Parrotta Elvira Immacolata, Lucchino Valeria, Zannino Clara, Valente Desirèe, Scalise Stefania, Bressan Davide, Benedetto Giorgia Lucia, Iazzetta Maria Roberta, Talarico Mariagrazia, Gagliardi Monica, Conforti Francesco, Di Agostino Silvia, Fiorenzano Alessandro, Quattrone Aldo, Cuda Giovanni, Quattrone Andrea
Laboratory of Stem Cells, Department of Medical and Surgical Sciences, University "Magna Graecia", Catanzaro, Italy.
Department of Experimental and Clinical Medicine, University "Magna Graecia", Catanzaro, Italy.
Ann Neurol. 2025 May;97(5):845-859. doi: 10.1002/ana.27172. Epub 2025 Jan 28.
Progressive Supranuclear Palsy (PSP) is a severe neurodegenerative disease characterized by tangles of hyperphosphorylated tau protein and tufted astrocytes. Developing treatments for PSP is challenging due to the lack of disease models reproducing its key pathological features. This study aimed to model sporadic PSP-Richardson's syndrome (PSP-RS) using multi-donor midbrain organoids (MOs).
The MOs were generated by pooling induced pluripotent stem cells (iPSCs) from 4 patients with sporadic probable PSP-RS and compared them with MOs from 3 healthy control (HC) subjects. We performed comprehensive analyses of MOs over 120 days to assess neuronal death, reactive gliosis, and the accumulation of 4R-tau and hyperphosphorylated tau forms (pThr231, pSer396, pThr181, and pSer202/pThr205 [AT8]) using immunofluorescence microscopy and Western blot. On day 90, immunohistochemical analysis using pSer396 and AT8 antibodies was conducted to assess disease pathology.
PSP-derived MOs showed progressive size reduction compared with HC-derived MOs, linked to upregulated apoptosis-related mRNA markers. Dopaminergic neuron degeneration was marked by decreased tyrosine hydroxylase (TH) and increased neurofilament light chain (NfL). Immunofluorescence and Western blot revealed accumulation of all investigated tau forms with a peak at 90 days, along with a significant rise in GFAP-positive cells in PSP-derived MOs. Immunochemistry confirmed typical PSP histological alterations, such as neurofibrillary tangles and tufted-shaped astrocytes, absent in HC-derived organoids.
We developed a robust in vitro PSP model reproducing the key molecular and histologic features of the disease. This result holds promise for advancing basic and clinical research in PSP, paving the way for in vitro molecular diagnosis and identification of novel therapeutic targets. ANN NEUROL 2025;97:845-859.
进行性核上性麻痹（PSP）是一种严重的神经退行性疾病，其特征为过度磷酸化的tau蛋白缠结和簇状星形胶质细胞。由于缺乏能够重现其关键病理特征的疾病模型，开发PSP的治疗方法具有挑战性。本研究旨在使用多供体中脑类器官（MOs）建立散发性PSP-理查森综合征（PSP-RS）模型。
通过汇集4例散发性可能PSP-RS患者的诱导多能干细胞（iPSCs）生成MOs，并将其与3名健康对照（HC）受试者的MOs进行比较。我们对MOs进行了超过120天的综合分析，使用免疫荧光显微镜和蛋白质印迹法评估神经元死亡、反应性胶质增生以及4R-tau和过度磷酸化tau形式（pThr231、pSer396、pThr181和pSer202/pThr205 [AT8]）的积累。在第90天，使用pSer396和AT8抗体进行免疫组织化学分析以评估疾病病理。
与HC来源的MOs相比，PSP来源的MOs显示出逐渐缩小的尺寸，这与凋亡相关mRNA标志物的上调有关。多巴胺能神经元变性表现为酪氨酸羟化酶（TH）减少和神经丝轻链（NfL）增加。免疫荧光和蛋白质印迹显示，所有研究的tau形式均有积累，在90天时达到峰值，同时PSP来源的MOs中GFAP阳性细胞显著增加。免疫组织化学证实了典型的PSP组织学改变，如神经原纤维缠结和簇状星形胶质细胞，而HC来源的类器官中不存在这些改变。
我们建立了一个强大的体外PSP模型，重现了该疾病的关键分子和组织学特征。这一结果有望推动PSP的基础和临床研究，为体外分子诊断和新型治疗靶点的鉴定铺平道路。《神经病学纪事》2025年；97：845 - 859。
