Costamagna Domiziana, Casters Valérie, Beltrà Marc, Sampaolesi Maurilio, Van Campenhout Anja, Ortibus Els, Desloovere Kaat, Duelen Robin
Stem Cell and Developmental Biology, Department of Development and Regeneration, KU Leuven, 3000 Leuven, Belgium.
Research Group for Neurorehabilitation, Department of Rehabilitation Sciences, KU Leuven, 3000 Leuven, Belgium.
Cells. 2022 Oct 24;11(21):3351. doi: 10.3390/cells11213351.
Hereditary spastic paraplegia (HSP) is a heterogeneous group of genetic neurodegenerative disorders, characterized by progressive lower limb spasticity and weakness resulting from retrograde axonal degeneration of motor neurons (MNs). Here, we generated in vitro human neuromuscular junctions (NMJs) from five HSP patient-specific induced pluripotent stem cell (hiPSC) lines, by means of microfluidic strategy, to model disease-relevant neuropathologic processes. The strength of our NMJ model lies in the generation of lower MNs and myotubes from autologous hiPSC origin, maintaining the genetic background of the HSP patient donors in both cell types and in the cellular organization due to the microfluidic devices. Three patients characterized by a mutation in the gene, encoding the ATLASTIN GTPase 1 protein, and two patients with a mutation in the gene, encoding the SPASTIN protein, were included in this study. Differentiation of the HSP-derived lines gave rise to lower MNs that could recapitulate pathological hallmarks, such as axonal swellings with accumulation of Acetyl-α-TUBULIN and reduction of SPASTIN levels. Furthermore, NMJs from HSP-derived lines were lower in number and in contact point complexity, denoting an impaired NMJ profile, also confirmed by some alterations in genes encoding for proteins associated with microtubules and responsible for axonal transport. Considering the complexity of HSP, these patient-derived neuronal and skeletal muscle cell co-cultures offer unique tools to study the pathologic mechanisms and explore novel treatment options for rescuing axonal defects and diverse cellular processes, including membrane trafficking, intracellular motility and protein degradation in HSP.
遗传性痉挛性截瘫（HSP）是一组异质性的遗传性神经退行性疾病，其特征是运动神经元（MNs）逆行轴突变性导致进行性下肢痉挛和无力。在此，我们通过微流控策略，从5个HSP患者特异性诱导多能干细胞（hiPSC）系中生成了体外人神经肌肉接头（NMJ），以模拟与疾病相关的神经病理过程。我们的NMJ模型的优势在于从自体hiPSC来源生成下运动神经元和肌管，由于微流控装置，在两种细胞类型以及细胞组织中都保持了HSP患者供体的遗传背景。本研究纳入了3名以编码ATLASTIN GTPase 1蛋白的基因突变为特征的患者，以及2名以编码SPASTIN蛋白的基因突变的患者。HSP来源系细胞的分化产生了能重现病理特征的下运动神经元，如轴突肿胀伴乙酰-α-微管蛋白积聚以及SPASTIN水平降低。此外，HSP来源系的神经肌肉接头数量减少且接触点复杂性降低，表明神经肌肉接头形态受损，这也通过一些与微管相关且负责轴突运输的蛋白质编码基因的改变得到证实。考虑到HSP的复杂性，这些患者来源的神经元和骨骼肌细胞共培养物为研究病理机制以及探索挽救轴突缺陷和多种细胞过程（包括HSP中的膜运输、细胞内运动和蛋白质降解）的新治疗选择提供了独特的工具。
