Journal of Stem Cell Research & Therapy

Abstract

Establishment and Characterization of Novel Porcine Induced Pluripotent Stem Cells Expressing hrGFP

Yu-Jing Liao, Chia-Hsin Liao, Jiunn-Wang Liao, Kuo Yuan, Yu-Zhan Liu, Yi-Shiou Chen, Lih-Ren Chen and Jenn-Rong Yang

Induced pluripotent stem (iPS) cells have been established in various animal species since 2006. The pig is a potentially useful model in human regenerative medicine, and the characters of porcine embryonic stem (pES) cells were much similar with human embryonic stem (hES) cells. In present study, the traceable humanized recombinant green fluorescent protein expressing porcine induced pluripotent stem (piPS/hrGFP+) cells were generated from porcine ear fibroblasts (pEF) by introducing four human transcription factors (Sox2, Oct4, Klf4, and c-Myc) constructed in lentivirus vectors. The piPS/hrGFP+ cells continuously and steadily expressed hrGFP signal continuously and steadily for more than 90 passages over 20 months despite repeated subcultures. They also retained the typical defined characteristics including continuous proliferation with undifferentiated status, expression of ES pluripotent pluripotency markers (Oct4, AP, SSEA-3, SSEA-4, TRA-1-60, and TRA-1-81), and maintenance of a normal karyotype (36+XY). Three embryonic germ layers were also successfully revealed from in vitro differentiation by EB formation. Various histological analysis and immunohistochemical staining of the teratomas revealed various tissues derived from three embryonic germ layers, including neural tissues, keratin-containing epidermal tissues, skeletal muscle, smooth muscle, cartilage, adipose tissues, and glandular structures. These results support that piPS/hrGFP+ cells can be generated from pEF by direct reprogramming, and these traceable piPS/hrGFP+ cells would be beneficial for future application on cell transplantation and tissue regeneration.