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Highly purified hepatocytes derived from human pluripotent stem cells using EpCAM and indomonocarbocyanine

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한국발생생물학회 (The Korea Society Of Developmental Biology)
초록

Hepatocytes and hepatic progenitors derived from human ES cells may be a useful source for clinical application. Therefore, identification and purification of these cell types would be following important issues. There are very few candidate surface markers that can be used to identify and purify hepatic progenitor cells. In addition, indocyanine-green can be uptaken by mature hepatocytes, but cannot be applied for fluorescence activated cell sorting (FACS) due to its long emission wavelength. In the present study, we tested EpCAM as a potential marker for magnetic-activated cell sorting (MACS) of hepatic progenitors and also modified indocyanine-green into fluorescent indomonocarbocyanine for FACS-mediated sorting of mature hepatocytes after differentiation of human ES cells. Hepatic progenitor cells were sorted by MACS after incubation with anti-human EpCAM antibodies. After the final differentiation, the differentiated cells and mouse primary hepatocytes (control group) were incubated with indomonocarbocyanine and were sorted by FACS. MACS and immunocytochemistry data showed that approximately 45% of differentiated cells were EpCAM-positive cells. EpCAM-positive cells expressed α-fetoprotein, FOXa2, HnF4a, and CK18. Differentiation efficiency into albumin-positive cells was significantly higher in EpCAM-positive cells, compared to EpCAM-negative cells. Importantly, indomonocarbocyanine successfully stained cells that expressed ALB. Furthermore, FACS analysis data showed that the purity of hepatocytes that expressed albumin was significantly increased after purification of indomonocarbocyanine-positive cells. Our data demonstrated that human ES cell-derived hepatic progenitors can be efficiently isolated by MACS using EpCAM antibody. In addition, we also showed that indomonocarbocyanine can be successfully used to identify and purify mature hepatocytes using FACS.

저자
  • Ji Young Park(Laboratory of Stem Cells & Tissue Regeneration, Division of Biotechnology, College of Life Sciences & Biotechnology, Korea University)
  • Jiyou Han(Laboratory of Stem Cells & Tissue Regeneration, Division of Biotechnology, College of Life Sciences & Biotechnology, Korea University)
  • Soo Jin Uhm(Department of Chemistry, Korea University)
  • Jong Hyun Kim(Laboratory of Stem Cells & Tissue Regeneration, Division of Biotechnology, College of Life Sciences & Biotechnology, Korea University)
  • Jeong Sang Son(Laboratory of Stem Cells & Tissue Regeneration, Division of Biotechnology, College of Life Sciences & Biotechnology, Korea University)
  • Yu Jin Jang(Laboratory of Stem Cells & Tissue Regeneration, Division of Biotechnology, College of Life Sciences & Biotechnology, Korea University)
  • Jae Hun Lee(Laboratory of Stem Cells & Tissue Regeneration, Division of Biotechnology, College of Life Sciences & Biotechnology, Korea University)
  • Gyunggyu Lee(Laboratory of Stem Cells & Tissue Regeneration, Division of Biotechnology, College of Life Sciences & Biotechnology, Korea University)
  • Jong Hoon Kim(Laboratory of Stem Cells & Tissue Regeneration, Division of Biotechnology, College of Life Sciences & Biotechnology, Korea University, NEXEL Co., Ltd.)
  • Jong Seung Kim(Department of Chemistry, Korea University)