milk yield (2.5 kg/d) and parity (2) were randomly selected and subjected for the present study, divided into two groups with six goats in each group. The goats in the control group received rice hulls (10 g/ day) only, and did not receive Se-Lys; goats in the treatment group were fed 0.06 g of Se-Lys with 10 g of rice hulls every day before feeding roughage for six weeks. The milk sample was collected every week, and its compositions were analyzed. The results of the present study showed that there is no significantly increased milk production in Se-Lys treated group goats when compared with control group goats. But, Se-Lys treatment significantly increased the milk protein content (3.98±0.16%), fat (3.72±0.27%), lactose (4.07±0.14%), total solids (12.51±0.28%) and urea (14.42±1.45 mg/dl) content as compared to the control group goats (p<0.05). The somatic cell counts (207,740±28.81 cells/ml) were significantly lower in the Se-Lys treated group than in the control group (p<0.05). Also, the results of the current study showed that supplementation of Se-Lys were significantly decreased the blood biochemical indices of IL-6 (34.34±6.04 pg/ml), TNT-α (0.56±0.22 ng/ml), MDA (5.07±1.03 ng/ml), GPx-1 (9.07±5.17 ng/ml), sCD4 (2.64±1.02 ng/ml) and sCD8 (5.08±2.08 ng/ml) level when compared with without addition of Se-Lys group dairy goats (p<0.05). On the other hand, the selenoprotein P (1,580.18±127.62 ng/ml) level was significantly higher in Se-Lys supplemented group than in the control group (p<0.05). Based on the study results, it was concluded that feed Se-Lys supplementation may improve milk yield with positively improved protein, fat, lactose, total solids, urea content, and biochemical indices without negative effects on milk production traits.

Autophagy is an intracellular degradation and recycling system. Oocyte maturation is dynamic process, in which various proteins should be synthesized and degraded. In our previous study, we reported the loci of autophagosome and dynamics of autophagic activity in porcine oocytes during In Vitro maturation. In this study, we verified loci of autophagosome in porcine follicular cumulus-oocyte complex by detection of microtubule-associated protein 1A/1B-light chain 3 (LC3) which is the reliable marker of autophagosome. Porcine ovary including various sizes of follicles was fixed within 1 hour after collection from slaughterhouse. After fixation, immunohistochemistry was conducted on sliced ovary tissue containing various sizes of follicles by using LC3 antibody. As a result, LC3 signal was clearly detected in both cumulus and oocytes of various sizes of follicles. We also found ring shaped signal which represent autophagosome near oocyte membrane. Most of the signals in oocytes were localized nearby cellular membrane while evenly dispersed in cumulus cells. Therefore, this result suggests that autophagy occurs in porcine COCs (cumulus-oocyte complexes) at follicular stage.

Tissue engineering (TE) has been developed to create functional organs and tissue by combining 3D matrix and cells in vitro. Vascularization and angiogenesis are utmost important for supply of nutrients and oxygen in tissue engineered organs. The present study was performed to isolate and characterize primary endothelial cells (EC) from aorta of alpha 1, 3-enzyme galactosyltransferase knock out (GalT KO) pig, to minimize immune rejection and analyze body immune system for future xenotransplantation studies. Isolation of primary EC from aorta were performed by incubation with dispase for 8-10 min at 37°C. Primary EC were cultured in EC growth medium on different extra cellular matrix (ECM), either collagen or gelation. Primary EC exhibits morphological characteristics and showed positive expressions of EC specific marker proteins i.e. PECAM1, KDR and VWF despite of their ECM surface; however, on collagen based surface they showed increase in mRNA level analyzed by qPCR. Primary EC cultured on collagen were sorted by flow cytometer using KDR marker and cultured as KDR positive cells and KDR negative cells, respectively. KDR positive cells showed dramatically increased in PECAM1 and VWF level as compared to KDR negative cells. Based on the above results, primary EC derived from GalT KO are successfully isolated and survived continuously in culture without becoming overgrown by fibroblast. Therefore, they can be utilize for xeno organ transfer, tissue engineering, and immune rejection study in future.

There is a growing interest in the application of primary hepatocytes for treatment of liver diseases in humans and for drug development. Several studies have focused on long-term survival and di-differentiation blocking of primary hepatocytes in an in vitro culture system. Therefore, the present study also aimed to optimize an in vitro culture system using primary rat hepatocytes. Primary rat hepatocytes from 6-week-old male Crl:CD rats were isolated using a modified two-step collagenase perfusion. Healthy 3.5 × 106 primary rat hepatocytes were seeded into a 2 dimensional (2D) culture in a 25T culture flask coated with collagen type I or into a 3D culture in a 125-ml spinner flask for 7 days. Production of plasma protein (ALB and TF), apoptosis (BAX and BCL2), and CYP (CYP3A1) related genes were compared between the 2D and 3D culture systems. The 3D culture system had an advantage over the 2D system because of the relatively high expression of ALB and low expression of BAX in the 3D system. However, the level of CYP3A1 did not improve in the 3D culture with and without the presence of a dexamethasone inducer. Therefore, 3D culture has an advantage for albumin production and primary rat hepatocyte survivability, but a low expression of CYP3A1 indicated that primary rat hepatocytes require a high–density culture for stress reduction by continuous flow.

The occurrence of mastitis in diary cattle has been caused by genetic, physiological, managemental and environment factors accounted for the highest percentage of worldwide disease in dairy cattle. The purpose of this study was to analyze the occurrences and causative bacteria of subclinical mastitis in milking cows and also examine the distribution of bacteria in milk by isolating and identifying bacteria both in whole milk and quarter milk. 31.4% of the milking cows suffered subclinical mastitis, and 9.5% had it in terms of quarter milk. According to the results of analyzing bacteria in quarter milk of which somatic cell count (SCC) was over 500 thousand, 15 kinds of bacteria were isolated, and among them, Pantoea spp. formed the biggest part as 15.8%. From whole milk, 37 kinds of bacteria were identified, and among them, Klebsiella oxytoca showed the highest identification rate as 30.1%. According to the results of bacteria analyzed from the quarter milk of entire milking cows, 52 kinds of bacteria were identified. Among them, 17 kinds of Staphylococci were isolated, and CNS (Coagulase-Negative Staphylococci) formed a large part as 44.9%. The findings of this study showed that various kinds of bacteria were isolated from cows having subclinical mastitis; therefore, when managing specifications about milking or such, dairy farm will have to take proper action like performing sanitary control or counting somatic cells regularly in order to do their best for reducing mastitis.

The objective of this study was to analyze the accuracy of estrus detection of heat detector and analysis of estrus behavior (mounting and mounted), and the evaluation of conditions required for improving reproductive efficiency in Holstein dairy cows fitted with a estrous detector. The heat detection system consists of estrous detector based on wireless sensor and an electric bulletin board displayed estrus behavior data. When cow mounting other cows, the accuracy of estrus behavior displayed an electric bulletin board were 87.5% (mounting other cows only), 100% (mounting other cows but not standing), 80.0% (mounting other cows with standing for 1∼4 seconds), 90.0% (mounting other cows but not standing for 1∼4 seconds), 80% (mounting other cows with standing for more than 5 seconds) and 90.0% (mounting other cows but not standing for more than 5 seconds). When cow mounted other cows, the accuracy of estrus behavior displayed an electric bulletin board were 100% (mounted other cows but not standing), 100% (mounted other cows with standing for 1∼4 seconds), 100% (mounted other cows but not standing for 1∼4 seconds) and 100% (mounted other cows with standing for more than 5 seconds). Circadian distribution of first observed in estrus were 59.1% (am 8∼pm 6) and 40.9% (pm 6∼am 8). Distribution for the number of estrus behavior were 40.9% (less than 3 times), 36.4% (4∼6 times) and 22.7% (more than 4 times). The conception rates relative to interval from first estrus behavior to insemination for estrus periods were 23.1% (less than 11 hours) and 55.6% (12∼20 hours).

This study aimed to improve the reproductive efficiency of dairy herds by comparison and analyzing estrous appearance rate, conception and non-conception rate according to the stage of lactation using the lactation and reproductive records of average (less than 10,000 liters milk in 305 days) and high yielding (more than 10,000 liters milk in 305 days) Holstein cows (n=102). Milk production and reproduction data were collected between January 2010 and December 2012 from Holstein cows kept in the commercial dairy farms. Average (n=32) and high yielding (n=24) Holstein cows used to analyze the relationship between milk yield and reproductive performance. Our results showed that estrous appearance rate according to the stage of lactation was 25.0% (30∼59d), 40.6% (60∼ 89d), 25% (90∼110d) and 9.4% (>111d) in average yielding cows and 16.7% (30∼59d), 20.8% (60∼89d), 12.5% (90 ∼110d) and 50.0% (>111d) in high yielding cows, respectively. Conception rate according to the stage of lactation was 87.5% (30∼59d), 61.5% (60∼ 89d), 75.0% (90∼110d) and 66.7% (>111d) in average yielding cows and 25.0% (30∼59d), 0% (60∼89d), 33.3% (90∼ 110d) and 50.0% (>111d) in high yielding cows, respectively. Days between parturition and conception was 23.7% (<149d), 0% (150∼209d) and 0% (>210 d) in average yielding cows and 69.0% (<149 d), 77.8% (150∼209d) and 38.9% (>210d) in high yielding cows, respectively. Conception rate from 110 days postpartum in high yielding cows was 41.7% (110∼150d), 50.0% (151∼180d) and 50.0% (>181d). Body condition score (BCS) in 120 days postpartum was 2.64±0.1 in average yielding cows and 2.28±0.1 in high yielding cows, respectively.

The objective of this work was to analyze the concentrations of progesterone (P4) and estrogen (E2) hormones changed during estrus synchronization in dairy heifers. Estrus synchronization was carried out with CIDRⓇ (Controlled Intravaginal Drug Release) devices. Corpus luteum (CL) was classified into three grades based on its size and palpable characteristics. The concentrations of P4 and E2 were measured by enzyme-amplified chemiluminescence. Serum P4 concentration was markedly low at the estrus stage (36 hrs after removal of CIDR) compared to other stages, while E2 concentration was kept high during estrus stage. The serum P4 concentration was highest in the CL classified into gradeⅠ. These results indicate that P4 concentration could be used as a criteria for determining recipients for artificial insemination or embryo transfer in dairy cattle.

This study was performed to analyze the causative agents of disease occurred in Korean black goat. Bovine viral diarrhea (BVD) and infectious bovine rhinotracheitis (IBR) known as main causative agents of diarrhea and abortion in cattle were founded in Korean black goats. Escherichia coli and Clostridium perfringens are the main causes underlying diarrhea in Korean black goats as well as other animals. In addition, the orf virus was found as a causative agent of contagious ecthyma that shows frequent occurrence in Korean black goats. Leptospirosis was found in Korean black goats with low occurrence rate. However, unlike the reports from many other countries, we could not detect the antibodies for Johne’s disease and Neospora caninum in Korean black goats. These results are likely to contribute to improving the productivity of raising black goats as a field of livestock farming.

Artificial insemination and embryo transfer is one of the most important factors affecting to the production of fawn from deer nuclear transfer in the field of deer farms. This study* was conducted to establish the production technology of nuclear transfered embryo in deer. For estrus synchronization or superovulation tretments in flower deer and elk, each 10 does were inserted into the vagina for 14 days with CIDR (Pfizer New Zealand Ltd., NZ) for elk and Ring-CIDR (Bioculture Co., Ltd., Korea) for flower deer, and then those inserted devices were removed. The estrus synchronization of each 6 does were induced by the intramuscular injection of PGF2α (25 mg/head) and PG600 (hCG 200IU + PMSG 400IU, Intevet, Holland). Then, the superovulation of each 4 does of flower deer and elk was induced by additional injection of FSH (200 mg/ head) twice with an interval of 24 hours , respectively. Follicular oocytes were collected from each 2 does superovulated after 48 hours since the injection of PG600 and FSH. In the meantime, the ovarian response and the number of the collected ovarian follicles were investigated with the surgical operations. As a result, the average number of the collected ovarian follicles were 8.5 and 9.0 in flower deer and elk, respectively. The ovarian follicles collected from each two does were cultured in vitro for 48 hours with m-DMEM medium, and then the cell fusion was carried out after the nuclear transfer by the antler cell. As a result, 5 out of 18 ovarian follicles collected from 2 elk does were reached on the MII stage, but there was no generation resulting from the nuclear transferred embryos by the antler cell after enucleation. In 2 flower does, 7 out of 17 ovarian follicles were reached to the MII stage, but one of them was developed to parthenogenetic embryo as well despite a case of fusion from the nuclear transferred embryo. Embryos were collected in a surgical way on the 7th day after artificial insemination, numbers of average embryos collected were 2.5 and 3.0 in each 2 flower deer and elk does superovulated, respectively. The collected two embryos were transplanted to each 2 does synchronized. As a result, a head of fawn was produced from only one elk doe, where as a head of fawn were delivered from one out of 4 elk does artificial inseminated. Given these findings, we consider that more or less of problems might have occurred in vitro culture system of ovarian follicles in the production of nuclear transfered deer embryos. In addition, the greatest reason why both the aetificial insemination and embryo transfer failed was considered attributable to stress due to anesthesia and catching.