• International Journal of Stem Cells
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• 제16권2호
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• pp.234-243
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• 2023

The recent advances in human pluripotent stem cells (hPSCs) enable to precisely edit the desired bases in hPSCs to be used for the establishment of isogenic disease models and autologous ex vivo cell therapy. The knock-in approach based on the homologous directed repair with Cas9 endonuclease, causing DNA double-strand breaks (DSBs), produces not only insertion and deletion (indel) mutations but also deleterious large deletions. On the contrary, due to the lack of Cas9 endonuclease activity, base editors (BEs) such as adenine base editor (ABE) and cytosine base editor (CBE) allow precise base substitution by conjugated deaminase activity, free from DSB formation. Despite the limitation of BEs in transition substitution, precise base editing by BEs with no massive off-targets is suggested to be a prospective alternative in hPSCs for clinical applications. Considering the unique cellular characteristics of hPSCs, a few points should be considered. Herein, we describe an updated and optimized protocol for base editing in hPSCs. We also describe an improved methodology for CBE-based C to T substitutions, which are generally lower than A to G substitutions in hPSCs.

• Molecules and Cells
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• 제20권3호
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• pp.305-314
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• 2005

Investigation of chemically synthesized inositol 1,4,5-trisphosphate [$Ins(1,4,5)P_3$] analogs has led to the isolation of a novel binding protein with a molecular size of 130 kDa, characterized as a molecule with similar domain organization to phospholipase C-${\delta}1$ (PLC-${\delta}1$) but lacking the enzymatic activity. An isoform of the molecule was subsequently identified, and these molecules have been named PRIP (PLC-related, but catalytically inactive protein), with the two isoforms named PRIP-1 and -2. Regarding its ability to bind $Ins(1,4,5)P_3$ via the pleckstrin homology domain, the involvement of PRIP-1 in $Ins(1,4,5)P_3$-mediated $Ca^{2+}$ signaling was examined using COS-1 cells overexpressing PRIP-1 and cultured neurons prepared from PRIP-1 knock-out mice. Yeast two hybrid screening of a brain cDNA library using a unique N-terminus as bait identified GABARAP ($GABA_A$ receptor associated protein) and PP1 (protein phosphatase 1), which led us to examine the possible involvement of PRIP in $GABA_A$ receptor signaling. For this purpose PRIP knock-out mice were analyzed for $GABA_A$ receptor function in relation to the action of benzodiazepines from the electrophysiological and behavioral aspects. During the course of these experiments we found that PRIP also binds to the b-subunit of $GABA_A$ receptors and PP2A (protein phosphtase 2A). Here, we summarize how PRIP is involved in $Ins(1,4,5)P_3$-mediated $Ca^{2+}$ signaling and $GABA_A$ receptor signaling based on the characteristics of binding molecules.

• 디자인학연구
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• 16호
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• pp.213-223
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• 1996

원시시대부터 인간의 커뮤니케이션 수단으로 이용되어 온 일러스트레이션은 이제 우리 생활 속에서 생활의 단편적이거나 혹은 심층적인 면을 함축하여 표현하는 수단이 되었다. 즉 그것은 피터 노크(Peter Knock)의 한 잡지 일러스트레이션을 보고 어떤 예술가가 '여태껏 읽은 것 중 가자 훌륭한 기사였다'고 말 한데서도 알 수 있듯이, 일러스트레이터의 임무는 하루의 일상생활에서 아니 순간 순간의 사물의 지각을 통한 비구체적이든, 구체적이든 일러스트레이션의 소스(source)가 우리의 잠재의식 속에 남아있다고 가정할 때 이들을 추출해서 사실적으로 묘사하는 작업을 하고 있다고 할 수 있고, 그것은 어떤 미사여구를 붙인 문자 언어보다도 강한 지각작용을 불러일으킬 수 있다는 것이다. 뿐만 아니라 일러스트레이션의 활용분야는 나날이 증가하고 있으며 그 대상도 인물, 자연, 인공, 과학 등 다양해지고 따라서 이러한 시점의 일러스트레이션에서 가장 요구되는 것은 얼마나 창조적이며 참신한 아이디어를 지녔냐 하는 것이다. 왜냐하면 복잡 다양한 사회 속에서 일반인의 마음속에 자리잡기 위해서는 보다 독특한 이미지와 호소력을 가져야 하기 때문이다. 따라서 일러스트레이션의 교육도 이젠 단순한 기술연마가 아닌 시넥틱스(Synetics)적 사고에 바탕을 둔 작품으로 유도되어야하며 기능 위주의 도식적인 일러스트이션 교육에서 벗어난 보다 자유롭고 풍부한 개성을 지닌 사고를 할 수 있도록 되어야 한다.

• 한국수정란이식학회지
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• 제32권3호
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• pp.87-94
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• 2017

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 $37^{\circ}C$. 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.

• 한국자동차공학회논문집
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• 제15권2호
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• pp.8-14
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• 2007

Compression ignition of homogeneous charges in IC engines indicates possibilities of achieving the high efficiency of DI diesel engines with low level of NOx and particulate emissions. The objectives of this study are to further understand the characteristics of the HCCI(Homogeneous charge compression ignition) combustion and to find ways of extending the rich HCCI operation limit in an engine-like environment. DME fuel is supplied either in the form of premixture with air or directly injected in the combustion chamber of a rapid compression and expansion machine under the conditions of various equivalence ratio and injection timing. The cylinder pressure is measured and the rate of heat release is computed from the measured pressure for the analysis of the combustion characteristics. The experimental data show that the RCEM can operate without knock on mixtures of higher equivalence ratio, when DME is directly injected in the combustion chamber than introduced as a fraction of a perfect or nearly perfect premixture. Very early fuel injection timings usually employed in HCCI operation are seen to have only insignificant effects in control of ignition timing.

• 연구논문집
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• 통권33호
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• pp.5-16
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• 2003

Fuel distribution, combustion, and flame propagation characteristics of heavy duty engine with the liquid phase LPG injection(LPLI) were studied in a single cylinder engine. Optically accessible single cylinder engine and laser diagnostics system were built for quantifying fuel concentration by acetone PLIF(planar laser induced fluorescence) measurements. In case of Otto cycle engine with large bore size, the engine knock and thermal stress of exhaust manifold are so critical that lean burn operation is needed to reduce the problems. It is generally known that fuel stratification is one of the key technologies to extend the lean misfire limit. The formation of rich mixture in the spark plug vicinity was achieved by open valve injection. With higher swirl strength(Rs=3.4) and open valve injection, the cloud of fuel followed the flow direction and the radial air/fuel mixing was limited by strong swirl flow. It was expected that axial stratification was maintained with open-valve injection if the radial component of the swirling motion was stronger than the axial components. The axial fuel stratification and concentration were sensitive to fuel injection timing in case of Rs=3.4 while those were relatively independent of the injection timing in case of Rs2.3. Thus, strong swirl flow could promote desirable axial fuel stratification and, in result, may make flame propagation stable in the early stage of combustion.

• Korean Chemical Engineering Research
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• 제49권6호
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• pp.823-828
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• 2011

오일-물 혼합물에서 물을 제거하기 위하여 FWKO(Free Water Knock Out) vessel에서의 물 분리특성에 대해 연구하였다. 오일을 모사하기 위해 데칸, 톨루엔, 아스팔트를 사용하였다. 데칸을 이용하여 프로토타입 vessel에서 기초실험을 수행하였으며, 기초실험결과를 토대로 최적 water cut이 0.8인 것을 알게 되었다. 이에 이어 톨루엔을 이용하여 성능평가를 실행한 결과로 최적 체류시간이 1,200 sec인 것을 알 수 있었고 아스팔트를 사용하여 방해판의 개수, 실험온도를 변수로 FWKO vessel에서의 최적의 물 분리조건에 대해 조사하였다. 최적의 분리조건은 방해판 3개, 온도 $45^{\circ}C$였다.

• Reproductive and Developmental Biology
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• 제33권3호
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• pp.157-162
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• 2009

This study was performed to comprehend the developmental characteristics of cloned embryos knocked out (KO) of $\alpha$-1,3-galactosyltransferase (GalT) gene. Immature oocytes were collected and cultured for 40 hrs (1-step) or 20hrs (with hormone) + 20hrs (without hormone) (2-step). The embryos transferred with miniature pig ear fibroblast cell were used as control. The reconstructed embryos were cultured in PZM-3 with 5% $CO_2$ in air at $38.5^{\circ}C$ for 6 days. To determine the quality of the blstocysts, TUNEL and quantitative realtime RT-PCR were performed. The embryos were transferred to a surrogate (Landrace) at an earlier stage of the estrus cycle. The maturation rate was significantly higher in 2-step method than that of 1-step (p<0.05). The blastocyst development of GalT KO embryos was significantly lower than that of normal cloned embryos (p<0.05). The total and apoptotic cell number of GalT KO blastocysts was not different statistically from control. The relative abundance of Bax-$\alpha$/Bcl-xl ratio was significantly higher in both cloned blastocysts than that of in vivo blastocysts (p<0.05). Taken together, it can be postulated that the lower developmental potential and higher expression of apoptosis related genes in GalT KO SCNT embryos might be a cause of a low efficiency of GalT KO cloned miniature pig production.

• Parasites, Hosts and Diseases
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• 제50권1호
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• pp.1-6
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• 2012

$Toxoplasma$ $gondii$ penetrates all kinds of nucleated eukaryotic cells but modulates host cells differently for its intracellular survival. In a previous study, we found out that serine protease inhibitors B3 and B4 (SERPIN B3/B4 because of their very high homology) were significantly induced in THP-1-derived macrophages infected with $T.$ $gondii$ through activation of STAT6. In this study, to evaluate the effects of the induced SERPIN B3/B4 on the apoptosis of $T.$ $gondii$-infected THP-1 cells, we designed and tested various small interfering (si-) RNAs of SERPIN B3 or B4 in staurosporine-induced apoptosis of THP-1 cells. Anti-apoptotic characteristics of THP-1 cells after infection with $T.$ $gondii$ disappeared when SERPIN B3/B4 were knock-downed with gene specific si-RNAs transfected into THP-1 cells as detected by the cleaved caspase 3, poly-ADP ribose polymerase and DNA fragmentation. This anti-apoptotic effect was confirmed in SERPIN B3/B4 overexpressed HeLa cells. We also investigated whether inhibition of STAT6 affects the function of SERPIN B3/B4, and vice versa. Inhibition of SERPIN B3/B4 did not influence STAT6 expression but SERPIN B3/B4 expression was inhibited by STAT6 si-RNA transfection, which confirmed that SERPIN B3/B4 was induced under the control of STAT6 activation. These results suggest that $T.$ $gondii$ induces SERPIN B3/B4 expression via STAT6 activation to inhibit the apoptosis of infected THP-1 cells for longer survival of the intracellular parasites themselves.

• 한국자동차공학회논문집
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• 제19권2호
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• pp.35-41
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• 2011

This work investigates the potential of in-cylinder thermal stratification and fuel stratification for extending the operating ranges in HCCI engines, and the coupling between thermal stratification and fuel stratification. Computational results areemployed. The computations were conducted using both a custom multi-zone version and the standard single-zone version of the Senkin application of the CHEMKINII kinetics rate code, and kinetic mechanism for di-methyl ether (DME). This study shows that the potential of thermal stratification and fuels stratification for extending the high-load operating limit by a staged combustion event with reduced pressure-rise rates is very large. It was also found that those stratification offers good potential to extend low-load limit by a same mechanism in high-load. However, a combination of thermal stratification and fuel stratification is not more effective than above stratification techniques for extending the operating ranges showing similar results of fuel stratification. Sufficient condition for combustion (enough temperature for) turns misfire in low-load limit to operate engines, which also leads to knock in high-load limit abruptly due to the too high temperature with high. DME shows a potential for maximizing effect of stratification to lower pressure-rise rate due to the characteristics of low-temperature heat release.