中国组织工程研究
中國組織工程研究
중국조직공정연구
Journal of Clinical Rehabilitative Tissue Engineering Research
2014年
29期
4663-4668
,共6页
尹寿新%周震%马美雪%杨景波%羊东晔
尹壽新%週震%馬美雪%楊景波%羊東曄
윤수신%주진%마미설%양경파%양동엽
组织构建%组织工程%卵圆细胞%长期培养%表皮生长因子%白血病抑制因子%国家自然科学基金
組織構建%組織工程%卵圓細胞%長期培養%錶皮生長因子%白血病抑製因子%國傢自然科學基金
조직구건%조직공정%란원세포%장기배양%표피생장인자%백혈병억제인자%국가자연과학기금
hepatocytes%epidermal growth factor%leukemia inhibitory factor%cells,cultured
背景：肝卵圆细胞是目前公认的成体肝干/祖细胞，但其体外长期培养时会不可避免地丢失干细胞的活性。
<br> 目的：探索大鼠肝卵圆细胞体外长期培养的方法。
<br> 方法：构建2-乙酰胺基芴/部分肝切除肝再生大鼠模型，通过酶消化和Percol 梯度离心分离纯化大鼠异质卵圆细胞并进行免疫染色鉴定，用含表皮生长因子、白血病抑制因子的培养基体外长期培养，后撤去表皮生长因子、白血病抑制因子，通过形态学的观察和分子标志物的检测判断其能否保持干/祖细胞活性。
<br> 结果与结论：采用含表皮生长因子、白血病抑制因子的培养基体外培养卵圆细胞4个月后，大鼠异质卵圆细胞仍能表达肝细胞标志物ALB、胆管上皮细胞标志物CK-19，经不含表皮生长因子、白血病抑制因子的培养液继续培养后，卵圆细胞胎肝标志AFP表达量迅速下降。该研究结果表明大鼠肝卵圆细胞在表皮生长因子、白血病抑制因子等培养条件下可长期增殖并保持干细胞活性。
揹景：肝卵圓細胞是目前公認的成體肝榦/祖細胞，但其體外長期培養時會不可避免地丟失榦細胞的活性。
<br> 目的：探索大鼠肝卵圓細胞體外長期培養的方法。
<br> 方法：構建2-乙酰胺基芴/部分肝切除肝再生大鼠模型，通過酶消化和Percol 梯度離心分離純化大鼠異質卵圓細胞併進行免疫染色鑒定，用含錶皮生長因子、白血病抑製因子的培養基體外長期培養，後撤去錶皮生長因子、白血病抑製因子，通過形態學的觀察和分子標誌物的檢測判斷其能否保持榦/祖細胞活性。
<br> 結果與結論：採用含錶皮生長因子、白血病抑製因子的培養基體外培養卵圓細胞4箇月後，大鼠異質卵圓細胞仍能錶達肝細胞標誌物ALB、膽管上皮細胞標誌物CK-19，經不含錶皮生長因子、白血病抑製因子的培養液繼續培養後，卵圓細胞胎肝標誌AFP錶達量迅速下降。該研究結果錶明大鼠肝卵圓細胞在錶皮生長因子、白血病抑製因子等培養條件下可長期增殖併保持榦細胞活性。
배경：간란원세포시목전공인적성체간간/조세포，단기체외장기배양시회불가피면지주실간세포적활성。
<br> 목적：탐색대서간란원세포체외장기배양적방법。
<br> 방법：구건2-을선알기물/부분간절제간재생대서모형，통과매소화화Percol 제도리심분리순화대서이질란원세포병진행면역염색감정，용함표피생장인자、백혈병억제인자적배양기체외장기배양，후철거표피생장인자、백혈병억제인자，통과형태학적관찰화분자표지물적검측판단기능부보지간/조세포활성。
<br> 결과여결론：채용함표피생장인자、백혈병억제인자적배양기체외배양란원세포4개월후，대서이질란원세포잉능표체간세포표지물ALB、담관상피세포표지물CK-19，경불함표피생장인자、백혈병억제인자적배양액계속배양후，란원세포태간표지AFP표체량신속하강。해연구결과표명대서간란원세포재표피생장인자、백혈병억제인자등배양조건하가장기증식병보지간세포활성。
BACKGROUND:Hepatic oval cells are recognized as stem/progenitor cells currently, however, long-term culture of hepatic oval cells can inevitably result in the loss of cellactivity.
<br> OBJECTIVE:To explore a long-term culture method of hepatic oval cells in vitro.
<br> METHODS:Partial y hepatectomized rat model was established by using 2AAF/PH. The regenerated liver was digested by col agenase, and hepatic oval cells were isolated and purified by density gradient centrifugation and identified by immunocytochemistry. Hepatic oval cells were cultured in a medium containing epidermal growth factor and leukaemia inhibitory factor. Then epidermal growth factor and leukaemia inhibitory factor were removed after several months, and its ability of maintaining stem/progenitor cellactivities was determined based on morphology and molecular markers.
<br> RESULTS AND CONCLUSION:Both hepatocyte marker ALB and biliary epithelial cellmarker CK-19 were found after hepatic oval cells were cultured in a medium containing epidermal growth factor and leukaemia inhibitory factor for 4 months. While in the absence of epidermal growth factor and leukaemia inhibitory factor, the expression of fetal liver marker AFP was decreased quickly. These results indicated that hepatic oval cells could expand in a medium containing epidermal growth factor and leukaemia inhibitory factor and maintain stem/progenitor cellactivities for a long time.
