Thomas A Vo1, Sina Abedi1, Kevin Schneider1, Marilyn Chwa1, M Cristina Kenney2. 1. Gavin Herbert Eye Institute, University of California, Irvine, CA, 92697, USA. 2. Gavin Herbert Eye Institute, University of California, Irvine, CA, 92697, USA; Department of Pathology and Laboratory Medicine, University of California Irvine, Irvine, CA, USA. Electronic address: mkenney@uci.edu.
Abstract
PURPOSE: A critical biological function of retina pigment epithelium (RPE) cells is phagocytosis of photoreceptor outer segment (POS) disc membranes. Mitochondrial damage and dysfunction are associated with RPE cells of age-related macular degeneration (AMD) retinas. In this study, we use a transmitochondrial cybrid model to compare the phagocytic properties of RPE cells that contain AMD mitochondria versus age-matched normal mitochondria and their response to treatment with anti-vascular endothelial growth factor (VEGF) drugs: bevacizumab, ranibizumab, and aflibercept. METHODS: Cybrids, which are cell lines with identical nuclei but mitochondria (mt) from different subjects, are created by fusing mtDNA depleted ARPE-19 cells with platelets from AMD or age-matched normal patients. AMD (n = 5) and normal (n = 5) cybrids were treated with 1 μm fluorescent latex beads (1.52 × 107 beads/mL) and either 2.09 μM of bevacizumab, 2.59 μM of ranibizumab, or 5.16 μM of aflibercept. These doses of anti-VEGF drugs are equivalent to intravitreal injections given to AMD patients with choroidal neovascularization. Flow cytometry was performed using the ImageStreamX Mark II to assess phagocytic bead-uptake. The average fold values for bead-uptake and SEM were calculated using GraphPad Prism software. RESULTS: Normal cybrids showed decreased bead-uptake with a fold value of 0.65 ± 0.103 (p = 0.01) after treatment with bevacizumab, 0.80 ± 0.034 (p = 0.0003) with ranibizumab, and 0.81 ± 0.053 (p = 0.007) with aflibercept compared to the untreated normal cybrids (baseline fold of 1). The bevacizumab-treated, ranibizumab-treated, and aflibercept-treated AMD cybrids had decreased bead-uptake with a fold value of 0.71 ± 0.061 (p = 0.001), 0.70 ± 0.101 (p = 0.02), and 0.74 ± 0.125 (p = 0.07), respectively, compared to the untreated AMD cybrids (baseline fold of 1). CONCLUSIONS: Our initial findings showed that when treated with bevacizumab and ranibizumab, both AMD cybrids and age-matched normal cybrids had a significant decrease in bead-uptake. A similar decrease in bead-uptake was found in normal cybrids treated with aflibercept and while the AMD values trended lower, they were not significant. This data suggests that anti-VEGF drugs can cause loss of phagocytic function.
PURPOSE: A critical biological function of retina pigment epithelium (RPE) cells is phagocytosis of photoreceptor outer segment (POS) disc membranes. Mitochondrial damage and dysfunction are associated with RPE cells of age-related macular degeneration (AMD) retinas. In this study, we use a transmitochondrial cybrid model to compare the phagocytic properties of RPE cells that contain AMD mitochondria versus age-matched normal mitochondria and their response to treatment with anti-vascular endothelial growth factor (VEGF) drugs: bevacizumab, ranibizumab, and aflibercept. METHODS: Cybrids, which are cell lines with identical nuclei but mitochondria (mt) from different subjects, are created by fusing mtDNA depleted ARPE-19 cells with platelets from AMD or age-matched normal patients. AMD (n = 5) and normal (n = 5) cybrids were treated with 1 μm fluorescent latex beads (1.52 × 107 beads/mL) and either 2.09 μM of bevacizumab, 2.59 μM of ranibizumab, or 5.16 μM of aflibercept. These doses of anti-VEGF drugs are equivalent to intravitreal injections given to AMDpatients with choroidal neovascularization. Flow cytometry was performed using the ImageStreamX Mark II to assess phagocytic bead-uptake. The average fold values for bead-uptake and SEM were calculated using GraphPad Prism software. RESULTS: Normal cybrids showed decreased bead-uptake with a fold value of 0.65 ± 0.103 (p = 0.01) after treatment with bevacizumab, 0.80 ± 0.034 (p = 0.0003) with ranibizumab, and 0.81 ± 0.053 (p = 0.007) with aflibercept compared to the untreated normal cybrids (baseline fold of 1). The bevacizumab-treated, ranibizumab-treated, and aflibercept-treated AMD cybrids had decreased bead-uptake with a fold value of 0.71 ± 0.061 (p = 0.001), 0.70 ± 0.101 (p = 0.02), and 0.74 ± 0.125 (p = 0.07), respectively, compared to the untreated AMD cybrids (baseline fold of 1). CONCLUSIONS: Our initial findings showed that when treated with bevacizumab and ranibizumab, both AMD cybrids and age-matched normal cybrids had a significant decrease in bead-uptake. A similar decrease in bead-uptake was found in normal cybrids treated with aflibercept and while the AMD values trended lower, they were not significant. This data suggests that anti-VEGF drugs can cause loss of phagocytic function.
Authors: Nitin Udar; Shari R Atilano; Masood Memarzadeh; David S Boyer; Marilyn Chwa; Stephanie Lu; Barak Maguen; Jonathan Langberg; Pinar Coskun; Douglas C Wallace; Anthony B Nesburn; Nikan Khatibi; Dieter Hertzog; Khoi Le; Daniel Hwang; M Cristina Kenney Journal: Invest Ophthalmol Vis Sci Date: 2009-01-17 Impact factor: 4.799
Authors: M Cristina Kenney; Marilyn Chwa; Shari R Atilano; Payam Falatoonzadeh; Claudio Ramirez; Deepika Malik; Mohamed Tarek; Javier Cáceres-del-Carpio; Anthony B Nesburn; David S Boyer; Baruch D Kuppermann; Marquis Vawter; S Michal Jazwinski; Michael Miceli; Douglas C Wallace; Nitin Udar Journal: Hum Mol Genet Date: 2014-02-28 Impact factor: 6.150
Authors: Sonali Nashine; Marilyn Chwa; Mina Kazemian; Kunal Thaker; Stephanie Lu; Anthony Nesburn; Baruch D Kuppermann; M Cristina Kenney Journal: PLoS One Date: 2016-08-03 Impact factor: 3.240