Enhanced hepatic differentiation in the subpopulation of human amniotic stem cells under 3D multicellular microenvironment

Table 1 List of primers

Name		Primer sequence
SOX2	F	TGCACCGCTACGACGTGA
	R	GGAGCCAAGAGCCATGCC
NANOG	F	TGCTGAGATGCCTCACACG
	R	TGCAGAAGTGGGTTGTTTGC
OCT4	F	GAAACCCACACTGCAGCAG
	R	GACCCAGCAGCCTCAAAATC
LIN28	F	GGATGTCTTTGTGCACCAGAGTA
	R	TGGATTCCAGACCCTTGGCT
ALB	F	TGCCTGTTGCCAAAGCTCG
	R	GCTACTGCCCATGCTTTGAAAG
AFP	F	GCAAACGATGAAGCAAGAGTTTC
	R	GCAGCATTTCTCCAACAGGC
AAT	F	ACTGGAACCTATGATCTGAAGAGC
	R	GCCTTATGCACGGCCTTGG
CYP3A4	F	GGATGAAAGAAAGTCGCCTCGA
	R	TCCAGATCGGACAGAGCTTTG
ACTB	F	CCTCGCCTTTGCCGATCC
	R	CATGCCGGAGCCGTTGT

Table 2 RNAseq data sources used in the bioinformatics analysis

Sample name	SRA number	Instrument	Memo	Ref.
AEC-1	SRR9643783	Illumina HiSeq 2500	Primary AEC
AEC-2	SRR9643784	Illumina HiSeq 2500	Primary AEC
MSC-1	SRR6431450	Illumina HiSeq 2000		[44]
MSC-2	SRR6431451	Illumina HiSeq 2000		[44]
hESC-1	SRR4241924	Illumina HiSeq 4000	H9	[45]
hESC-2	SRR4241926	Illumina HiSeq 4000	H9	[45]
NiPS-1	SRR7592168	Illumina HiSeq 2500	Normal human iPSC	[46]
NiPS-2	SRR7592169	Illumina HiSeq 2500	Normal human iPSC	[46]
DE-1	SRR771468	Illumina HiSeq 2000	Definitive endoderm induced from H9	[47]
DE-2	SRR771469	Illumina HiSeq 2000	Definitive endoderm induced from H9	[47]
hiHep-1	SRR5974291	Illumina HiSeq 2000	Umbilical cord fibroblast derived hepatocyte-like like cell	[8]
hiHep-2	SRR5974292	Illumina HiSeq 2000	Umbilical cord fibroblast derived hepatocyte-like cell	[8]
iPSCHLC-1	SRR5974295	Illumina HiSeq 2500	iPSC-derived Hepatocyte-like cell	[8]
iPSCHLC-2	SRR5974296	Illumina HiSeq 2500	iPSC-derived hepatocyte-like cell	[8]
Hepa-1	SRR6176953	Illumina HiSeq 2500	Hepatocyte from clinical sample of adult	[48]
Hepa-2	SRR6176948	Illumina HiSeq 2500	Hepatocyte from clinical sample of adult	[48]
Hepa-3	SRR5974298	Illumina HiSeq 2000	Primary human hepatocyte 2 d	[8]
Hepa-4	SRR5974299	Illumina HiSeq 2000	Primary human hepatocyte 4 d	[8]

Table 3 Stemness (upper)- and hepatic (lower) genes expressed in amniotic epithelial cells.

Gene symbol	Explanation	Ref.
CD9	CD9 belongs to the transmembrane 4 superfamily. It is associated with cell proliferation, motility, and adhesion and regulates hematopoietic differentiation	[49]
TJP1	Tight junction protein 1 is a determinant of plasma cell proteasome and is associated with EGFR, JAK1, and STAT3. It is regulated by TGF-b and involved with cell motility	[30,50]
IGF2BP2	This member of the insulin–like growth factor 2 mRNA-binding protein family participates in normal embryonic growth and development. It is expressed in the pancreas and associated with type 2 diabetes mellitus	[51]
KRT19	Keratin 19 is a cytoplasmic intermediate filament protein and belongs to the type 1 keratin family. It is used as a cholangiocyte marker and is not expressed in hepatocytes. It is associated with the progression of several cancers	[31,32]
GRB7	Growth factor receptor-bound protein 7 mediates signal transduction and cell migration. It is associated with the metastasis of several cancers	[52]
KRT8	Keratin 8 is a cytoplasmic intermediate filament protein of the type 2 keratin family. It is expressed in single layered epithelial cells. In cancer cells, it is associated with progression in the form of migration and adhesion	[33]
Gene symbol	Explanation	Ref.
ApoM	Apolipoprotein M (apoM) belongs to the lipocalin family. It is expressed in the liver and kidney. Hepatic apoM controls HDL metabolism	[53,54]
CPS1	Carbamoyl phosphate synthase 1 is a mitochondrial enzyme and participates in the first step of the urea cycle in the liver.	[55]
SLC51A	Organic solute transporter subunit alpha is a bile acid transporter in the liver, small intestine, and kidney. It prevents the bile acid reflux	[56]
IL6ST	Interleukin 6 signal transducer controls IL-6 and other cytokines such as IL-11, IL-27, oncostatin M, and leukemia inhibitory factor	[57]
ACOX1	Acyl-CoA oxidase 1 is a rate-limiting enzyme in fatty acid β-oxidation	[58]
RXRa	The nuclear hormone receptor retinoid X receptor belongs to the steroid hormone receptor family. It is a key factor of cholesterol synthesis	[59]
MET	MET encodes the hepatocyte growth factor (HGF) receptor and the key factor of hepatic regeneration. It activates epithelial migration and 3D morphogenesis	[35,36]
ABCC2	ATP binding cassette subfamily C member 2 is expressed in the hepatocytes and is a biliary transporter. It is also related to drug elimination and multidrug resistance in several cancers	[60]
CYP1A1	This member of the cytochrome P450 enzyme superfamily participates in fatty acid and steroid metabolism. It is associated with the detoxification of anthropogenic chemicals such as polycyclic aromatic hydrocarbons	[61]