Genotyping, generation and proteomic profiling of the first human autosomal dominant osteopetrosis type II-specific induced pluripotent stem cells

作者：Minglin Ou, Chunhong Li, Donge Tang, Wen Xue, Yong Xu, Peng Zhu, Bo Li, Jiansheng Xie, Jiejing Chen, Weiguo Sui, Lianghong Yin, Yong Dai · 发表于：Stem Cell Research & Therapy · 年份：2019 · DOI：10.1186/s13287-019-1369-8 · 被引用次数：90 · 研究领域：Bone Metabolism and Diseases、Dermatological and Skeletal Disorders、Hypertrophic osteoarthropathy and related conditions

BACKGROUND: Autosomal dominant osteopetrosis type II (ADO2) is a rare human genetic disease that has been broadly studied as an important osteopetrosis model; however, there are no disease-specific induced pluripotent stem cells (ADO2-iPSCs) that may be valuable for understanding the pathogenesis and may be a potential source of cells for autologous cell-based therapies. METHODS: To generate the first human ADO2-iPSCs from a Chinese family with ADO2 and to identify their characteristics, blood samples were collected from the proband and his parents and were used for genotyping by whole-exome sequencing (WES); the urine-derived cells of the proband were reprogrammed with episomal plasmids that contained transcription factors, such as KLF4, OCT4, c-MYC, and SOX2. The proteome-wide protein quantification and lysine 2-hydroxyisobutyrylation detection of the ADO2-iPSCs and normal control iPSCs (NC-iPSCs) were performed by high-resolution LC-MS/MS and bioinformatics analysis. RESULTS: WES with filtering strategies identified a mutation in CLCN7 (R286W) in the proband and his father, which was absent in the proband's mother and the healthy controls; this was confirmed by Sanger sequencing. The ADO2-iPSCs were successfully generated, which carried a normal male karyotype (46, XY) and the mutation of CLCN7 (R286W); the ADO2-iPSCs positively expressed alkaline phosphatase and other surface markers; and no vector and transgene were detected. The ADO2-iPSCs could differentiate into all t...