Initiation of a zebrafish blastula cell line on rainbow trout stromal cells and subsequent development under feeder-free conditions into a cell line, ZEB2J.
Initiation of a zebrafish blastula cell line on rainbow trout stromal cells and subsequent development under feeder-free conditions into a cell line, ZEB2J.
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在虹鳟基质细胞上启动斑马鱼囊胚细胞系，并随后在无饲养条件下发育成细胞系ZEB2J。
DOI：
10.1089/zeb.2007.0512
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发表时间：
2008
期刊：
影响因子：
2
通讯作者：
Bols,NielsC
中科院分区：
文献类型：
--
作者：
Xing,JerryG;Lee,LucyEJ;Fan,Lianchun;Collodi,Paul;Holt,ShawnE;Bols,NielsC
从斑马鱼囊胚期胚胎中建立了表达增强型绿色荧光蛋白（GFP）的连续细胞系ZEB 2。最初，虹鳟鱼脾细胞系RTS 34 st用作饲养细胞，以通过几代来启动和维持细胞。ZEB 2在没有饲养层的情况下在含有15%胎牛血清（FBS）的L-15中生长2年，用于120个群体倍增。这种新的细胞系ZEB 2 J是异倍体的，具有可检测的端粒酶活性，并且是粘附的。在生长成单层后，一些细胞继续生长成土丘。培养物表达Pou-2 mRNA，并含有许多碱性磷酸酶和一些阶段特异性胚胎抗原-1阳性细胞。在用磷脂聚合物（2-甲基丙烯酰氧基乙基磷酰胆碱，MPC）包被的皿中，ZEB 2 J形成球形聚集体。聚集体附着在常规培养塑料上，并且从聚集体中出现的大多数细胞具有典型的上皮样形状的ZEB 2 J，这表明ZEB 2 J具有有限的分化潜力，尽管表达一些干细胞特性。ZEB 2 J的荧光允许与饲养细胞的关系进行研究。在MPC培养皿中，ZEB 2 J与RTS 34 st形成混合球状体。在贴壁共培养物中，RTS 34 st和其他鱼细胞系强烈刺激ZEB 2 J生长，这可以具体地定量，因为ZEB 2 J表达GFP。ZEB 2 J可用于优化斑马鱼胚胎干细胞的培养条件。
A continuous cell line, ZEB2, was developed from zebrafish blastula-stage embryos expressing enhanced green fluorescent protein (GFP). Originally the rainbow trout spleen cell line, RTS34st, was used as feeders to initiate and maintain the cells through several passages. ZEB2 was then grown for 2 years without feeders in L-15 with 15% fetal bovine serum (FBS) for 120 population doublings. This new cell line, ZEB2J, was heteroploid, had detectable telomerase activity, and was adherent. After growing into monolayers, some cells continued to grow into mounds. Cultures expressed Pou-2 mRNA and contained many alkaline phosphatase and a few stage-specific embryonic antigen-1–positive cells. In dishes coated with a phospholipid polymer (2-methacryloxyloxyethyl phosphorylcholine, MPC), ZEB2J formed spherical aggregates. Aggregates attached to conventional culture plastic, and most cells that emerged from aggregates had typical epithelial-like shapes of ZEB2J, which suggests that ZEB2J had limited differentiation potential, despite expressing some stem cell properties. The fluorescence of ZEB2J allowed relationships with feeder cells to be studied. In MPC dishes, ZEB2J formed mixed spheroids with RTS34st. In adherent cocultures, RTS34st and other fish cell lines strongly stimulated the ZEB2J growth, which could be quantified specifically because ZEB2J expressed GFP. ZEB2J should be useful for optimizing culture conditions for zebrafish embryonic stem cells.