{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,15]],"date-time":"2026-07-15T22:27:47Z","timestamp":1784154467567,"version":"3.55.0"},"reference-count":50,"publisher":"Public Library of Science (PLoS)","issue":"1","license":[{"start":{"date-parts":[[2021,1,12]],"date-time":"2021-01-12T00:00:00Z","timestamp":1610409600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"PRIN-MIUR","award":["2015MJBEM2_005"],"award-info":[{"award-number":["2015MJBEM2_005"]}]},{"DOI":"10.13039\/501100007423","name":"Duchenne Parent Project","doi-asserted-by":"publisher","award":["DPP-NL 2015"],"award-info":[{"award-number":["DPP-NL 2015"]}],"id":[{"id":"10.13039\/501100007423","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS ONE"],"abstract":"<jats:p>The <jats:italic>mdx<\/jats:italic> mouse model of Duchenne muscular dystrophy is characterized by functional and structural alterations of the diaphragm since early stages of pathology, closely resembling patients\u2019 condition. In recent years, ultrasonography has been proposed as a useful longitudinal non-invasive technique to assess <jats:italic>mdx<\/jats:italic> diaphragm dysfunction and evaluate drug efficacy over time. To date, only a few preclinical studies have been conducted. Therefore, an independent validation of this method by different laboratories is needed to increase results reliability and reduce biases. Here, we performed diaphragm ultrasonography in 3- and 6-month-old <jats:italic>mdx<\/jats:italic> mice, the preferred age-window for pharmacology studies. The alteration of diaphragm function over time was measured as diaphragm ultrasound movement amplitude. At the same time points, a first-time assessment of diaphragm echodensity was performed, as an experimental index of progressive loss of contractile tissue. A parallel evaluation of other <jats:italic>in vivo<\/jats:italic> and <jats:italic>ex vivo<\/jats:italic> dystrophy-relevant readouts was carried out. Both 3- and 6-month-old <jats:italic>mdx<\/jats:italic> mice showed a significant decrease in diaphragm amplitude compared to wild type (wt) mice. This index was well-correlated either with <jats:italic>in vivo<\/jats:italic> running performance or <jats:italic>ex vivo<\/jats:italic> isometric tetanic force of isolated diaphragm. In addition, diaphragms from 6-month-old dystrophic mice were also highly susceptible to eccentric contraction <jats:italic>ex vivo<\/jats:italic>. Importantly, we disclosed an age-dependent increase in echodensity in <jats:italic>mdx<\/jats:italic> mice not observed in wt animals, which was independent from abdominal wall thickness. This was accompanied by a notable increase of pro-fibrotic TGF-\u03b21 levels in the <jats:italic>mdx<\/jats:italic> diaphragm and of non-muscle tissue amount in diaphragm sections stained by hematoxylin &amp; eosin. Our findings corroborate the usefulness of diaphragm ultrasonography in preclinical drug studies as a powerful tool to monitor <jats:italic>mdx<\/jats:italic> pathology progression since early stages.<\/jats:p>","DOI":"10.1371\/journal.pone.0245397","type":"journal-article","created":{"date-parts":[[2021,1,12]],"date-time":"2021-01-12T23:18:22Z","timestamp":1610493502000},"page":"e0245397","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":13,"title":["Ultrasonography validation for early alteration of diaphragm echodensity and function in the mdx mouse model of Duchenne muscular dystrophy"],"prefix":"10.1371","volume":"16","author":[{"given":"Antonietta","family":"Mele","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Paola","family":"Mantuano","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Adriano","family":"Fonzino","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Francesco","family":"Rana","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Roberta Francesca","family":"Capogrosso","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Francesca","family":"Sanarica","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jean-Francois","family":"Rolland","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ornella","family":"Cappellari","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5652-7341","authenticated-orcid":true,"given":"Annamaria","family":"De Luca","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"340","published-online":{"date-parts":[[2021,1,12]]},"reference":[{"issue":"1","key":"pone.0245397.ref001","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1016\/j.nbd.2008.03.008","article-title":"Towards developing standard operating procedures for pre-clinical testing in the mdx mouse model of Duchenne muscular dystrophy","volume":"31","author":"MD Grounds","year":"2008","journal-title":"Neurobiol Dis"},{"issue":"1","key":"pone.0245397.ref002","doi-asserted-by":"crossref","first-page":"43","DOI":"10.1016\/j.nmd.2011.04.012","article-title":"TREAT-NMD Neuromuscular Network. 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