Tauroursodeoxycholic acid (TUDCA)-induced increase in ectopic muscle mineralization occurs exclusively in dystrophic muscles and is independent of endoplasmic reticulum stress

Tauroursodeoxycholic acid (TUDCA)-induced increase in ectopic muscle mineralization occurs exclusively in dystrophic muscles and is independent of endoplasmic reticulum stress | 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 Tauroursodeoxycholic acid (TUDCA)-induced increase in ectopic muscle mineralization occurs exclusively in dystrophic muscles and is independent of endoplasmic reticulum stress Justyna Róg, Dorota Dymkowska, Bernadeta Michalska, Olga Krupska, and 8 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6401076/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 28 Oct, 2025 Read the published version in Scientific Reports → Version 1 posted 13 You are reading this latest preprint version Abstract Calcification of dystrophic skeletal muscles was described previously and attributed, among others, to ER-stress, elevated phosphate concentration and chronic inflammation. Tauroursodeoxycholic acid (TUDCA) has been broadly tested as a potential drug in therapy of various diseases. In particular it is considered an artificial chaperone protecting cells against ER-stress which could prevent an ectopic mineralisation of soft tissues . We hypothesised that TUDCA treatment should reduce muscle mineral deposits. The goal of this study was to test this concept using two mouse models of DMD. Four-week old mdx, mdx betageo and w/t mice were administered TUDCA in drinking water for 4 weeks. Then, following tissue-clearing and calcium minerals staining with alizarin, animal bodies were evaluated using whole body scanning Additionally, isolated skeletal muscles were analysed by Western blotting for ER-stress and calcification markers, and using various microscopic methods. Enzymatic activity of alkaline phosphatase was also assayed. Unexpectedly, TUDCA enhanced calcification of dystrophic but not dystrophin-positive muscles. TUDCA did not affect the elevated ER-stress markers found in dystrophic muscles nor impact pro-calcifying proteins RUNX2, Osterix and BMP2/4, which were overexpressed in dystrophic muscles. The alkaline phosphatase levels, which were reduced in dystrophic muscles, were not affected by this treatment. The increase in ectopic calcification in dystrophic muscles induced by TUDCA is specific to muscles lacking dystrophin. This effect is not linked to the alleviation of ER stress or the overexpression of proteins directly involved in calcium mineral accumulation, both of which are elevated in dystrophic muscles. Biological sciences/Biochemistry Biological sciences/Cell biology Health sciences/Pathogenesis Full Text Additional Declarations No competing interests reported. Supplementary Files SupplementaryforSR.pdf Cite Share Download PDF Status: Published Journal Publication published 28 Oct, 2025 Read the published version in Scientific Reports → Version 1 posted Editorial decision: Revision requested 12 Jun, 2025 Reviews received at journal 05 Jun, 2025 Reviews received at journal 27 May, 2025 Reviews received at journal 12 May, 2025 Reviewers agreed at journal 10 May, 2025 Reviewers agreed at journal 08 May, 2025 Reviewers agreed at journal 07 May, 2025 Reviewers agreed at journal 06 May, 2025 Reviewers invited by journal 05 May, 2025 Editor assigned by journal 25 Apr, 2025 Editor invited by journal 23 Apr, 2025 Submission checks completed at journal 22 Apr, 2025 First submitted to journal 08 Apr, 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-6401076","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":453456843,"identity":"acf57024-45af-4070-a1cd-70ded3826e80","order_by":0,"name":"Justyna Róg","email":"","orcid":"","institution":"Nencki Institute of Experimental Biology PAS","correspondingAuthor":false,"prefix":"","firstName":"Justyna","middleName":"","lastName":"Róg","suffix":""},{"id":453456844,"identity":"b634bba6-9e40-483a-a754-1f3c9e118866","order_by":1,"name":"Dorota Dymkowska","email":"","orcid":"","institution":"Nencki Institute of Experimental Biology PAS","correspondingAuthor":false,"prefix":"","firstName":"Dorota","middleName":"","lastName":"Dymkowska","suffix":""},{"id":453456845,"identity":"a06199bb-fbc5-4c3c-b99b-5bed26fbfe6d","order_by":2,"name":"Bernadeta Michalska","email":"","orcid":"","institution":"Nencki Institute of Experimental Biology PAS","correspondingAuthor":false,"prefix":"","firstName":"Bernadeta","middleName":"","lastName":"Michalska","suffix":""},{"id":453456846,"identity":"143d1b62-c44c-4da7-b555-a9d9c0c09159","order_by":3,"name":"Olga Krupska","email":"","orcid":"","institution":"Nencki Institute of Experimental Biology PAS","correspondingAuthor":false,"prefix":"","firstName":"Olga","middleName":"","lastName":"Krupska","suffix":""},{"id":453456847,"identity":"e560c608-e3ea-4217-8ea3-91e606b79e4c","order_by":4,"name":"Krzysztof Milewski","email":"","orcid":"","institution":"Nencki Institute of Experimental Biology PAS","correspondingAuthor":false,"prefix":"","firstName":"Krzysztof","middleName":"","lastName":"Milewski","suffix":""},{"id":453456848,"identity":"05698f21-7626-42f9-9b7e-77ea12d3b806","order_by":5,"name":"Paweł Matryba","email":"","orcid":"","institution":"Medical University of Warsaw","correspondingAuthor":false,"prefix":"","firstName":"Paweł","middleName":"","lastName":"Matryba","suffix":""},{"id":453456849,"identity":"643cb633-ced0-46b5-b85b-f73c9ca8c913","order_by":6,"name":"Artur Wolny","email":"","orcid":"","institution":"Nencki Institute of Experimental Biology PAS","correspondingAuthor":false,"prefix":"","firstName":"Artur","middleName":"","lastName":"Wolny","suffix":""},{"id":453456850,"identity":"e1258862-b0ae-411f-88e0-c81e9f0aeaed","order_by":7,"name":"Monika Pawłowska","email":"","orcid":"","institution":"Nencki Institute of Experimental Biology PAS","correspondingAuthor":false,"prefix":"","firstName":"Monika","middleName":"","lastName":"Pawłowska","suffix":""},{"id":453456851,"identity":"dcc69ef2-652c-4ffe-b278-77f64d1df73c","order_by":8,"name":"Łukasz Bożycki","email":"","orcid":"","institution":"Nencki Institute of Experimental Biology PAS","correspondingAuthor":false,"prefix":"","firstName":"Łukasz","middleName":"","lastName":"Bożycki","suffix":""},{"id":453456852,"identity":"a7c95eac-3dc3-4956-91ac-beae94427437","order_by":9,"name":"Dawid Stępnik","email":"","orcid":"","institution":"Nencki Institute of Experimental Biology PAS","correspondingAuthor":false,"prefix":"","firstName":"Dawid","middleName":"","lastName":"Stępnik","suffix":""},{"id":453456853,"identity":"bf0cf9bb-965c-4f64-8bfc-e88d32f0bb57","order_by":10,"name":"Dariusz C. 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Tauroursodeoxycholic acid (TUDCA) has been broadly tested as a potential drug in therapy of various diseases. In particular it is considered an artificial chaperone protecting cells against ER-stress which could prevent an ectopic mineralisation of soft\u0026nbsp; tissues . We hypothesised that TUDCA treatment should reduce muscle mineral deposits. The goal of this study was to test this concept using two mouse models of DMD.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eFour-week old mdx, mdx\u003csup\u003ebetageo\u003c/sup\u003e and w/t mice were administered TUDCA in drinking water for 4 weeks. Then, following tissue-clearing and calcium minerals staining with alizarin, animal bodies were evaluated using whole body scanning Additionally, isolated skeletal muscles were analysed by Western blotting for ER-stress and calcification markers, and using various microscopic methods. Enzymatic activity of alkaline phosphatase was also assayed. Unexpectedly, TUDCA enhanced calcification of dystrophic but not dystrophin-positive muscles. TUDCA did not affect the elevated ER-stress markers found in dystrophic muscles nor impact pro-calcifying proteins RUNX2, Osterix and BMP2/4, which were overexpressed in dystrophic muscles. The alkaline phosphatase levels, which were reduced in dystrophic muscles, were not affected by this treatment.\u0026nbsp;\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe increase in ectopic calcification in dystrophic muscles induced by TUDCA is specific to muscles lacking dystrophin. This effect is not linked to the alleviation of ER stress or the overexpression of proteins directly involved in calcium mineral accumulation, both of which are elevated in dystrophic muscles. \u0026nbsp;\u003c/p\u003e","manuscriptTitle":"Tauroursodeoxycholic acid (TUDCA)-induced increase in ectopic muscle mineralization occurs exclusively in dystrophic muscles and is independent of endoplasmic reticulum stress","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-05-08 05:39:27","doi":"10.21203/rs.3.rs-6401076/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-06-12T06:00:04+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-06-05T20:34:57+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-05-27T07:43:52+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-05-12T14:12:10+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"174371699725253717308687560646442073663","date":"2025-05-10T15:33:42+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"47512496238349592813421518743610552367","date":"2025-05-08T07:13:56+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"196309085026268927935193115113873403013","date":"2025-05-07T21:21:59+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"216661452206802928000794486004800600369","date":"2025-05-06T18:54:39+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-05-05T14:28:51+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-04-25T12:12:06+00:00","index":"","fulltext":""},{"type":"editorInvited","content":"","date":"2025-04-23T16:26:23+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-04-22T11:29:57+00:00","index":"","fulltext":""},{"type":"submitted","content":"Scientific Reports","date":"2025-04-08T08:25:01+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"ec2deec8-121b-4ed2-9ade-e4b260b6cff2","owner":[],"postedDate":"May 8th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[{"id":48223816,"name":"Biological sciences/Biochemistry"},{"id":48223817,"name":"Biological sciences/Cell biology"},{"id":48223818,"name":"Health sciences/Pathogenesis"}],"tags":[],"updatedAt":"2025-11-03T16:06:31+00:00","versionOfRecord":{"articleIdentity":"rs-6401076","link":"https://doi.org/10.1038/s41598-025-21534-0","journal":{"identity":"scientific-reports","isVorOnly":false,"title":"Scientific Reports"},"publishedOn":"2025-10-28 15:57:42","publishedOnDateReadable":"October 28th, 2025"},"versionCreatedAt":"2025-05-08 05:39:27","video":"","vorDoi":"10.1038/s41598-025-21534-0","vorDoiUrl":"https://doi.org/10.1038/s41598-025-21534-0","workflowStages":[]},"version":"v1","identity":"rs-6401076","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6401076","identity":"rs-6401076","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}