Bei Jia1, Shen Chen2, Zhiju Zhao2, Pengfei Liu3, Jinglei Cai2, Dajiang Qin2, Juan Du2, Changwei Wu2, Qianyu Chen2, Xiujuan Cai2, Hui Zhang2, Yanhong Yu1, Duanqing Pei2, Mei Zhong4, Guangjin Pan5. 1. The Center for Prenatal and Hereditary Disease Diagnosis, Department of Obstetrics and Gynecology, Nanfang Hospital, Southern Medical University, Guangzhou, P. R. China. 2. Key Laboratory of Regenerative Biology, Guangdong Provincial Key Laboratory of Stem Cell and Regenerative Medicine, Guangzhou Institute of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, P. R. China. 3. Key Laboratory of Regenerative Biology, Guangdong Provincial Key Laboratory of Stem Cell and Regenerative Medicine, Guangzhou Institute of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, P. R. China; Department of Regeneration Medicine, School of Pharmaceutical Science, Jilin University, Changchun, P. R. China. 4. The Center for Prenatal and Hereditary Disease Diagnosis, Department of Obstetrics and Gynecology, Nanfang Hospital, Southern Medical University, Guangzhou, P. R. China. Electronic address: zhongmei@smu.edu.cn. 5. Key Laboratory of Regenerative Biology, Guangdong Provincial Key Laboratory of Stem Cell and Regenerative Medicine, Guangzhou Institute of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, P. R. China. Electronic address: pan_guangjin@gibh.ac.cn.
Abstract
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, SV40LT 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.
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, SV40LT 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.
Authors: Christopher Goldring; Daniel J Antoine; Frank Bonner; Jonathan Crozier; Chris Denning; Robert J Fontana; Neil A Hanley; David C Hay; Magnus Ingelman-Sundberg; Satu Juhila; Neil Kitteringham; Beatriz Silva-Lima; Alan Norris; Chris Pridgeon; James A Ross; Rowena Sison Young; Danilo Tagle; Belen Tornesi; Bob van de Water; Richard J Weaver; Fang Zhang; B Kevin Park Journal: Hepatology Date: 2016-11-30 Impact factor: 17.425
Authors: Bárbara C M F Paes; Luiza C J R Stabeli; Péricles N M Costa; Maristela Delgado Orellana; Simone Kashima; Dimas Tadeu Covas; Virgínia Picanço-Castro Journal: Hematol Transfus Cell Ther Date: 2020-03-06