DISSECTING MICROGLIA-ASTROCYTE CROSSTALK AND CCL2-MEDIATED NEURONAL VULNERABILITY IN PARKINSON’S DISEASE USING IPSC-DERIVED MODELS

Parkinson’s disease (PD) is a common, incurable neurodegenerative disorder marked by progressive loss of dopamine-producing neurons (DAn) in the substantia nigra and accumulation of misfolded α-synuclein in surviving neurons. Growing evidence points to neuroinflammation as a central driver of PD onset and progression. However, the mechanisms by which disease-associated mutations shape intrinsic microglial inflammatory states over time remain unclear. To address this, we used human iPSC-derived microglia (MG) to investigate inflammatory responses in both monoculture and multicellular co-culture systems. We first performed bulk RNA sequencing of MG carrying the familial PD mutation LRRK2-G2019S (LRRK2-PD) and their isogenic-corrected counterparts (ISO-PD), under basal conditions and following lipopolysaccharide (LPS) stimulation. Remarkably, expression of microglial homeostatic genes was observed only in ISO-PD MG after LPS treatment. Next, to study inflammation in a more disease-relevant context, we integrated LRRK2-PD MG into a human iPSC-derived co-culture with healthy DAn and either ISO-PD or LRRK2-PD astrocytes, and performed single-cell RNA sequencing. At early timepoints, the pro-inflammatory chemokine CCL2 was elevated in both ISO-PD and LRRK2-PD MG. By later timepoints, CCL2 expression in MG co-cultured with wild-type astrocytes was significantly reduced, indicating a shift toward a less reactive microglial state. Importantly, pharmacological inhibition of CCL2 rescued DAn degeneration, demonstrating that CCL2-dependent pathways drive neuronal vulnerability in this system. Future studies will dissect astrocyte-induced transcriptional programs that promote microglial homeostasis. Comparing these signatures with monoculture profiles will reveal how astrocyte-mediated cues restore homeostatic gene expression and constrain inflammatory activation.