Human iPSC-based models highlight defective glial and neuronal differentiation from neural progenitor cells in metachromatic leukodystrophy

Abstract

The pathological cascade leading from primary storage to neural cell dysfunction and death in metachromatic leukodystrophy (MLD) has been poorly elucidated in human-derived neural cell systems. In the present study, we have modeled the progression of pathological events during the differentiation of patient-specific iPSCs to neuroepithelial progenitor cells (iPSC-NPCs) and mature neurons, astrocytes, and oligodendrocytes at the morphological, molecular, and biochemical level. We showed significant sulfatide accumulation and altered sulfatide composition during the differentiation of MLD iPSC-NPCs into neuronal and glial cells. Changes in sulfatide levels and composition were accompanied by the expansion of the lysosomal compartment, oxidative stress, and apoptosis. The neuronal and glial differentiation capacity of MLD iPSC-NPCs was significantly impaired. We showed delayed appearance and/or reduced levels of oligodendroglial and astroglial markers as well as reduced number of neurons and disorganized neuronal network. Restoration of a functional Arylsulfatase A (ARSA) enzyme in MLD cells using lentiviral-mediated gene transfer normalized sulfatide levels and composition, globally rescuing the pathological phenotype. Our study points to MLD iPSC-derived neural progeny as a useful in vitro model to assess the impact of ARSA deficiency along NPC differentiation into neurons and glial cells. In addition, iPSC-derived neural cultures allowed testing the impact of ARSA reconstitution/overexpression on disease correction and, importantly, on the biology and functional features of human NPCs, with important therapeutic implications.

Addressing variability in iPSC-derived models of human disease: guidelines to promote reproducibility

“…Disease Models & Mechanisms (2020) 13, dmm042317. doi:10.1242/dmm.042317 mutation favours the maintenance of immature oligodendroglial progenitors and impairs their differentiation with consequent reduction in neuronal function support and eventually neuron death (Frati et al, 2018). This mechanism has been confirmed to contribute to the early stages of pathology before neurodegeneration occurs, pointing to the importance of such models to investigate disease processes that are shaped early in brain development and cannot be properly assessed in brain tissues of patients at the late stages of the disease, with implications for the timing and efficacy of treatments.…”

Human iPSC-based neurodevelopmental models of globoid cell leukodystrophy uncover patient- and cell type-specific disease phenotypes

“…Human iPSCs have been differentiated into neurons and/or glial cells to model the neural defects associated to several lysosomal storage diseases (LSDs) including mucopolysaccharidosis (Bayó-Puxan et al, 2018;Canals et al, 2015;Kobolák et al, 2019;Lemonnier et al, 2011;Vallejo-Diez et al, 2018), Niemann−Pick type C (Maetzel et al, 2014;Ordoñez and Steele, 2016;Trilck et al, 2013;Yu et al, 2014), Pompe disease (Higuchi et al, 2014), Gaucher disease (Panicker et al, 2012;Sun et al, 2015;Tiscornia et al, 2013), GM1 and GM2 gangliosidosis (Son et al, 2015) (Allende et al, 2018), NCL (Lojewski et al, 2014), and MLD (Frati et al, 2018;Meneghini et al, 2017). To our knowledge this is the first report describing a human iPSC-based neural model of GLD.…”

“…Therefore, we envisaged that differentiation of iPSC-NPCs into mixed neuronal/glial cultures would better model the composite GLD pathology. We adapted a published protocol (Frati et al, 2018) to generate cell populations containing neurons, astrocytes, and oligodendrocytes from ND, GLD, GLD GALC , and ND GALC iPSC-NPCs. Cultures were evaluated at day 0 (d0; NPCs), d7, d14, and d24 of differentiation ( Figure 3E), to track the time-course GALC expression/activity and the glial/neuronal cell commitment and maturation by means of biochemical and molecular analysis.…”

“…We applied a published protocol (Frati et al, 2018) with some modifications. Cells were detached using Accutase and plated on Matrigel-coated dishes (20,000 cells/cm 2 ) in N2 medium with 20 ng/ml bFGF, 20…”

Harnessing the Potential of Stem Cells for Disease Modeling: Progress and Promises

“…In this regard, genome editing tools enable creating isogenic controls iPSCs that differ from the original counterpart only on the gene/s responsible for the disease [43,44]. Despite these limitations, iPSCs have shown to be very promising for recapitulating complex genetically disorders especially when combined with gene editing techniques [33,45,46]. Timothy syndrome Patient-derived fibroblasts [77,78] The selective modification of cells' genetic information was the groundbreaking discovery that revolutionized the concept of cell models.…”