{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,11]],"date-time":"2026-09-11T12:29:12Z","timestamp":1789129752817,"version":"build-2803163510"},"reference-count":27,"publisher":"MDPI AG","issue":"4","license":[{"start":{"date-parts":[[2023,2,9]],"date-time":"2023-02-09T00:00:00Z","timestamp":1675900800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000090","name":"Congressionally Directed Medical Research Programs","doi-asserted-by":"publisher","award":["DM160252"],"award-info":[{"award-number":["DM160252"]}],"id":[{"id":"10.13039\/100000090","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Cells"],"abstract":"<jats:p>Ischemia-reperfusion (I\/R) injury is a complication impacting multiple organs and tissues in clinical conditions ranging from peripheral arterial disease to musculoskeletal trauma and myocardial infarction. Stem cell-derived extracellular vesicles (EVs) may represent one therapeutic resource for preventing the tissue damage associated with I\/R injury. Here we tested the hypothesis that lyophilized extracellular vesicles derived from adipose stem cells could serve as an \u201coff-the-shelf\u201d treatment modality for I\/R injury in a mouse hindlimb ischemia model. Ischemia was induced for 90 min using a rubber band tourniquet and extracellular vesicles (0, 50, or 100 \u00b5g) administered via tail vein injection immediately prior to reperfusion. Perfusion was measured prior to, during, and after ischemia using laser Doppler imaging. Serum and tissue were collected 24 h after reperfusion. Mass spectrometry (MS)-based proteomics was used to characterize the EV cargo and proteins from the ischemic and non-ischemic hindlimb. Inflammatory cytokines were measured in muscle and serum using a multiplex array. Results indicate that EVs significantly increase reperfusion and significantly increase expression of the anti-inflammatory factor annexin a1 in skeletal muscle; however, the increased reperfusion was also associated with a marked decrease in muscle structural proteins such as dystrophin, plectin, and obscurin. Circulating inflammatory cytokines TNF-alpha and IL-6 were increased with EV treatment, and serum TNF-alpha showed a significant, positive correlation with reperfusion level. These findings suggest that, while EVs may enhance reperfusion, the increased reperfusion can negatively impact muscle tissue and possibly remote organs. Alternative approaches, such as targeting mitochondrial permeability, may be more effective at mitigating I\/R injury.<\/jats:p>","DOI":"10.3390\/cells12040557","type":"journal-article","created":{"date-parts":[[2023,2,9]],"date-time":"2023-02-09T03:21:55Z","timestamp":1675912915000},"page":"557","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":15,"title":["Lyophilized Extracellular Vesicles from Adipose-Derived Stem Cells Increase Muscle Reperfusion but Degrade Muscle Structural Proteins in A Mouse Model of Hindlimb Ischemia-Reperfusion Injury"],"prefix":"10.3390","volume":"12","author":[{"given":"Bharati","family":"Mendhe","sequence":"first","affiliation":[{"name":"Medical College of Georgia, Augusta University, Augusta, GA 30912, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mohammad B.","family":"Khan","sequence":"additional","affiliation":[{"name":"Medical College of Georgia, Augusta University, Augusta, GA 30912, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Damon","family":"Dunwody","sequence":"additional","affiliation":[{"name":"Medical College of Georgia, Augusta University, Augusta, GA 30912, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Khairat Bahgat Youssef","family":"El Baradie","sequence":"additional","affiliation":[{"name":"Medical College of Georgia, Augusta University, Augusta, GA 30912, USA"},{"name":"Faculty of Science, Tanta University, Tanta 31111, Egypt"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kathryn","family":"Smith","sequence":"additional","affiliation":[{"name":"Medical College of Georgia, Augusta University, Augusta, GA 30912, USA"},{"name":"College of Medicine, University of Arkansas for Medical Sciences, Little Rock, AR 72205, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wenbo","family":"Zhi","sequence":"additional","affiliation":[{"name":"Medical College of Georgia, Augusta University, Augusta, GA 30912, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9597-4374","authenticated-orcid":false,"given":"Ashok","family":"Sharma","sequence":"additional","affiliation":[{"name":"Medical College of Georgia, Augusta University, Augusta, GA 30912, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tae Jin","family":"Lee","sequence":"additional","affiliation":[{"name":"Medical College of Georgia, Augusta University, Augusta, GA 30912, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mark W.","family":"Hamrick","sequence":"additional","affiliation":[{"name":"Medical College of Georgia, Augusta University, Augusta, GA 30912, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2023,2,9]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"101","DOI":"10.1016\/j.biocel.2014.02.013","article-title":"Mitochondria: Mitochondrial participation in ischemia-reperfusion injury in skeletal muscle","volume":"50","author":"Lejay","year":"2014","journal-title":"Int. J. Biochem. Cell Biol."},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Pizzimenti, M., Riou, M., Charles, A.-L., Talha, S., Meyer, A., Andres, E., Chakf\u00e9, N., Lejay, A., and Geny, B. (2019). The Rise of Mitochondria in Peripheral Arterial Disease Physiopathology: Experimental and Clinical Data. J. Clin. Med., 8.","DOI":"10.3390\/jcm8122125"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1126","DOI":"10.1089\/ars.2012.5149","article-title":"Reactive Oxygen Species in Inflammation and Tissue Injury","volume":"20","author":"Mittal","year":"2014","journal-title":"Antioxid. Redox Signal."},{"key":"ref_4","doi-asserted-by":"crossref","unstructured":"Garbaisz, D., Turoczi, Z., Aranyi, P., Fulop, A., Rosero, O., Hermesz, E., Ferencz, A., Lotz, G., Harsanyi, L., and Szijarto, A. (2014). Attenuation of Skeletal Muscle and Renal Injury to the Lower Limb following Ischemia-Reperfusion Using mPTP Inhibitor NIM-811. PLoS ONE, 9.","DOI":"10.1371\/journal.pone.0101067"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"6152","DOI":"10.1038\/s41598-021-85753-x","article-title":"The cyclophilin inhibitor NIM-811 increases muscle cell survival with hypoxia in vitro and improves gait performance following ischemia-reperfusion in vivo","volume":"11","author":"Khan","year":"2021","journal-title":"Sci Rep."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"301","DOI":"10.1016\/j.scr.2013.01.002","article-title":"Mesenchymal stem cell-derived exosomes increase ATP levels, decrease oxidative stress and activate Pi3K\/Akt pathway to enhance myocardial viability and prevent adverse remodeling after myocardial ischemia\/reperfusion injury","volume":"10","author":"Arslan","year":"2013","journal-title":"Stem Cell Res."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"e008344","DOI":"10.1161\/JAHA.117.008344","article-title":"Extracellular Vesicle Injection Improves Myocardial Function and Increases Angiogenesis in a Swine Model of Chronic Ischemia","volume":"7","author":"Potz","year":"2018","journal-title":"J. Am. Hear. Assoc."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"H765","DOI":"10.1152\/ajpheart.00247.2019","article-title":"Exosomes derived from induced vascular progenitor cells promote angiogenesis in vitro and in an in vivo rat hindlimb ischemia model","volume":"317","author":"Johnson","year":"2019","journal-title":"Am. J. Physiol. Circ. Physiol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"3289","DOI":"10.1089\/scd.2012.0095","article-title":"Microvesicles Derived from Human Umbilical Cord Mesenchymal Stem Cells Stimulated by Hypoxia Promote Angiogenesis Both In Vitro and In Vivo","volume":"21","author":"Zhang","year":"2012","journal-title":"Stem Cells Dev."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"181","DOI":"10.3389\/fcell.2020.00181","article-title":"Freeze-Dried Extracellular Vesicles from Adipose-Derived Stem Cells Prevent Hypoxia-Induced Muscle Cell Injury","volume":"8","author":"Nouh","year":"2020","journal-title":"Front. Cell Dev. Biol."},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Bari, E., Perteghella, S., Di Silvestre, D., Sorlini, M., Catenacci, L., Sorrenti, M., Marrubini, G., Rossi, R., Tripodo, G., and Mauri, P. (2018). Pilot Production of Mesenchymal Stem\/Stromal Freeze-Dried Secretome for Cell-Free Regenerative Nanomedicine: A Validated GMP-Compliant Process. Cells, 7.","DOI":"10.3390\/cells7110190"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1568","DOI":"10.3390\/molecules19021568","article-title":"miRNA in Plasma Exosome is Stable under Different Storage Conditions","volume":"19","author":"Ge","year":"2014","journal-title":"Molecules"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"H830","DOI":"10.1152\/ajpheart.00581.2006","article-title":"A novel model of acute murine hindlimb ischemia","volume":"292","author":"Crawford","year":"2007","journal-title":"Am. J. Physiol. Circ. Physiol."},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Drysch, M., Wallner, C., Schmidt, S.V., Reinkemeier, F., Wagner, J.M., Lehnhardt, M., and Behr, B. (2019). An optimized low-pressure tourniquet murine hind limb ischemia reperfusion model: Inducing acute ischemia reperfusion injury in C57BL\/6 wild type mice. PLoS ONE, 14.","DOI":"10.1371\/journal.pone.0210961"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"1907","DOI":"10.1016\/j.jprot.2010.06.006","article-title":"Exosomes: Extracellular organelles important in intercellular communication","volume":"73","author":"Mathivanan","year":"2010","journal-title":"J. Proteom."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"796","DOI":"10.1016\/j.jprot.2011.02.024","article-title":"Protein profile of exosomes from trabecular meshwork cells","volume":"74","author":"Stamer","year":"2011","journal-title":"J. Proteom."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"6094562","DOI":"10.1155\/2020\/6094562","article-title":"Proteomic Analysis of Exosomes from Adipose-Derived Mesenchymal Stem Cells: A Novel Therapeutic Strategy for Tissue Injury","volume":"2020","author":"Xing","year":"2020","journal-title":"BioMed Res. Int."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1435","DOI":"10.1161\/01.ATV.0000134300.87476.d1","article-title":"Heath shock protein 90 transfection reduces ischemia-reperfusion-induced myocardial dysfunction via reciprocal endothelial NO synthase serine 1177 phosphorylation and threonine 495 dephosphorylation","volume":"24","author":"Kupatt","year":"2004","journal-title":"Arterioscler. Thromb. Vasc. Biol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"C1281","DOI":"10.1152\/ajpcell.00550.2007","article-title":"Activation of Hsp90-eNOS and increased NO generation attenuate respiration of hypoxia-treated endothelial cells","volume":"295","author":"Presley","year":"2008","journal-title":"Am. J. Physiol. Cell Physiol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"3389","DOI":"10.3389\/fimmu.2019.00938","article-title":"Annexin-A1: Therapeutic potential in microvascular disease","volume":"10","author":"Purvis","year":"2019","journal-title":"Front. Immunol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"1165","DOI":"10.1006\/jmcc.2001.1380","article-title":"Ischemic loss of sarcolemmal dystrophin and spectrin: Correlation with myocardial injury","volume":"33","author":"Armstrong","year":"2001","journal-title":"J. Mol. Cell Cardiol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"723","DOI":"10.1016\/j.yjmcc.2005.02.019","article-title":"Ischemia depletes dystrophin and inhibits protein synthesis in the canine heart: Mechanisms of myocardial ischemic injury","volume":"38","author":"Cai","year":"2005","journal-title":"J. Mol. Cell. Cardiol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"631","DOI":"10.1002\/cphy.c170023","article-title":"Thick Filament Protein Network, Functions, and Disease Association","volume":"8","author":"Wang","year":"2018","journal-title":"Compr. Physiol."},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Zrelski, M.M., Kustermann, M., and Winter, L. (2021). Muscle-Related Plectinopathies. Cells, 10.","DOI":"10.3390\/cells10092480"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"C709","DOI":"10.1152\/ajpcell.00485.2019","article-title":"Synemin-related skeletal and cardiac myopathies: An overview of pathogenic variants","volume":"318","author":"Paulin","year":"2020","journal-title":"Am. J. Physiol. Physiol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"e501","DOI":"10.1002\/ams2.501","article-title":"Therapeutic strategies for ischemia reperfusion injury in emergency medicine","volume":"7","author":"Naito","year":"2020","journal-title":"Acute Med. Surg."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"504","DOI":"10.1177\/10742484211046674","article-title":"Controlling Reperfusion Injury with Controlled Reperfusion: Historical Perspectives and New Paradigms","volume":"26","author":"Fischesser","year":"2021","journal-title":"J. Cardiovasc. Pharmacol. Ther."}],"container-title":["Cells"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4409\/12\/4\/557\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T18:28:13Z","timestamp":1760120893000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4409\/12\/4\/557"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,2,9]]},"references-count":27,"journal-issue":{"issue":"4","published-online":{"date-parts":[[2023,2]]}},"alternative-id":["cells12040557"],"URL":"https:\/\/doi.org\/10.3390\/cells12040557","relation":{},"ISSN":["2073-4409"],"issn-type":[{"value":"2073-4409","type":"electronic"}],"subject":[],"published":{"date-parts":[[2023,2,9]]}}}