Efficient hepatic differentiation and regeneration potential under xeno-free conditions using mass-producible amnion-derived mesenchymal stem cells

Stem Cell Research & Therapy

Ji-Yoon Lee¹  Soo Jin Oh¹  Gyeong-Nam Kim²  Young Hoon Sung²  Seongjun So³  Jiwan Choi⁴  Seoon Kang⁴  Jongsuk Han⁴  Eunju Kang⁴  Sung Hoon Kim⁵  Mi-Young Lee⁵  Seonae Roh⁶  Yeonmi Lee⁷  Bitnara Kim⁷ 

[1] Asan Institute for Life Sciences, Asan Medical Center and Department of Convergence Medicine, College of Medicine, University of Ulsan, 05505, Seoul, South Korea;Department of Medical Science, Asan Medical Institute of Convergence Science and Technology (AMIST), University of Ulsan College of Medicine, Asan Medical Center, 05505, Seoul, South Korea;Convergence Medicine Research Center, Asan Institute for Life Sciences, Asan Medical Center, 05505, Seoul, South Korea;Department of Medical Science, Asan Medical Institute of Convergence Science and Technology (AMIST), University of Ulsan College of Medicine, Asan Medical Center, 05505, Seoul, South Korea;Stem Cell Center, Asan Institute for Life Sciences, Asan Medical Center, 05505, Seoul, South Korea;Department of Medical Science, Asan Medical Institute of Convergence Science and Technology (AMIST), University of Ulsan College of Medicine, Asan Medical Center, 05505, Seoul, South Korea;Stem Cell Center, Asan Institute for Life Sciences, Asan Medical Center, 05505, Seoul, South Korea;Present Address: Center for Embryo & Stem Cell Research, CHA Advanced Research Institute and Department of Biomedical Science, CHA University, 13488, Pocheon-si, Gyeonggi, South Korea;Department of Obstetrics and Gynecology, University of Ulsan College of Medicine, Asan Medical Center, 05505, Seoul, South Korea;Stem Cell Center, Asan Institute for Life Sciences, Asan Medical Center, 05505, Seoul, South Korea;Stem Cell Center, Asan Institute for Life Sciences, Asan Medical Center, 05505, Seoul, South Korea;Present Address: Center for Embryo & Stem Cell Research, CHA Advanced Research Institute and Department of Biomedical Science, CHA University, 13488, Pocheon-si, Gyeonggi, South Korea;

关键词: Amnion-derived mesenchymal stem cells;    Differentiation;    Hepatic progenitor;    Xeno-free;    GSK3 inhibitor;    Polyvinyl alcohol;   

DOI  :  10.1186/s13287-021-02470-y

来源: Springer

【 摘 要 】

BackgroundAmnion-derived mesenchymal stem cells (AM-MSCs) are an attractive source of stem cell therapy for patients with irreversible liver disease. However, there are obstacles to their use due to low efficiency and xeno-contamination for hepatic differentiation.MethodsWe established an efficient protocol for differentiating AM-MSCs into hepatic progenitor cells (HPCs) by analyzing transcriptome-sequencing data. Furthermore, to generate the xeno-free conditioned differentiation protocol, we replaced fetal bovine serum (FBS) with polyvinyl alcohol (PVA). We investigated the hepatocyte functions with the expression of mRNA and protein, secretion of albumin, and activity of CYP3A4. Finally, to test the transplantable potential of HPCs, we transferred AM-MSCs along with hepatic progenitors after differentiated days 11, 12, and 13 based on the expression of hepatocyte-related genes and mitochondrial function. Further, we established a mouse model of acute liver failure using a thioacetamide (TAA) and cyclophosphamide monohydrate (CTX) and transplanted AM-HPCs in the mouse model through splenic injection.ResultsWe analyzed gene expression from RNA sequencing data in AM-MSCs and detected downregulation of hepatic development-associated genes including GATA6, KIT, AFP, c-MET, FGF2, EGF, and c-JUN, and upregulation of GSK3. Based on this result, we established an efficient hepatic differentiation protocol using the GSK3 inhibitor, CHIR99021. Replacing FBS with PVA resulted in improved differentiation ability, such as upregulation of hepatic maturation markers. The differentiated hepatocyte-like cells (HLCs) not only synthesized and secreted albumin, but also metabolized drugs by the CYP3A4 enzyme. The best time for translation of AM-HPCs was 12 days from the start of differentiation. When the AM-HPCs were transplanted into the liver failure mouse model, they settled in the damaged livers and differentiated into hepatocytes.ConclusionThis study offers an efficient and xeno-free conditioned hepatic differentiation protocol and shows that AM-HPCs could be used as transplantable therapeutic materials. Thus, we suggest that AM-MSC-derived HPCs are promising cells for treating liver disease.

【 授权许可】

CC BY