Pluripotent stem cells derived from mouse primordial germ cells by small molecule compounds

Area of Science:

• Stem Cell Biology
• Developmental Biology
• Epigenetics

Background:

• Primordial germ cells (PGCs) naturally form embryonic germ cells (EGCs) with specific growth factors.
• Somatic cells can be reprogrammed into induced pluripotent stem cells (iPSCs) using transcription factors or small molecules.
• Small molecules like TGFβR inhibitor and Kempaullone can mimic reprogramming factors Sox2 and Klf4.

Purpose of the Study:

• To investigate the effect of TGFβR inhibitor and Kempaullone on deriving pluripotent stem cells from PGCs.
• To determine if these small molecules can generate induced EGCs (iEGCs) under simplified culture conditions.
• To analyze the DNA methylation patterns of imprinted and germline genes in the newly derived iEGCs.

Main Methods:

• Treatment of PGCs with TGFβR inhibitor and/or Kempaullone.
• Culture of treated PGCs under standard embryonic stem cell (ESC) conditions without bFGF and SCF.
• Analysis of iEGC derivation efficiency based on PGC differentiation stage and sex.
• Assessment of DNA methylation levels in imprinted genes (e.g., H19) and germline development genes.

Main Results:

• Pluripotent stem cells, termed iEGCs, were successfully generated from PGCs using TGFβR inhibitor and Kempaullone.
• iEGC derivation efficiency varied with PGC differentiation stage and sex.
• DNA methylation was reduced in most imprinted genes in iEGCs, except for H19.
• While germline development gene promoters were generally hypomethylated in PGCs, three genes showed similar methylation levels across iEGCs, ESCs, and iPSCs.

Conclusions:

• PGCs can be reprogrammed into a pluripotent state using small molecule compounds, creating iEGCs.
• This small molecule-driven reprogramming eliminates the need for specific growth factors like bFGF and SCF.
• DNA methylation patterns of key germline genes are not fully maintained during small molecule-induced reprogramming of PGCs into iEGCs.