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Molecular characteristics of Escherichia coli from bulk tank milk in Korea

  • Yoon, Sunghyun;Lee, Young Ju
    • Journal of Veterinary Science
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    • 제23권1호
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    • pp.9.1-9.11
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    • 2022
  • Background: Escherichia coli, which causes subclinical or clinical mastitis in cattle, is responsible for transmitting antimicrobial resistance via human consumption of raw milk or raw milk products. Objectives: The objective of this study was to investigate the molecular characteristics of 183 E. coli from bulk tank milk of five different dairy factories in Korea. Methods: The molecular characteristics of E. coli such as serogroup, virulence, antimicrobial resistance, and integron genes were detected using polymerase chain reaction and antimicrobial susceptibility were tested using the disk diffusion test. Results: In the distribution of phylogenetic groups, group D was the most prevalent (59.6%) and followed by group B1 (25.1%). The most predominant serogroup was O173 (15.3%), and a total of 46 different serotypes were detected. The virulence gene found most often was fimH (73.2%), and stx1, fimH, incC, fyuA, and iutA genes were significantly higher in isolates of phylogenetic group B1 compared to phylogenetic groups A, B2, and D (p < 0.05). Among 64 E. coli isolates that showed resistance to at least one antimicrobial, the highest resistance rate was observed for tetracyclines (37.5%). All 18 integron-positive E. coli carried the integron class I (int1) gene, and three different gene cassette arrangements, dfrA12+aadA2 (2 isolates), aac(6')-Ib3+aac(6')-Ib-cr+aadA4 (2 isolates), and dfrA17+aadA5 (1 isolate) were detected. Conclusions: These data suggest that the E. coli from bulk tank milk can be an indicator for dissemination of antimicrobial resistance and virulence factors via cross-contamination.

Chemoquiescence with Molecular Targeted Ablation of Cancer Stem Cells in Gastrointestinal Cancers

  • Jong-Min Park;Young-Min Han;Migyeong Jeong;Eun Jin Go;Napapan Kangwan;Woo Sung Kim;Ki Baik Hahm
    • Journal of Digestive Cancer Research
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    • 제4권1호
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    • pp.1-9
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    • 2016
  • The abundance of multi-drug resistance ATPase binding cassette and deranged self-renewal pathways shown in cancer stem cells (CSCs) played a crucial role in tumorigenesis, tumor resistance, tumor recurrence, and tumor metastasis. Therefore, elucidation of CSCs biology can improve diagnosis, enable targeted treatment, and guide the follow up of GI cancer patients. In order to achieve chemoquiescence, seizing cancer through complete ablation of CSCs, CSCs are rational targets for the design of interventions that will enhance responsiveness to traditional therapeutic strategies and contribute in the prevention of local recurrence as well as metastasis. However, current cancer treatment strategies fail to either detect or differentiate the CSCs from their non-tumorigenic progenies mostly due to the absence of specific biomarkers and potent agents to kill CSCs. Recent advances in knowledge of CSCs enable to produce several candidates to ablate CSCs in gastrointestinal (GI) cancers, especially cancers originated from inflammation-driven mutagenesis such as Barrett's esophagus (BE), Helicobacter pylori-associated gastric cancer, and colitis-associated cancer (CAC). Our research teams elucidated through revisiting old drugs that proton pump inhibitor (PPI) and potassium competitive acid blocker (p-CAB) beyond authentic acid suppression, chloroquine for autophage inhibition, sonic hedgehog (SHH) inhibitors, and Wnt/β-catenin/NOTCH inhibitor can ablate CSCs specifically and efficiently. Furthermore, nanoformulations of these molecules could provide an additional advantage for more selective targeting of the pathways existing in CSCs just like current molecular targeted therapeutics and sustained action, while normal stem cells intact. In this review article, the novel approach specifically to ablate CSCs existing in GI cancers will be introduced with the introduction of explored mode of action.

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Ellagic acid, a functional food component, ameliorates functionality of reverse cholesterol transport in murine model of atherosclerosis

  • Sin-Hye Park;Min-Kyung Kang;Dong Yeon Kim;Soon Sung Lim;Il-Jun Kang;Young-Hee Kang
    • Nutrition Research and Practice
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    • 제18권2호
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    • pp.194-209
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    • 2024
  • BACKGROUND/OBJECTIVES: High levels of plasma low-density lipoprotein (LDL) cholesterol are an important determinant of atherosclerotic lesion formation. The disruption of cholesterol efflux or reverse cholesterol transport (RCT) in peripheral tissues and macrophages may promote atherogenesis. The aim of the current study was to examine whether bioactive ellagic acid, a functional food component, improved RCT functionality and high-density lipoprotein (HDL) function in diet-induced atherogenesis of apolipoproteins E (apoE) knockout (KO) mice. MATERIALS/METHODS: Wild type mice and apoE KO mice were fed a high-cholesterol Paigen diet for 10 weeks to induce hypercholesterolemia and atherosclerosis, and concomitantly received 10 mg/kg ellagic acid via gavage. RESULTS: Supplying ellagic acid enhanced induction of apoE and ATP-binding cassette (ABC) transporter G1 in oxidized LDL-exposed macrophages, facilitating cholesterol efflux associated with RCT. Oral administration of ellagic acid to apoE KO mice fed on Paigen diet improved hypercholesterolemia with reduced atherogenic index. This compound enhanced the expression of ABC transporters in peritoneal macrophages isolated from apoE KO mice fed on Paigen diet, indicating increased cholesterol efflux. Plasma levels of cholesterol ester transport protein and phospholipid transport protein involved in RCT were elevated in mice lack of apoE gene, which was substantially reduced by supplementing ellagic acid to Paigen diet-fed mice. In addition, ellagic acid attenuated hepatic lipid accumulation in apoE KO mice, evidenced by staining of hematoxylin and eosin and oil red O. Furthermore, the supplementation of 10 mg/kg ellagic acid favorably influenced the transcriptional levels of hepatic LDL receptor and scavenger receptor-B1 in Paigen diet-fed apoE KO mice. CONCLUSION: Ellagic acid may be an athero-protective dietary compound encumbering diet-induced atherogenesis though improving the RCT functionality.

Glucosamine increases macrophage lipid accumulation by regulating the mammalian target of rapamycin signaling pathway

  • Sang-Min Kim;Dong Yeol Kim;Jiwon Park;Young-Ah Moon;Inn-Oc Han
    • BMB Reports
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    • 제57권2호
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    • pp.92-97
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    • 2024
  • Elevated blood glucose is associated with an increased risk of atherosclerosis. Data from the current study showed that glucosamine (GlcN), a normal glucose metabolite of the hexosamine biosynthetic pathway (HBP), promoted lipid accumulation in RAW264.7 macrophage cells. Oleic acid- and lipopolysaccharide (LPS)-induced lipid accumulation was further enhanced by GlcN in RAW264.7 cells, although there was no a significant change in the rate of fatty acid uptake. GlcN increased acetyl CoA carboxylase (ACC), fatty acid synthase (FAS), scavenger receptor class A, liver X receptor, and sterol regulatory element-binding protein-1c (SREBP-1c) mRNA expression, and; conversely, suppressed ATP-binding cassette transporter A1 (ABCA-1) and ABCG-1 expression. Additionally, GlcN promoted O-GlcNAcylation of nuclear SREBP-1 but did not affect its DNA binding activity. GlcN stimulated phosphorylation of mammalian target of rapamycin (mTOR) and S6 kinase. Rapamycin, a mTOR-specific inhibitor, suppressed GlcN-induced lipid accumulation in RAW264.7 cells. The GlcN-mediated increase in ACC and FAS mRNA was suppressed, while the decrease in ABCA-1 and ABCG-1 by GlcN was not significantly altered by rapamycin. Together, our results highlight the importance of the mTOR signaling pathway in GlcN-induced macrophage lipid accumulation and further support a potential link between mTOR and HBP signaling in lipogenesis.

Why A Multimedia Approach to English Education\ulcorner

  • Keem, Sung-uk
    • 대한음성학회:학술대회논문집
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    • 대한음성학회 1997년도 7월 학술대회지
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    • pp.176-178
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    • 1997
  • To make a long story short I made up my mind to experiment with a multimedia approach to my classroom presentations two years ago because my ways of giving instructions bored the pants off me as well as my students. My favorite ways used to be sometimes referred to as classical or traditional ones, heavily dependent on the three elements: teacher's mouth, books, and chalk. Some call it the 'MBC method'. To top it off, I tried audio-visuals such as tape recorders, cassette players, VTR, pictures, and you name it, that could help improve my teaching method. And yet I have been unhappy about the results by a trial and error approach. I was determined to look for a better way that would ensure my satisfaction in the first place. What really turned me on was a multimedia CD ROM title, ELLIS (English Language Learning Instructional Systems) developed by Dr. Frank Otto. This is an integrated system of learning English based on advanced computer technology. Inspired by the utility and potential of such a multimedia system for regular classroom or lab instructions, I designed a simple but practical multimedia language learning laboratory in 1994 for the first time in Korea(perhaps for the first time in the world). It was high time that the conventional type of language laboratory(audio-passive) at Hahnnam be replaced because of wear and tear. Prior to this development, in 1991, I put a first CALL(Computer Assisted Language Learning) laboratory equipped with 35 personal computers(286), where students were encouraged to practise English typing, word processing and study English grammar, English vocabulary, and English composition. The first multimedia language learning laboratory was composed of 1) a multimedia personal computer(486DX2 then, now 586), 2) VGA multipliers that enable simultaneous viewing of the screen at control of the instructor, 3) an amplifIer, 4) loud speakers, 5)student monitors, 6) student tables to seat three students(a monitor for two students is more realistic, though), 7) student chairs, 8) an instructor table, and 9) cables. It was augmented later with an Internet hookup. The beauty of this type of multimedia language learning laboratory is the economy of furnishing and maintaining it. There is no need of darkening the facilities, which is a must when an LCD/beam projector is preferred in the laboratory. It is headset free, which proved to make students exasperated when worn more than- twenty minutes. In the previous semester I taught three different subjects: Freshman English Lab, English Phonetics, and Listening Comprehension Intermediate. I used CD ROM titles like ELLIS, Master Pronunciation, English Tripple Play Plus, English Arcade, Living Books, Q-Steps, English Discoveries, Compton's Encyclopedia. On the other hand, I managed to put all teaching materials into PowerPoint, where letters, photo, graphic, animation, audio, and video files are orderly stored in terms of slides. It takes time for me to prepare my teaching materials via PowerPoint, but it is a wonderful tool for the sake of presentations. And it is worth trying as long as I can entertain my students in such a way. Once everything is put into the computer, I feel relaxed and a bit excited watching my students enjoy my presentations. It appears to be great fun for students because they have never experienced this type of instruction. This is how I freed myself from having to manipulate a cassette tape player, VTR, and write on the board. The student monitors in front of them seem to help them concentrate on what they see, combined with what they hear. All I have to do is to simply click a mouse to give presentations and explanations, when necessary. I use a remote mouse, which prevents me from sitting at the instructor table. Instead, I can walk around in the room and enjoy freer interactions with students. Using this instrument, I can also have my students participate in the presentation. In particular, I invite my students to manipulate the computer using the remote mouse from the student's seat not from the instructor's seat. Every student appears to be fascinated with my multimedia approach to English teaching because of its unique nature as a new teaching tool as we face the 21st century. They all agree that the multimedia way is an interesting and fascinating way of learning to satisfy their needs. Above all, it helps lighten their drudgery in the classroom. They feel other subjects taught by other teachers should be treated in the same fashion. A multimedia approach to education is impossible without the advent of hi-tech computers, of which multi functions are integrated into a unified system, i.e., a personal computer. If you have computer-phobia, make quick friends with it; the sooner, the better. It can be a wonderful assistant to you. It is the Internet that I pay close attention to in conjunction with the multimedia approach to English education. Via e-mail system, I encourage my students to write to me in English. I encourage them to enjoy chatting with people all over the world. I also encourage them to visit the sites where they offer study courses in English conversation, vocabulary, idiomatic expressions, reading, and writing. I help them search any subject they want to via World Wide Web. Some day in the near future it will be the hub of learning for everybody. It will eventually free students from books, teachers, libraries, classrooms, and boredom. I will keep exploring better ways to give satisfying instructions to my students who deserve my entertainment.

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Molecular Epidemiologic Study of a Methicillin-resistant Staphylococcus aureus Outbreak at a Newborn Nursery and Neonatal Intensive Care Unit

  • Kang, Hyun Mi;Park, Ki Cheol;Lee, Kyung-Yil;Park, Joonhong;Park, Sun Hee;Lee, Dong-Gun;Kim, Jong-Hyun
    • Pediatric Infection and Vaccine
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    • 제26권3호
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    • pp.148-160
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    • 2019
  • 목적: 본 연구에서는 신생아실과 신생아 중환자실에서 발생한 methicillin-resistant Staphylococcus aureus (MRSA) 유행에서 환자와 보균자에서 분리된 MRSA의 분자역학적 연관성을 조사하여 유행의 감염원과 전파경로를 파악하고자 하였다. 방법: MRSA 유행기간인 2017년 8월부터 9월까지 피부감염 및 패혈증 환자들과 보균자로부터 분리된 MRSA 균주를 대상으로 유전형 및 병원성 인자를 분석하고 항생제 감수성 결과를 수집하였다. 결과: 연구기간 동안 신생아실(n=27)과 신생아 중환자실(n=14)에 총 41명의 신생아들이 입원하였다. 그 중, 7명(피부감염[n=6], 패혈증[n=1])에서 MRSA 감염이 확진되었고, 보균자 4명이 발견되었다. 신생아와 접촉이 있는 의료진 32명 중 3명이 MRSA를 비강에 보균하였다. 피부감염 유행 원인 균주는 Staphylococcal chromosomal cassette mec (SCCmec) type II, sequence type (ST) 89, spa type t375였고, 뮤피로신 저농도 내성을 포함하여 항생제 다제내성을 보였다. 패혈증을 일으킨 균주는 SCCmec type IVa, ST 72, 새로운 spa type인 t17879였다. 신생아 4명에게 집락된 MRSA 균주들은 다양하였으나 SCCmec type IVa, ST 72, spa type t664가 의료진과 신생아 2명에서 공통적으로 분리되었다. Panton-Valentine leukocidin (PVL) toxin 유전자가 신생아에게 집락된 모든 균주에서 발견되었다. 결론: 피부감염 유행을 일으킨 MRSA 균주는 항생제 다제내성을 보이는 균주였다. 신생아 MRSA 보균자에게서 분리된 균주는 모두 PVL 독소 유전자를 보유하였다. 유행기간 동안 다양한 MRSA 균주가 신생아들에게서 분리되기 때문에, 효과적인 감염 관리 및 추가 환자발생의 차단을 위하여 분자역학조사를 통하여 원인균을 확인하고 전파경로를 파악하는 것이 중요하다.

메탄올 자화효모 Hansenula polymorpha에서의 재조합 단백질 분비발현을 위한 인체 혈청 알부민 융합단편의 활용 (Use of Human Serum Albumin Fusion Tags for Recombinant Protein Secretory Expression in the Methylotrophic Yeast Hansenula polymorpha)

  • 송지혜;황동현;오두병;이상기;권오석
    • 한국미생물·생명공학회지
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    • 제41권1호
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    • pp.17-25
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    • 2013
  • 메탄올 자화효모 Hansenula polymorpha에서 분비 발현이 잘 된다고 보고된 인체 혈청 알부민(human serum albumin, HSA)을 융합단편으로 사용하여 외래 재조합 단백질을 효과적으로 분비 발현할 수 있는 발현시스템을 개발하고자 하였다. 이때 조작의 용이성 및 발현 효율 제고를 위하여 전장의 HSA 뿐만 아니라 세 종류의 각기 다른 크기의 HSA 단편을 설계하여 융합단편으로 사용하였다. 즉 HSA의 N-말단으로 부터 각기 137, 172, 320, 608개 아미노산을 갖는 융합단편 HSAft (1-4)를 제작하였다. 아울러 발현되는 HSA 단편의 검출 및 분리정제를 위한 His8-tag, HSA 융합단편과 외래 단백질간의 유연성을 부여하기 위한 2조의 $Gly_4Ser_1$ linker, 융합 발현된 타겟 단백질을 HSA 단편으로부터 용이하게 분리하기 위한 담배식각바이러스 단백질분해효소(tobacco etch virus protease, Tev) 인지 서열, 타겟 단백질 유전자를 클로닝하기 위한 멀티 클로닝 사이트(multiple cloning site, MCS)서열, 그리고 타겟 재조합 단백질의 발현 검출 및 정제를 위한 Strep-tag을 포함하는 작용기 도메인을 발현카세트 기본 골격에 포함시켰다. 이렇게 구축된 4종의 HSA 융합단편 분비발현 벡터를 H. polymorpha에 형질전환한 후 각 융합단편의 발현을 조사한 결과 HSAft 단편 3, 4의 발현을 확인할 수 있었다. 녹색형광단백질 유전자 ($GFP_{uv}$)를 상기 벡터에 클로닝한 후 H. polymorpha에 도입한 결과 형질 전환체 모두에서 녹색형광단백질의 발현을 관찰 할 수 있었다. 해당 세포로부터 분비되거나 세포내에 발현되는 HSA 단편 융합 형광단백질의 발현양을 비교한 결과 HSAft 단편 4에 융합된 경우를 제외하고 나머지 경우 모두에서 세포 파쇄액과 세포 배양액 양쪽에서 해당 HSA 단편 융합 형광단백질의 발현을 확인 할 수 있었다. 한편 HSA 융합단편의 크기에 따라 자체 혹은 타겟 단백질과 융합된 형태의 단백질 분비 발현 정도가 달라지는 것은 해당 단백질의 접힘이나 단백질 분해효소에 대한 민감성 등 여러 변수에 의한 것으로 사료되며 따라서 본 연구에서 개발한 H. polymorpha용 HSA 융합단편 분비발현 시스템은 특정 외래 재조합 단백질의 효율적인 분비발현 융합단편의 선별 및 과발현 시스템 구축에 유용하게 활용될 수 있을 것으로 기대된다.

골반강 내 방사선 치료 환자에서 Electronic Portal Imaging Device(EPID)를 이용한 Portal Image의 유용성에 관한 연구 (Evaluation of Usefulness of Portal Image Using Electronic Portal Imaging Device (EPID) in the Patients Who Received Pelvic Radiation Therapy)

  • 김우철;박원;김현정;박성용;조영갑;노준규;서창옥;김귀언
    • Radiation Oncology Journal
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    • 제16권4호
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    • pp.497-504
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    • 1998
  • 목적 : matrix ion chamber type의 EPID와 video camera based EPID를 이용한 portal image와 기존의 film을 이용한 port film의 영상의 질을 객관적으로 비교 평가하여 EPID의 유용성을 알아보고자 본 연구를 계획하였다. 대상 및 방법 : 1997년 4월부터 10월까지의 인하대 병원과 세브란스 병원에서 방사선 치료를 받은 골반강내 치료 환자 각 10명씩을 대상으로 환자 1명 당 5-10회의 port film과 EPID를 이용한 portal image를 동시에 얻어 비교하였다. 환자의 나이는 32세에서 79세이었고 2명의 AP영상을 제외하고는 모두 PA영상을 얻었다. 환자의 두께는 17cm에서 20cm으로 비교적 균일하였다. beam energy는 10MV X-ray를 사용하였고 dose rate은 100-300MU/min으로 2-10MU을 주어 영상을 얻었다. port film은 Kodak diagnostic film을 사용하였고 film을 넣는 cassette는 납을 전후에 부착한 것을 이용하였다. source to detector(film) distance는 140cm으로 하였다. 영상의 판독은 4명의 치료방사선과 의사에 의해서 시행되었으며 pelvic brim, sacrum, acetabulum, iliopectineal line, symphysis, ischium, obturator foramen, sacroiliac joint를 각각 very clear(1), clear(2), visible(3), not clear(4), not visible(5) 다섯 단계로 나누어 점수를 주었다. 결과 : video camera based EPID를 이용하여 얻은 영상을 비교하여 보았을 때 film을 이용한 port film과 enhancement를 시행하지 않은 portal image는 각 해부학적 구조에서는 차이를 보이지 않았다. 그러나 portal image를 window level로 영상의 변화를 주었을 때는 sacrum과 obturator는 영상의 판독에 도움이 되었다. 또한 portal image를 CLAHE로 enhance를 하였을 때는 모든 해부학적 구조물의 판독이 film보다 용이한 것으로 나타났다. matrix ion chamber type의 EPID를 이용하여 얻은 영상에서도 역시 port film과 영상의 변화를 주지 않은 portal image간에는 커다란 차이를 보이지 않았으나, portal image를 window level로 변화를 주었을 경우는 portal film에 비하여 영상의 질이 더욱 좋아지을 알 수 있었다. 결론 : 방사선 치료를 받는 환자 중에서 골반강의 영상에서는 EPID의 영상의 질은 기존의 port film과 비교하여 차이가 없었으며, window level로 영상에 변화를 주거나 enhance를 하였을 경우는 port film보다 더 나은 영상을 얻을 수 있어 기존의 port film을 대체 할 수 있을 것으로 생각된다.

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DV에서 MPEG-2로의 주파수 영역 변환 부호화 (A Frequency Domain DV-to-MPEG-2 Transcoding)

  • 김도년;윤범식;최윤식
    • 대한전자공학회논문지SP
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    • 제38권2호
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    • pp.138-148
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    • 2001
  • 디지털 캠코더에서 이용하는 영상 압축 방식인 DY 부호화 방식은 DCT와 가변장 부호화 방식을 이용한다. DV 방식은 하드웨어 복잡도가 낮은 반면 압축된 비트율이 악 26Mb/s로 높은 편이다. 따라서 스튜디오에서 낮은 복잡도로 영상을 부호화 한 후 VOD 시스템에 이용하기 위하여 MPEG-2로 변환부호화 할 필요가 있다. 이때의 두 압축방식이 DCT를 이용하므로, DCT 영역에서 변환부호화 하면 중간 과정을 줄일 수 있어서 계산상의 복잡도를 줄일 수 있다. DV 방식에서 MPEG-2 인트라로 변환부호화 시에, DV 방식의 4:1:1 색차 포맷을 MPEG-2의 4:2:2 영상 포맷으로 변환할 때 와 2-4-8 DCT 모드에서 8-8 DCT 모드로 변환 시 변환 영역에 있는 데이터에 미리 계산된 행렬을 곱함으로써 병렬처리가 가능하게 하였다. MPEG-2 율제어 시에 서브 블록의 분산을 완전히 DCT 영역에서 계산하였다. 실험을 통하여 제안한 방식들을 검증하였다. MPEG-2 인터 프레임 부호화로 변환 부호화 할 때 DCT 계수를 이용하여 계층적으로 움직임을 추정하였다. 먼저 4개의 서브 블록에 있는 4개의 DC 값으로 하나의 매크로 블록에 대한 움직임을 추정한 다음 각 서브 블록의 저주파수에 해당하는 2×2에 IDCT를 취하여 16 포인트로 구성된 매크로 블록을 만든 후 이에 대한 움직임을 추정하며, 다섯 번째 단계에서 서브 화소에 대한 움직임을 추정함으로써 움직임 추정을 마친다. 탐색영역을 겹치는 방식이 겹치지 않는 방식보다 좋은 PSNR값을 보여 주었다.

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alpha 1,3-galactosyltransferase 기능 제거 및 MCP 발현 형질전환 돼지의 대동맥 혈관내피세포에 CD37/CD73 발현 세포주 개발 (Development of aortic endothelial cells to express CD37 and CD73 isolated from alpha 1,3-galactosyltransferase knock-out and MCP expressing pig)

  • 노진구;변승준;양현;옥선아;우제석;이휘철;황인설;김지윤;박상현;이주영;오건봉
    • 한국수정란이식학회지
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    • 제33권3호
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    • pp.129-137
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    • 2018
  • Acute vascular rejection has been known as a main barrier occurring in a xenograted tissue of alpha 1,3-galactosyltransferase knock-out (GalT KO) pig into a non-human primate (NHP). Adenosine which is a final metabolite following sequential hydrolysis of nucleotide by ecto-nucleotidases such as CD39 and CD73, act as a regulator of coagulation, and inflammation. Thus xenotransplantation of CD39 and CD73 expressing pig under the GalT KO background could lead to enhanced survival of recipient NHP. We constructed a human CD39 and CD73 expression cassette designed for endothelial cell-specific expression using porcine Icam2 promoter (pIcam2-hCD39/hCD73). We performed isolation of endothelial cells (pAEC) from aorta of 4 week-old GalT KO and membrane cofactor protein expressing pig ($GalT^{-MCP/-MCP}$). We were able to verify that isolated cells were endothelial-like cells using immunofluorescence staining analysis with von Willebrand factor antibody, which is well known as an endothelial maker, and tubal formation assay. To find optimal condition for efficient transfection into pAEC, we performed transfection with GFP expression vector using four programs of nucleofection, M-003, U-023, W-023 and Y-022. We were able find that the program W-023 was optimal for pAEC with regard to viability and transfection efficiency by flow cytometry and fluorescent microscopy analyses. Finally, we were able to obtain $GalT^{-MCP/-MCP}/CD39/CD73$ pAEC expressing CD39 and CD73 at levels of 33.3% and 26.8%, respectively. We suggested that pACE isolated from $GalT^{-MCP/-MCP}$ pig might be provided as a basic resource to understand biochemical and molecular mechanisms of the rejections and as an alternative donor cells to generate $GalT^{-MCP/-MCP}/CD39/CD73$ pig expressing CD39 and CD73 at endothelial cells.