Comparative analysis of porcine iPSCs derived from Sertoli cells and fibroblasts

Abstract

Porcine embryonic fibroblasts (PEFs) can be directly reprogrammed into porcine induced pluripotent stem cells (piPSCs). However, the reprogramming process is generally lengthy and inefficient. Here, we established a fast and efficient induction system of piPSCs from porcine Sertoli cells (SCs) via forced expression of pig Yamanaka factors. The alkaline phosphatase (AP)-positive colonies from SCs developed on Day 3 after lentivirus infection, and were expanded and then picked up on Day 7, whereas reprogramming process from PEFs did not show any colonies in the same period. The picked piPSCs strongly expressed pluripotent genes, had the differentiation capacity to three germ layers, and could be also induced into primordial germ cell-like cells. Screening for transcription factor combinations showed that POU class 5 homeobox 1 (OCT4) is the core factor for AP-positive colony formation, and two factors (OCT4 and c-MYC) could successfully reprogram SCs into piPSCs. We then compared the RNA-sequencing data of piPSCs derived from SCs and PEFs, and found that the most significant difference was the activation of Transforming Growth Factor β signaling pathway. We also compared the RNA levels of SCs and PEFs, and found that SCs exhibited higher Wnt signaling activity and Bone Morphogenetic Protein 4 expression than PEFs, which might be correlated with higher cell proliferation rate and reprogramming efficiency. In summary, the data demonstrated that starting cell sources of piPSCs significantly affect reprogramming dynamics and SCs could serve as cell sources for efficient reprogramming.

The progress of induced pluripotent stem cells derived from pigs: a mini review of recent advances

“…These cell lines displayed conventional PSC properties and could be differentiated into three germ layers in vitro and form teratomas. Integrative reprogramming strategies have proven effective for efficiently making piPSC-like colonies from porcine somatic cells and have been used for many applications related to xenotransplantation and immunogenicity ( Park et al, 2013 ; Liu et al, 2013 ), understanding key developmental signaling ( Arai et al, 2013 ; Xu et al, 2020 ; Yang et al, 2022 ; Yuan et al, 2019 ), and deriving disease-relevant cell types ( Gu et al, 2012 ; Aravalli, Cressman, and Steer, 2012 ; Yang et al, 2013 ; Park et al, 2016 ; Liao et al, 2018 ; Yu et al, 2022 ; Liao et al, 2023 ) ( Table 1 ). However, an inevitable drawback of using integrating methods for introducing reprogramming factors is that they compromise the integrity of the host cell genome, raising their oncogenic potential ( Prigione et al, 2011 ; Chen et al, 2014 ) and limiting their translational applications ( Fan et al, 2013 ; Kang et al, 2015 ).…”