The use of induced pluripotent stem cells to reveal pathogenic gene mutations and explore treatments for retinitis pigmentosa.
The use of induced pluripotent stem cells to reveal pathogenic gene mutations and explore treatments for retinitis pigmentosa.
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DOI：
10.1186/1756-6606-7-45
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发表时间：
2014-06-16
期刊：
影响因子：
3.6
通讯作者：
Okano H
中科院分区：
文献类型：
--
作者：
Yoshida T;Ozawa Y;Suzuki K;Yuki K;Ohyama M;Akamatsu W;Matsuzaki Y;Shimmura S;Mitani K;Tsubota K;Okano H
色素性视网膜炎（RP）是一种遗传性的人类视网膜疾病，导致进行性感光细胞丧失，导致严重的视力障碍或失明。然而，迄今为止还没有有效的治疗方法。虽然已经确定了基因突变，但现有的临床数据并不总是足以阐明这些突变在疾病发病机制中的作用，部分原因是遗传背景的差异。我们从一名携带视紫红质突变（E181K）的RP患者身上获得了诱导多能干细胞（iPSCs）。使用辅助依赖腺病毒载体（HDAdV）基因转移，在患者的iPSCs中纠正了突变，并将其引入对照iPSCs。然后将这些细胞进行视网膜分化；培养5周后，用Nrl启动子驱动的增强型绿色荧光蛋白（EGFP）携带腺病毒标记杆状光感受器细胞，用流式细胞术纯化。通过这种方法，我们发现E181K突变的感光细胞存活率降低，这与内质网（ER）应激和凋亡标志物的表达增加有关。治疗试剂的筛选表明，雷帕霉素、PP242、AICAR、NQDI-1和salubrinal可促进患者ipsc来源的光感受器细胞的存活，同时内质网应激和凋亡标志物减少。此外，自噬标志物被发现与内质网应激相关，表明自噬通过抑制内质网应激诱导的凋亡变化而减少。RP患者来源的iPSCs与基因组编辑相结合，提供了一个通用的细胞系统，用于定义具有或不具有突变的等基因iPSCs中基因突变的作用，也提供了一个可用于探索候选治疗方法的系统。
Retinitis pigmentosa (RP) is an inherited human retinal disorder that causes progressive photoreceptor cell loss, leading to severe vision impairment or blindness. However, no effective therapy has been established to date. Although genetic mutations have been identified, the available clinical data are not always sufficient to elucidate the roles of these mutations in disease pathogenesis, a situation that is partially due to differences in genetic backgrounds. We generated induced pluripotent stem cells (iPSCs) from an RP patient carrying a rhodopsin mutation (E181K). Using helper-dependent adenoviral vector (HDAdV) gene transfer, the mutation was corrected in the patient’s iPSCs and also introduced into control iPSCs. The cells were then subjected to retinal differentiation; the resulting rod photoreceptor cells were labeled with an Nrl promoter-driven enhanced green fluorescent protein (EGFP)-carrying adenovirus and purified using flow cytometry after 5 weeks of culture. Using this approach, we found a reduced survival rate in the photoreceptor cells with the E181K mutation, which was correlated with the increased expression of endoplasmic reticulum (ER) stress and apoptotic markers. The screening of therapeutic reagents showed that rapamycin, PP242, AICAR, NQDI-1, and salubrinal promoted the survival of the patient’s iPSC-derived photoreceptor cells, with a concomitant reduction in markers of ER stress and apoptosis. Additionally, autophagy markers were found to be correlated with ER stress, suggesting that autophagy was reduced by suppressing ER stress-induced apoptotic changes. The use of RP patient-derived iPSCs combined with genome editing provided a versatile cellular system with which to define the roles of genetic mutations in isogenic iPSCs with or without mutation and also provided a system that can be used to explore candidate therapeutic approaches.
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影响因子：
30.8
作者：
Mears, AJ;Kondo, M;Swaroop, A
通讯作者：
Swaroop, A
影响因子：
3.7
作者：
Jin ZB;Okamoto S;Osakada F;Homma K;Assawachananont J;Hirami Y;Iwata T;Takahashi M
通讯作者：
Takahashi M
影响因子：
10.5
作者：
Nishitoh, H;Matsuzawa, A;Ichijo, H
通讯作者：
Ichijo, H
DOI：
10.1073/pnas.0508214103
发表时间：
2006-03-07
影响因子：
11.1
作者：
Akimoto, M;Cheng, H;Swaroop, A
通讯作者：
Swaroop, A
DOI：
10.1073/pnas.0905245106
发表时间：
2009-09-29
影响因子：
11.1
作者：
Meyer, Jason S.;Shearer, Rebecca L.;Gamm, David M.
通讯作者：
Gamm, David M.