Generating high-quality porcine iPSCs with the new medium cocktail LACID.

Area of Science:

• Stem Cell Biology
• Developmental Biology
• Genetics

Background:

• Pigs are crucial models for disease research, xenotransplantation, and organogenesis.
• Generating porcine induced pluripotent stem cells (piPSCs) with desired attributes like feeder-free culture and developmental potential has been challenging.

Purpose of the Study:

• To develop an improved strategy for generating piPSCs that meet key criteria for advanced research applications.
• To establish piPSCs suitable for feeder-free culture, robust developmental potential, and nuclear transfer-mediated blastocyst development.

Main Methods:

• Utilized chemically defined medium 3 to promote epithelium-like colony formation during porcine reprogramming.
• Employed a novel medium cocktail, LACID, for subsequent reprogramming steps.
• Cultured piPSCs under feeder-free conditions and assessed their pluripotency and developmental capacity.

Main Results:

• Successfully generated piPSCs exhibiting flat morphology and capable of feeder-free culture.
• Demonstrated robust teratoma and blastoid formation, and the generation of chimeric blastocysts.
• Showcased successful CRISPR-Cas9 editing and blastocyst development following nuclear transfer with the generated piPSCs.

Conclusions:

• The newly established LACID piPSCs possess key features making them ideal for regenerative medicine applications.
• The developed method offers a significant advancement in generating functional piPSCs.
• Future improvements may lead to the generation of naive or totipotent porcine stem cells.