Establishment of Bovine-Induced Pluripotent Stem Cells

Pluripotent stem cells (PSCs) have been successfully developed in many species. However, the establishment of bovine-induced pluripotent stem cells (biPSCs) has been challenging. Here we report the generation of biPSCs from bovine mesenchymal stem cells (bMSCs) by overexpression of lysine-specific demethylase 4A (KDM4A) and the other reprogramming factors OCT4, SOX2, KLF4, cMYC, LIN28, and NANOG (KdOSKMLN). These biPSCs exhibited silenced transgene expression at passage 10, and had prolonged self-renewal capacity for over 70 passages. The biPSCs have flat, primed-like PSC colony morphology in combined media of knockout serum replacement (KSR) and mTeSR, but switched to dome-shaped, naive-like PSC colony morphology in mTeSR medium and 2i/LIF with single cell colonization capacity. These cells have comparable proliferation rate to the reported primed- or naive-state human PSCs, with three-germ layer differentiation capacity and normal karyotype. Transcriptome analysis revealed a high similarity of biPSCs to reported bovine embryonic stem cells (ESCs) and embryos. The naive-like biPSCs can be incorporated into mouse embryos, with the extended capacity of integration into extra-embryonic tissues. Finally, at least 24.5% cloning efficiency could be obtained in nuclear transfer (NT) experiment using late passage biPSCs as nuclear donors. Our report represents a significant advance in the establishment of bovine PSCs.

Automated summary

This study successfully generated biPSCs with long-term self-renewal capacity and pluripotent characteristics from bovine mesenchymal stem cells. The cells exhibited morphological changes under different culture conditions, and transcriptome analysis showed high similarity to bovine ESCs and embryos. The naive-like biPSCs also demonstrated the ability to integrate into extra-embryonic tissues.