DEVELOPMENT OF BRAIN ORGANOIDS FROM HUMAN INDUCED PLURIPOTENT STEM CELLS FOR STUDYING THE RARE LEUKODYSTROPHY MEGALENCEPHALIC LEUKOENCEPHALOPATHY WITH SUBCORTICAL CYSTS

Megalencephalic leukoencephalopathy with subcortical cysts (MLC) is a rare leukodystrophy caused by primary astrocyte dysfunction, characterized by macrocephaly, cerebral edema, subcortical cysts, and myelin vacuolation. The disease is mainly associated with mutations in the MLC1 gene, which encodes an astrocyte-specific protein highly expressed at perivascular end-feet. Although the precise function of this protein remains unclear, MLC1 plays a key role in regulating astrocyte volume, proliferation, and activation in response to injury. Using astrocytes differentiated from patient-derived induced pluripotent stem cells (iPSCs), we showed that MLC1 mutations lead to astrocyte maturation defects. In this study, brain organoids were generated from iPSCs of two healthy controls and four MLC patients to investigate whether the impaired astrocyte maturation also affects oligodendrocyte differentiation/maturation. Organoids were analyzed at days 50 and 70 using Western blotting, quantitative PCR, and immunofluorescence. Both control and MLC organoids developed the major brain cell populations. However, some MLC patient-derived organoids showed a reduced expression of mature astrocyte markers and a marked decrease in oligodendrocyte markers compared to controls. These findings highlight maturation defects in MLC astrocytes that may negatively affect oligodendrocyte development, emphasizing organoids as a valuable model for studying MLC and testing therapies.