Human Umbilical Cord Blood-Derived Serum for Culturing the Supportive Feeder Cells of Human Pluripotent Stem Cell Lines

Figure 5

In vitro and in vivo differentiation of human pluripotent stem cell lines cocultured with inactivated HFF-hUCS. The differentiation capacities of hPSC lines cocultured with inactivated HFF-hUCS and inactivated HFF-FBS were determined through the formation of embryoid bodies (EBs) and teratoma formation. hPSC lines cocultured with inactivated HFF-hUCS formed EBs in suspension culture similar to hPSC lines cocultured with inactivated HFF-FBS (a). After the EBs were plated into culture dishes and continuously cultured for 21 days, the EBs differentiated to embryonic germ layers, including the ectoderm (NESTIN, PAX6), mesoderm (BRACHYURY, SMA), endoderm (AFP), and the trophoblast markers (CDX2) were detected through immunostaining (b) and RT-PCR (c). hPSCs lines cocultured with inactivated HFF-hUCS formed teratoma tissue and differentiated into ectoderm (neural rosette-like structure; arrowhead), mesoderm (cartilage; star), and endoderm (gut-like structure; triangle) similar to those cocultured with inactivated HFF-FBS (d). AFP = alpha-fetoprotein, smooth muscle actin (SMA), HFF = human foreskin fibroblasts, FBS = fetal bovine serum, hUCS = human cord blood-derived serum, hPSCs = human pluripotent stem cells, DAPI = 4′,6-diamidino-2-phenylindole, and RT-PCR = reverse transcription polymerase chain reaction. Lane 1 = EB Chula2.hES-HFF-hUCS, Lane 2 = EB Chula2.hES-HFF-FBS, Lane 3 = EB HFSK#11.hiPS-HFF-hUCS, Lane 4 = EB HFSK#11.hiPS-HFF-FBS, Lane 5 = undifferentiated Chula2.hES-HFF-hUCS, Lane 6 = undifferentiated Chula2.hES-HFF-FBS, Lane 7 = undifferentiated HFSK#11.hiPS-HFF-hUCS, and Lane 8 = undifferentiated HFSK#11.hiPS-HFF-FBS. Scale bars (a) and (d) = 200 μm and (b) = 40 μm.

(a)

(b)

(c)

(d)