@article{19593,
  abstract     = {Prenatal immune challenges pose significant risks to human embryonic brain and eye development. However, our knowledge about the safe usage of anti-inflammatory drugs during pregnancy is still limited. While human induced pluripotent stem cells (hIPSC)-derived brain organoid models have started to explore functional consequences upon viral stimulation, these models commonly lack microglia, which are susceptible to and promote inflammation. Furthermore, microglia are actively involved in neuronal development. Here, we generate hIPSC-derived microglia precursor cells and assemble them into retinal organoids. Once the outer plexiform layer forms, these hIPSC-derived microglia (iMG) fully integrate into the retinal organoids. Since the ganglion cell survival declines by this time in 3D-retinal organoids, we adapted the model into 2D and identify that the improved ganglion cell number significantly decreases only with iMG presence. In parallel, we applied the immunostimulant POLY(I:C) to mimic a fetal viral infection. While POLY(I:C) exposure alters the iMG phenotype, it does not hinder their interaction with ganglion cells. Furthermore, iMG significantly enhance the supernatant’s inflammatory secretome and increase retinal cell proliferation. Simultaneous exposure with the non-steroidal anti-inflammatory drug (NSAID) ibuprofen dampens POLY(I:C)-mediated changes of the iMG phenotype and ameliorates cell proliferation. Remarkably, while POLY(I:C) disrupts neuronal calcium dynamics independent of iMG, ibuprofen rescues this effect only if iMG are present. Mechanistically, ibuprofen targets the enzymes cyclooxygenase 1 and 2 (COX1/PTGS1 and COX2/PTGS2) simultaneously, from which iMG mainly express COX1. Selective COX1 blockage fails to restore the calcium peak amplitude upon POLY(I:C) stimulation, suggesting ibuprofen’s beneficial effect depends on the presence and interplay of COX1 and COX2. These findings underscore the importance of microglia in the context of prenatal immune challenges and provide insight into the mechanisms by which ibuprofen exerts its protective effects during embryonic development.},
  author       = {Hübschmann, Verena and Korkut, Medina and Venturino, Alessandro and Maya-Arteaga, Juan Pablo and Siegert, Sandra},
  issn         = {1742-2094},
  journal      = {Journal of Neuroinflammation},
  number       = {1},
  publisher    = {Springer Nature},
  title        = {{Microglia determine an immune-challenged environment and facilitate ibuprofen action in human retinal organoids}},
  doi          = {10.1186/s12974-025-03366-x},
  volume       = {22},
  year         = {2025},
}

@article{12117,
  abstract     = {To understand how potential gene manipulations affect in vitro microglia, we provide a set of short protocols to evaluate microglia identity and function. We detail steps for immunostaining to determine microglia identity. We describe three functional assays for microglia: phagocytosis, calcium response following ATP stimulation, and cytokine expression upon inflammatory stimuli. We apply these protocols to human induced-pluripotent-stem-cell (hiPSC)-derived microglia, but they can be also applied to other in vitro microglial models including primary mouse microglia.
For complete details on the use and execution of this protocol, please refer to Bartalska et al. (2022).1},
  author       = {Hübschmann, Verena and Korkut, Medina and Siegert, Sandra},
  issn         = {2666-1667},
  journal      = {STAR Protocols},
  keywords     = {General Immunology and Microbiology, General Biochemistry, Genetics and Molecular Biology, General Neuroscience},
  number       = {4},
  publisher    = {Elsevier},
  title        = {{Assessing human iPSC-derived microglia identity and function by immunostaining, phagocytosis, calcium activity, and inflammation assay}},
  doi          = {10.1016/j.xpro.2022.101866},
  volume       = {3},
  year         = {2022},
}

@article{11478,
  abstract     = {Cerebral organoids differentiated from human-induced pluripotent stem cells (hiPSC) provide a unique opportunity to investigate brain development. However, organoids usually lack microglia, brain-resident immune cells, which are present in the early embryonic brain and participate in neuronal circuit development. Here, we find IBA1+ microglia-like cells alongside retinal cups between week 3 and 4 in 2.5D culture with an unguided retinal organoid differentiation protocol. Microglia do not infiltrate the neuroectoderm and instead enrich within non-pigmented, 3D-cystic compartments that develop in parallel to the 3D-retinal organoids. When we guide the retinal organoid differentiation with low-dosed BMP4, we prevent cup development and enhance microglia and 3D-cysts formation. Mass spectrometry identifies these 3D-cysts to express mesenchymal and epithelial markers. We confirmed this microglia-preferred environment also within the unguided protocol, providing insight into microglial behavior and migration and offer a model to study how they enter and distribute within the human brain.},
  author       = {Bartalska, Katarina and Hübschmann, Verena and Korkut, Medina and Cubero, Ryan J and Venturino, Alessandro and Rössler, Karl and Czech, Thomas and Siegert, Sandra},
  issn         = {2589-0042},
  journal      = {iScience},
  number       = {7},
  publisher    = {Elsevier},
  title        = {{A systematic characterization of microglia-like cell occurrence during retinal organoid differentiation}},
  doi          = {10.1016/j.isci.2022.104580},
  volume       = {25},
  year         = {2022},
}

@article{11995,
  abstract     = {G protein-coupled receptors (GPCRs) regulate processes ranging from immune responses to neuronal signaling. However, ligands for many GPCRs remain unknown, suffer from off-target effects or have poor bioavailability. Additionally, dissecting cell type-specific responses is challenging when the same GPCR is expressed on different cells within a tissue. Here, we overcome these limitations by engineering DREADD-based GPCR chimeras that bind clozapine-N-oxide and mimic a GPCR-of-interest. We show that chimeric DREADD-β2AR triggers responses comparable to β2AR on second messenger and kinase activity, post-translational modifications, and protein-protein interactions. Moreover, we successfully recapitulate β2AR-mediated filopodia formation in microglia, an immune cell capable of driving central nervous system inflammation. When dissecting microglial inflammation, we included two additional DREADD-based chimeras mimicking microglia-enriched GPR65 and GPR109A. DREADD-β2AR and DREADD-GPR65 modulate the inflammatory response with high similarity to endogenous β2AR, while DREADD-GPR109A shows no impact. Our DREADD-based approach allows investigation of cell type-dependent pathways without known endogenous ligands.},
  author       = {Schulz, Rouven and Korkut, Medina and Venturino, Alessandro and Colombo, Gloria and Siegert, Sandra},
  issn         = {2041-1723},
  journal      = {Nature Communications},
  publisher    = {Springer Nature},
  title        = {{Chimeric GPCRs mimic distinct signaling pathways and modulate microglia responses}},
  doi          = {10.1038/s41467-022-32390-1},
  volume       = {13},
  year         = {2022},
}