{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,21]],"date-time":"2026-07-21T04:16:55Z","timestamp":1784607415071,"version":"3.55.0"},"reference-count":38,"publisher":"MDPI AG","issue":"9","license":[{"start":{"date-parts":[[2021,9,16]],"date-time":"2021-09-16T00:00:00Z","timestamp":1631750400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Cells"],"abstract":"<jats:p>Ischemia\/reperfusion (I\/R) events are involved in the pathophysiology of numerous ocular diseases. The purpose of this study was to test the hypothesis that betulinic acid protects from I\/R injury in the mouse retina. Ocular ischemia was induced in mice by increasing intraocular pressure (IOP) to 110 mm Hg for 45 min, while the fellow eye served as a control. One group of mice received betulinic acid (50 mg\/kg\/day p.o. once daily) and the other group received the vehicle solution only. Eight days after the I\/R event, the animals were killed and the retinal wholemounts and optic nerve cross-sections were prepared and stained with cresyl blue or toluidine blue, respectively, to count cells in the ganglion cell layer (GCL) of the retina and axons in the optic nerve. Retinal arteriole responses were measured in isolated retinas by video microscopy. The levels of reactive oxygen species (ROS) were assessed in retinal cryosections and redox gene expression was determined in isolated retinas by quantitative PCR. I\/R markedly reduced cell number in the GCL and axon number in the optic nerve of the vehicle-treated mice. In contrast, only a negligible reduction in cell and axon number was observed following I\/R in the betulinic acid-treated mice. Endothelial function was markedly reduced and ROS levels were increased in retinal arterioles of vehicle-exposed eyes following I\/R, whereas betulinic acid partially prevented vascular endothelial dysfunction and ROS formation. Moreover, betulinic acid boosted mRNA expression for the antioxidant enzymes SOD3 and HO-1 following I\/R. Our data provide evidence that betulinic acid protects from I\/R injury in the mouse retina. Improvement of vascular endothelial function and the reduction in ROS levels appear to contribute to the neuroprotective effect.<\/jats:p>","DOI":"10.3390\/cells10092440","type":"journal-article","created":{"date-parts":[[2021,9,16]],"date-time":"2021-09-16T21:38:12Z","timestamp":1631828292000},"page":"2440","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":31,"title":["Betulinic Acid Protects from Ischemia-Reperfusion Injury in the Mouse Retina"],"prefix":"10.3390","volume":"10","author":[{"given":"Aytan","family":"Musayeva","sequence":"first","affiliation":[{"name":"Department of Ophthalmology, University Medical Center, Johannes Gutenberg University Mainz, Langenbeckstrasse 1, 55131 Mainz, Germany"},{"name":"Laboratory of Corneal Immunology, Transplantation and Regeneration, Schepens Eye Research Institute, Massachusetts Eye and Ear, Department of Ophthalmology, Harvard Medical School, Boston, MA 02114, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Johanna C.","family":"Unkrig","sequence":"additional","affiliation":[{"name":"Department of Ophthalmology, University Medical Center, Johannes Gutenberg University Mainz, Langenbeckstrasse 1, 55131 Mainz, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mayagozel B.","family":"Zhutdieva","sequence":"additional","affiliation":[{"name":"Department of Ophthalmology, University Medical Center, Johannes Gutenberg University Mainz, Langenbeckstrasse 1, 55131 Mainz, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5891-1155","authenticated-orcid":false,"given":"Caroline","family":"Manicam","sequence":"additional","affiliation":[{"name":"Department of Ophthalmology, University Medical Center, Johannes Gutenberg University Mainz, Langenbeckstrasse 1, 55131 Mainz, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yue","family":"Ruan","sequence":"additional","affiliation":[{"name":"Department of Ophthalmology, University Medical Center, Johannes Gutenberg University Mainz, Langenbeckstrasse 1, 55131 Mainz, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Panagiotis","family":"Laspas","sequence":"additional","affiliation":[{"name":"Department of Ophthalmology, University Medical Center, Johannes Gutenberg University Mainz, Langenbeckstrasse 1, 55131 Mainz, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7524-2117","authenticated-orcid":false,"given":"Panagiotis","family":"Chronopoulos","sequence":"additional","affiliation":[{"name":"Department of Ophthalmology, University Medical Center, Johannes Gutenberg University Mainz, Langenbeckstrasse 1, 55131 Mainz, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marie L.","family":"G\u00f6bel","sequence":"additional","affiliation":[{"name":"Department of Ophthalmology, University Medical Center, Johannes Gutenberg University Mainz, Langenbeckstrasse 1, 55131 Mainz, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Norbert","family":"Pfeiffer","sequence":"additional","affiliation":[{"name":"Department of Ophthalmology, University Medical Center, Johannes Gutenberg University Mainz, Langenbeckstrasse 1, 55131 Mainz, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6761-0247","authenticated-orcid":false,"given":"Christoph","family":"Brochhausen","sequence":"additional","affiliation":[{"name":"Institute of Pathology, University Medical Center, Johannes Gutenberg University Mainz, Langenbeckstrasse 1, 55131 Mainz, Germany"},{"name":"Institute of Pathology, University of Regensburg, Franz-Josef-Strau\u00df-Allee 11, 93053 Regensburg, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2769-0094","authenticated-orcid":false,"given":"Andreas","family":"Daiber","sequence":"additional","affiliation":[{"name":"Department of Cardiology 1, Laboratory of Molecular Cardiology, University Medical Center, Johannes Gutenberg University Mainz, Building 605, Langenbeckstrasse 1, 55131 Mainz, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Matthias","family":"Oelze","sequence":"additional","affiliation":[{"name":"Department of Cardiology 1, Laboratory of Molecular Cardiology, University Medical Center, Johannes Gutenberg University Mainz, Building 605, Langenbeckstrasse 1, 55131 Mainz, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3458-7391","authenticated-orcid":false,"given":"Huige","family":"Li","sequence":"additional","affiliation":[{"name":"Department of Pharmacology, University Medical 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S., Musayeva, A., and Gericke, A. (2020). Oxidative Stress and Vascular Dysfunction in the Retina: Therapeutic Strategies. Antioxidants, 9.","DOI":"10.3390\/antiox9080761"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"733","DOI":"10.1016\/S0002-9394(99)00359-1","article-title":"Aggressive systematic treatment for central retinal artery occlusion","volume":"128","author":"Rumelt","year":"1999","journal-title":"Am. J. Ophthalmol."},{"key":"ref_3","first-page":"RA138","article-title":"Ocular ischemic syndrome\u2014A systematic review","volume":"18","author":"Skonieczna","year":"2012","journal-title":"Med. Sci. Monit."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"950","DOI":"10.1007\/s00417-005-0140-2","article-title":"Multicenter study of the European Assessment Group for Lysis in the Eye (EAGLE) for the treatment of central retinal artery occlusion: Design issues and implications. EAGLE Study report no. 1: EAGLE Study report no. 1","volume":"244","author":"Feltgen","year":"2006","journal-title":"Graefes Arch. Clin. Exp. Ophthalmol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"1367","DOI":"10.1016\/j.ophtha.2010.03.061","article-title":"Central Retinal Artery Occlusion: Local Intra-arterial Fibrinolysis versus Conservative Treatment, a Multicenter Randomized Trial","volume":"117","author":"Schumacher","year":"2010","journal-title":"Ophthalmology"},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Zadeh, J.K., Garcia-Bardon, A., Hartmann, E.K., Pfeiffer, N., Omran, W., Ludwig, M., Patzak, A., Xia, N., Li, H., and Gericke, A. (2019). Short-Time Ocular Ischemia Induces Vascular Endothelial Dysfunction and Ganglion Cell Loss in the Pig Retina. Int. J. Mol. Sci., 20.","DOI":"10.3390\/ijms20194685"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"152","DOI":"10.1016\/j.exer.2019.04.021","article-title":"Responses of retinal arterioles and ciliary arteries in pigs with acute respiratory distress syndrome (ARDS)","volume":"184","author":"Zadeh","year":"2019","journal-title":"Exp. Eye Res."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"8123","DOI":"10.1167\/iovs.11-8318","article-title":"Neuroprotection from Retinal Ischemia\/Reperfusion Injury by NOX2 NADPH Oxidase Deletion","volume":"52","author":"Yokota","year":"2011","journal-title":"Investig. Opthalmol. Vis. Sci."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"20667","DOI":"10.1039\/C9NR07171G","article-title":"Tetrahedral framework nucleic acids prevent retina ischemia-reperfusion injury from oxidative stress via activating the Akt\/Nrf2 pathway","volume":"11","author":"Qin","year":"2019","journal-title":"Nanoscale"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"2016","DOI":"10.3390\/molecules14062016","article-title":"Pentacyclic Triterpene Distribution in Various Plants\u2014Rich Sources for a New Group of Multi-Potent Plant Extracts","volume":"14","author":"Trojan","year":"2009","journal-title":"Molecules"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"1407","DOI":"10.1002\/jps.2600650938","article-title":"Tumor Inhibitory Agents from Vauquelinia corymbosa (Rosaceae)","volume":"65","author":"Trumbull","year":"1976","journal-title":"J. Pharm. Sci."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"111990","DOI":"10.1016\/j.biopha.2021.111990","article-title":"Betulinic acid in the treatment of tumour diseases: Application and research progress","volume":"142","author":"Jiang","year":"2021","journal-title":"Biomed. Pharmacother."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"657","DOI":"10.2174\/0929867053202214","article-title":"Betulinic Acid and Its Derivatives: A Review on their Biological Properties","volume":"12","author":"Yogeeswari","year":"2005","journal-title":"Curr. Med. Chem."},{"key":"ref_14","first-page":"1","article-title":"Structure and Activity of Pentacyclic Triterpenes Codrugs. A Review","volume":"21","author":"Olender","year":"2021","journal-title":"Mini-Rev. Med. Chem."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"132","DOI":"10.1016\/j.niox.2011.01.007","article-title":"Betulinic acid protects against cerebral ischemia\u2013reperfusion injury in mice by reducing oxidative and nitrosative stress","volume":"24","author":"Lu","year":"2011","journal-title":"Nitric Oxide"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1155\/2014\/573745","article-title":"Cardioprotective Effect of Betulinic Acid on Myocardial Ischemia Reperfusion Injury in Rats","volume":"2014","author":"Xia","year":"2014","journal-title":"Evid.-Based Complement. Altern. Med."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"325","DOI":"10.1002\/ptr.2929","article-title":"Betulinic acid protects against ischemia\/reperfusion-induced renal damage and inhibits leukocyte apoptosis","volume":"24","author":"Bilgin","year":"2010","journal-title":"Phytother. Res."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"744","DOI":"10.1007\/s11596-017-1798-5","article-title":"Down-regulation of NOX4 by betulinic acid protects against cerebral ischemia-reperfusion in mice","volume":"37","author":"Lu","year":"2017","journal-title":"Curr. Med. Sci."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"2829","DOI":"10.1021\/acschemneuro.1c00198","article-title":"Betulinic Acid Ameliorates Cerebral Injury in Middle Cerebral Artery Occlusion Rats through Regulating Autophagy","volume":"12","author":"Zhao","year":"2021","journal-title":"ACS Chem. Neurosci."},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Hartsock, M.J., Cho, H., Wu, L., Chen, W.-J., Gong, J., and Duh, E.J. (2016). A Mouse Model of Retinal Ischemia-Reperfusion Injury Through Elevation of Intraocular Pressure. J. Vis. Exp., e54065.","DOI":"10.3791\/54065-v"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"30","DOI":"10.1167\/iovs.11-8753","article-title":"Acute Retinal Ischemia Inhibits Endothelium-Dependent Nitric Oxide\u2013Mediated Dilation of Retinal Arterioles via Enhanced Superoxide Production","volume":"53","author":"Hein","year":"2012","journal-title":"Investig. Opthalmol. Vis. Sci."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"5222","DOI":"10.1038\/s41598-019-41425-5","article-title":"The M1 muscarinic acetylcholine receptor subtype is important for retinal neuron survival in aging mice","volume":"9","author":"Laspas","year":"2019","journal-title":"Sci. Rep."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"7479","DOI":"10.1167\/iovs.11-7370","article-title":"Identification of the Muscarinic Acetylcholine Receptor Subtype Mediating Cholinergic Vasodilation in Murine Retinal Arterioles","volume":"52","author":"Gericke","year":"2011","journal-title":"Investig. Opthalmol. Vis. Sci."},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Gericke, A., Goloborodko, E., Pfeiffer, N., and Manicam, C. (2018). Preparation Steps for Measurement of Reactivity in Mouse Retinal Arterioles Ex Vivo. J. Vis. Exp., e56199.","DOI":"10.3791\/56199"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"9736047","DOI":"10.1155\/2019\/9736047","article-title":"Elevated Intraocular Pressure Causes Abnormal Reactivity of Mouse Retinal Arterioles","volume":"2019","author":"Gericke","year":"2019","journal-title":"Oxidative Med. Cell. Longev."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"5181429","DOI":"10.1155\/2019\/5181429","article-title":"Apolipoprotein E Deficiency Causes Endothelial Dysfunction in the Mouse Retina","volume":"2019","author":"Zadeh","year":"2019","journal-title":"Oxidative Med. Cell. Longev."},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Musayeva, A., Jiang, S., Ruan, Y., Zadeh, J., Chronopoulos, P., Pfeiffer, N., M\u00fcller, W., Ackermann, M., Xia, N., and Li, H. (2021). Aged Mice Devoid of the M3 Muscarinic Acetylcholine Receptor Develop Mild Dry Eye Disease. Int. J. Mol. Sci., 22.","DOI":"10.3390\/ijms22116133"},{"key":"ref_28","first-page":"285","article-title":"Ganglion cell distribution in the retina of the mouse","volume":"20","author":"Olsen","year":"1981","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_29","first-page":"1387","article-title":"Evaluation of the percentage of ganglion cells in the ganglion cell layer of the rodent retina","volume":"19","author":"Schlamp","year":"2013","journal-title":"Mol. Vis."},{"key":"ref_30","first-page":"1","article-title":"Systemic and Ocular Hemodynamic Risk Factors in Glaucoma","volume":"2015","author":"Choi","year":"2015","journal-title":"BioMed. Res. Int."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"S162","DOI":"10.1016\/j.survophthal.2007.08.012","article-title":"What is the present pathogenetic concept of glaucomatous optic neuropathy?","volume":"52","author":"Flammer","year":"2007","journal-title":"Surv. Ophthalmol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"4","DOI":"10.1111\/j.1755-3768.2007.01167.x","article-title":"Endothelial dysfunction in glaucoma","volume":"87","author":"Resch","year":"2009","journal-title":"Acta Ophthalmol."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"165","DOI":"10.1007\/s10787-019-00622-4","article-title":"Betulinic acid attenuates lipopolysaccharide-induced vascular hyporeactivity in the rat aorta by modulating Nrf2 antioxidative function","volume":"28","author":"Bai","year":"2020","journal-title":"Inflammopharmacology"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"588","DOI":"10.4103\/0253-7613.100382","article-title":"Betulinic acid inhibits superoxide anion-mediated impairment of endothelium-dependent relaxation in rat aortas","volume":"44","author":"Xia","year":"2012","journal-title":"Indian J. Pharmacol."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"385","DOI":"10.1016\/j.ejps.2011.08.025","article-title":"Betulinic acid ameliorates endothelium-dependent relaxation in l-NAME-induced hypertensive rats by reducing oxidative stress","volume":"44","author":"Fu","year":"2011","journal-title":"Eur. J. Pharm. Sci."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"71","DOI":"10.1016\/j.exer.2017.06.004","article-title":"Rapid repeatable in vivo detection of retinal reactive oxygen species","volume":"161","author":"Fan","year":"2017","journal-title":"Exp. Eye Res."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"408","DOI":"10.1016\/j.freeradbiomed.2018.06.024","article-title":"Extracellular superoxide dismutase (SOD3) regulates oxidative stress at the vitreoretinal interface","volume":"124","author":"Wert","year":"2018","journal-title":"Free. Radic. Biol. Med."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"3884","DOI":"10.2174\/1381612823666170413122439","article-title":"Therapeutic Potential of Heme Oxygenase-1\/carbon Monoxide System Against Ischemia-Reperfusion Injury","volume":"23","author":"Cheng","year":"2017","journal-title":"Curr. Pharm. Des."}],"container-title":["Cells"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4409\/10\/9\/2440\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T07:00:44Z","timestamp":1760166044000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4409\/10\/9\/2440"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,9,16]]},"references-count":38,"journal-issue":{"issue":"9","published-online":{"date-parts":[[2021,9]]}},"alternative-id":["cells10092440"],"URL":"https:\/\/doi.org\/10.3390\/cells10092440","relation":{},"ISSN":["2073-4409"],"issn-type":[{"value":"2073-4409","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,9,16]]}}}