Niceforo Alessia, Marioli Chiara, Colasuonno Fiorella, Petrini Stefania, Massey Keith, Tartaglia Marco, Bertini Enrico, Moreno Sandra, Compagnucci Claudia
Department of Science, LIME, University Roma Tre, Rome 00146, Italy.
Department of Neuroscience, Unit of Neuromuscular and Neurodegenerative Diseases, Laboratory of Molecular Medicine, IRCCS Ospedale Pediatrico Bambino Gesù, Rome 00146, Italy.
Dis Model Mech. 2021 Jan 19;14(2). doi: 10.1242/dmm.046391.
The cytoskeletal network plays a crucial role in differentiation, morphogenesis, function and homeostasis of the nervous tissue, so that alterations in any of its components may lead to neurodegenerative diseases. Riboflavin transporter deficiency (RTD), a childhood-onset disorder characterized by degeneration of motor neurons (MNs), is caused by biallelic mutations in genes encoding the human riboflavin (RF) transporters. In a patient- specific induced Pluripotent Stem Cells (iPSCs) model of RTD, we recently demonstrated altered cell-cell contacts, energy dysmetabolism and redox imbalance.The present study focusses on cytoskeletal composition and dynamics associated to RTD, utilizing patients' iPSCs and derived MNs. Abnormal expression and distribution of α- and β-tubulin (α- and β-TUB), as well as imbalanced tyrosination of α-TUB, accompanied by impaired ability to repolymerize after nocodazole treatment, were found in RTD patient-derived iPSCs. Following differentiation, MNs showed consistent changes in TUB content, which was associated with abnormal morphofunctional features, such as neurite length and Ca homeostasis, suggesting impaired differentiation.Beneficial effects of RF supplementation, alone or in combination with the antioxidant molecule N-acetyl-cystine (NAC), were assessed. RF administration resulted in partially improved cytoskeletal features in patients' iPSCs and MNs, suggesting that redundancy of transporters may rescue cell functionality in the presence of adequate concentrations of the vitamin. Moreover, supplementation with NAC was demonstrated to be effective in restoring all the considered parameters, when used in combination with RF, thus supporting the therapeutic use of both compounds.
细胞骨架网络在神经组织的分化、形态发生、功能及内稳态中发挥着关键作用，因此其任何组分的改变都可能导致神经退行性疾病。核黄素转运蛋白缺乏症（RTD）是一种儿童期发病的疾病，其特征为运动神经元（MNs）退化，由编码人类核黄素（RF）转运蛋白的基因双等位基因突变引起。在RTD患者特异性诱导多能干细胞（iPSCs）模型中，我们最近证明了细胞间接触改变、能量代谢紊乱和氧化还原失衡。本研究利用患者的iPSCs和分化得到的MNs，聚焦于与RTD相关的细胞骨架组成和动力学。在RTD患者来源的iPSCs中发现了α-和β-微管蛋白（α-和β-TUB）的异常表达和分布，以及α-TUB酪氨酸化失衡，同时在诺考达唑处理后再聚合能力受损。分化后，MNs的TUB含量出现一致变化，这与异常的形态功能特征相关，如神经突长度和钙稳态，提示分化受损。评估了单独补充RF或与抗氧化分子N-乙酰胱氨酸（NAC）联合补充的有益效果。给予RF可部分改善患者iPSCs和MNs的细胞骨架特征，表明在维生素浓度充足时，转运蛋白的冗余可能挽救细胞功能。此外，当与RF联合使用时，补充NAC被证明可有效恢复所有考虑的参数，从而支持这两种化合物的治疗用途。
