--- _id: '11478' abstract: - lang: eng text: 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. acknowledged_ssus: - _id: Bio - _id: LifeSc acknowledgement: We thank the scientific service units at ISTA, specifically the lab support facility and imaging & optics facility for their support; Nicolas Armel for performing the Mass Spectrometry. We thank Alexandra Lang and Tanja Peilnsteiner for their help in human brain tissue collection, Rouven Schulz for his insights into the functional assays We thank all members of the Siegert group for constant feedback on the project and Margaret Maes, Rouven Schulz, and Marco Benevento for feedback on the manuscript. This project has received funding from the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation program (grant No. 715571 to S.S.) and from the Gesellschaft für Forschungsförderung Niederösterreich (grant No. Sc19-017 to V.H.). article_number: '104580' article_processing_charge: Yes article_type: original author: - first_name: Katarina full_name: Bartalska, Katarina id: 4D883232-F248-11E8-B48F-1D18A9856A87 last_name: Bartalska - first_name: Verena full_name: Hübschmann, Verena id: 32B7C918-F248-11E8-B48F-1D18A9856A87 last_name: Hübschmann - first_name: Medina full_name: Korkut, Medina id: 4B51CE74-F248-11E8-B48F-1D18A9856A87 last_name: Korkut orcid: 0000-0003-4309-2251 - first_name: Ryan J full_name: Cubero, Ryan J id: 850B2E12-9CD4-11E9-837F-E719E6697425 last_name: Cubero orcid: 0000-0003-0002-1867 - first_name: Alessandro full_name: Venturino, Alessandro id: 41CB84B2-F248-11E8-B48F-1D18A9856A87 last_name: Venturino orcid: 0000-0003-2356-9403 - first_name: Karl full_name: Rössler, Karl last_name: Rössler - first_name: Thomas full_name: Czech, Thomas last_name: Czech - first_name: Sandra full_name: Siegert, Sandra id: 36ACD32E-F248-11E8-B48F-1D18A9856A87 last_name: Siegert orcid: 0000-0001-8635-0877 citation: ama: Bartalska K, Hübschmann V, Korkut M, et al. A systematic characterization of microglia-like cell occurrence during retinal organoid differentiation. iScience. 2022;25(7). doi:10.1016/j.isci.2022.104580 apa: Bartalska, K., Hübschmann, V., Korkut, M., Cubero, R. J., Venturino, A., Rössler, K., … Siegert, S. (2022). A systematic characterization of microglia-like cell occurrence during retinal organoid differentiation. IScience. Elsevier. https://doi.org/10.1016/j.isci.2022.104580 chicago: Bartalska, Katarina, Verena Hübschmann, Medina Korkut, Ryan J Cubero, Alessandro Venturino, Karl Rössler, Thomas Czech, and Sandra Siegert. “A Systematic Characterization of Microglia-like Cell Occurrence during Retinal Organoid Differentiation.” IScience. Elsevier, 2022. https://doi.org/10.1016/j.isci.2022.104580. ieee: K. Bartalska et al., “A systematic characterization of microglia-like cell occurrence during retinal organoid differentiation,” iScience, vol. 25, no. 7. Elsevier, 2022. ista: Bartalska K, Hübschmann V, Korkut M, Cubero RJ, Venturino A, Rössler K, Czech T, Siegert S. 2022. A systematic characterization of microglia-like cell occurrence during retinal organoid differentiation. iScience. 25(7), 104580. mla: Bartalska, Katarina, et al. “A Systematic Characterization of Microglia-like Cell Occurrence during Retinal Organoid Differentiation.” IScience, vol. 25, no. 7, 104580, Elsevier, 2022, doi:10.1016/j.isci.2022.104580. short: K. Bartalska, V. Hübschmann, M. Korkut, R.J. Cubero, A. Venturino, K. Rössler, T. Czech, S. Siegert, IScience 25 (2022). date_created: 2022-07-03T22:01:33Z date_published: 2022-07-15T00:00:00Z date_updated: 2023-11-02T12:21:33Z day: '15' ddc: - '610' department: - _id: SaSi doi: 10.1016/j.isci.2022.104580 ec_funded: 1 external_id: isi: - '000830428500005' file: - access_level: open_access checksum: a470b74e1b3796c710189c81a4cd4329 content_type: application/pdf creator: cchlebak date_created: 2022-07-04T08:19:25Z date_updated: 2022-07-04T08:19:25Z file_id: '11480' file_name: 2022_iScience_Bartalska.pdf file_size: 19400048 relation: main_file success: 1 file_date_updated: 2022-07-04T08:19:25Z has_accepted_license: '1' intvolume: ' 25' isi: 1 issue: '7' language: - iso: eng month: '07' oa: 1 oa_version: Published Version project: - _id: 25D4A630-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '715571' name: Microglia action towards neuronal circuit formation and function in health and disease - _id: B67AFEDC-15C9-11EA-A837-991A96BB2854 name: IST Austria Open Access Fund - _id: 9B99D380-BA93-11EA-9121-9846C619BF3A grant_number: SC19-017 name: How human microglia shape developing neurons during health and inflammation publication: iScience publication_identifier: eissn: - 2589-0042 publication_status: published publisher: Elsevier quality_controlled: '1' related_material: record: - id: '12117' relation: other status: public scopus_import: '1' status: public title: A systematic characterization of microglia-like cell occurrence during retinal organoid differentiation tmp: image: /images/cc_by.png legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0) short: CC BY (4.0) type: journal_article user_id: 3E5EF7F0-F248-11E8-B48F-1D18A9856A87 volume: 25 year: '2022' ... --- _id: '12117' abstract: - lang: eng text: "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.\r\nFor complete details on the use and execution of this protocol, please refer to Bartalska et al. (2022).1" acknowledged_ssus: - _id: Bio acknowledgement: This project has received funding from the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation program (grant No. 715571 to S.S.) and from the Gesellschaft für Forschungsförderung Niederösterreich (grant No. Sc19-017 to V.H.). We thank Rouven Schulz and Alessandro Venturino for their insights into functional assays and data analysis, Verena Seiboth for insights into necessary institutional permission, and ISTA imaging & optics facility (IOF) especially Bernhard Hochreiter for their support. article_number: '101866' article_processing_charge: No article_type: letter_note author: - first_name: Verena full_name: Hübschmann, Verena id: 32B7C918-F248-11E8-B48F-1D18A9856A87 last_name: Hübschmann - first_name: Medina full_name: Korkut, Medina id: 4B51CE74-F248-11E8-B48F-1D18A9856A87 last_name: Korkut orcid: 0000-0003-4309-2251 - first_name: Sandra full_name: Siegert, Sandra id: 36ACD32E-F248-11E8-B48F-1D18A9856A87 last_name: Siegert orcid: 0000-0001-8635-0877 citation: ama: Hübschmann V, Korkut M, Siegert S. Assessing human iPSC-derived microglia identity and function by immunostaining, phagocytosis, calcium activity, and inflammation assay. STAR Protocols. 2022;3(4). doi:10.1016/j.xpro.2022.101866 apa: Hübschmann, V., Korkut, M., & Siegert, S. (2022). Assessing human iPSC-derived microglia identity and function by immunostaining, phagocytosis, calcium activity, and inflammation assay. STAR Protocols. Elsevier. https://doi.org/10.1016/j.xpro.2022.101866 chicago: Hübschmann, Verena, Medina Korkut, and Sandra Siegert. “Assessing Human IPSC-Derived Microglia Identity and Function by Immunostaining, Phagocytosis, Calcium Activity, and Inflammation Assay.” STAR Protocols. Elsevier, 2022. https://doi.org/10.1016/j.xpro.2022.101866. ieee: V. Hübschmann, M. Korkut, and S. Siegert, “Assessing human iPSC-derived microglia identity and function by immunostaining, phagocytosis, calcium activity, and inflammation assay,” STAR Protocols, vol. 3, no. 4. Elsevier, 2022. ista: Hübschmann V, Korkut M, Siegert S. 2022. Assessing human iPSC-derived microglia identity and function by immunostaining, phagocytosis, calcium activity, and inflammation assay. STAR Protocols. 3(4), 101866. mla: Hübschmann, Verena, et al. “Assessing Human IPSC-Derived Microglia Identity and Function by Immunostaining, Phagocytosis, Calcium Activity, and Inflammation Assay.” STAR Protocols, vol. 3, no. 4, 101866, Elsevier, 2022, doi:10.1016/j.xpro.2022.101866. short: V. Hübschmann, M. Korkut, S. Siegert, STAR Protocols 3 (2022). date_created: 2023-01-12T11:56:38Z date_published: 2022-12-16T00:00:00Z date_updated: 2023-11-02T12:21:32Z day: '16' ddc: - '570' department: - _id: SaSi - _id: GradSch doi: 10.1016/j.xpro.2022.101866 ec_funded: 1 file: - access_level: open_access checksum: 3c71b8a60633d42c2f77c49025d5559b content_type: application/pdf creator: dernst date_created: 2023-01-23T09:50:51Z date_updated: 2023-01-23T09:50:51Z file_id: '12340' file_name: 2022_STARProtocols_Huebschmann.pdf file_size: 6251945 relation: main_file success: 1 file_date_updated: 2023-01-23T09:50:51Z has_accepted_license: '1' intvolume: ' 3' issue: '4' keyword: - General Immunology and Microbiology - General Biochemistry - Genetics and Molecular Biology - General Neuroscience language: - iso: eng month: '12' oa: 1 oa_version: Published Version project: - _id: 25D4A630-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '715571' name: Microglia action towards neuronal circuit formation and function in health and disease - _id: 9B99D380-BA93-11EA-9121-9846C619BF3A grant_number: SC19-017 name: How human microglia shape developing neurons during health and inflammation publication: STAR Protocols publication_identifier: issn: - 2666-1667 publication_status: published publisher: Elsevier quality_controlled: '1' related_material: record: - id: '11478' relation: other status: public scopus_import: '1' status: public title: Assessing human iPSC-derived microglia identity and function by immunostaining, phagocytosis, calcium activity, and inflammation assay tmp: image: /images/cc_by_nc_nd.png legal_code_url: https://creativecommons.org/licenses/by-nc-nd/4.0/legalcode name: Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0) short: CC BY-NC-ND (4.0) type: journal_article user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87 volume: 3 year: '2022' ... --- _id: '11995' abstract: - lang: eng text: 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. acknowledged_ssus: - _id: PreCl - _id: Bio - _id: LifeSc acknowledgement: The authors thank the Scientific Service Units at ISTA, in particular the Molecular Biology Service of the Lab Support Facility, Imaging & Optics Facility, and the Preclinical Facility, and the Novarino group, Harald Janoviak, and Marco Benevento for sharing reagents and expertise. This research was supported by a DOC Fellowship (24979) awarded to R.S. by the Austrian Academy of Sciences. article_number: '4728' article_processing_charge: No article_type: original author: - first_name: Rouven full_name: Schulz, Rouven id: 4C5E7B96-F248-11E8-B48F-1D18A9856A87 last_name: Schulz orcid: 0000-0001-5297-733X - first_name: Medina full_name: Korkut, Medina id: 4B51CE74-F248-11E8-B48F-1D18A9856A87 last_name: Korkut orcid: 0000-0003-4309-2251 - first_name: Alessandro full_name: Venturino, Alessandro id: 41CB84B2-F248-11E8-B48F-1D18A9856A87 last_name: Venturino orcid: 0000-0003-2356-9403 - first_name: Gloria full_name: Colombo, Gloria id: 3483CF6C-F248-11E8-B48F-1D18A9856A87 last_name: Colombo orcid: 0000-0001-9434-8902 - first_name: Sandra full_name: Siegert, Sandra id: 36ACD32E-F248-11E8-B48F-1D18A9856A87 last_name: Siegert orcid: 0000-0001-8635-0877 citation: ama: Schulz R, Korkut M, Venturino A, Colombo G, Siegert S. Chimeric GPCRs mimic distinct signaling pathways and modulate microglia responses. Nature Communications. 2022;13. doi:10.1038/s41467-022-32390-1 apa: Schulz, R., Korkut, M., Venturino, A., Colombo, G., & Siegert, S. (2022). Chimeric GPCRs mimic distinct signaling pathways and modulate microglia responses. Nature Communications. Springer Nature. https://doi.org/10.1038/s41467-022-32390-1 chicago: Schulz, Rouven, Medina Korkut, Alessandro Venturino, Gloria Colombo, and Sandra Siegert. “Chimeric GPCRs Mimic Distinct Signaling Pathways and Modulate Microglia Responses.” Nature Communications. Springer Nature, 2022. https://doi.org/10.1038/s41467-022-32390-1. ieee: R. Schulz, M. Korkut, A. Venturino, G. Colombo, and S. Siegert, “Chimeric GPCRs mimic distinct signaling pathways and modulate microglia responses,” Nature Communications, vol. 13. Springer Nature, 2022. ista: Schulz R, Korkut M, Venturino A, Colombo G, Siegert S. 2022. Chimeric GPCRs mimic distinct signaling pathways and modulate microglia responses. Nature Communications. 13, 4728. mla: Schulz, Rouven, et al. “Chimeric GPCRs Mimic Distinct Signaling Pathways and Modulate Microglia Responses.” Nature Communications, vol. 13, 4728, Springer Nature, 2022, doi:10.1038/s41467-022-32390-1. short: R. Schulz, M. Korkut, A. Venturino, G. Colombo, S. Siegert, Nature Communications 13 (2022). date_created: 2022-08-28T22:01:59Z date_published: 2022-08-15T00:00:00Z date_updated: 2024-02-21T12:34:51Z day: '15' ddc: - '570' department: - _id: SaSi doi: 10.1038/s41467-022-32390-1 external_id: isi: - '000840984400032' pmid: - '35970889' file: - access_level: open_access checksum: 191d9db0266e14a28d3a56dc7f65da84 content_type: application/pdf creator: cchlebak date_created: 2022-08-29T06:44:30Z date_updated: 2022-08-29T06:44:30Z file_id: '12002' file_name: 2022_NatComm_Schulz.pdf file_size: 7317396 relation: main_file success: 1 file_date_updated: 2022-08-29T06:44:30Z has_accepted_license: '1' intvolume: ' 13' isi: 1 language: - iso: eng month: '08' oa: 1 oa_version: Published Version pmid: 1 project: - _id: 267F75D8-B435-11E9-9278-68D0E5697425 name: Modulating microglia through G protein-coupled receptor (GPCR) signaling publication: Nature Communications publication_identifier: eissn: - 2041-1723 publication_status: published publisher: Springer Nature quality_controlled: '1' related_material: link: - description: News on ISTA website relation: press_release url: https://ista.ac.at/en/news/dreaddful-mimicry/ record: - id: '11945' relation: part_of_dissertation status: public - id: '11542' relation: research_data status: public scopus_import: '1' status: public title: Chimeric GPCRs mimic distinct signaling pathways and modulate microglia responses tmp: image: /images/cc_by.png legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0) short: CC BY (4.0) type: journal_article user_id: 4359f0d1-fa6c-11eb-b949-802e58b17ae8 volume: 13 year: '2022' ...