GSDMD gene knockout alleviates hyperoxia-induced hippocampal brain injury in neonatal mice

GSDMD gene knockout alleviates hyperoxia-induced hippocampal brain injury in neonatal mice | 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 Research Article GSDMD gene knockout alleviates hyperoxia-induced hippocampal brain injury in neonatal mice Naga Venkata Divya Challa, Shaoyi Chen, Huijun Yun, Matthew R Duncan, and 7 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-3055085/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 07 Sep, 2023 Read the published version in Journal of Neuroinflammation → Version 1 posted 7 You are reading this latest preprint version Abstract Background: Neonatal hyperoxia exposure is associated with brain injury and poor neurodevelopment outcomes in preterm infants. Our previous studies in neonatal rodent models have shown that hyperoxia stimulates the brain’s inflammasome pathway, leading to the activation of gasdermin D (GSDMD), a key executor of pyroptotic inflammatory cell death. Moreover, we found inhibition of GSDMD activation attenuates hyperoxia-induced brain injury in neonatal mice. We hypothesized that GSDMD plays a pathogenic role in hyperoxia-induced neonatal brain injury and that GSDMD gene knockout (KO) will alleviate hyperoxia-induced brain injury. Methods: Newborn GSDMD knockout mice and their wildtype (WT) littermates were randomized within 24 h after birth to be exposed to room air or hyperoxia (85% O2) from postnatal day 1 to 14. Hippocampal brain inflammatory injury was assessed in brain sections by immunohistology for allograft inflammatory factor 1 (AIF1), a marker of microglial activation. Cell proliferation was evaluated by Ki-67 staining, and cell death was determined by TUNEL assay. RNA sequencing of the hippocampus was performed to identify the transcriptional effects of hyperoxia and GSDMD-KO, and qRT-PCR was performed to confirm some of the significantly regulated genes. Results: Hyperoxia-exposed WT mice had increased microglia consistent with activation, which was associated with decreased cell proliferation and increased cell death in the hippocampal area. Conversely, hyperoxia-exposed GSDMD-KO mice exhibited considerable resistance to hyperoxia as O2 exposure failed to increase either AIF1+ or TUNEL+ cell numbers, nor decrease cell proliferation. Hyperoxia exposure differentially regulated 258 genes in WT and only 16 in GSDMD-KO mice compared to room air- exposed WT and GSDMD-KO, respectively. Gene set enrichment analysis showed that in the WT brain, hyperoxia differentially regulated genes associated with neuronal and vascular development and differentiation, axonogenesis, glial cell differentiation, and core development pathways hypoxia-induced factor 1, and neuronal growth factor pathways. These changes were prevented by GSDMD-KO. Conclusion: GSDMD-KO alleviates hyperoxia-induced inflammatory injury, cell survival and death, and alterations of transcriptional gene expression of pathways involved in neuronal growth, development, and differentiation in the hippocampus of neonatal mice. This suggests that GSDMD plays a pathogenic role in preterm brain injury, and targeting GSDMD may be beneficial in preventing and treating brain injury and poor neurodevelopmental outcomes in preterm infants. GSDMD knockout neonatal brain injury pyroptosis microglial gene transcription Full Text Additional Declarations No competing interests reported. Cite Share Download PDF Status: Published Journal Publication published 07 Sep, 2023 Read the published version in Journal of Neuroinflammation → Version 1 posted Editorial decision: Major revision 04 Jul, 2023 Reviews received at journal 27 Jun, 2023 Reviewers agreed at journal 17 Jun, 2023 Reviewers invited by journal 17 Jun, 2023 Editor assigned by journal 13 Jun, 2023 Submission checks completed at journal 13 Jun, 2023 First submitted to journal 12 Jun, 2023 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. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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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-3055085","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":209402822,"identity":"cfa3e6a8-f94b-420e-bfca-63903de60a5f","order_by":0,"name":"Naga Venkata Divya Challa","email":"","orcid":"","institution":"University of Miami","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Naga","middleName":"Venkata Divya","lastName":"Challa","suffix":""},{"id":209402826,"identity":"8e1d0f43-22cf-4ab6-80cf-bc39fd4d9f59","order_by":1,"name":"Shaoyi Chen","email":"","orcid":"","institution":"University of 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Our previous studies in neonatal rodent models have shown that hyperoxia stimulates the brain’s inflammasome pathway, leading to the activation of gasdermin D (GSDMD), a key executor of pyroptotic inflammatory cell death. Moreover, we found inhibition of GSDMD activation attenuates hyperoxia-induced brain injury in neonatal mice. We hypothesized that GSDMD plays a pathogenic role in hyperoxia-induced neonatal brain injury and that GSDMD gene knockout (KO) will alleviate hyperoxia-induced brain injury. \u0026nbsp;\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethods:\u003c/strong\u003e Newborn GSDMD knockout mice and their wildtype (WT) littermates were randomized within 24 h after birth to be exposed to room air or hyperoxia (85% O2) from postnatal day 1 to 14. Hippocampal brain inflammatory injury was assessed in brain sections by immunohistology for allograft inflammatory factor 1 (AIF1), a marker of microglial activation. Cell proliferation was evaluated by Ki-67 staining, and cell death was determined by TUNEL assay. RNA sequencing of the hippocampus was performed to identify the transcriptional effects of hyperoxia and GSDMD-KO, and qRT-PCR was performed to confirm some of the significantly regulated genes. \u0026nbsp;\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults: \u003c/strong\u003eHyperoxia-exposed WT mice had increased microglia consistent with activation, which was associated with decreased cell proliferation and increased cell death in the hippocampal area. Conversely, hyperoxia-exposed GSDMD-KO mice exhibited considerable resistance to hyperoxia as O2 exposure failed to increase either AIF1+ or TUNEL+ cell numbers, nor decrease cell proliferation. Hyperoxia exposure differentially regulated 258 genes in WT and only 16 in GSDMD-KO mice compared to room air- exposed WT and GSDMD-KO, respectively. Gene set enrichment analysis showed that in the WT brain, hyperoxia differentially regulated genes associated with neuronal and vascular development and differentiation, axonogenesis, glial cell differentiation, and core development pathways hypoxia-induced factor 1, and neuronal growth factor pathways. These changes were prevented by GSDMD-KO. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConclusion:\u003c/strong\u003e GSDMD-KO alleviates hyperoxia-induced inflammatory injury, cell survival and death, and alterations of transcriptional gene expression of pathways involved in neuronal growth, development, and differentiation in the hippocampus of neonatal mice. This suggests that GSDMD plays a pathogenic role in preterm brain injury, and targeting GSDMD may be beneficial in preventing and treating brain injury and poor neurodevelopmental outcomes in preterm infants.\u003c/p\u003e","manuscriptTitle":"GSDMD gene knockout alleviates hyperoxia-induced hippocampal brain injury in neonatal mice","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2023-06-15 01:39:39","doi":"10.21203/rs.3.rs-3055085/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Major revision","date":"2023-07-04T04:24:47+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2023-06-27T11:22:41+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"a4bde209-5c7a-444c-97d3-8b7530f6e27e","date":"2023-06-17T12:43:41+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2023-06-17T12:40:09+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2023-06-13T21:27:25+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2023-06-13T09:05:13+00:00","index":"","fulltext":""},{"type":"submitted","content":"Journal of Neuroinflammation","date":"2023-06-12T21:37:28+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"journal-of-neuroinflammation","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"jneu","sideBox":"Learn more about [Journal of Neuroinflammation](http://jneuroinflammation.biomedcentral.com)","snPcode":"12974","submissionUrl":"https://submission.nature.com/new-submission/12974/3","title":"Journal of Neuroinflammation","twitterHandle":"@bmc","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"3bd048a9-43c1-446a-8bfd-22a702970f00","owner":[],"postedDate":"June 15th, 2023","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2023-09-11T15:08:51+00:00","versionOfRecord":{"articleIdentity":"rs-3055085","link":"https://doi.org/10.1186/s12974-023-02878-8","journal":{"identity":"journal-of-neuroinflammation","isVorOnly":false,"title":"Journal of Neuroinflammation"},"publishedOn":"2023-09-07 15:02:03","publishedOnDateReadable":"September 7th, 2023"},"versionCreatedAt":"2023-06-15 01:39:39","video":"","vorDoi":"10.1186/s12974-023-02878-8","vorDoiUrl":"https://doi.org/10.1186/s12974-023-02878-8","workflowStages":[]},"version":"v1","identity":"rs-3055085","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-3055085","identity":"rs-3055085","version":["v1"]},"buildId":"cBFmMYwuxLRRLfASyISRj","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}