Functional stability of the second mitotic spindle in blastomeres and its association with multinucleation repair

Functional stability of the second mitotic spindle in blastomeres and its association with multinucleation repair | 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 Functional stability of the second mitotic spindle in blastomeres and its association with multinucleation repair Taichi Sakaguchi, Yukihiro Terada, Hiromitsu Shirasawa, Yuki Ono, and 15 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4770513/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract During human embryonic development, early cleavage divisions are prone to errors, with abnormal division, non-segregation of chromosomes, and multinucleation often observed during the first mitosis. Some studies suggest that the nuclear status at the 2 cell stage does not affect blastocyst progression or implantation; however, chromosomal aneuploidy at the blastocyst stage is linked to the nuclear status at the 4-cell stage. We fluorescently labelled DNA and microtubules in 21 blastomeres of human 2-cell embryos and used confocal microscopy to study spindle morphology, chromosomal dynamics, and nuclear status during the second mitosis. We found significantly fewer multinucleated blastomeres in 4-cell embryos than in 2-cell embryos (22% vs. 56%: P=0.03). The second mitotic spindles were bipolar in 14 and multipolar in 7 of the 21 blastomeres. Most daughter cells from bipolar spindles were mononuclear. Further, the second mitotic bipolar spindles showed no defocusing poles. Pre-implantation genetic testing for aneuploidy analysis showed that multinucleation at the 4-cell stage did not necessarily reflect aneuploidy at the blastocyst stage. Errors in the first mitosis might be repaired in the second, suggesting that understanding early mitosis can improve embryo evaluation methods. Biological sciences/Developmental biology/Embryology Biological sciences/Developmental biology/Embryogenesis Full Text Additional Declarations There is NO Competing Interest. Supplementary Files S1.mp4 Supplemental Movie 1 S2.mp4 Supplemental Movie 2 S3.mp4 Supplemental Movie 3 S4.mp4 Supplemental Movie 4 S5.mp4 Supplemental Movie 5 Cite Share Download PDF Status: Posted Version 1 posted 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. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. 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-4770513","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":331856388,"identity":"4a1ff147-7627-4d57-8dc0-a9bdb3dae26b","order_by":0,"name":"Taichi Sakaguchi","email":"data:image/png;base64,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","orcid":"","institution":"Akita University","correspondingAuthor":true,"prefix":"","firstName":"Taichi","middleName":"","lastName":"Sakaguchi","suffix":""},{"id":331856389,"identity":"cb808d39-fed5-46e6-8442-b58b20fae48c","order_by":1,"name":"Yukihiro Terada","email":"","orcid":"https://orcid.org/0000-0003-2392-5498","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Yukihiro","middleName":"","lastName":"Terada","suffix":""},{"id":331856390,"identity":"84cb5968-1d15-4b38-91b5-1bbc1bc00f36","order_by":2,"name":"Hiromitsu Shirasawa","email":"","orcid":"","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Hiromitsu","middleName":"","lastName":"Shirasawa","suffix":""},{"id":331856391,"identity":"42bec771-a918-4c86-b16f-d71c0f15db9e","order_by":3,"name":"Yuki Ono","email":"","orcid":"https://orcid.org/0009-0003-8083-7555","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Yuki","middleName":"","lastName":"Ono","suffix":""},{"id":331856392,"identity":"7d7d1a4a-f94d-4962-81d3-a07ed4066a24","order_by":4,"name":"Kazumasa Takahashi","email":"","orcid":"https://orcid.org/0000-0001-5424-7736","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Kazumasa","middleName":"","lastName":"Takahashi","suffix":""},{"id":331856393,"identity":"cdb904d1-1e89-470f-8848-8b25abb202a9","order_by":5,"name":"Mayumi Goto","email":"","orcid":"","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Mayumi","middleName":"","lastName":"Goto","suffix":""},{"id":331856394,"identity":"465f5b67-8635-4c64-ae92-b53d70b1d1f7","order_by":6,"name":"Motonari Okabe","email":"","orcid":"","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Motonari","middleName":"","lastName":"Okabe","suffix":""},{"id":331856395,"identity":"fd4e75aa-9cef-4895-a703-732f97ff81b7","order_by":7,"name":"Chika Ariake","email":"","orcid":"","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Chika","middleName":"","lastName":"Ariake","suffix":""},{"id":331856396,"identity":"e8a639ff-4f5b-4196-ae05-1b873076078a","order_by":8,"name":"Akari Tuya","email":"","orcid":"","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Akari","middleName":"","lastName":"Tuya","suffix":""},{"id":331856397,"identity":"efef2853-5217-42e3-8e7f-f0c9f3cf47cf","order_by":9,"name":"Takeo Hirakawa","email":"","orcid":"https://orcid.org/0000-0003-2085-3927","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Takeo","middleName":"","lastName":"Hirakawa","suffix":""},{"id":331856398,"identity":"11827700-7e12-4a8a-9ee4-b0a2a2b2f5e8","order_by":10,"name":"Takuya Iwasawa","email":"","orcid":"","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Takuya","middleName":"","lastName":"Iwasawa","suffix":""},{"id":331856399,"identity":"0e898a2d-2031-4454-a9e5-75e3e7b16f24","order_by":11,"name":"Akiko Fujishima","email":"","orcid":"","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Akiko","middleName":"","lastName":"Fujishima","suffix":""},{"id":331856400,"identity":"49fcda9d-78b2-46a2-81fe-f5fdee0f96ab","order_by":12,"name":"Yohei Onodera","email":"","orcid":"","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Yohei","middleName":"","lastName":"Onodera","suffix":""},{"id":331856401,"identity":"1245b4f6-28d1-4571-91e8-fc566d9a9280","order_by":13,"name":"Tae Sugawara","email":"","orcid":"","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Tae","middleName":"","lastName":"Sugawara","suffix":""},{"id":331856404,"identity":"a26ec815-9e3f-41c9-ad9e-edda696f433c","order_by":14,"name":"Kenichi Makino","email":"","orcid":"","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Kenichi","middleName":"","lastName":"Makino","suffix":""},{"id":331856406,"identity":"d2688111-ffb6-42e7-b40d-ddb18cdb73c9","order_by":15,"name":"Hiroshi Miura","email":"","orcid":"","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Hiroshi","middleName":"","lastName":"Miura","suffix":""},{"id":331856409,"identity":"b20233fa-2c07-40c3-b5a7-915bc993756b","order_by":16,"name":"Yukiyo Kumazawa","email":"","orcid":"","institution":"Akita University","correspondingAuthor":false,"prefix":"","firstName":"Yukiyo","middleName":"","lastName":"Kumazawa","suffix":""},{"id":331856410,"identity":"144db690-11de-4091-accc-db60d18cb2be","order_by":17,"name":"Noritaka Fukunaga","email":"","orcid":"https://orcid.org/0000-0003-2161-3319","institution":"Asada Ladies Clinic","correspondingAuthor":false,"prefix":"","firstName":"Noritaka","middleName":"","lastName":"Fukunaga","suffix":""},{"id":331856412,"identity":"c2b982cc-fa7b-43be-8461-87d1ce392a55","order_by":18,"name":"Yoshimasa Asada","email":"","orcid":"","institution":"Asada Institute for Reproductive Medicine, Asada Ladies Clinic","correspondingAuthor":false,"prefix":"","firstName":"Yoshimasa","middleName":"","lastName":"Asada","suffix":""}],"badges":[],"createdAt":"2024-07-19 23:10:14","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4770513/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4770513/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":62989293,"identity":"ff104cb6-2729-4990-8407-18845cfd816f","added_by":"auto","created_at":"2024-08-21 21:03:32","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":4378162,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4770513/v1_covered_02994b83-da56-45b8-a977-524a2f62484f.pdf"},{"id":61378778,"identity":"88d51374-e061-4cff-b6fd-18a51269fe1c","added_by":"auto","created_at":"2024-07-30 05:07:05","extension":"mp4","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":9676894,"visible":true,"origin":"","legend":"Supplemental Movie 1","description":"","filename":"S1.mp4","url":"https://assets-eu.researchsquare.com/files/rs-4770513/v1/2afd0e1407d6dd2e34b17ae1.mp4"},{"id":61378776,"identity":"f533b995-983c-4918-978c-937d4bface5e","added_by":"auto","created_at":"2024-07-30 05:07:05","extension":"mp4","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":8556076,"visible":true,"origin":"","legend":"Supplemental Movie 2","description":"","filename":"S2.mp4","url":"https://assets-eu.researchsquare.com/files/rs-4770513/v1/083b82b93420d0dc69a0e18d.mp4"},{"id":61378777,"identity":"b65f960f-19d4-486e-a075-c4696039e9f9","added_by":"auto","created_at":"2024-07-30 05:07:05","extension":"mp4","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":11873062,"visible":true,"origin":"","legend":"Supplemental Movie 3","description":"","filename":"S3.mp4","url":"https://assets-eu.researchsquare.com/files/rs-4770513/v1/92feed0eed3f755c08dd1167.mp4"},{"id":61378779,"identity":"595dace5-86af-4f0b-aa1c-ad6a3f4dc245","added_by":"auto","created_at":"2024-07-30 05:07:06","extension":"mp4","order_by":4,"title":"","display":"","copyAsset":false,"role":"supplement","size":24814129,"visible":true,"origin":"","legend":"\u003cp\u003eSupplemental Movie 4\u003c/p\u003e","description":"","filename":"S4.mp4","url":"https://assets-eu.researchsquare.com/files/rs-4770513/v1/6c35e0bba90232ba35009fcf.mp4"},{"id":61378775,"identity":"6cf3b8fd-ebef-41aa-96b5-2e7e38776afb","added_by":"auto","created_at":"2024-07-30 05:07:05","extension":"mp4","order_by":5,"title":"","display":"","copyAsset":false,"role":"supplement","size":11581064,"visible":true,"origin":"","legend":"\u003cp\u003eSupplemental Movie 5\u003c/p\u003e","description":"","filename":"S5.mp4","url":"https://assets-eu.researchsquare.com/files/rs-4770513/v1/2a3d858ebb0897894b8a55bb.mp4"}],"financialInterests":"There is \u003cb\u003eNO\u003c/b\u003e Competing Interest.","formattedTitle":"Functional stability of the second mitotic spindle in blastomeres and its association \r\nwith multinucleation repair","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-4770513/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4770513/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"During human embryonic development, early cleavage divisions are prone to errors, \r\nwith abnormal division, non-segregation of chromosomes, and multinucleation often \r\nobserved during the first mitosis. Some studies suggest that the nuclear status at the 2\r\ncell stage does not affect blastocyst progression or implantation; however, chromosomal \r\naneuploidy at the blastocyst stage is linked to the nuclear status at the 4-cell stage. We \r\nfluorescently labelled DNA and microtubules in 21 blastomeres of human 2-cell \r\nembryos and used confocal microscopy to study spindle morphology, chromosomal \r\ndynamics, and nuclear status during the second mitosis. We found significantly fewer \r\nmultinucleated blastomeres in 4-cell embryos than in 2-cell embryos (22% vs. 56%: \r\nP=0.03). The second mitotic spindles were bipolar in 14 and multipolar in 7 of the 21 \r\nblastomeres. Most daughter cells from bipolar spindles were mononuclear. Further, the \r\nsecond mitotic bipolar spindles showed no defocusing poles. Pre-implantation genetic \r\ntesting for aneuploidy analysis showed that multinucleation at the 4-cell stage did not \r\nnecessarily reflect aneuploidy at the blastocyst stage. Errors in the first mitosis might be \r\nrepaired in the second, suggesting that understanding early mitosis can improve embryo \r\nevaluation methods.","manuscriptTitle":"Functional stability of the second mitotic spindle in blastomeres and its association \nwith multinucleation repair","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-07-30 05:07:00","doi":"10.21203/rs.3.rs-4770513/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"7c53b13b-cba7-491f-9b79-d1d1bea7a319","owner":[],"postedDate":"July 30th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":35139298,"name":"Biological sciences/Developmental biology/Embryology"},{"id":35139299,"name":"Biological sciences/Developmental biology/Embryogenesis"}],"tags":[],"updatedAt":"2024-12-04T10:53:15+00:00","versionOfRecord":[],"versionCreatedAt":"2024-07-30 05:07:00","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-4770513","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4770513","identity":"rs-4770513","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}