期刊
LIFE SCIENCES
卷 108, 期 1, 页码 22-29出版社
PERGAMON-ELSEVIER SCIENCE LTD
DOI: 10.1016/j.lfs.2014.05.004
关键词
Hemophilia A; Clotting factor VIII; Induced pluripotent stem cells; Directed differentiation in vitro
资金
- National Basic Research Program of China
- 973 Program of China [2012CB966500, 2011CB965204]
- President's Fund of Nanfang Hospital
- Southern Medical University [2013C013]
- Strategic Priority Research Program of the Chinese Academy of Sciences [XDA01020202, XDA01020401]
- National S&T Major Special Project on Major New Drug Innovation [2011ZX09102-010]
- National Natural Science Foundation of China [31200970]
- Open Project of Key Laboratory of Regenerative Biology, Chinese Academy of Sciences [KLRB201106]
- 100 Talents Project of the Chinese Academy of Sciences, China
Aims: Hemophilia A (HA) is a severe, congenital bleeding disorder caused by the deficiency of clotting factor VIII (FVIII). For years, traditional laboratory animals have been used to study HA and its therapies, although animal models may not entirely mirror the human pathophysiology. Human induced pluripotent stem cells (iPSCs) can undergo unlimited self-renewal and differentiate into all cell types. This study aims to generate hemophilia A (HA) patient-specific iPSCs that differentiate into disease-affected hepatocyte cells. These hepatocytes are potentially useful for in vitro disease modeling and provide an applicable cell source for autologous cell therapy after genetic correction. Main methods: In this study, we mainly generated iPSCs from urine collected from HA patients with integration-free episomal vectors PEP4-EO2S-ET2K containing human genes OCT4, SOX2, SV4OLT and KLF4, and differentiated these iPSCs into hepatocyte-like cells. We further identified the genetic phenotype of the FVIII genes and the FVIII activity in the patient-specific iPSC derived hepatic cells. Key findings: HA patient-specific iPSCs (HA-iPSCs) exhibited typical pluripotent properties evident by immunostaining, in vitro assays and in vivo assays. Importantly, we showed that HA-iPSCs could differentiate into functional hepatocyte-like cells and the HA-iPSC-derived hepatocytes failed to produce FVIII, but otherwise functioned normally, recapitulating the phenotype of HA disease in vitro. Significance: HA-iPSCs, particular those generated from the urine using a non-viral approach, provide an efficient way for modeling HA in vitro. Furthermore, HA-iPSCs and their derivatives serve as an invaluable cell source that can be used for gene and cell therapy in regenerative medicine. (C) 2014 Elsevier Inc. All rights reserved.
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