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.

The transcription factor POU5F1, also known as OCT4 plays critical roles in maintaining pluripotency during early mammalian embryonic development and in embryonic stem cells. It is important to establish an OCT4 promoter region-based reporter system to study pluripotency. However, there is still a lack of information about the porcine OCT4 upstream region. To improve our understanding of the porcine OCT4 regulatory region, we identified conserved regions in the porcine OCT4 promoter upstream region by sequence-based comparative analysis using various mammalian genome sequences. The similarity of nucleotide sequences in the 5' upstream region was low among mammalian species. However, the OCT4 promoter and four regulatory regions, including distal and proximal enhancer elements, had high similarity. The putative transcription factor binding sites in the Oct4 5' upstream region nucleotide sequences from mice and pigs also differed. Some of these genes are related to pluripotency, and further research will allow us to better understand the differences in species-specific pluripotency. Next, a functional analysis of the porcine OCT4 promoter region was conducted. Luciferase reporter assay results indicated that the porcine OCT4 distal enhancer and proximal enhancer were highly activated in mouse embryonic stem cells and embryonic carcinoma cells, respectively. Similar to OCT4 upstream-based reporter systems derived from other species, the porcine OCT4 upstream region-based reporter constructs showed exclusive expression patterns depending on the state of pluripotency. This work provides basic information about the porcine OCT4 upstream region and various porcine OCT4 fluorescence reporter constructs, which can be applied to study species-specific pluripotency in early embryo development and the establishment of embryonic stem cells in pigs. This research was supported by Basic Science Research Program through the National Research Foundation of Korea(NRF) funded by the Ministry of Education (NRF-2017R1D1A1B03032256).