An In Vitro Differentiation Protocol for Human Embryonic Bipotential Gonad and Testis Cell Development

Area of Science:

• Developmental Biology
• Stem Cell Biology
• Reproductive Biology

Background:

• A fully functional in vitro model of the human embryonic gonad is currently unavailable.
• Understanding human gonadal development is crucial for addressing disorders of sex development.

Purpose of the Study:

• To establish a feeder-free protocol for generating human testis organoids from induced pluripotent stem cells.
• To create a reliable in vitro model for studying early human gonadogenesis.

Main Methods:

• Utilized a stepwise protocol with small molecules to mimic embryonic development.
• Induced bipotential gonad markers (LHX9, EMX2, GATA4, WT1) by day 10.
• Induced testis Sertoli cell markers (SOX9, WT1, AMH) by day 15.
• Aggregated cells into 3D structures and cultured as organoids on Transwell filters.

Main Results:

• Successfully generated early testis-like cells from human induced pluripotent stem cells.
• Achieved organoids with defined tissue structures and distinct Sertoli cell marker expression.
• Demonstrated upregulation of key gonad and Sertoli cell markers at specific time points.

Conclusions:

• The developed protocol provides a significant advancement in creating human gonad organoids.
• The findings offer insights into human gonadal development, suggesting lateral mesodermal origins for precursor cells.
• This model is essential for future research into disorders/differences of sex development.