Molecular Medicine for Neurodegenerative and Neuromuscular Diseases Unit, IRCCS Fondazione Stella Maris, Via dei Giacinti 2, Calambrone, 56128 Pisa, Italy.
Department of Neurosciences, Psychology, Drug Research and Child Health (NEUROFARBA), University of Florence, Viale Pieraccini, 6, 50139 Florence, Italy.
Int J Mol Sci. 2024 Oct 2;25(19):10614. doi: 10.3390/ijms251910614.
Hereditary spastic paraplegias are rare genetic disorders characterized by corticospinal tract impairment. Spastic paraplegia 83 (SPG83) is associated with biallelic mutations in the gene, leading to varied severities from neonatal to juvenile onset. The function of HPDL is unclear, though it is speculated to play a role in alternative coenzyme Q10 biosynthesis. Here, we report the generation of hiPS lines from primary skin fibroblasts derived from three SPG83 patients with different mutations, using episomal reprogramming. The patients' clinical characteristics are carefully listed. The hiPS lines were meticulously characterized, demonstrating typical pluripotent characteristics through immunofluorescence assays for stemness markers (OCT4, TRA1-60, NANOG, and SSEA4) and RT-PCR for endogenous gene expression. Genetic integrity and identity were confirmed via Sanger sequencing and short tandem repeat analysis. These hiPS cells displayed typical pluripotent characteristics and were able to differentiate into neocortical neurons via a dual SMAD inhibition protocol. In addition, HPDL mutant neurons assessed via long-term culturing were able to achieve effective maturation, similarly to their wild-type counterparts. The HPDL hiPS lines we generated will provide a valuable model for studying SPG83, offering insights into its molecular mechanisms and potential for developing targeted therapies.
遗传性痉挛性截瘫是一种罕见的遗传疾病，其特征是皮质脊髓束损伤。SPG83 与 基因的双等位基因突变有关，导致从新生儿到青少年发病的不同严重程度。HPDL 的功能尚不清楚，但据推测它在辅酶 Q10 的替代生物合成中发挥作用。在这里，我们使用附加体重编程技术，从三个具有不同 突变的 SPG83 患者的原代皮肤成纤维细胞中生成 hiPS 系。详细列出了患者的临床特征。精心表征了 hiPS 系，通过干性标志物（OCT4、TRA1-60、NANOG 和 SSEA4）的免疫荧光检测和内源性基因表达的 RT-PCR 证明了典型的多能性特征。通过 Sanger 测序和短串联重复分析确认了遗传完整性和同一性。这些 hiPS 细胞表现出典型的多能性特征，并能够通过双 SMAD 抑制方案分化为新皮层神经元。此外，通过长期培养评估的 HPDL 突变神经元能够实现有效成熟，与野生型神经元相似。我们生成的 HPDL hiPS 系将为研究 SPG83 提供有价值的模型，深入了解其分子机制并为开发靶向治疗方法提供可能。
