Efficient mRNA delivery with graphene oxide-polyethylenimine for generation of footprint-free human induced pluripotent stem cells.
Clinical applications of induced pluripotent stem cells (iPSCs) require development of technologies for the production of "footprint-free" (gene integration-free) iPSCs, which avoid the potential risk of insertional mutagenesis in humans. Previously, several studies have shown that mRNA transfer can generate "footprint-free" iPSCs, but these studies did not use a delivery vehicle and thus repetitive daily transfection was required because of mRNA degradation. Here, we report an mRNA delivery system employing graphene oxide (GO)-polyethylenimine (PEI) complexes for the efficient generation of "footprint-free" iPSCs. GO-PEI complexes were found to be very effective for loading mRNA of reprogramming transcription factors and protection from mRNA degradation by RNase. Dynamic suspension cultures of GO-PEI/RNA complexes-treated cells dramatically increased the reprogramming efficiency and successfully generated rat and human iPSCs from adult adipose tissue-derived fibroblasts without repetitive daily transfection. The iPSCs showed all the hallmarks of pluripotent stem cells including expression of pluripotency genes, epigenetic reprogramming, and differentiation into the three germ layers. These results demonstrate that mRNA delivery using GO-PEI-RNA complexes can efficiently generate "footprint-free" iPSCs, which may advance the translation of iPSC technology into the clinical settings.
Department of Animal Biotechnology (Stem Cell & Regenerative Biotechnology), Animal Resources Research Center, Incurable Disease Animal Model and Stem Cell Institute (IDASI), Konkuk University, 120 Neungdong-ro, Gwangjin-gu, Seoul 05029, Republic of Korea.
Hye Yeon
HY
Choi
School of Chemical and Biological Engineering, Seoul National University, 1 Gwanak-ro, Gwanak-gu, Seoul 08826, Republic of Korea; Department of Pathology and Immunology, Washington University School of Medicine, MO, USA.
Tae-Jin
TJ
Lee
Department of Animal Biotechnology (Stem Cell & Regenerative Biotechnology), Animal Resources Research Center, Incurable Disease Animal Model and Stem Cell Institute (IDASI), Konkuk University, 120 Neungdong-ro, Gwangjin-gu, Seoul 05029, Republic of Korea.
Gwang-Mo
GM
Yang
School of Chemical and Biological Engineering, Seoul National University, 1 Gwanak-ro, Gwanak-gu, Seoul 08826, Republic of Korea.
Jaesur
J
Oh
Department of Animal Biotechnology (Stem Cell & Regenerative Biotechnology), Animal Resources Research Center, Incurable Disease Animal Model and Stem Cell Institute (IDASI), Konkuk University, 120 Neungdong-ro, Gwangjin-gu, Seoul 05029, Republic of Korea.
Jihye
J
Won
Department of Animal Biotechnology (Stem Cell & Regenerative Biotechnology), Animal Resources Research Center, Incurable Disease Animal Model and Stem Cell Institute (IDASI), Konkuk University, 120 Neungdong-ro, Gwangjin-gu, Seoul 05029, Republic of Korea.
Jihae
J
Han
School of Chemical and Biological Engineering, Seoul National University, 1 Gwanak-ro, Gwanak-gu, Seoul 08826, Republic of Korea.
Gun-Jae
GJ
Jeong
Department of Biomedical Engineering, Dongguk University, Seoul 100-715, Republic of Korea.
Jongpil
J
Kim
Department of Animal Biotechnology (Stem Cell & Regenerative Biotechnology), Animal Resources Research Center, Incurable Disease Animal Model and Stem Cell Institute (IDASI), Konkuk University, 120 Neungdong-ro, Gwangjin-gu, Seoul 05029, Republic of Korea.
Jin-Hoi
JH
Kim
School of Chemical and Biological Engineering, Seoul National University, 1 Gwanak-ro, Gwanak-gu, Seoul 08826, Republic of Korea; Institute of Chemical Processes, Seoul National University, 1 Gwanak-ro, Gwanak-gu, Seoul 08826, Republic of Korea. Electronic address: byungskim@snu.ac.kr.
Byung-Soo
BS
Kim
Department of Animal Biotechnology (Stem Cell & Regenerative Biotechnology), Animal Resources Research Center, Incurable Disease Animal Model and Stem Cell Institute (IDASI), Konkuk University, 120 Neungdong-ro, Gwangjin-gu, Seoul 05029, Republic of Korea. Electronic address: ssangoo@konkuk.ac.kr.
Ssang-Goo
SG
Cho
D000273:Adipose Tissue / Q000166:cytology; D000469:Alkaline Phosphatase / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D005347:Fibroblasts / Q000166:cytology; D006108:Graphite / Q000008:administration & dosage*; D057809:HEK293 Cells; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology*; D010087:Oxides / Q000008:administration & dosage*; D011094:Polyethyleneimine / Q000008:administration & dosage*; D012333:RNA, Messenger / Q000008:administration & dosage*
D010087:Oxides; D012333:RNA, Messenger; D006108:Graphite; D011094:Polyethyleneimine; D000469:Alkaline Phosphatase
Footprint-free transgene-free; Gene delivery; Graphene oxide-polyethylenimine complex; Human induced pluripotent stem cells; Integration-free; RNA delivery; iPSC