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안현희 ( Hyun Hee Ahn ),김혜수 ( Hye Soo Kim ),이혜경 ( Hye Kyung Lee ),신소영 ( So Young Shin ),이형진 ( Hyung Jin Lee ),강길선 ( Gil Son Khang ),이해방 ( Hai Bang Lee ),이일우 ( Il Woo Lee ) 한국조직공학과 재생의학회 2005 조직공학과 재생의학 Vol.2 No.4
Adipose derived stromal cells (ADSCs) are very similar to MSCs and can differentiate into mesenchymal lineage and nonmesenchymal lineage. ADSCs are very easily obtained than MSCs and no pain to acquire cells. Recently researches revealed that MSCs have nonspecific immune suppressive effect on lymphocyte proliferation. We studied about whether nonself ADSCs have lymphocyte suppressive effect on mixed lymphocyte culture (MLR) like MSCs. First we obtained 1st passaged ADSCs and tested whether these cells have stem cell characteristics. We confirmed induced cell by ALP and Oil red O staining. For immune suppressive effect test, ADSC were co-cultured with reaction lymphocyte and stimulator lymphocyte. We defined nonself ADSCs could suppression lymphocyte proliferation in MLR and two more cell number treatment could suppression lymphocyte proliferation 5-fold. Accordingly we found out ADSCs can lymphocyte suppression by dose-dependent pattern. In nonspecific immune suppression test by PHA as non specific lymphocyte stimulator, ADSCs can suppression PHA induced lymphocyte proliferation. In conclusion, it is our belief that ADSCs cells can alternative sources of MSCs in clinically usage and also remain carefully treatment.
섬유아세포에서의 DNA/폴리에틸렌이민 나노복합체를 이용한 유전자 전달
이정화 ( Jung Hwa Lee ),안현희 ( Hyun Hee Ahn ),신유나 ( Yu Na Shin ),김문석 ( Moon Suk Kim ),이봉 ( Bong Lee ),강길선 ( Gil Son Khang ),이일우 ( Il Woo Lee ),이해방 ( Hai Bang Lee ) 한국조직공학·재생의학회 2007 조직공학과 재생의학 Vol.4 No.4
The aim of this study is to evaluate transfection efficiency in NIH/3T3cell using DNA/polyethyleneimine (PEI) complexes. The formation of complexes from DNA and PEI was performed by the adding of the PEI solution to the DNA solution. Gel retardation assay showed the optimal N/P charge ratio where PEI completely binds DNA. The particle size of DNA/PEI complexes measured by dynamic light scattering showed diameters of up to 500 nm. The viability of NIH/3T3 fibroblast cells in the presence of DNA/PEI complexes was evaluated by the MTT assay. After gene transfection, enhanced green fluorescence protein was monitored by flow cytometry. From AFM measurement, DNA/PEI complexes showed round-shape. These results indicated that DNA was sufficiently condensed by PEI to give DNA/PEI complexes. As the relative amount of PEI was increased, its cytotoxicity to NIH/3T3cells increased. By increasing culture time, fluorescence microscopy showed a more green fluorescence cells. Naked DNA showed no EGFP expression, whereas delivery of DNA/PEI complexes to cells showed EGFP expression and resulted in 2-15% transfection. Transfection efficiencies of DNA/PEI complexes increased with increasing N/P charge ratio.
Taewoo Kang(강태우),Hyun-Hee An(안현희),Sul-Gi Park(박슬기),Sun-Nyoung Yu(유선녕),You-Lim Hwang(황유림),Ji-Won Kim(김지원),Soon-Cheol Ahn(안순철) 한국생명과학회 2019 생명과학회지 Vol.29 No.8
전립선암은 전이성 종양 중의 하나로 치료를 위해 호르몬 요법이나 외과 적 거세 방법이 주로 수행되지만 많은 부작용을 나타내었다. 최근 많은 연구자들이 이러한 상황을 해결하기 위해 종양 미세 환경을 연구하고 있으며 그 중 면역 세포, 특히 대식세포는 종양 미세 환경의 중요한 구성요소이다. 정상적인 조건에서 대식세포는 여러 암세포에 대해 약한 종양 살균 활성을 갖으나 interferon-γ 또는 lipopolysaccharide에 의해 활성화되면, 염증성 사이토카인 및 케모카인을 분비함으로써 암세포를 직접 또는 간접적으로 사멸 시키게 된다. 본 연구에서는, 마우스 대식세포인 RAW 264.7 세포에 Phellinus linteus 추출물을 처리하여 산화질소의 방출과 pro-inflammatory cytokine들을 real-time PCR과 ELISA 방법으로 분석하였다. RAW 264.7의 조정 배지는 48시간 동안 전립선 암세포처리하여 상피간엽세포전이 관련 유전자의 발현을 측정 하였다. 그 때에 mesenchymal 관련 유전자들인 N-cadherin, snail, twist, slug 및 cadherin 11이 감소했을 뿐만 아니라 epithelial 관련 유전자인 E-cadherin은 증가하였다. 또한 암 전이 및 신생 혈관 형성에 관여하는 vimentin, ccl2 및 vegfa가 감소되었는데, 이는 EMT가 암세포의 이동과 침범에 밀접한 관련이 있기 때문이다. 따라서 Phellinus linteu에 의해 자극된 RAW 264.7 세포의 조정 배지는 인간 전립선 암세포주인 PC-3 세포의 이동과 전이를 억제하고 EMT 경로를 조절한다는 것을 나타낸다. Prostate cancer (PCa) is one of the most metastatic tumor. Although hormone therapy or surgical castration is mostly conducted to treat PCa, it has a lot of side effects. Recently, many researchers have been exploring the tumor microenvironment to remedy these circumstances. Immune cells, especially macrophages, are an important composition of the tumor microenvironment. Under normal conditions, macrophages exhibit mild tumoricidal activity against tumors. However, once activated by interferon gamma or lipopolysaccharides, macrophages can kill cancer cells directly or indirectly by secreting cytokines and chemokines. In this study, murine macrophage RAW 264.7 cells were treated with Phellinus linteus extract. To analyze their pro-inflammatory phenotype, we were used several assays such as a real-time polymerase chain reaction, an enzyme-linked immunosorbent and nitric oxide assay. Prostate cancer cells were treated with the RAW 264.7-conditioned media, which was identified as a pro-inflammatory nature, for 48 h, and the expression of epithelial-mesenchymal transition (EMT)-related genes was determined. Not only N-cadherin, Snail, Twist, Slug, and Cadherin 11, which are mechenchymal-related proteins, were decrease, but epithelial marker of E-cadherin was increased. In addition, the mRNA level of vimentin, ccl2, and vegfa were decreased, as the EMT is closely related to the migration and invasion of cancer cells. In conclusion, the RAW 264.7-conditioned media stimulated with P. linteus extract inhibited migration and invasion and regulated the EMT pathway in human prostate cancer cells.
견우자의 생리활성 분석과 추출물로부터 항암 활성물질의 분리
최현덕(Hyeun Deok Choi),유선녕(Sun Nyoung Yu),박슬기(Sul-Gi Park),김영욱(Young Wook Kim),남효원(Hyo Won Nam),안현희(Hyun Hee An),김상헌(Sang Hun Kim),김광연(Kwang-Youn Kim),안순철(Soon Cheol Ahn) 한국생명과학회 2017 생명과학회지 Vol.27 No.2
견우자는 한의학에서 사하 작용 및 이뇨 작용 등의 치료 목적으로 사용되어 왔다. 본 연구에서는 견우자를 methanol로 추출하고(PNM), 이를 기본으로 ethyl acetate 분획물(PNE), butanol 분획물(PNB), water 분획물(PNW)로 나누었으며 각 분획물을 이용하여 mushroom tyrosinase 저해활성, pancreatic lipase 저해활성, DPPH free radical 소거활성 및 암세포주의 증식 저해활성 등을 조사하였다. 특히, 전립선암 세포주 PC-3 세포에서 강력한 세포 증식 저해활성을 보였으며, 용매 분획물 중, butanol 분획에서 가장 높은 암세포의 증식 저해활성을 보였다. 따라서 견우자의 methanol 추출물을 Diaion HP-20 column chromatography, reverse phase column chromatography, Sephadex LH-20 column chromatography를 순차적으로 실시하여 최종적으로 저해활성이 가장 높은 compound PN을 얻었으며 HPLC를 통해 그 순도를 확인하였다. Compound PN을 이용하여 cell cycle arrest과 annexin V/PI를 측정한 결과, 세포의 sub-G1기를 축적되면서 apoptosis가 유발되어 전립선암 세포주 PC-3 세포의 세포사멸이 유도되는 것을 확인하였다. 따라서 본 연구를 통하여 견우자에 대한 항산화 활성, 미백 활성 등의 효과 뿐 만 아니라 항암 활성에도 활용할 수 있을 것으로 사료된다. This study aimed to evaluate several biological activities of Pharbitis nil and to isolate an anticancer agent from its methanol extract. Pharbitis nil seeds were extracted with methanol (PNM). Then, PNM was fractionated into solvent layers such as ethyl acetate fraction (PNE), butanol fraction (PNB), and water fraction (PNW). The biological activities of the fractions were analyzed for tyrosinase inhibition, lipase inhibition, DPPH-free radical scavenging, and cell growth inhibition. PNM showed strong growth inhibition of prostate cancer PC-3 cells. PNM was subjected to Diaion HP-20 and eluted stepwise with 50%, 80%, and 100% methanol. Then, for activity-guided fraction, each fraction was analyzed for growth inhibition of prostate cancer PC-3 cells by using an MTT assay. Because the 100% fraction showed significantly strong inhibitory activity, the fraction was further separated in the reverse phase C18, which was eluted with 80% and 90% methanol. The 90% fraction was further subjected to Sephadex LH-20 using a mobile solvent of 100% methanol. Finally, the compound PN was partially purified for HPLC analysis. PN showed cell growth inhibitory activity and induced the apoptosis and cell cycle arrest of prostate cancer PC-3 cells, as measured by flow cytometry. The results together suggest that Pharbitis nil possesses various biological activities, especially the inhibitory activity for the proliferation of prostate cancer PC-3 cells, suggesting the possibility of its use as an anticancer agent.