3T3-L1 脂肪細胞培養及分化平台建立及其 應用於活性藻類的篩選

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题名:	3T3-L1 脂肪細胞培養及分化平台建立及其應用於活性藻類的篩選
作者:	洪綉茹;Hsiu-Ju Hung
贡献者:	生命科學研究所
日期:	2005-07-11
上传时间:	2009-09-22 10:18:10 (UTC+8)
出版者:	國立中央大學圖書館
摘要:	本實驗建立3T3-L1前脂肪細胞培養及利用分化劑誘導其分化的機制，作為篩選藻類可能存在抑制脂肪細胞分化活性物質的實驗工作平台。利用黃豆異黃酮genistein 100 ?M ( = 27 ?g/ ml)為正反應對照組，分別測試藍綠藻的NP01、NP02、NP03、CR02、FE01、FE02、FE03，紅藻的Ｈa01、Gc01、Grs04、Pd01，及褐藻Ec01等12株不同藻細胞的甲醇萃出物在27、270和2700??g/ml的濃度下對3T3-L1前脂肪細胞株的分化抑制活性。將3T3-L1前脂肪細胞培養於含DMEM-FBS的24孔培養盤，待其長滿後兩天，在培養液中添加insulin, dexamethasone和3-isobutyl-1-methylxathine作為分化誘發劑，隨後每兩日更換含有insulin的新鮮培養液至接種後的第十四天，分析其脂肪含量，而分化抑制劑與藻類萃出物的添加處理設定於接種後第六天。利用Oil redＯ染色法，比較各實驗組經分化劑誘導後之細胞培養，其細胞中脂質含量的差異，結果並未能發現測試的各藻株甲醇萃出物含有類似genistein之分化抑制效果，而且也不具備強化或減低genistein之抑制細胞脂質含量的作用。同時也發現實驗所採的各藻株甲醇萃出物，除FE02有稍許抑制細胞生長的活性外，並不會影響細胞的存活率。雖然本研究未能發現到具有抑制脂肪細胞分化的藻株萃出物，但已能充分掌握此細胞分化培養的實驗條件，並證明這樣的實驗平台可應用於天然物對脂肪細胞分化抑制活性篩選與測試。 The study tried to establish a 3T3-L1 preadipocyte culture and differentiation system for the screening of inhibiting agents of lipid accumulation in algal natural products. Inoculated preadipocytes were growing in 24-well plate of FBS-DMEM medium. After reaching confluent, culture medium were added insulin, dexamethasone and 3 isobutyl-1-mehtyl-xathine, and refreshed with new the medium containing insulin every two days afterwards. This culture system used 100 ?M genistein (27??g/ml) as positive control in comparison with the MeOH extracts of the algal cells including NP01, NP02, NP03, CR02, FE01, FE02, and FE03 of blue green algae, Ha01, Gc01, Grs04 and Pd01 of red algae, and Ec01 of brown algae. Concentrations of the algal extract applied for the screening were 27, 270 and 2700??g/ml. Lipid contents in the differentiated cells were analyzed by Oil red O staining. All the algal extracts tested were found to be of no effect like genistein in inhibition of cell differentiation and lipid accumulation. It was also found that the algal extracts showed no cytotoxicity or growth inhibition to the cells, except FE02 that showed slight growth inhibition to the cells but no cytotoxicity. Although we did not find any inhibition activity of lipid accumulation in the algal extracts, we found this culture and differentiable system of 3T3-L1 cell line were reliable and practicable for the screening of anti-obesity natural products.
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