Lyu Cuicui, Shen Jun, Zhang Jianping, Xue Feng, Liu Xiaofan, Liu Wei, Fu Rongfeng, Zhang Liyan, Li Huiyuan, Zhang Donglei, Zhang Xiaobing, Cheng Tao, Yang Renchi, Zhang Lei
1 State Key Laboratory of Experimental Hematology, Institute of Hematology and Blood Disease Hospital , Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin, China .
2 Department of Hematology, The First Central Hospital of Tianjin , Tianjin, China .
Stem Cells Dev. 2017 Jul 1;26(13):1003-1011. doi: 10.1089/scd.2016.0323. Epub 2017 Mar 22.
Skewed X chromosome inactivation (XCI) is a rare reason for hemophilia B in females. It is indefinite whether X chromosome reactivation (XCR) would occur when cells of hemophilia B patients with skewed XCI were reprogrammed into induced pluripotent stem cells (iPSCs). In this study, we investigated a female hemophilia B patient with a known F9 gene mutation: c.676C>T, p.Arg226Trp. We demonstrated that skewed XCI was the pathogenesis of the patient, and we successfully generated numerous iPSC colonies of the patient from peripheral blood mononuclear cells (PBMNCs), which was the first time for generating hemophilia-specific iPSCs from PBMNCs. Then we detected the XCI state of these iPSCs. Ninety-two iPSC lines were picked for XCI analysis. All of them retained an inactive X chromosome, which could be proved by amplification of the androgen receptor gene and XIST (X inactivation-specific transcript), expression of H3K27me3, and existence of XIST clouds in XIST RNA fluorescence in situ hybridization (FISH) analysis. We attempted to obtain iPSC lines with the wild-type F9 gene on the active X chromosome for further disease treatment. But it turned out that the patient's iPSCs were still skewed such as the somatic cells with 92 iPSC lines having mutant F9 on the active X chromosome. In conclusion, skewed XCI is one reason for hemophilia in females. PBMNCs are excellent somatic cell resources for hemophilia patients to do reprogramming. More attentions should be paid to generate naive iPSCs with two active X chromosomes for further clinical disease treatment. The state of skewed XCI is retained in the iPSCs from a female with hemophilia B.
X染色体失活偏倚（XCI）是女性患B型血友病的罕见原因。对于XCI偏倚的B型血友病患者的细胞重编程为诱导多能干细胞（iPSC）时是否会发生X染色体重新激活（XCR）尚不确定。在本研究中，我们调查了一名已知F9基因突变（c.676C>T，p.Arg226Trp）的女性B型血友病患者。我们证明XCI偏倚是该患者的发病机制，并且我们成功地从外周血单个核细胞（PBMNC）中生成了该患者的大量iPSC集落，这是首次从PBMNC中生成血友病特异性iPSC。然后我们检测了这些iPSC的XCI状态。挑选了92个iPSC系进行XCI分析。所有这些系均保留了一条失活的X染色体，这可以通过雄激素受体基因和XIST（X失活特异性转录本）的扩增、H3K27me3的表达以及XIST RNA荧光原位杂交（FISH）分析中XIST云的存在来证明。我们试图获得活性X染色体上具有野生型F9基因的iPSC系用于进一步的疾病治疗。但结果表明，患者的iPSC仍然存在偏倚，例如92个iPSC系的体细胞在活性X染色体上具有突变的F9。总之，XCI偏倚是女性患血友病的一个原因。PBMNC是血友病患者进行重编程的优良体细胞资源。为了进一步的临床疾病治疗，应更加关注生成具有两条活性X染色体的原始iPSC。来自B型血友病女性的iPSC中保留了XCI偏倚状态。
