Characterization and Activity of Endothelin Converting Enzyme-1 in Human Non-Pigmented Ciliary Epithelial Cells

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dc.contributor.advisorThomas Yorio
dc.contributor.committeeMemberS. Dan Dimitrijevich
dc.contributor.committeeMemberVictoria J. Rudick
dc.creatorFinkley, Alvin
dc.date.accessioned2019-08-22T20:52:48Z
dc.date.available2019-08-22T20:52:48Z
dc.date.issued1999-01-01
dc.date.submitted2014-01-14T06:24:04-08:00
dc.description.abstractFinkley, Alvin, Characterization and Activity of Endothelin Converting Enzyme-1 in Human Non-Pigmented Ciliary Epithelial Cells. Master of Science (Biomedical Sciences). Endothelins (ETs) are potent vasoactive peptides, that are present in many ocular tissues including the ciliary epithelium where active ET-1 is produced from the precursor Big ET-1 by a membrane-bound metalloprotease, endothelin-converting enzyme (ECE). Although the role of ocular ET’s are uncertain, ETs have been shown to lower the intraocular pressure. In the current study, ET-1 and Big-ET-1 were detected in SV-40 transformed human ciliary epithelial (HNPE) cells by immunofluorescence suggesting the presence of ECE activity. The presence of ECE was confirmed by Western blotting using polyclonal antibodies against ECE-1 which detected a 124 KDa protein in the membrane fraction and not in the cytosol. Further characterization of the enzymatic activity of ECE (conversion of Big ET-1 to ET-1) was performed using a novel assay involving 121I-Big ET-1 (substrate; 2fmloe) and polyclonal antibodies specific for Big ET-1. Mean ECE-1 activity (expressed as the ratio of 121^1-ET-1 produced to the total 125^I-Big ET-1 incubated X 100) was measured and corresponded to: 26% (0.5 3±0.02 fmole, 1 hr), 63% (1.26±0.07 fmole, 3hr) and 66% (1.33±0.11 fmole, 24 hr) compared to blank controls at 13% (0.25±0.03 fmole). Thiorphan (2mM), an inhibitor of ECE, abolished ECE-1 activity. These results suggest that ECE-1 is localized in HNPE cells and is essential for the production of ET-1. The physiological importance of the proteolytic processing by ECE-1 in ocular tissue may reflect on how ET regulates intraocular pressure. Key Words: endothelin converting enzyme-1; endothelin-1; Big endothelin-1; ciliary epithelium; aqueous humor dynamics; intraocular pressure, Western blotting, ECE-1
dc.format.mimetypeapplication/pdf
dc.identifier.urihttps://hdl.handle.net/20.500.12503/28869
dc.language.isoen
dc.provenance.legacyDownloads0
dc.subjectCell Anatomy
dc.subjectCell Biology
dc.subjectCells
dc.subjectLife Sciences
dc.subjectMedical Cell Biology
dc.subjectMedicine and Health Sciences
dc.subjectMembrane Science
dc.subjectOphthalmology
dc.subjectOptometry
dc.subjectOther Cell and Developmental Biology
dc.subjectSense Organs
dc.subjectVision Science
dc.subjectendothelin converting enzyme-1
dc.subjectendothelin-1
dc.subjectBig endothelin-1
dc.subjectciliary epithelium
dc.subjectaqueous humor dynamics
dc.subjectintraocular pressure
dc.subjectWestern blotting
dc.subjectECE-1
dc.titleCharacterization and Activity of Endothelin Converting Enzyme-1 in Human Non-Pigmented Ciliary Epithelial Cells
dc.typeThesis
dc.type.materialtext
thesis.degree.departmentGraduate School of Biomedical Sciences
thesis.degree.disciplineBiomedical Sciences
thesis.degree.grantorUniversity of North Texas Health Science Center at Fort Worth
thesis.degree.nameMaster of Science

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