{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,29]],"date-time":"2026-07-29T05:16:20Z","timestamp":1785302180003,"version":"3.55.0"},"reference-count":22,"publisher":"MDPI AG","issue":"7","license":[{"start":{"date-parts":[[2023,3,29]],"date-time":"2023-03-29T00:00:00Z","timestamp":1680048000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000053","name":"SPORT Program","doi-asserted-by":"publisher","award":["NIH RO1 EY 030443"],"award-info":[{"award-number":["NIH RO1 EY 030443"]}],"id":[{"id":"10.13039\/100000053","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000053","name":"SPORT Program","doi-asserted-by":"publisher","award":["NIA (030443-03S1)"],"award-info":[{"award-number":["NIA (030443-03S1)"]}],"id":[{"id":"10.13039\/100000053","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000053","name":"SPORT Program","doi-asserted-by":"publisher","award":["R56-AI172487"],"award-info":[{"award-number":["R56-AI172487"]}],"id":[{"id":"10.13039\/100000053","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Optos, PLC","award":["NIH RO1 EY 030443"],"award-info":[{"award-number":["NIH RO1 EY 030443"]}]},{"name":"Optos, PLC","award":["NIA (030443-03S1)"],"award-info":[{"award-number":["NIA (030443-03S1)"]}]},{"name":"Optos, PLC","award":["R56-AI172487"],"award-info":[{"award-number":["R56-AI172487"]}]},{"name":"NIAID","award":["NIH RO1 EY 030443"],"award-info":[{"award-number":["NIH RO1 EY 030443"]}]},{"name":"NIAID","award":["NIA (030443-03S1)"],"award-info":[{"award-number":["NIA (030443-03S1)"]}]},{"name":"NIAID","award":["R56-AI172487"],"award-info":[{"award-number":["R56-AI172487"]}]},{"name":"Belfast Association for the Blind: purchase of FLIM","award":["NIH RO1 EY 030443"],"award-info":[{"award-number":["NIH RO1 EY 030443"]}]},{"name":"Belfast Association for the Blind: purchase of FLIM","award":["NIA (030443-03S1)"],"award-info":[{"award-number":["NIA (030443-03S1)"]}]},{"name":"Belfast Association for the Blind: purchase of FLIM","award":["R56-AI172487"],"award-info":[{"award-number":["R56-AI172487"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>We have shown that all sub-retinal pigment epithelial (sub-RPE) deposits examined contain calcium phosphate minerals: hydroxyapatite (HAP), whitlockite (Wht), or both. These typically take the form of ca. 1 \u03bcm diameter spherules or &gt;10 \u03bcm nodules and appear to be involved in the development and progression of age-related macular degeneration (AMD). Thus, these minerals may serve as useful biomarkers the for early detection and monitoring of sub-RPE changes in AMD. We demonstrated that HAP deposits could be imaged in vitro by fluorescence lifetime imaging microscopy (FLIM) in flat-mounted retinas using legacy tetracycline antibiotics as selective sensors for HAP. As the contrast on a FLIM image is based on the difference in fluorescence lifetime and not intensity of the tetracycline-stained HAP, distinguishing tissue autofluorescence from the background is significantly improved. The focus of the present pilot study was to assess whether vascular perfusion of the well tolerated and characterized chlortetracycline (widely used as an orally bioavailable antibiotic) can fluorescently label retinal HAP using human cadavers. We found that the tetracycline delivered through the peripheral circulation can indeed selectively label sub-RPE deposits opening the possibility for its use for ophthalmic monitoring of a range of diseases in which deposit formation is reported, such as AMD and Alzheimer disease (AD).<\/jats:p>","DOI":"10.3390\/ijms24076421","type":"journal-article","created":{"date-parts":[[2023,3,30]],"date-time":"2023-03-30T02:23:46Z","timestamp":1680143026000},"page":"6421","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":6,"title":["Fluorescence Lifetime Imaging of Human Sub-RPE Calcification In Vitro Following Chlortetracycline Infusion"],"prefix":"10.3390","volume":"24","author":[{"given":"Kavita R.","family":"Hegde","sequence":"first","affiliation":[{"name":"Department of Natural Sciences, Coppin State University, Baltimore, MD 21216, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Adam C.","family":"Puche","sequence":"additional","affiliation":[{"name":"Department of Anatomy and Neurobiology, School of Medicine, University of Maryland, Baltimore, MD 21201, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Henryk","family":"Szmacinski","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Molecular Biology, School of Medicine, University of Maryland, Baltimore, MD 21201, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kristina","family":"Fuller","sequence":"additional","affiliation":[{"name":"Department of Anatomy and Neurobiology, School of Medicine, University of Maryland, Baltimore, MD 21201, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Krishanu","family":"Ray","sequence":"additional","affiliation":[{"name":"Institute for Human Virology, School of Medicine, University of Maryland, Baltimore, MD 21201, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nikita","family":"Patel","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Molecular Biology, School of Medicine, University of Maryland, Baltimore, MD 21201, USA"},{"name":"Department of Medicine, Renaissance School of Medicine at Stony Brook University, Stony Brook, NY 11794, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7467-2174","authenticated-orcid":false,"given":"Imre","family":"Lengyel","sequence":"additional","affiliation":[{"name":"The Wellcome-Wolfson Institute for Experimental Medicine, School of Medicine, Dentistry and Biomedical Science, Queen\u2019s University Belfast, Belfast BT9 7BL, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Richard B.","family":"Thompson","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Molecular Biology, School of Medicine, University of Maryland, Baltimore, MD 21201, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2023,3,29]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1474","DOI":"10.1056\/NEJMra062326","article-title":"Age-related macular degeneration","volume":"355","year":"2006","journal-title":"N. Engl. J. Med."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"1565","DOI":"10.1073\/pnas.1413347112","article-title":"Identification of hydroxyapatite spherules provides new insight into subretinal pigment epithelial deposit formation in the aging eye","volume":"112","author":"Thompson","year":"2015","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"100053","DOI":"10.1016\/j.xops.2021.100053","article-title":"Characterization of Calcium Phosphate Spherical Particles in the Sub\u2013Retinal Pigment Epithelium\u2013Basal Lamina Space in Aged Human Eyes","volume":"1","author":"Pilgrim","year":"2021","journal-title":"Ophthalmol. Sci."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"705","DOI":"10.1016\/S1350-9462(01)00010-6","article-title":"An integrated hypothesis that considers drusen as biomarkers of immune-mediated processes at the RPE-Bruch\u2019s membrane interface in aging and age-related macular degeneration","volume":"20","author":"Hageman","year":"2001","journal-title":"Prog. Retin. Eye Res."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"443","DOI":"10.1586\/17469899.2.3.443","article-title":"Genetic markers and biomarkers for age-related macular degeneration","volume":"2","author":"Ross","year":"2007","journal-title":"Expert Rev. Ophthalmol."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"eaat4544","DOI":"10.1126\/scitranslmed.aat4544","article-title":"Calcified nodules in retinal drusen are associated with disease progression in age-related macular degeneration","volume":"10","author":"Tan","year":"2018","journal-title":"Sci. Transl. Med."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"182","DOI":"10.1159\/000487053","article-title":"Peripheral Retinal Imaging Biomarkers for Alzheimer\u2019s Disease: A Pilot Study","volume":"59","author":"Csincsik","year":"2018","journal-title":"Ophthalm. Res."},{"key":"ref_8","first-page":"e12232","article-title":"Retinal phenotyping of variants of Alzheimer\u2019s disease using ultra-widefield retinal images","volume":"13","author":"Csincsik","year":"2021","journal-title":"Alzheimer\u2019s Dement. Diagn. Assess. Dis. Monit."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"83","DOI":"10.1016\/j.preteyeres.2018.10.001","article-title":"The clinical relevance of visualising the peripheral retina","volume":"68","author":"Quinn","year":"2019","journal-title":"Prog. Retin. Eye Res."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"138","DOI":"10.1016\/j.actbio.2022.03.003","article-title":"Nuclear and cellular, micro and nano calcification in Alzheimer\u2019s disease patients and correlation to phosphorylated Tau","volume":"143","author":"Tsolaki","year":"2022","journal-title":"Acta Biomater."},{"key":"ref_11","first-page":"377","article-title":"Tetracyclines and mineralized tissues: Review and perspectives","volume":"48","author":"Skinner","year":"1975","journal-title":"Yale J. Biol. Med."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"167","DOI":"10.1016\/j.ab.2011.05.011","article-title":"Near-infrared-labeled tetracycline derivative is an effective marker of bone deposition in mice","volume":"416","author":"Kovar","year":"2011","journal-title":"Anal. Biochem."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1148","DOI":"10.1038\/nbt1201-1148","article-title":"In vivo near-infrared fluorescence imaging of osteoblastic activity","volume":"19","author":"Zaheer","year":"2001","journal-title":"Nat. Biotech."},{"key":"ref_14","first-page":"151","article-title":"Red and near-infrared fluorometry","volume":"Volume 4","author":"Lakowicz","year":"1994","journal-title":"Topics in Fluorescence Spectroscopy Volume 4: Probe Design and Chemical Sensing"},{"key":"ref_15","first-page":"2435","article-title":"Spectral profiling of autofluorescence associated with lipofuscin, Bruch\u2019s membrane, and sub-RPE deposits in normal and AMD eyes","volume":"43","author":"Marmorstein","year":"2002","journal-title":"Investig. Ophthalmol. Vis. Sci."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"256","DOI":"10.1093\/jac\/dkl224","article-title":"Pharmacokinetics and pharmacodynamics of the tetracyclines including glycylcyclines","volume":"58","author":"Agwuh","year":"2006","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"76","DOI":"10.1159\/000458761","article-title":"Phototoxicity of Doxycycline: A Systematic Review on Clinical Manifestations, Frequency, Cofactors, and Prevention","volume":"30","author":"Goetze","year":"2017","journal-title":"Ski. Pharmacol. Physiol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"316","DOI":"10.1016\/0003-2697(92)90112-K","article-title":"Fluorescence lifetime imaging","volume":"202","author":"Lakowicz","year":"1992","journal-title":"Anal. Biochem."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1117\/1.JBO.25.4.047001","article-title":"Imaging hydroxyapatite in sub-retinal pigment epithelial deposits by fluorescence lifetime imaging microscopy with tetracycline staining","volume":"25","author":"Szmacinski","year":"2020","journal-title":"J. Biomed. Opt."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"714","DOI":"10.1007\/s00347-009-1975-4","article-title":"Comparison of parameters of time-resolved autofluorescence between healthy subjects and patients suffering from early AMD","volume":"106","author":"Schweitzer","year":"2009","journal-title":"Der Ophthalmol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"120","DOI":"10.1016\/j.preteyeres.2017.06.005","article-title":"Fluorescence lifetime imaging ophthalmoscopy","volume":"60","author":"Dysli","year":"2017","journal-title":"Prog. Retin. Eye Res."},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Zinkernagel, M., and Dysli, C. (2019). Fluorescence Lifetime Imaging Ophthalmoscopy, Springer.","DOI":"10.1007\/978-3-030-22878-1"}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/24\/7\/6421\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T19:05:58Z","timestamp":1760123158000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/24\/7\/6421"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,3,29]]},"references-count":22,"journal-issue":{"issue":"7","published-online":{"date-parts":[[2023,4]]}},"alternative-id":["ijms24076421"],"URL":"https:\/\/doi.org\/10.3390\/ijms24076421","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2023,3,29]]}}}