• Asian-Australasian Journal of Animal Sciences
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• 제20권12호
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• pp.1820-1826
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• 2007

In this study, we conducted various experiments in order to develop enhanced cultural conditions for in vitro-produced porcine embryos. All embryos were produced by in vitro maturation (IVM) and fertilization (IVF) of immature oocytes from abattoir-derived ovaries. In experiment 1, we cultured IVF embryos in 4 different groups, namely, 0% bovine serum albumin (BSA), 3% BSA, 0.05% Polyvinyl alcohol (PVA), and 0.5% Polyvinylpyrrolidone (PVP) added to the basal fluid cultural medium, Porcine zygote medium 3 (PZM-3). The rates of embryo development were higher in the group where the PZM-3 media had been supplemented with 3% BSA than the other groups. While not statistically significant, the percent of blastocysts and hatched blastocytes were 6.9% and 25.0% in the 3% BSA group vs. 1.2-6.4% and 0-16.7% in the other groups, respectively. In experiment 2, we added 10% fetal bovine serum (FBS) to PZM-3 on day 0 of culture and observed the development rate of blastocysts per day of culture from days 0 to 5. The development rate of blastocysts was higher at 15.6% on day 4 than on any other day, and was significantly higher than on day 0 or day 1 (p<0.05). The development rate of hatched blastocysts was 26.7% on day 4, and was higher than on any other day. In experiment 3, we cultured IVF embryos with different fluid culture media, grouped as 1) PZM-3+0.3% BSA (day0-day7); 2) PZM-3+0.3% BSA${\rightarrow}$day-4) PZM-3+10% FBS; 3) PZM-3+0.3% BSA${\rightarrow}$PZM-3+0.3% BSA+(day-4) FBS 10%; and 4) PZM-3+0.3% BSA+10% FBS (day0-day7). The development rates of blastocysts and hatched blastocysts were 21.5% and 53.1% in group 3, respectively, which was significantly higher than group 4 with respect to blastocyst development (5.2%, p<0.05) but not hatched blastocysts (14.3%). The total cell number (TCN) of blastocysts in group 3 was higher at $37.8{\pm}16.1$ than the other groups at $16.8{\pm}4.4$ - $30.1{\pm}10.9$; however, this was not significantly different. The results of this study showed that PZM-3 containing 0.3% BSA and supplemented with FBS during the later stage of culture on day 4 resulted in better TCNs and an increased rate of hatched blastocysts.

• 한국수정란이식학회지
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• 제19권3호
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• pp.191-199
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• 2004

본 연구는 돼지 태아 섬유아세포유래 공여세포를 미세주입에 의해 주입 후 재 조합한 핵 이식 배에 대한 배양액, 세포주기의 동기화, 배양시간 및 난자의 활성화에 따른 융합율과 체외발생율에 대해 조사하였다. 핵 이식 배를 NCSU-23, TL Hepes 및 TZM-3 배양액으로 1시간 및 8시간 배양하였을 때 배반포로의 분할율은 각각 15.6%, 14.0%, 15.0% 및 13.9%, 10.5%, 13.3%로서 배양액 및 시간에 따른 분할율의 유의적인 차이는 없었다. 공여핵원용 세포를 0, 8, 15시간 배양했을 때 G2/M기로의 체외발달율은 12.0%, 18.0%, 48.0%였다(p<0.01). 공여핵원용 세포를 12-24시간 배양했을 때 G2/M기로의 체외발달율은 유의한 증가를 나타내지 않았다. 공여핵원용 세포를 10% FBS + NCSU-23 배양액으로 1-2, 6-8, 12-14일간 배양 후 핵 이식한 배의 융합율은 각각 60.0%, 73.3%, 62.5%였으며, 분할율은 각각 36.0%, 56.7%, 50.0%였다. 0.5% FBS + NCSU-23, 0.5% + TL-Heaps 및 0.5% + TZM-3 배양액으로 5% $O_2$조건 하에서 배양하였을 때 핵 이식배의 $\geq$2 cell 및 배반포로의 발생율은 각각 12.5$\pm$1.6%, 11.1$\pm$1.8%, 11.7$\pm$1.0%였으며, 10% $O_2$조건 하에서 배양하였을 때 핵 이식배의 $\geq$2 cell 및 배반포로의 발생율은 각각 10.5$\pm$1.5%, 9.8$\pm$1.4%, 10.0$\pm$0.8%였다 배양액과 $O_2$ 조건에 따른 유의한 발생율에 차이는 인정되지 않았다.

• Asian-Australasian Journal of Animal Sciences
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• 제18권1호
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• pp.107-112
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• 2005

A high environmental temperature affects the economic performance of pigs. Heat shock protein 70 (HSP70) has been reported to participate importantly in thermotolerance. This study aims to produce transgenic pigs overexpressing porcine HSP70.2, the highly inducible one of HSP70 members, and to prove the cellular thermotolerance in the primary fibroblasts from the transgenics. A recombinant plasmid in which the sequence that encodes the porcine HSP70.2 gene is fused to green fluorescence protein (GFP) was constructed under the control of cytomegalovirus (CMV) enhancer and promoter. Two transgenic pigs were produced by microinjecting pCMV-HSP70-GFP DNA into the pronucleus of fertilized eggs. Immunoblot assay revealed the varied overexpression level (6.4% and 1.4%) of HSP70-GFP in transgenic pigs. After heating at $45^{\circ}C$ for 3 h, the survival rate (78.1%) of the primary fibroblast cells from the highly expressing transgenic pig exceeded that from the non-transgenic pig (62.9%). This result showed that primary fibroblasts overexpressing HSP70-GFP confer cell thermotolerance. We suggest that transgenic pigs overexpressing HSP70 might improve their thermotolerance in summer and therefore reduce the economic loss in animal production.

• 한국임상수의학회지
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• 제9권1호
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• pp.333-366
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• 1992

Safety of recombinant porcine somatotropin administration on pig was studied using 32 Landrace x Yorkshire crossbred pigs. The starting body weight ranged from 55.5kg to 65.3kg. Eight pigs were allotted to each low dose group of sustained releasing rPST(SL), high dose group of sustained releasing rPST(SH), daily injection group of rPST(DI), and control group(C). Pigs in SL group and SH group were injected subcutaneously twice in 3 week-interval with 1000$\mu\textrm{g}$ and 2000$\mu\textrm{g}$ of sustained releasing rPST per kg body weight, respectively. Pigs in DI group were injected intramuscularly with 100$\mu\textrm{g}$ of rPST everyday for 6 weeks. Blood was collected from anterior vena cava just before the first treatment, and at four weeks and six weeks of experiment. Hematological parameters and blood chemical parameters indicating liver function, kidney function, electrolyte metabolism, mineral metabolism and lipid metabolism were determined. Necropsy and urinalysis were performed after final blood collection. The results were summarized as follows, and it is concluded that rPST administration does not affect pig health negatively. 1. rPST administration did not affect kidney function as manisfested by BUN, creatinine and urinalysis. 2. rPST administration did not affect liver function as manisfested by total protein, albumin, serum AST activity serum ALT activity serum ALP activity, serum LDH activity, serum GGT activity and serum SDH activity. 3. rPST administration did not affect skeletal muscle, cardiac muscle and brain as manifasted by serum AST activity and serum LDH activity. 4. rPST administration increased blood glucose level within normal range. 5. rPST administration did not affect lipid metabolism as manisfested by triglyceride, cholesterol, and phospholipid concentrati on. 6. rPst administration dia not affect mineral metabolism as manisfested by calcium, phosphorus, magnesium and iron concentration. 7. rPST administration did not affect electrolyte metabolism as manisfested by Na, K, chloride concentration. 8. rPST administration did not affect erythrocyte count, leukocyte count, thrombocyte count, and plasma fibrinogen level.

• Asian-Australasian Journal of Animal Sciences
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• 제20권9호
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• pp.1349-1353
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• 2007

Calcineurin is a calmodulin dependent protein that functions as a regulator of muscle cell growth and function. Agents capable of interacting with calcineurin could have important applications in muscle disease treatment as well as in the improvement of livestock production. Calsarcins comprise a family of muscle-specific calcineurin binding proteins which play an important role in modulating the function of calcineurin in muscle cells. Recently, we described the first two members of the calsarcin family (calsarcin-1 and calsarcin-2) in the pig. Here, we characterized the third member of the calsarcin family, calsarcin-3, which is also expressed specifically in skeletal muscle. However, unlike calsarcin-1 and calsarcin-2, the calsarcin-3 mRNA expression in skeletal muscle kept rising throughout the prenatal and postnatal development periods. In addition, radiation hybrid mapping indicated that porcine calsarcin-3 mapped to the distal end of the q arm of pig chromosome 2 (SSC2). A C/T single nucleotide polymorphism site in exon 5 was genotyped using the denaturing high performance liquid chromatography (DHPLC) method and the allele frequencies at this locus were significantly different among breeds.

• 한국동물위생학회지
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• 제30권3호
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• pp.353-362
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• 2007

A total of 1,354 samples was collected from bovine and porcine carcass from January 2005 to December 2006 in a slaughter house. Twenty five strains(1.8%) of Listeria monocytogenes were isolated from 1,354 samples using selective media. Ten(1.4%) L monocytogenes were isolated from the 677 of bovine carcasses, and 15(2.2%) were isolated from the 677 of porcine carcasses. Among 15 L mono-cytogenes from porcine, 11 siolates were serovars 1/2c, followed by 1/2b (3 strains, 20.0%) and 1/2a(1 strain) Out of 10 bovine samples, positive cases in 1/2a were 9 strains (90.0%), 1/2b were 1 strains(10.0%). PCR primers were selected to amplify a 520-base pair(bp) DNA fragment from the listeolysin O gene (hlyA) of L mono-cytogenes. A 520-bp product was detected in PCR with DNA from L monocytogenes, but not from the other Listeria species tested. A total of 25 L monocytogenes strains were analysed by PFGE after digestion with Apa I. PFGE analysis of genomic DNA showed the $14{\sim}18$ fragments ranging in size from 30 to 550 kb, resulting in 14 patterns.

• 한국수정란이식학회지
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• 제21권3호
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• pp.169-175
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• 2006

본 연구에서 돼지 난포란에서 채취된 난모세포들을 체외 성숙 후 세포 손상이 없이 성숙 난모세포의 발생능을 알아낼 수 있는 마커로 극체의 방출이 효과적으로 활용될 수 있는지를 알아보았다. 난모세포를 48시간 성숙 배양 후 극체의 방출 유무를 검사하고, 핵염색하여 염색체의 형태를 검사하였다. 확인된 난모세포들을 $16{\sim}18$시간 추가 배양한 후 7% ethanol로 활성화시키고 $5{\mu}g/ml$ cytochalasin B에 5시간 노출 후 NCSU23 배양액으로 7일간 배양하였다. 극체 방출율은 반복에 따라 $9.9{\sim}52.4%$, 퇴행율은 $21.4{\sim}61.8%$로 변이가 크게 나타났다(p<0.01). 극체를 방출한 난모세포의 핵상은 모두 극체와 19개의 염색체를 가진 제 2 감수분열 중기 핵상을 보여주었으며, 극체를 방출하지 못한 난모세포의 핵상은 핵이 팽화된 상태인 핵형이 39.1%, PCC 형태의 핵상이 19.6%, MI 형태의 핵상이 10.9%, MII이지만 극체가 관찰되지 않거나 매우 작은 상태인 경우가 13%, 핵이 응축된 형태인 경우가 6.5%, 핵이 없는 경우가 8.7%로 나타났다. 퇴행란으로 판단한 난모세포들은 핵염색을 한 결과 역시 세포질 상태가 정상적이지 못한 염색 상태를 보여주었다. 극체 방출 유무를 확인하지 않고 활성화 처리 후 배양하였을 때 분할율은 45.0%, 배반포기까지 발달율은 11.3%였으나, 극체 방출란만을 모아서 활성화처리를 하였을 때 분할율은 94.2%, 배반포기까지 발달율은 42.5%로 급격하게 향상되었다. 이상의 결과로 퇴행란과 극체 미방출란을 제거하고 실험에 활용한다면 배양 효과를 확인하거나 배아 생명 공학에 활용할 때 좀더 유리 할 것으로 사료된다.

• Asian-Australasian Journal of Animal Sciences
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• 제27권2호
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• pp.266-277
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• 2014

Somatic cell nuclear transfer (SCNT) has generally demonstrated that a differentiated cell can convert into a undifferentiated or pluripotent state. In the SCNT experiment, nuclear reprogramming is induced by exposure of introduced donor nuclei to the recipient cytoplasm of matured oocytes. However, because the efficiency of SCNT still remains low, a combination of SCNT technique with the ex-ovo method may improve the normal development of SCNT embryos. Here we hypothesized that treatment of somatic cells with extracts prepared from the germinal vesicle (GV) stage Siberian sturgeon oocytes prior to their use as nuclear donor for SCNT would improve in vitro development. A reversible permeability protocol with $4{\mu}g/mL$ of digitonin for 2 min at $4^{\circ}C$ in order to deliver Siberian sturgeon oocyte extract (SOE) to porcine fetal fibroblasts (PFFs) was carried out. As results, the intensity of H3K9ac staining in PFFs following treatment of SOE for 7 h at $18^{\circ}C$ was significantly increased but the intensity of H3K9me3 staining in PFFs was significantly decreased as compared with the control (p<0.05). Additionally, the level of histone acetylation in SCNT embryos at the zygote stage was significantly increased when reconstructed using SOE-treated cells (p<0.05), similar to that of IVF embryos at the zygote stage. The number of apoptotic cells was significantly decreased and pluripotency markers (Nanog, Oct4 and Sox2) were highly expressed in the blastocyst stage of SCNT embryos reconstructed using SOE-treated cells as nuclear donor (p<0.05). And there was observed a better development to the blastocyst stage in the SOE-treated group (p<0.05). Our results suggested that pre-treatment of cells with SOE could improve epigenetic reprogramming and the quality of porcine SCNT embryos.

• 대한의용생체공학회:학술대회논문집
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• 대한의용생체공학회 1995년도 춘계학술대회
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• pp.23-26
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• 1995

The objectives of the present study are 1)to find the effect of the diameter of transpedicular screws on their fixational strength in pedicles under static pull-out loading, 2)to determine the biomechanical correlation between the pedicle diameter and the screw diameter, and 3)to find the effects of other factors in the screw design, such as materials, screw pitch, thread height and shape on their fixational strength. Biomechanical tests (Test I) were performed to evaluate the effect of the screw diameter on pull-out strength by using 60 porcine pedicls and six groups of custom-made pedicle screws with different diameters (the major and the minor diameter of the screws used in the testing varied from 4mm upto 9mm and from 3mm upto 8mm, respectively) while all other factors (materials, screw pitch, thread height and shape etc.) were fixed. In Test II, by using 61 porcine pedicles, the relationship between the ratio of the pedicle diameter and the screw diameter(=aspect ratio) of the custum-made screw and the pull-out strength of the screw was investigated. Test III was performed with 94 porcine pedicles and 8 different types of the commercial screws from 6 major productors in order to determine the effect of the screw diameter, pitch and the thread shape on the pull-out strength of the screw, respectively. The results of Test I showed that the axial pull-out resistance of the screw could be increased prportionaly to the screw diameter(P<0.05). But this increase in the pull-out resistance did not found when the screws of 4mm or 9mm in the diameter were employed. It was found from the results of Test II that the screws had its maximum pull-out resistant force when the aspect ratio ranging 40 - 69% (P<0.05). based on the results for the major diameter against the minor diameter of screw, the maximal pull-out resistance was found at 60-65% (P<0.05). According to these biomechanical testing results, it seems that the screw with a moderately large pitch is more desirable and the buttress-shaped screw can provide stronger fixation than the V-shape one can, if other designal factor and conditions were fixed.

• 한국수정란이식학회지
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• 제19권1호
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• pp.19-25
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• 2004

본 연구는 돼지 체외수정란의 생산에 있어서 체외배양체계를 확립하기 위해 정액의 차이와 보관시간에 따른 체외성숙난자의 체외수정 및 체외발달율을 조사하였고, NCSU23와 G1.3/G2.3 배양액으로 체외배양 시 분할율과 체외발달율을 조사하였고, 체외배양액에 GSH를 첨가 시 배발달율을 조사하였다. 본 연구에서 얻은 결과는 다음과 같다. 1. 액상정액과 동결정액을 사용하여 정액에 따른 체외수정율은 각각 46.2 및 39.7%로써 액상정액을 이용한 것이 수정율이 높았지만 유의적(P<0.05)인 차이는 없었다. 한편 배반포로의 발달율은 액상정액이 동결정액보다 유의적으로 높았다.(P< 0.05) 2. 정소상체미부 정액을 day 1, 2, 3 보관후 체외수정을 유도하였을 때 수정율은 각각 60.5, 61.0 및 56.8%로서 2일 동안 보관후 사용하였을 때가 높았지만 유의적(P<0.05)인 차이가 없었다. 또한 상실배와 배반포기까지의 배발달율에서도 2일 동안 보관 후 사용하였을 때가 높았지만 유의적(P<0.05)인 차이는 없었다. 3. NCSU-23와 G1.3/G2.3으로 나눠 배양 시 분할율과 상실배기까지의 배발달율은 각각 52.8, 62.1%와 16.0, 28.9%로서 G1.3/G2.3에서 유의적(P<0.05)인 차이로 높았다. 그러나 배반포기로의 배발달율을 각각 11.6와 4.7%로서 NCSU-23에서 유의적으로 높았다.(P<0.05) 4. NCSU-23을 기본 배양액으로 하여 1 mM GSH가 첨가된 군의 분할율은 62.3%로서 GSH가 첨가되지 않은 군 53.5%보다 유의적(P<0.05)인 차로 높았지만, 상실배나 배반포기까지의 배발달율은 GSH가 첨가되지 않은 군보다 높았지만 유의적(P<0.05)인 차는 존재하지 않았다.ne을 처리하는 것이 수태율을 향상시키는 것으로 나타났다.25^{\circ}C$에서 발현되었고 이 중 79.1%가 3일령, 4일령과 5일령에 집중적으로 나타났다. 계분의 경우 $25^{\circ}C$ 처리구에서 35.12%(56.95mg/kg), $37^{\circ}C$에서 45.89%(74.40mg/kg)로 나타나 주로 $25^{\circ}C$ 이상에서 발현한 것으로 특징지어졌다. 우분의 경우 $10^{\circ}C$ 처리구에서 28.21%(43.86mg/kg), $25^{\circ}C$에서 49.30% (76.66mg/kg)로 나타나 주로 $25^{\circ}C$ 이하에서 발현한 것으로 나타났다. 3. 배양온도에서 검지 된 뷰틸산의 량은 6일 동안 돈분에서 1,463.87mg/kg, 계분에서 96.72mg/kg, 우분에서 129.18mg/kg이 발현되었으며 돈분의 경우 93.31%(1,365.95mg/kg)가 $25^{\circ}C$에서 발현되었고 이 중 87.92%가 3일령, 4일령과 6일령에 집중적으로 나타났다. 계분의 경우 $37^{\circ}C$ 처리구에서 76.60%(74.09 mg/kg)로 발현되었고 이 중 88%가 1일령, 2일령과 5일령에 집중적으로 나타났다. 우분의 경우 61.55%(79.51mg/kg)가 $25^{\circ}C$에서 발현되었고 이 중 89.6%가 1일령, 3일령과 4일령에 집중적으로 나타났다. 4. 배양온도에서 검지된 이소밸릭산의 량은 6일 동안 돈분에서 6,885.99mg/kg, 계분에서 307.47mg/kg,