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Table 1 Frequently used growth media constituents, their working mechanisms, effects and applications

From: Organoid technology in female reproductive biomedicine

   Mechanism of action Function Significance in reproductive organoid cultures Ref
Frequently used factors Noggin Binds and inactivates members of the TGF-β superfamily signaling proteins, such as BMP4. Allows for long-term expansion of organoids by preventing differentiation. The lack of Noggin resulted in reduced numbers and/or smaller human EMO.
The lack of Noggin growth markedly reduced the expansion and passageability of mouse EMO.
Limited differentiation and promoted trophoblast survival in CTB-orgs.
[3,4,5]
RSPO1 Interacts with WNT4 in the process of female sex development.
Potentiates the cellular response to Wnts.
Facilitates the growth, expansion, and long-term culture of organoids.
Plays a crucial role in formation and stem cell maintenance of FTO.
The lack of RSPO1 resulted in reduced numbers and/or smaller human EMO.
Removal of RSPO1 alone did not affect mouse EMO formation efficiency.
The lack of RSPO1 markedly reduced growth, expansion and passageability of mouse EMO.
RSPO1 was required for efficient, long-term human EMO expansion.
In the absence of RSPO1, human organoids could no longer be passaged after P3.
Addition of RSPO1 increased the FTO size.
RSPO1 is not absolutely required for long-term maintenance of mouse FTE organoids.
Addition of RSPO1 increased differentiation of FTE organoids toward the ciliated lineage.
Withdrawal of RSPO1 promoted trophoblast outgrowth from the outer CTB layers and expression of HLA-G at distal sites from the CTB-orgs.
Critical for maintenance of CTB-orgs.
[4, 5, 22, 23]
WNT3A The ligand of the canonical Wnt signaling pathway,
Interacts with the LRP6/Frizzled receptor complex
Crucial for maintenance of stable growth over time. The lack of WNT3A markedly reduced growth, expansion and passageability of mouse EMO.
The presence of WNT3A (alone or together with RSPO1) enhanced the efficiency of mouse EMO formation.
WNT3A was not needed for further expansion and passaging of human EMO.
[4]
HGF Activates a tyrosine kinase signaling cascade after binding to the proto-oncogene c-Met receptor Epithelial-cell mitogen. The lack of HGF resulted in reduced numbers and/or smaller human EMO.
Limit differentiation and promote trophoblast survival in CTB-orgs
[4, 5]
EGF Activates the RAS/RAF/MEK/ERK signaling pathway Epithelial-cell mitogen.
Crucial for maintenance of stable growth over time.
Supports proliferation and differentiation of FTE cells.
The lack of EGF resulted in reduced numbers and/or smaller human EMO.
The lack of EGF resulted in markedly reduced growth, expansion, and passageability of mouse EMO.
Required for long-term expansion of CTB-orgs.
[4, 5]
FGF10 Acts mostly on the epithelium via Fgfr2b. Epithelial-cell mitogens.   
Prostaglandin E2 Binding and activation of the prostaglandin E2 receptor.   Critical for maintenance of CTB-orgs. [5]
Nicotinamide A form of vitamin B3.   Withdrawal of nicotinamide had the strongest effect on the numbers of EMO that formed. [3]
Molecule inhibitors Y27632 Inhibits anoikis of dissociated cells.   Important for long-term culture and passaging of organoids.  
A-83-01 Alk3/4/5 inhibitor.
Blocks the TGF-β pathway.
Maintains epithelial-cell features The lack of A-83-01 resulted in reduced numbers and/or smaller human EMO.
Critical for maintenance of CTB-ORGs.
Required for long-term expansion of CTB-orgs.
[4, 5]
SB202190 p38 inhibitor.   Decreasing concentrations of p38i were beneficial for long-term expansion of endometrial cancer organoids. [24]
SB431542 TGF-β R kinase inhibitor IV. Crucial for quasi-indefinite expansion of FTO. Without TGF-β, RK inhibitor FTO had slower expansion and finally growth arrest by four to six passages (three to four months).
Important for formation and maintenance of large FTE organoids.
[23, 25]
CHIR99021 Inhibitor of GSK3.   Critical for maintenance of CTB-orgs. [5]
  1. EMO Endometrial organoid, FTO Fallopian tube organoid, CTO Cytotrophoblast, BMP4 Bone morphogenetic protein-4, RSPO1 R-spondin-1, Fgfr2b Fibroblast growth factor receptor 2b, FTE Fallopian tube epithelium, HGF Hepatocyte growth factor