Astrocytic Ceruloplasmin Deficiency Triggers Iron Toxicity and Neurodegeneration in a LRRK2 Parkinson's Tri-Culture Model

Testa, V.; Montero-Munoz, J.; Baruffi, V.; Alzeer, M.; Mularoni, L.; Fernandez-Carasa, I.; Simmini, G.; Comellas-Comaposada, N.; Ake, F. A.; Richaud, Y.; Weissleder, C.; Amengual, J.; Fabregat, I.; Deleidi, M.; Plass, M.; Pluchino, S.; Raya, A.; Consiglio, A.

doi:10.1101/2025.08.01.668099

bioRxiv · 10.1101/2025.08.01.668099

Astrocytic Ceruloplasmin Deficiency Triggers Iron Toxicity and Neurodegeneration in a LRRK2 Parkinson's Tri-Culture Model

Testa, V.·Montero-Munoz, J.·Baruffi, V.·Alzeer, M.·Mularoni, L.·Fernandez-Carasa, I.·Simmini, G.·Comellas-Comaposada, N.·Ake, F. A.·Richaud, Y.·Weissleder, C.·Amengual, J.·Fabregat, I.·Deleidi, M.·Plass, M.·Pluchino, S.·Raya, A.·Consiglio, A.

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Abstract

Astrocytes and microglia carrying the LRRK2-G2019S mutation contribute to non-cell- autonomous dopaminergic neuron (DAn) degeneration in Parkinsons disease (PD), but the mechanisms underlying their interplay remain unclear. Here, we developed a novel induced pluripotent stem cell (iPSC)-derived tri-culture system comprising healthy DAn and either LRRK2-mutant or isogenic control iPSC-derived astrocytes and microglia. Using integrated functional assays and transcriptomic profiling, we found that mutant astrocytes adopt a hyperreactive state, driving microglial activation and subsequent DAn degeneration. Mechanistically, we identified a selective downregulation of ceruloplasmin (CP), a copper-dependent ferroxidase, in mutant astrocytes, leading to disrupted iron homeostasis with accumulation of Fe2+ and ROS. This iron dysregulation mediated both microglial reactivity and neurodegeneration. Notably, pharmacological restoration of CP re-established iron homeostasis, reduced microglial activation, and protected DAn from degeneration. Our findings uncover a novel astrocyte-microglia-neuron axis driving PD pathogenesis and showcase the power of our unique stem cell tri-culture platform for dissecting disease mechanisms and discovering therapeutic targets.

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Testa, V., Montero-Munoz, J., Baruffi, V., Alzeer, M., Mularoni, L., Fernandez-Carasa, I., Simmini, G., Comellas-Comaposada, N., Ake, F. A., Richaud, Y., Weissleder, C., Amengual, J., Fabregat, I., Deleidi, M., Plass, M., Pluchino, S., Raya, A., Consiglio, A.. 2025-08-02. Astrocytic Ceruloplasmin Deficiency Triggers Iron Toxicity and Neurodegeneration in a LRRK2 Parkinson's Tri-Culture Model. https://doi.org/10.1101/2025.08.01.668099

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