As a preclinical study, many researchers have been attempted to convert the porcine PSCs into several differentiated cells with transplantation of the differentiated cells into the pigs. Here, we attempted to derive neuronal progenitor cells from pig embryonic germ cells (EGCs). As a result, neuronal progenitor cells could be derived directly from pig embryonic germ cells through the serum-free floating culture of EB-like aggregates (SFEB) method. Treating retinoic acid was more efficient for inducing neuronal lineages from EGCs rather than inhibiting SMAD signaling. The differentiated cells expressed neuronal markers such as PAX6, NESTIN, and SOX1 as determined by qRT-PCR and immunostaining. These data indicated that pig EGCs could provide valid models for human therapy. Finally, it is suggested that developing transgenic pig for disease models as well as differentiation methods will provide basic preclinical data for human regenerative medicine and lead to the success of stem cell therapy.

Stem cells are progenitor cells that are capable of self-renewal and differentiation into various cells. Especially, pluripotent stem cells (PSCs) have in vivo and in vitro differentiation capacity into three germ layers and can proliferate infinitely. The differentiation ability of PSCs can be applied for regenerative medicine and tissue engineering. In domestic animals, their PSCs have a potential for preclinical therapy as well as the production of transgenic animals and agricultural usage such as cultured meat. Among several domestic animals, a pig is considered as an ideal model for biomedical and agricultural purposes mentioned above. In this reason, studies for pig PSCs including embryonic stem cells (ESCs), embryonic germ cells (EGCs) and induced pluripotent stem cells (iPSCs) have been conducted for decades. Therefore, this review will discuss the history of PSCs derived from various origins and recent progress in pig PSC research field.

Because the pig is a valuable candidate for a preclinical model of human cell therapy as well as an important food source, understanding a physiology of pig myogenic progenitors such as skeletal muscle satellite cells and myoblasts is required for cure of muscular diseases and improvement of meat production. For these reasons, we tried to isolate and culture the pig progenitor cells from skeletal muscle. Pig satellite cells, known as muscle stem cells, were isolated from biceps femoris of neonatal pigs by enzymatic digestion method. Muscle satellite cells are quiescent in uninjured muscle. Upon injury, they are activated into proliferating state, known as myoblasts, by growth factors and, in turn, differentiated forward to myocytes and myotubes. To trigger proliferation in vitro, the isolated satellite cells were cultured with epidermal growth factor (EGF) and dexamethasone (BMP4 inhibitor). As a result, the pig satellite cells were efficiently converted into proliferating myoblasts and stably maintained over an extended period. The myoblasts were confirmed by markers of PAX7, MYF5, and MYOD1. Our finding showed that modulating EGF and BMP4 signaling are essential for maintaining muscle stem cells. This culture method could be applied for a production of cultured meat and further basic research of muscle development. This work was supported by the Korea Institute of Planning and Evaluation for Technology in food, agriculture, forestry, and fisheries (IPET) through the Development of High Value-Added Food Technology Program funded by the Ministry of Agriculture, Food, and Rural Affairs (MAFRA; 118042-03-1-HD020).

Pluripotent cells are categorized as either "naive" or "primed" based upon their pluripotent status. According to previous studies, embryonic stem cells and embryonic germ cells are identified as naive pluripotent stem cells and epiblast stem cells are identified as primed pluripotent stem cells. In a permissive species such as the mouse, naive and primed pluripotent stem cells can be derived from embryos without genetic manipulations. In non-permissive species such as humans and pigs, primed pluripotent cells are only established from embryos. However, previous studies have shown that the embryonic germ cells of non-permissive species share similar morphology and features with naive pluripotent cells. For these reasons porcine embryonic germ cells (pEGCs) may provide a useful cell source for comparative studies on naive pluripotent cells in non-permissive species. In this study, we attempted to establish and characterize porcine embryonic germ cells. Consequently, an embryonic germ cell line was derived from the genital ridges of a porcine dpc 30 fetus in media containing LIF and bFGF. After establishment, this cells were cultured and stabilized in LIF or bFGF contained media. This cell lines displayed a dome-shaped colony morphology in both culture condition. The cell lines were maintained in both condition over an extended time period and were able to differentiate into the three germ layers in vitro. Interestingly, cell lines cultured in LIF or bFGF expressed different pluripotency markers. LIF-dependent pEGCs expressed naive-pluripotency markers such as OCT4, SOX2, NANOG and SSEA1, while bFGF-dependent pEGCs expressed primed-pluripotency markers such as OCT4, SOX2, NANOG and SSEA4. However, as a result of analysis of XCI, two cell lines showed hemi-methylated pattern similarly in XIST promoter regions. In conclusion, we were able to successfully derive embryonic germ cells from genital ridges of a porcine fetus. Pluripotent state of pEGCs were regulated by modulation of culture condition. In LIF supplement, pEGCs showed naive-pluripotency expressing SSEA1, while pEGCs show primed-pluripotency expressing SSEA4 in bFGF condition. This cell line could potentially be used as naive pluripotent cell source for comparative study with porcine embryonic stem cells and other pluripotent cell lines. As porcine pluripotent cells, pEGCs could be useful candidates for preliminary studies of human disease as well as a source for generating transgenic animals.

Pluripotent cells are categorized as either "naive" or "primed" based upon their pluripotent status. According to previous studies, embryonic stem cells and embryonic germ cells are identified as naive pluripotent stem cells and epiblast stem cells are identified as primed pluripotent stem cells. In a permissive species such as the mouse, naive and primed pluripotent stem cells can be derived from embryos without genetic manipulations. In non-permissive species such as humans and pigs, primed pluripotent cells are only established from embryos. However, previous studies have shown that the embryonic germ cells of non-permissive species share similar morphology and features with naive pluripotent cells. For these reasons porcine embryonic germ cells (pEGCs) may provide a useful cell source for comparative studies on naive pluripotent cells in non-permissive species. In this study, we attempted to establish and characterize porcine embryonic germ cells. Consequently, an embryonic germ cell line was derived from the genital ridges of a porcine dpc 30 fetus in media containing bFGF. This cell line displayed a dome-shaped colony morphology. The cell line was maintained in a stable condition over an extended time period and was able to differentiate into the three germ layers in vitro. Pluripotency markers such as OCT4, SOX2, NANOG and SSEA4 were expressed in these pEGCs. Similar with pESCs, Mek/Erk signaling pathway were activated by bFGF in the cultured pEGCs. In conclusion, we were able to successfully derive embryonic germ cells from genital ridges of a porcine fetus. Unlikely naive pluripotent cells such as mESCs, pluripotency of pEGCs were regulated by Mek/Erk signaling pathway activated by bFGF. This cell line could potentially be used as naive pluripotent cell source for comparative study with porcine embryonic stem cells and other pluripotent cell lines. As porcine pluripotent cells, pEGCs could be useful candidates for preliminary studies of human disease as well as a source for generating transgenic animals.