P19细胞体外向脂肪细胞诱导分化的实验

目的 探讨应用96孔板悬浮法诱导P19细胞体外向脂肪细胞分化的可行性和有效性。方法 P19细胞使用96孔板悬浮培养法制备拟胚体（EBs），其中第3天至第5天使用诱导剂全反式维甲酸（RA）。第7天挑取EBs用胰岛素和三碘甲状腺原氨酸诱导，继续贴壁培养20d后，油红O染色鉴定分化细胞的特征。
结果 使用96孔板悬浮培养法可以形成大小均一的EBs，诱导结束后EBs生长晕周围部分细胞分化为脂肪细胞。细胞形态趋于类圆或圆形，胞质中出现小脂滴，可被油红O染色清晰显示。 结论 使用96孔板悬浮法获得的P19细胞EBs在体外可诱导分化为脂肪细胞。

Differentiation of P19 cells into adipocytes in vitro

Objective To investigate the feasibility and effectiveness of the 96-well plate suspension method for differentiation of P19 cells into adipocytesin vitro.Methods P19 cells were cultivated in aggregates termed embryoid bodies (EBs) in the 96-well plate containing a thin layer of low-gelling temperature agarose for 7 days with the cultivation medium, supplemented with all-trans retinoic acid (RA) between the 2nd and the 5th day. The EBs were transferred into gelatin-coated culture dishes and cultivated for another 20 days in the presence of insulin and triiodothyronine (T3). Oil-Red-O staining was applied to indicate the generation of adipocytes. Results The 96-well plate suspending culture facilitated the formation of the EBs in similar size. At the end of this period, part of cells present in the EBs outgrowth formed adipocyte colonies, which were characterized by round in shape and small lipid droplets scattered throughout cytoplasm. Adipocytes were stained with Oil Red O for fat droplets. Conclusion P19 cell-derived EBs formed in 96-well plate suspension could undergo adipocyte differentiation in vitro.