Supplementary Data
Materials and Methods
Microarray analysis
Based on the BeadArray technology CD34+ stem cells from UCB were analyzed with a
Sentrix HumanRef8 v2 Exoression BeadChip (Illumina, USA) and a selected marker profile.
Endodermal/hepatic and cell-cell communication related genes were determined after a
cultivation period of 10 days. RNA isolation Illumina TotalPrep Amplification Kit was used
concerning manufacturing instructions for RNA preparation. Generation of biotin labeld
cRNA, hybridisation and Cy3 streptavidin labelling are performed concerning available
protocols from Illumina.
Video imaging of co-culture
UCBC and AML12 cells were labeled as described in Material and Methods (Dye Transfer).
3x105 CalceinAM labelled AML12 cells were seeded out in a 24well plate. Co-culture was
initiated after adhesion period of 24h by adding 1,5x105 pkH26 labelled UCBC. Co-culture
was visualized by using an inverse microscope and AxioCamMR camera (Carl Zeiss
Microimaging) and analyzed with AxioCamMR software. Co-cultured was monitored by
video imaging over 24 h using a time interval of 5 min.
Tables
Table S1. Applied Biosystems TaqMan Gene Expression Assays® for real time
PCR.
Assay ID
Gene name
Gene
symbol
Tissue/ Specificity
(Applied
Biosystem)
Hs00173490_m1
Hs00609411_m1
AFP, hCG15197
ALB, hCG14967
Hs00702141_s1
GJB1, CMTX, CMTX1 Cx32
Hs00748445_s1
GJA1, AVSD3,
DFNB38, GJAL
Cx43
Hs00156373_m1
CD34, hCG21280
CD34+
human CD34+ adult stem cells
Hs00269972_s1
C/EBPA, hCG20142
C/EBP
adult liver, hepatocytes
CK8
epithelial tissue, cell differentiation,
structural integrity of cells
CK18
early epithelial tissue,
Hs01630795_s1
Hs01920599_gh
KRT8, CARD2, CK-8,
CK8, CYK8
KRT18, CYK18, K18,
PIG46
AFP
ALB
Hs00761767_s1
KRT19, CK19, K19,
K1CS
Hs00170423_m1
CDH1, cadherin 1, type
E-Cadherin
1
Hs00232764_m1
FOXA2
Hs00171403_m1
GATA 4, ASD2, VSD1 GATA 4
Hs00230853_m1
Hs01895061_u1
Hs00751752_s1
Hs99999905_m1
HNF4A
POU5F1
SOX17
GAPDH, hCG2005673
CK19
HNF3
HNF4
Oct-4
Sox17
GAPDH
fetal liver, endodermspecific
adult liver, hepatocytes
gap junction protein, tumorigenesis and
liver regeneration
gap junction protein, heart
development, cell-cell communication
structural integrity of epithelial cells,
cell differentiation involved in
embryonic placenta development
intracellular communication,
cell-cell adhesion (epithelial tissue),
tumorigenesis
fetal liver, definitive endoderm
embryogenesis, myocardial
differentiation and function, lineage
determination
adult liver, hepatocytes
pluripotency
fetal liver, definitive endoderm
house keeping gene
The following cycling conditions were used for all of the above primer assays.
95°C 10 min.
Denaturation: 95°C 15 sec
} 45 cycles
Amplification: 60°C
Figures
Figure S1. Video imaging of co-cultured AML12 and umbilical cord blood cells (CD34+
stem cells).
Over 24h co-culture of CalceinAM labelled murine AML12 (green) and pkH26 labelled
UCBC (red) was monitored by video imaging using a time interval of 5 min (20x
magnification). The video demonstrated the rare and sporadic cell-cell contact between the
two different cell types.