Human and mouse iPSC-derived astrocyte subtypes reveal vulnerability in Vanishing White Matter

Abstract

Astrocytes gained attention as important players in neurological disease, including a number of leukodystrophies. Several studies explored the generation of induced pluripotent stem cell-derived astrocytes for drug screening and regenerative studies. Developing robust models of patient induced pluripotent stem cells is challenged by high variability due to diverse genetic backgrounds and long-term culture procedures. While human models are of special interest, mouse-based models have the advantage that for them these issues are less pronounced. Here we present astrocyte differentiation protocols for both human and mouse induced pluripotent stem cells to specifically induce grey and white matter astrocytes. Both subtypes expressed astrocyte-associated markers, had typical astrocyte morphologies, and gave a reactive response to stress. Importantly, the grey and white matter-like astrocytes differed in size, complexity of processes, and expression profile, conform primary grey and white matter astrocytes. The newly presented mouse and human stem cell-based models for the leukodystrophy Vanishing White Matter replicated earlier findings, such as increased proliferation, decreased OPC maturation and modulation by hyaluronidase. We studied intrinsic astrocyte subtype vulnerability in Vanishing White Matter in both human and mouse cells. Oligodendrocyte maturation was specifically inhibited in cultures with Vanishing WhiteMatter white matter-like astrocytes. By performing RNA sequencing, we found more differentially regulated genes in the white than in the grey matter-like astrocytes. Human and mouse astrocytes showed the same affected pathways, although human white matter-like astrocytes presented human-specific disease mechanisms involved in Vanishing White Matter. Using both human and mouse induced pluripotent stem cells, our study presents protocols to generate white and grey matter-like astrocytes, and shows astrocyte subtypespecific defects in Vanishing White Matter. While mouse induced pluripotent stem cell-based cultures may be less suitable to mimic human astrocyte subtype-or patient-specific changes, they might more robustly represent disease mutation-related cellular phenotypes as the cells are derived from inbred mice and the protocols are faster. The presented models give new tools to generate astrocyte subtypes for in vitro disease modeling and in vivo regenerative applications.

Identification of repurposable cytoprotective drugs for Vanishing White Matter Disease

“…A previous study identified that white matter-derived astrocytes, generated using a CNTFbased differentiation protocol, showed a more vulnerable phenotype to stress, compared to grey matter astrocytes generated using FBS (7). Consequently, we generated astrocytes from iPSCs using a CNTF-based method ( Figure S2).…”

“…EIF2B mutant mouse models and induced pluripotent stem cell (iPSC) models (7,8) have identified a central role for dysfunctional astrocytes in the development of VWMD with evidence for astrocytic apoptosis (8) and an inability to promote oligodendrocyte maturation (6). A key driver of cellular pathogenesis in VWMD involves the ISR, with alterations in responses to endoplasmic reticulum stress, proteasomal stress and oxidative stress (3).…”