Extrinsic Apoptosis and Necroptosis in Telencephalic Development: A Single-Cell Mass Cytometry Study

Extrinsic Apoptosis and Necroptosis in Telencephalic Development: A Single-Cell Mass Cytometry Study | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article Extrinsic Apoptosis and Necroptosis in Telencephalic Development: A Single-Cell Mass Cytometry Study Christopher Deppmann, JIACHEN SHI, Weile Liu, Alison Song, Timi Sanni, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6221176/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 21 Oct, 2025 Read the published version in Cell Death & Differentiation → Version 1 posted 9 You are reading this latest preprint version Abstract Regulated cell death is integral to sculpting the developing brain, yet the relative contributions of extrinsic apoptosis and necroptosis remain unclear. Here, we leverage single-cell mass cytometry (CyTOF) to characterize the cellular landscape of the mouse telencephalon in wild-type (WT), RIPK3 knockout (RIPK3 KO), and RIPK3/Caspase-8 double knockout (DKO) mice. Strikingly, combined deletion of RIPK3 and Caspase-8 leads to a 12.6% increase in total cell count, challenging the prevailing notion that intrinsic apoptosis exclusively governs developmental cell elimination. Detailed subpopulation analysis reveals that DKO mice display selective enrichment of Tbr2⁺ intermediate progenitors and endothelial cells, underscoring distinct, cell type–specific roles for extrinsic apoptotic and necroptotic pathways. These findings provide a revised framework for understanding the coordinated regulation of cell number during telencephalic development and suggest potential mechanistic links to neurodevelopmental disorders characterized by aberrant cell death. Biological sciences/Neuroscience Biological sciences/Cell biology Caspase-8 RIPK3 Telencephalon Mass cytometry Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Full Text Additional Declarations There is no duality of interest Supplementary Files SupplementaryTable1.pdf Supplementary Table 1 SupplementaryFig1.png Supplementary Figure 1 SupplementaryFig2.png Supplementary Figure 2 SupplementaryFig3.png Supplementary Figure 3 SupplementaryFig4.png Supplementary Figure 4 SupplementaryFig5.png Supplementary Figure 5 SupplementaryFig6.png Supplementary Figure 6 SupplementaryFig7.png Supplementary Figure 7 SupplementaryFig8.png Supplementary Figure 8 SupplementaryFig9.png Supplementary Figure 9 SupplementaryFig10.png Supplementary Figure 10 Cite Share Download PDF Status: Published Journal Publication published 21 Oct, 2025 Read the published version in Cell Death & Differentiation → Version 1 posted Editorial decision: revise 24 Apr, 2025 Review # 2 received at journal 19 Apr, 2025 Reviewer # 2 agreed at journal 10 Apr, 2025 Review # 1 received at journal 03 Apr, 2025 Reviewer # 1 agreed at journal 21 Mar, 2025 Reviewers invited by journal 20 Mar, 2025 Submission checks completed at journal 14 Mar, 2025 Editor assigned by journal 13 Mar, 2025 First submitted to journal 13 Mar, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-6221176","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":431824427,"identity":"f38f8b35-7253-4f1b-a9e4-08346674cc4d","order_by":0,"name":"Christopher Deppmann","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA9ElEQVRIiWNgGAWjYDCCA1Can4GxgYHBgIEHxJEgSotkA8laDA4gCeLVwne899ijGzV38oxvJLd9/FJwR4a/gfngbR48WiTPnEs3zjn2rNjsRmLzbBmDZzwSB9iSrfFpMbiRYyadw3Y4cduZg83MEgaHeRgO8JhJ49Vy/w1Qy7/DiZt7oFrkD/B/w6/lBtDM3LbDiRvYG5sZPwC1GBzgYcOrRfJMXrpxbt/hxBnHG5uZGYBaDA+zGVvOwaOF7/jZY49zvh1O7G9mf8z4489he7njzQ9vvMGjhYGBhw3OZAa7hxmvcjQtjD8Iqh4Fo2AUjIKRCACmcFGSlPHbygAAAABJRU5ErkJggg==","orcid":"https://orcid.org/0000-0002-6591-1767","institution":"University of Virginia","correspondingAuthor":true,"prefix":"","firstName":"Christopher","middleName":"","lastName":"Deppmann","suffix":""},{"id":431824428,"identity":"79b6aaba-d905-4779-b631-6bdadeacb766","order_by":1,"name":"JIACHEN SHI","email":"","orcid":"","institution":"University of Virginia","correspondingAuthor":false,"prefix":"","firstName":"JIACHEN","middleName":"","lastName":"SHI","suffix":""},{"id":431824429,"identity":"1c8bee0d-2fb8-4d30-a411-bd65d1a43eb8","order_by":2,"name":"Weile Liu","email":"","orcid":"","institution":"University of Virginia","correspondingAuthor":false,"prefix":"","firstName":"Weile","middleName":"","lastName":"Liu","suffix":""},{"id":431824430,"identity":"f0009fb7-378e-434b-a829-a41c52b6c3a1","order_by":3,"name":"Alison Song","email":"","orcid":"https://orcid.org/0009-0001-6102-6867","institution":"University of Virginia","correspondingAuthor":false,"prefix":"","firstName":"Alison","middleName":"","lastName":"Song","suffix":""},{"id":431824431,"identity":"58e00f69-68b9-4e62-a541-b95c8328c259","order_by":4,"name":"Timi Sanni","email":"","orcid":"","institution":"University of Virginia","correspondingAuthor":false,"prefix":"","firstName":"Timi","middleName":"","lastName":"Sanni","suffix":""},{"id":431824432,"identity":"33f654a6-fe1c-4e07-a1cb-3e214daa0c43","order_by":5,"name":"Amy Van Deusen","email":"","orcid":"","institution":"University of Virginia","correspondingAuthor":false,"prefix":"","firstName":"Amy","middleName":"Van","lastName":"Deusen","suffix":""},{"id":431824433,"identity":"ec201785-61ad-41d1-a4af-2e1d67038de1","order_by":6,"name":"Eli Zunder","email":"","orcid":"","institution":"University of Virginia","correspondingAuthor":false,"prefix":"","firstName":"Eli","middleName":"","lastName":"Zunder","suffix":""}],"badges":[],"createdAt":"2025-03-13 14:56:17","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-6221176/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6221176/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1038/s41418-025-01594-5","type":"published","date":"2025-10-21T04:00:00+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":79065793,"identity":"1a062d9e-43cd-4751-a8b8-5ec0ff7b99cb","added_by":"auto","created_at":"2025-03-24 04:11:01","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":2134934,"visible":true,"origin":"","legend":"\u003cp\u003eCell Death, Proliferation, and Abundance in the Developing Telencephalon. (A) Schematic representation of the modified experimental workflow for including both live and \u0026nbsp;dying cells. Single-cell mass cytometry data from mouse telencephalon (E13-P4) were \u0026nbsp;reanalyzed to include both live and dying cells. Flow cytometry plots show the inclusion of \u0026nbsp;Cisplatin+ cells in the new analysis. (B) Temporal dynamics of CC3+%, Cisplatin+%, and Ki67+% cells throughout development (E13- P4). Individual replicates are shown along with Loess curve fitting of the data. \u0026nbsp;(C) UMAP visualization of 23 distinct cell clusters identified by Leiden clustering. Insets show \u0026nbsp;the expression patterns of CC3 (apoptosis), Cisplatin (plasma membrane integrity), and Ki67 \u0026nbsp;(proliferation) across the UMAP layout. Cell clusters identified as precursor and progenitor cell \u0026nbsp;types by their protein expression profiles are highlighted by a red dashed line in the Ki67 inset. (D) Detailed analysis of selected clusters representing immature neuronal, mature neuronal, \u0026nbsp;immature nonneuronal, and mature nonneuronal cell populations. Top: UMAP plots highlighting \u0026nbsp;the distribution of each cluster. Bottom: Temporal dynamics of cluster abundance% (gray), \u0026nbsp;CC3+ cells% (orange), Cisplatin+ cells% (green), and Ki67+ cells% (blue) from E13 to P4. \u0026nbsp;Individual replicates are shown along with Loess curve fitting of the data.\u003c/p\u003e","description":"","filename":"MainFig1.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/71307c4ea599fffe4761f957.png"},{"id":79066572,"identity":"1eb061ae-94f0-4067-b871-3ce1f16c10da","added_by":"auto","created_at":"2025-03-24 04:27:02","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":3900546,"visible":true,"origin":"","legend":"\u003cp\u003eInvestigation of Extrinsic Apoptosis and Necroptosis in Telencephalon \u0026nbsp;Development (A) Schematic representation of extrinsic apoptotic and necroptotic pathways in WT, RIPK3 KO, \u0026nbsp;Caspase-8 KO, and DKO mice. (B) Representative images of RNAscope in situ hybridization for Casp8 (magenta) and Ripk3 \u0026nbsp;(yellow) mRNA in P4 WT mouse cortex. DAPI (cyan) labels nuclei. Scale bar: 50 μm. (C) Quantification of total cell numbers in the telencephalon of P4 mice (n = 12 per genotype). \u0026nbsp;(D) Workflow schematic for mass cytometry analysis of P4 mouse telencephalon from WT, \u0026nbsp;RIPK3 KO and DKO mice, including tissue processing, antibody conjugation, sample staining, \u0026nbsp;and data acquisition and processing steps. (E) UMAP visualization of mass cytometry data showing 30 distinct cell clusters identified by \u0026nbsp;Leiden clustering. Major cell type categories are annotated. Insets show expression patterns of \u0026nbsp;CC3, Cisplatin, and Ki67 across the UMAP layout. (F) Violin plot showing relative expression of all 37 markers across the 30 identified cell clusters. \u0026nbsp;(G) Quantification of CC3+ , Cisplatin+ , and Ki67+ cells% across genotypes from mass cytometry \u0026nbsp;data. \u0026nbsp;For panels C and G, data presented as box plots where the middle line represents the median, \u0026nbsp;the box represents the interquartile range (IQR), and the whiskers extend to 1.5 times the IQR. \u0026nbsp;Individual data points are shown as dots. Statistical differences between groups are denoted by \u0026nbsp;asterisks. Statistical significance levels are as follows: p\u0026lt;0.05, * p\u0026lt;0.01, *** p\u0026lt;0.001, ns: not \u0026nbsp;significant (one-way ANOVA with Tukey's post-hoc test).\u003c/p\u003e","description":"","filename":"MainFig2.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/660bdf0ee449b15edf4ebd24.png"},{"id":79065795,"identity":"20f14848-5321-4f7a-883d-cd5d080b65ca","added_by":"auto","created_at":"2025-03-24 04:11:02","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":1786198,"visible":true,"origin":"","legend":"\u003cp\u003eDifferential Abundance of Telencephalic Neuronal Subpopulations (A) Initial clustering analysis showing UMAP visualization and quantification of four major \u0026nbsp;neuronal populations. For each cluster, relative abundance% is compared across WT, RIPK3 \u0026nbsp;KO, and DKO. (B) Refined clustering analysis showing UMAP visualization and quantification of key neuronal \u0026nbsp;subpopulations. Left UMAP plot displays 26 distinct neuronal subclusters (C1-C26) identified \u0026nbsp;through iterative Leiden clustering (colored overlay), with inset showing the analyzed clusters \u0026nbsp;from initial clustering (dark blue). Representative UMAP plots and quantification are shown for \u0026nbsp;seven key neuronal populations. For each cluster, relative abundance% is compared across \u0026nbsp;WT, RIPK3 KO, and DKO. (C) IHC validation of Tbr2+ cells. Left: Representative images of Tbr2 staining in the dentate \u0026nbsp;gyrus across genotypes. Right: Quantification of Tbr2+ cell numbers in the dentate gyrus. Scale \u0026nbsp;bar: 50 μm. \u0026nbsp;For all quantifications: Data are presented as box plots showing median (middle line), \u0026nbsp;interquartile range (box), and whiskers (1.5×IQR), with individual data points overlaid. Statistical \u0026nbsp;significance determined by one-way ANOVA with Tukey's post-hoc test (*p\u0026lt;0.05, **p\u0026lt;0.01, \u0026nbsp;***p\u0026lt;0.001, ns: not significant).\u003c/p\u003e","description":"","filename":"MainFig3.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/4c98ebe76fe7515254d46652.png"},{"id":79066045,"identity":"3de3d072-a1fa-457b-a891-43879edd91a6","added_by":"auto","created_at":"2025-03-24 04:19:02","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":2046673,"visible":true,"origin":"","legend":"\u003cp\u003eDifferential Abundance of Telencephalic Nonneuronal Subpopulations (A) Initial clustering analysis showing UMAP visualization and quantification of four major \u0026nbsp;nonneuronal populations. For each cluster, relative abundance% is compared across WT, \u0026nbsp;RIPK3 KO, and DKO.\u003c/p\u003e\n\u003cp\u003e(B) Refined clustering analysis showing UMAP visualization and quantification of key \u0026nbsp;endothelial, astrocytic, and nonneural subpopulations. Left UMAP plot displays 22 distinct \u0026nbsp;nonneuronal subclusters (C45-C66) identified through iterative Leiden clustering (colored \u0026nbsp;overlay), with inset showing the analyzed clusters from initial clustering (dark blue). \u0026nbsp;Representative UMAP plots and quantification are shown for seven key endothelial, astrocytic, \u0026nbsp;and nonneural subpopulations. For each cluster, relative abundance% is compared across WT, \u0026nbsp;RIPK3 KO, and DKO. C) IHC validation of PECAM+ area. Left: Representative images of PECAM staining in the \u0026nbsp;cortex across genotypes. Right: Quantification of PECAM+ Area% in the cortex. Scale bar: 50 \u0026nbsp;μm.\u003c/p\u003e","description":"","filename":"MainFig4.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/815b425f2853093f2962976b.png"},{"id":79066043,"identity":"363d169e-ab16-45fa-b95d-cdeab513eda4","added_by":"auto","created_at":"2025-03-24 04:19:02","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":1014748,"visible":true,"origin":"","legend":"\u003cp\u003eSubclustering Analysis Reveals Differential Glial Subpopulation Responses to \u0026nbsp;Necroptotic and Extrinsic Apoptotic Pathway Disruption (A) Refined clustering analysis showing UMAP visualization and quantification of key glial \u0026nbsp;lineage subpopulations. Left UMAP plot displays 22 distinct glial subclusters (C27-C44) \u0026nbsp;identified through iterative Leiden clustering (colored overlay), with inset showing the analyzed \u0026nbsp;clusters from initial clustering (dark blue). Representative UMAP plots and quantification are \u0026nbsp;shown for three key glial subpopulations. \u0026nbsp;(B) Refined clustering analysis showing UMAP visualization and quantification of key microglial, \u0026nbsp;blood, and monocyte subpopulations. Left UMAP plot displays 23 distinct microglial, blood, and \u0026nbsp;monocyte subclusters (C67-C89) identified through iterative Leiden clustering (colored overlay), \u0026nbsp;with inset showing the analyzed clusters from initial clustering (dark blue). Representative \u0026nbsp;UMAP plots and quantification are shown for three microglial, blood, and monocyte \u0026nbsp;subpopulations. For each cluster, relative abundance% is compared across WT, RIPK3 KO, and \u0026nbsp;DKO.\u003c/p\u003e","description":"","filename":"MainFig5.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/136308b4d2560bf5fe9a3c9e.png"},{"id":79066577,"identity":"70f3c0b4-abeb-4493-b18d-93f31e30c6fd","added_by":"auto","created_at":"2025-03-24 04:27:02","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":792624,"visible":true,"origin":"","legend":"\u003cp\u003eComprehensive Analysis of Death Pathway-Specific Effects on Telencephalic \u0026nbsp;Cell Population Dynamics (A) Model of developmental cell number regulation. Left: Cell numbers are balanced between \u0026nbsp;input processes (proliferation and differentiation into the population) and output mechanisms \u0026nbsp;(cell death and differentiation into other populations). Right: Potential mechanisms regulating \u0026nbsp;developmental cell reduction, including distinct death pathways (apoptosis, necroptosis), lineage \u0026nbsp;transitions, and compensatory responses.\u003c/p\u003e\n\u003cp\u003e(B) Summary of cell populations with distinct death signatures in the developing telencephalon \u0026nbsp;(Fig. 1, Supplementary Fig. S1, S2). UMAP visualization (top) and cluster identities (bottom) for \u0026nbsp;neuronal (left) and nonneuronal (right) populations. Clusters showing cleaved Caspase-3 \u0026nbsp;positivity are highlighted in orange, while those with Cisplatin uptake are shown in green. \u0026nbsp;Bidirectional arrows indicate populations positive for both markers. \u0026nbsp;(C) Summary of cell populations affected by death pathway deletion (Fig. 3-5). UMAP \u0026nbsp;visualization (top) and cluster identities (bottom) showing populations regulated by: extrinsic \u0026nbsp;apoptosis (left, purple, unique to DKO), necroptosis (middle, teal/green, RIPK3-dependent), or \u0026nbsp;both pathways (right, blue, sequential changes across genotypes). Arrows indicate increased \u0026nbsp;(↑) or decreased (↓) cluster abundance in knockout conditions.\u003c/p\u003e","description":"","filename":"MainFig6.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/559de43916ce7043a853ed86.png"},{"id":94062844,"identity":"b3101841-23b8-408a-928a-ed5aaeba39f7","added_by":"auto","created_at":"2025-10-22 07:19:02","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":3737863,"visible":true,"origin":"","legend":"Article File","description":"","filename":"ManuscriptCDDExtrinsicApoptosisandNecroptosisinTelencephalicDevelopment.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1_covered_8cf84e77-05b4-4c62-a284-ea2dfe6bac10.pdf"},{"id":79065794,"identity":"3388431e-6699-4242-ab6e-5bd55a73264d","added_by":"auto","created_at":"2025-03-24 04:11:01","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":38178,"visible":true,"origin":"","legend":"Supplementary Table 1","description":"","filename":"SupplementaryTable1.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/c032214ac0ab898a9c012847.pdf"},{"id":79066040,"identity":"96433d10-dd58-4fc8-94f6-34a7e732c6df","added_by":"auto","created_at":"2025-03-24 04:19:01","extension":"png","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":1311317,"visible":true,"origin":"","legend":"Supplementary Figure 1","description":"","filename":"SupplementaryFig1.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/a6a32ebc85c0832bb6398bdf.png"},{"id":79065799,"identity":"ba2b194d-8560-4ef3-b7a3-5f298eaa4ec2","added_by":"auto","created_at":"2025-03-24 04:11:02","extension":"png","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":939066,"visible":true,"origin":"","legend":"Supplementary Figure 2","description":"","filename":"SupplementaryFig2.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/99f14c8b16a17d296dadfd52.png"},{"id":79065811,"identity":"d3068157-50a3-4d2c-b205-3beb4b294fcc","added_by":"auto","created_at":"2025-03-24 04:11:02","extension":"png","order_by":4,"title":"","display":"","copyAsset":false,"role":"supplement","size":9650566,"visible":true,"origin":"","legend":"Supplementary Figure 3","description":"","filename":"SupplementaryFig3.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/25f10c99ce51894d9fbba758.png"},{"id":79065812,"identity":"9d34b87a-1b2a-4d16-a892-df255ce983ed","added_by":"auto","created_at":"2025-03-24 04:11:02","extension":"png","order_by":5,"title":"","display":"","copyAsset":false,"role":"supplement","size":3224662,"visible":true,"origin":"","legend":"Supplementary Figure 4","description":"","filename":"SupplementaryFig4.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/684c18b800834fb5974301c6.png"},{"id":79066052,"identity":"8294406a-ed5b-4589-85b6-6613e987364d","added_by":"auto","created_at":"2025-03-24 04:19:02","extension":"png","order_by":6,"title":"","display":"","copyAsset":false,"role":"supplement","size":1964124,"visible":true,"origin":"","legend":"Supplementary Figure 5","description":"","filename":"SupplementaryFig5.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/18850574df57f552fdd444b8.png"},{"id":79065817,"identity":"7e3ac8df-56a7-44e9-858d-dd84e6dfe461","added_by":"auto","created_at":"2025-03-24 04:11:03","extension":"png","order_by":7,"title":"","display":"","copyAsset":false,"role":"supplement","size":121212,"visible":true,"origin":"","legend":"Supplementary Figure 6","description":"","filename":"SupplementaryFig6.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/ffdb5157d22d2e46666f7c5e.png"},{"id":79065807,"identity":"0e257998-e4e8-4d0f-ad1a-83b3f8ade514","added_by":"auto","created_at":"2025-03-24 04:11:02","extension":"png","order_by":8,"title":"","display":"","copyAsset":false,"role":"supplement","size":783817,"visible":true,"origin":"","legend":"Supplementary Figure 7","description":"","filename":"SupplementaryFig7.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/e66f610cb3ab27157efdbd0a.png"},{"id":79065831,"identity":"7c1b9ae3-d026-4387-a922-8856573f7b14","added_by":"auto","created_at":"2025-03-24 04:11:03","extension":"png","order_by":9,"title":"","display":"","copyAsset":false,"role":"supplement","size":181892,"visible":true,"origin":"","legend":"Supplementary Figure 8","description":"","filename":"SupplementaryFig8.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/bf26a88aa0bca63a90ed3d90.png"},{"id":79066822,"identity":"abe05097-7633-410c-a6d7-a1b1817ca5b9","added_by":"auto","created_at":"2025-03-24 04:35:02","extension":"png","order_by":10,"title":"","display":"","copyAsset":false,"role":"supplement","size":2201996,"visible":true,"origin":"","legend":"Supplementary Figure 9","description":"","filename":"SupplementaryFig9.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/58a93a47ca5a1b3282bd36b6.png"},{"id":79066053,"identity":"51702880-e3ce-4b02-baa9-4c2af1bdad6c","added_by":"auto","created_at":"2025-03-24 04:19:03","extension":"png","order_by":11,"title":"","display":"","copyAsset":false,"role":"supplement","size":185155,"visible":true,"origin":"","legend":"Supplementary Figure 10","description":"","filename":"SupplementaryFig10.png","url":"https://assets-eu.researchsquare.com/files/rs-6221176/v1/f839f80d17415901d8a3e6d7.png"}],"financialInterests":"There is no duality of interest","formattedTitle":"Extrinsic Apoptosis and Necroptosis in Telencephalic Development: A Single-Cell Mass Cytometry Study","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"cell-death-and-differentiation","isNatureJournal":false,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"cdd","sideBox":"Learn more about [Cell Death \u0026 Differentiation](http://www.nature.com/cdd/)","snPcode":"41418","submissionUrl":"https://mts-cdd.nature.com/cgi-bin/main.plex","title":"Cell Death \u0026 Differentiation","twitterHandle":"@cddpress","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"ejp","reportingPortfolio":"Nature AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Caspase-8, RIPK3, Telencephalon, Mass cytometry","lastPublishedDoi":"10.21203/rs.3.rs-6221176/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6221176/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"Regulated cell death is integral to sculpting the developing brain, yet the relative contributions of extrinsic apoptosis and necroptosis remain unclear. Here, we leverage single-cell mass cytometry (CyTOF) to characterize the cellular landscape of the mouse telencephalon in wild-type (WT), RIPK3 knockout (RIPK3 KO), and RIPK3/Caspase-8 double knockout (DKO) mice. Strikingly, combined deletion of RIPK3 and Caspase-8 leads to a 12.6% increase in total cell count, challenging the prevailing notion that intrinsic apoptosis exclusively governs developmental cell elimination. Detailed subpopulation analysis reveals that DKO mice display selective enrichment of Tbr2⁺ intermediate progenitors and endothelial cells, underscoring distinct, cell type–specific roles for extrinsic apoptotic and necroptotic pathways. These findings provide a revised framework for understanding the coordinated regulation of cell number during telencephalic development and suggest potential mechanistic links to neurodevelopmental disorders characterized by aberrant cell death.","manuscriptTitle":"Extrinsic Apoptosis and Necroptosis in Telencephalic Development: A Single-Cell Mass Cytometry Study","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-03-24 04:10:57","doi":"10.21203/rs.3.rs-6221176/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"revise","date":"2025-04-24T09:21:18+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"This content is not available.","date":"2025-04-19T09:58:52+00:00","index":2,"fulltext":"This content is not available."},{"type":"reviewerAgreed","content":"This content is not available.","date":"2025-04-11T02:50:52+00:00","index":2,"fulltext":"This content is not available."},{"type":"editorInvitedReview","content":"This content is not available.","date":"2025-04-04T01:57:04+00:00","index":1,"fulltext":"This content is not available."},{"type":"reviewerAgreed","content":"This content is not available.","date":"2025-03-22T01:25:00+00:00","index":1,"fulltext":"This content is not available."},{"type":"reviewersInvited","content":"","date":"2025-03-21T01:41:00+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-03-14T12:24:51+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-03-13T14:52:44+00:00","index":"","fulltext":""},{"type":"submitted","content":"Cell Death \u0026 Differentiation","date":"2025-03-13T14:52:43+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"cell-death-and-differentiation","isNatureJournal":false,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"cdd","sideBox":"Learn more about [Cell Death \u0026 Differentiation](http://www.nature.com/cdd/)","snPcode":"41418","submissionUrl":"https://mts-cdd.nature.com/cgi-bin/main.plex","title":"Cell Death \u0026 Differentiation","twitterHandle":"@cddpress","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"ejp","reportingPortfolio":"Nature AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"ca3d5d99-88de-436d-b8f0-825bb3f56a6a","owner":[],"postedDate":"March 24th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[{"id":45996328,"name":"Biological sciences/Neuroscience"},{"id":45996329,"name":"Biological sciences/Cell biology"}],"tags":[],"updatedAt":"2025-10-22T07:18:47+00:00","versionOfRecord":{"articleIdentity":"rs-6221176","link":"https://doi.org/10.1038/s41418-025-01594-5","journal":{"identity":"cell-death-and-differentiation","isVorOnly":false,"title":"Cell Death \u0026 Differentiation"},"publishedOn":"2025-10-21 04:00:00","publishedOnDateReadable":"October 21st, 2025"},"versionCreatedAt":"2025-03-24 04:10:57","video":"","vorDoi":"10.1038/s41418-025-01594-5","vorDoiUrl":"https://doi.org/10.1038/s41418-025-01594-5","workflowStages":[]},"version":"v1","identity":"rs-6221176","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6221176","identity":"rs-6221176","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}