Objective: This study aims to determine how patient-specific microglia influence phospho-α‑synuclein aggregation and contribute to disease-relevant phenotypes within human midbrain organoids.

Background: Parkinson’s disease (PD) is characterized by dopaminergic neuron degeneration and the pathological aggregation of α‑synuclein into Lewy bodies and Lewy neurites. While neuronal contribution in PD has widely been studied, microglia, key regulators of CNS immunity and homeostasis, are increasingly recognized as active drivers of disease mechanisms. However, their role in initiating or amplifying α‑synuclein pathology in human midbrain tissue remains insufficiently understood.

Method: We generated human midbrain assembloids by integrating patient-specific microglia harbouring a triplication of the SNCA locus (3xSNCA) into midbrain organoids derived from human induced pluripotent stem cells (iPSC) [1,2]. Microglial and assembloid phenotypes were assessed using Immunofluorescence, Western blotting FACS, and bulk-RNA sequencing. We evaluated α‑synuclein expression and aggregation, phagocytic capacity, lysosomal function, inflammatory signatures, midbrain-specific neuronal markers, to determine microglia-mediated effects on organoid pathology.

Results: 3xSNCA microglia exhibited impaired lysosomal and phagocytic activity, altered inflammatory signalling, and metabolic dysregulation. When integrated into midbrain organoids, diseased microglia promoted the accumulation of phosphorylated α‑synuclein and additional disease-associated proteins, accompanied mitochondrial and oxidative stress alterations. Importantly, 3xSNCA microglia were sufficient to induce phospho-α‑synuclein pathology even within organoids generated from healthy control iPSCs, demonstrating a cell-autonomous capacity of diseased microglia to initiate PD-related pathology.

Conclusion: Our results demonstrate that the SNCA‑triplication microglia, which carry a pathogenic α‑synuclein mutation, directly contribute to the development and propagation of phospho‑α‑synuclein pathology in human midbrain organoids. Patient-specific microglia not only exacerbate PD-related phenotypes in genetically matched midbrain organoids but also initiate endogenous pathology in a healthy neuronal environment. These findings underscore microglia as key modulators of early PD pathogenesis and highlight them as promising therapeutic targets.

References: [1] Monzel AS, Smits LM, Hemmer K, Hachi S, Moreno EL, van Wuellen T, Jarazo J, Walter J, Brüggemann I, Boussaad I, Berger E, Fleming RMT, Bolognin S, Schwamborn JC. Derivation of Human Midbrain-Specific Organoids from Neuroepithelial Stem Cells. Stem Cell Reports. 2017 May 9;8(5):1144-1154. doi: 10.1016/j.stemcr.2017.03.010. Epub 2017 Apr 13. PMID: 28416282; PMCID: PMC5425618.
[2] Sabate-Soler S, Nickels SL, Saraiva C, Berger E, Dubonyte U, Barmpa K, Lan YJ, Kouno T, Jarazo J, Robertson G, Sharif J, Koseki H, Thome C, Shin JW, Cowley SA, Schwamborn JC. Microglia integration into human midbrain organoids leads to increased neuronal maturation and functionality. Glia. 2022 Jul;70(7):1267-1288. doi: 10.1002/glia.24167. Epub 2022 Mar 9. PMID: 35262217; PMCID: PMC9314680.

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MDS Abstracts - https://www.mdsabstracts.org/abstract/investigating-phospho-%ce%b1-synuclein-pathology-in-patient-specific-microglia-integrated-into-midbrain-organoids/