Autophagy and Mitochondrial Dysfunction in Tenon Fibroblasts from Exfoliation Glaucoma Patients

To test the hypothesis that autophagy dysfunction is involved in exfoliation syndrome (XFS), a systemic disorder of extracellular elastic matrices that causes a distinct form of human glaucoma. Fibroblasts derived from tenon tissue discards (TFs) from filtration surgery to relieve intraocular pressu...

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Bibliographic Details
Published in:PloS one 2016-07, Vol.11 (7), p.e0157404-e0157404
Main Authors: Want, Andrew, Gillespie, Stephanie R, Wang, Zheng, Gordon, Ronald, Iomini, Carlo, Ritch, Robert, Wolosin, J Mario, Bernstein, Audrey M
Format: Article
Language:English
Subjects:
Age
Age related diseases
Aged
Aged, 80 and over
Aging
Amino Acid Oxidoreductases - genetics
Analysis
Apoptosis
Autophagy
Autophagy (Cytology)
Biology and Life Sciences
Brain
Care and treatment
Cell culture
Cell death
Cell size
Child, Preschool
Comparative analysis
Cytometry
Diagnosis
Disease
Electron microscopy
Endosomes
Exfoliatins
Exfoliation
Exfoliation Syndrome - metabolism
Exfoliation Syndrome - surgery
Experiments
Extracellular matrix
Female
Fibroblasts
Fibroblasts - metabolism
Flow cytometry
Genetic aspects
Glaucoma
Glaucoma, Open-Angle - metabolism
Glaucoma, Open-Angle - surgery
Golgi apparatus
Humans
Immunocytochemistry
Intraocular Pressure
Light
Light scattering
Lysosomes
Male
Medicine
Medicine and Health Sciences
Membrane potential
Membrane Potentials
Microtubules
Middle Aged
Mitochondria
Mitochondrial Membranes - metabolism
Neurodegeneration
Organelles
Pathology
Patients
Phagocytosis
Photoreceptors
Physical Sciences
Proteins
Relocation
Retina
Scattering
Scattering, Radiation
Strabismus
Strabismus - surgery
Surgery
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Summary:To test the hypothesis that autophagy dysfunction is involved in exfoliation syndrome (XFS), a systemic disorder of extracellular elastic matrices that causes a distinct form of human glaucoma. Fibroblasts derived from tenon tissue discards (TFs) from filtration surgery to relieve intraocular pressure in XFS patients were compared against age-matched TFs derived from surgery in primary open-angle glaucoma (POAG) patients or from strabismus surgery. Differential interference contrast light, and electron microscopy were used to examine structural cell features. Immunocytochemistry was used to visualize LOXL1 and Fibulin-5, lysosomes, endosomes, Golgi, and microtubules. Light scatter, Cyto-IDTM and JC1 flow cytometry were used to measure relative cell size, autophagic flux rate and mitochondrial membrane potential (MMPT), respectively. Enhanced autophagy was induced by serum withdrawal. In culture, XFS-TFs were 1.38-fold larger (by light scatter ratio, p = 0.05), proliferated 42% slower (p = 0.026), and were morphologically distinct in 2D and 3D culture compared to their POAG counterparts. In extended 3D cultures, XFS-TFs accumulated 8-10 times more Fibulin-5 than the POAG-TFs, and upon serum withdrawal, there were marked deficiencies in relocation of endosomes and lysosomes to the perinuclear area. Correspondingly, the XFS-TFs displayed significant accumulation of the autophagasome marker LC3 II (3.9 fold increase compared to POAG levels, p = 0.0001) and autophagic flux rate as measured by Cyto-ID dye was 53% lower in XFS-TFs than in POAG-TFs (p = 0.01), indicating reduced clearance of autophagasomes. Finally the percent of cells with diminished MMPT was 3-8 times larger in the XFS-TFs than in POAG-TFs (p = 0.02). Our results provide for the first time a link between XFS pathology to autophagy dysfunction, a major contributor to multiple age related diseases systemically throughout the body, in the brain and in the retina. A diminished capacity for degradation of denatured protein and aging cellular organelles may underpin the development of extracellular protein aggregates in XFS.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0157404