Abstract
Although the pig is considered an important model of human disease and an ideal animal for the preclinical testing of cell transplantation, the utility of this model has been hampered by a lack of genuine porcine embryonic stem cells. Here, we derived a porcine pluripotent stem cell (pPSC) line from day 5.5 blastocysts in a newly developed culture system based on MXV medium and a 5% oxygen atmosphere. The pPSCs had been passaged more than 75 times over two years, and the morphology of the colony was similar to that of human embryonic stem cells. Characterization and assessment showed that the pPSCs were alkaline phosphatase (AKP) positive, possessed normal karyotypes and expressed classic pluripotent markers, including OCT4, SOX2 and NANOG. In vitro differentiation through embryonic body formation and in vivo differentiation via teratoma formation in nude mice demonstrated that the pPSCs could differentiate into cells of the three germ layers. The pPSCs transfected with fuw-DsRed (pPSC-FDs) could be passaged with a stable expression of both DsRed and pluripotent markers. Notably, when pPSC-FDs were used as donor cells for somatic nuclear transfer, 11.52% of the reconstructed embryos developed into blastocysts, which was not significantly different from that of the reconstructed embryos derived from porcine embryonic fibroblasts. When pPSC-FDs were injected into day 4.5 blastocysts, they became involved in the in vitro embryonic development and contributed to the viscera of foetuses at day 50 of pregnancy as well as the developed placenta after the chimeric blastocysts were transferred into recipients. These findings indicated that the pPSCs were porcine pluripotent cells; that this would be a useful cell line for porcine genetic engineering and a valuable cell line for clarifying the molecular mechanism of pluripotency regulation in pigs.
Highlights
The pig is an important farm animal and a useful experimental model for human disease due to its obvious physiological and immunological similarity with humans [1,2,3]
Efforts have been made on establishing porcine ESCs since the first group of reports about porcine ESC-like cell lines in 1990 [4,5,6,7,8], but no bona fide embryonic stem cell (ESC) lines that could fulfil all the characterization demands that mouse ESCs do have been established in pigs in past decades
To obtain porcine pluripotent stem cell lines from early embryos, provide an opportunity to clarify the molecular mechanism of porcine pluripotency regulation, and obtain materials for porcine genetic engineering, we used in vitro fertilization (IVF) blastocysts as an embryo resource, which had advantages in the selection of precise embryo development stages for seeding; we developed a new culture medium named MXV containing both hLIF and bFGF as a basic culture system in an atmosphere of 5% oxygen
Summary
The pig is an important farm animal and a useful experimental model for human disease due to its obvious physiological and immunological similarity with humans [1,2,3]. The pig holds great potential for testing the safety of clinical stem cell transfer and related techniques. Embryonic stem cells (ESCs) have offered a wide range of cellular resources for developmental research and clinical applications. The limited proliferation potency of most of the established porcine ESC-like cell lines prevented thorough characterization, except for the characterization of morphology and a few pluripotency-related markers, such as AKP, OCT4 and SOX2. This situation becomes even more complicated when the lack of validated antibodies and other related testing techniques is considered
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
