National Brain Research Project (NAP) Molecular Psychiatry Research Group, Hungarian Academy of Sciences and Semmelweis University, Budapest, Hungary.
Molecular Cell Biology Research Group, Institute of Enzymology, Research Center for Natural Sciences, 1117 Magyar tudósok körútja 2, Budapest, Hungary.
Stem Cell Res Ther. 2020 Nov 27;11(1):504. doi: 10.1186/s13287-020-01980-5.
De novo mutations (DNMs) have been implicated in the etiology of schizophrenia (SZ), a chronic debilitating psychiatric disorder characterized by hallucinations, delusions, cognitive dysfunction, and decreased community functioning. Several DNMs have been identified by examining SZ cases and their unaffected parents; however, in most cases, the biological significance of these mutations remains elusive. To overcome this limitation, we have developed an approach of using induced pluripotent stem cell (iPSC) lines from each member of a SZ case-parent trio, in order to investigate the effects of DNMs in cellular progenies of interest, particularly in dentate gyrus neuronal progenitors.
We identified a male SZ patient characterized by early disease onset and negative symptoms, who is a carrier of 3 non-synonymous DNMs in genes LRRC7, KHSRP, and KIR2DL1. iPSC lines were generated from his and his parents' peripheral blood mononuclear cells using Sendai virus-based reprogramming and differentiated into neuronal progenitor cells (NPCs) and hippocampal dentate gyrus granule cells. We used RNASeq to explore transcriptomic differences and calcium (Ca) imaging, cell proliferation, migration, oxidative stress, and mitochondrial assays to characterize the investigated NPC lines.
NPCs derived from the SZ patient exhibited transcriptomic differences related to Wnt signaling, neuronal differentiation, axonal guidance and synaptic function, and decreased Ca reactivity to glutamate. Moreover, we could observe increased cellular proliferation and alterations in mitochondrial quantity and morphology.
The approach of reprograming case-parent trios represents an opportunity for investigating the molecular effects of disease-causing mutations and comparing these in cell lines with reduced variation in genetic background. Our results are indicative of a partial overlap between schizophrenia and autism-related phenotypes in the investigated family.
Our study investigated only one family; therefore, the generalizability of findings is limited. We could not derive iPSCs from two other siblings to test for possible genetic effects in the family that are not driven by DNMs. The transcriptomic and functional assays were limited to the NPC stage, although these variables should also be investigated at the mature neuronal stage.
从头突变（DNMs）已被认为与精神分裂症（SZ）的病因有关，SZ 是一种慢性衰弱性精神疾病，其特征为幻觉、妄想、认知功能障碍和社交功能下降。通过检查 SZ 病例及其未受影响的父母，已经鉴定出了一些 DNMs；然而，在大多数情况下，这些突变的生物学意义仍然难以捉摸。为了克服这一限制，我们开发了一种方法，使用来自 SZ 病例-父母三人组的每个成员的诱导多能干细胞（iPSC）系，以研究 DNMs 在感兴趣的细胞后代中的影响，特别是在齿状回神经元祖细胞中。
我们鉴定了一名男性 SZ 患者，其疾病发病早且具有阴性症状，是 LRRC7、KHSRP 和 KIR2DL1 基因中 3 个非同义 DNMs 的携带者。我们使用基于 Sendai 病毒的重编程从他和他父母的外周血单核细胞中生成 iPSC 系，并将其分化为神经元祖细胞（NPC）和海马齿状回颗粒细胞。我们使用 RNA-seq 来探索转录组差异，并用钙（Ca）成像、细胞增殖、迁移、氧化应激和线粒体测定来表征所研究的 NPC 系。
来自 SZ 患者的 NPC 表现出与 Wnt 信号、神经元分化、轴突导向和突触功能相关的转录组差异，以及对谷氨酸的 Ca 反应性降低。此外，我们可以观察到细胞增殖增加和线粒体数量和形态的改变。
对病例-父母三人组进行重编程的方法为研究致病突变的分子影响并在遗传背景变异减少的细胞系中进行比较提供了机会。我们的结果表明，在所研究的家族中，SZ 与自闭症相关表型之间存在部分重叠。
我们的研究仅调查了一个家庭；因此，研究结果的普遍性有限。我们无法从另外两个兄弟姐妹中获得 iPSC 来测试家族中不是由 DNMs 驱动的可能遗传效应。转录组和功能测定仅限于 NPC 阶段，尽管这些变量也应在成熟神经元阶段进行研究。
