{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,11,6]],"date-time":"2026-11-06T21:30:20Z","timestamp":1794000620351,"version":"4.8.0"},"reference-count":43,"publisher":"MDPI AG","issue":"1","license":[{"start":{"date-parts":[[2023,12,19]],"date-time":"2023-12-19T00:00:00Z","timestamp":1702944000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100000925","name":"National Health and Medical Research Council Australia","doi-asserted-by":"publisher","award":["GNT2018194"],"award-info":[{"award-number":["GNT2018194"]}],"id":[{"id":"10.13039\/501100000925","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000925","name":"National Health and Medical Research Council Australia","doi-asserted-by":"publisher","award":["GNT2005791"],"award-info":[{"award-number":["GNT2005791"]}],"id":[{"id":"10.13039\/501100000925","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000925","name":"National Health and Medical Research Council Australia","doi-asserted-by":"publisher","award":["H21\/174585"],"award-info":[{"award-number":["H21\/174585"]}],"id":[{"id":"10.13039\/501100000925","id-type":"DOI","asserted-by":"publisher"}]},{"name":"NSW Office of Health and Medical Research","award":["GNT2018194"],"award-info":[{"award-number":["GNT2018194"]}]},{"name":"NSW Office of Health and Medical Research","award":["GNT2005791"],"award-info":[{"award-number":["GNT2005791"]}]},{"name":"NSW Office of Health and Medical Research","award":["H21\/174585"],"award-info":[{"award-number":["H21\/174585"]}]},{"name":"Therapeutic Innovation Australia","award":["GNT2018194"],"award-info":[{"award-number":["GNT2018194"]}]},{"name":"Therapeutic Innovation Australia","award":["GNT2005791"],"award-info":[{"award-number":["GNT2005791"]}]},{"name":"Therapeutic Innovation Australia","award":["H21\/174585"],"award-info":[{"award-number":["H21\/174585"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Improved human-relevant preclinical models of coronary artery disease (CAD) are needed to improve translational research and drug discovery. Mitochondrial dysfunction and associated oxidative stress contribute to endothelial dysfunction and are a significant factor in the development and progression of CAD. Endothelial colony-forming cells (ECFCs) can be derived from peripheral blood mononuclear cells (PBMCs) and offer a unique potentially personalised means for investigating new potential therapies targeting important components of vascular function. We describe the application of the high-throughput and confocal Opera Phenix\u00ae High-Content Screening System to examine mitochondrial superoxide (mROS) levels, mitochondrial membrane potential, and mitochondrial area in both established cell lines and patient-derived ECFCs simultaneously. Unlike traditional plate readers, the Opera Phenix\u00ae is an imaging system that integrates automated confocal microscopy, precise fluorescent detection, and multi-parameter algorithms to visualize and precisely quantify targeted biological processes at a cellular level. In this study, we measured mROS production in human umbilical vein endothelial cells (HUVECs) and patient-derived ECFCs using the mROS production probe, MitoSOXTM Red. HUVECs exposed to oxidized low-density lipoprotein (oxLDL) increased mROS levels by 47.7% (p &lt; 0.0001). A pooled group of patient-derived ECFCs from participants with CAD (n = 14) exhibited 30.9% higher mROS levels compared to patients with no CAD when stimulated with oxLDL (n = 14; p &lt; 0.05). When tested against a small group of candidate compounds, this signal was attenuated by PKT-100 (36.22% reduction, p = 0.03), a novel P2X7 receptor antagonist. This suggests the P2X7 receptor as a valid target against excess mROS levels. As such, these findings highlight the potential of the MitoSOX-Opera Phenix technique to be used for drug discovery efforts in CAD.<\/jats:p>","DOI":"10.3390\/ijms25010022","type":"journal-article","created":{"date-parts":[[2023,12,19]],"date-time":"2023-12-19T06:03:11Z","timestamp":1702965791000},"page":"22","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":8,"title":["High-Throughput Measure of Mitochondrial Superoxide Levels as a Marker of Coronary Artery Disease to Accelerate Drug Translation in Patient-Derived Endothelial Cells Using Opera Phenix\u00ae Technology"],"prefix":"10.3390","volume":"25","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-2232-1759","authenticated-orcid":false,"given":"Weiqian E.","family":"Lee","sequence":"first","affiliation":[{"name":"Kolling Institute, University of Sydney, Sydney, NSW 2006, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marie","family":"Besnier","sequence":"additional","affiliation":[{"name":"Kolling Institute, University of Sydney, Sydney, NSW 2006, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Elijah","family":"Genetzakis","sequence":"additional","affiliation":[{"name":"Kolling Institute, University of Sydney, Sydney, NSW 2006, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Owen","family":"Tang","sequence":"additional","affiliation":[{"name":"Kolling Institute, University of Sydney, Sydney, NSW 2006, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9923-2722","authenticated-orcid":false,"given":"Katharine A.","family":"Kott","sequence":"additional","affiliation":[{"name":"Kolling Institute, University of Sydney, Sydney, NSW 2006, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Stephen T.","family":"Vernon","sequence":"additional","affiliation":[{"name":"Kolling Institute, University of Sydney, Sydney, NSW 2006, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1154-3011","authenticated-orcid":false,"given":"Michael P.","family":"Gray","sequence":"additional","affiliation":[{"name":"Kolling Institute, University of Sydney, Sydney, NSW 2006, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2712-157X","authenticated-orcid":false,"given":"Stuart M.","family":"Grieve","sequence":"additional","affiliation":[{"name":"Kolling Institute, University of Sydney, Sydney, NSW 2006, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6655-0529","authenticated-orcid":false,"given":"Michael","family":"Kassiou","sequence":"additional","affiliation":[{"name":"Kolling Institute, University of Sydney, Sydney, NSW 2006, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Gemma A.","family":"Figtree","sequence":"additional","affiliation":[{"name":"Kolling Institute, University of Sydney, Sydney, NSW 2006, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2023,12,19]]},"reference":[{"key":"ref_1","unstructured":"(2023, April 24). 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