Figure 1

Transcriptional profile of PD-iMGL carrying LRRK2-G2019S and shared PD-microglial expression signature (a) Graphical presentation of the differentiation protocol of iPSC-derived microglial-like cells (iMGL). iPSCs are guided to iMGL by the introduction of different cytokines and environmental factors. iPSCs differentiate towards mesoderm (meso.) and hemangioblast (hemo.) in hypoxic conditions. Emerging erythromyeloid progenitors (EMP) are collected and further differentiated in ultra-low attachment (ULA) culture vessels. For experiments, iMGL progenitors are seeded on Poly-d-lysine (PDL) coated culture vessels and the cells are matured for one week before functional analysis. Impact of LRRK2-G2019S in iMGL was studied with RNAseq of vehicle and interferon-γ (IFNγ) stimulated iMGL. (b) Cell type signature analysis of our iMGL and (iPSC-)microglia and monocytes from Abud et al. (2017) dataset²⁹ showing high correlation of microglial cell type-specific expression profiles. (c) iMGL express LRRK2 and the expression is increased by IFNγ. Expression of LRRK2 in iMGL as Log2 CPM (count per million) and Log2 Fold change LRRK2 expression in IFNγ stimulated iMGL. (d) Western blot analysis of LRRK2 and phosphorylated RAB10 (pT73) in vehicle and IFNγ stimulated iMGL. n = 3 batches, data are presented as mean ± SD. Two-way Anova with Sidak ´s multiple comparison test, *p < 0.05, **p < 0.01, ***p < 0.001. (e) Ingenuity pathway analysis (IPA) how functional annotation of DEGs between f-PD1 and f-ISO1 iMGL. (f) Comparison of iMGL transcriptional profile to different cell types extracted from snRNA-SEQ of midbrain³⁰, showing that that the majority our iMGL resemble the native microglia at the transcriptional level. (g) Shared pathways describing a PD-microglial profile analysed from DEGs between f-PD1/f-ISO1 iMGL & PD and Control microglia. (h) Simplified neuroinflammation pathway identified by IPA. For iMGL, n = 3–4 batches of iMGL, with 2 million cells used for each batch (subpanel b–f), n = 2 independently collected batches for both iPSC genotypes (subpanel b). For snSEQ, more details in the original publications³⁰, n = 6 controls (22,433 nuclei in total), n = 5 idiopathic PD patients (19 002 nuclei in total), (subpanels e,f).