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Two Bioactive Molecular Weight Fractions of a Conditioned Medium Enhance RPE Cell Survival on Age-Related Macular Degeneration and Aged Bruch's Membrane
PURPOSE: To characterize molecular weight fractions of bovine corneal endothelial cell conditioned medium (CM) supporting retinal pigment epithelium (RPE) cell survival on aged and age-related macular degeneration (AMD) Bruch's membrane. METHODS: CM was subject to size separation using centrifu...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
The Association for Research in Vision and Ophthalmology
2016
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4771074/ https://www.ncbi.nlm.nih.gov/pubmed/26933521 http://dx.doi.org/10.1167/tvst.5.1.8 |
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author | Sugino, Ilene K. Sun, Qian Springer, Carola Cheewatrakoolpong, Noounanong Liu, Tong Li, Hong Zarbin, Marco A. |
author_facet | Sugino, Ilene K. Sun, Qian Springer, Carola Cheewatrakoolpong, Noounanong Liu, Tong Li, Hong Zarbin, Marco A. |
author_sort | Sugino, Ilene K. |
collection | PubMed |
description | PURPOSE: To characterize molecular weight fractions of bovine corneal endothelial cell conditioned medium (CM) supporting retinal pigment epithelium (RPE) cell survival on aged and age-related macular degeneration (AMD) Bruch's membrane. METHODS: CM was subject to size separation using centrifugal filters. Retentate and filtrate fractions were tested for bioactivity by analyzing RPE survival on submacular Bruch's membrane of aged and AMD donor eyes and behavior on collagen I-coated tissue culture wells. Protein and peptide composition of active fractions was determined by mass spectrometry. RESULTS: Two bioactive fractions, 3-kDa filtrate and a 10-50–kDa fraction, were necessary for RPE survival on aged and AMD Bruch's membrane. The 3-kDa filtrate, but not the 10-50–kDa fraction, supported RPE growth on collagen 1‐coated tissue culture plates. Mass spectrometry of the 10-50–kDa fraction identified 175 extracellular proteins, including growth factors and extracellular matrix molecules. Transforming growth factor (TGF)β-2 was identified as unique to active CM. Peptides representing 29 unique proteins were identified in the 3-KDa filtrate. CONCLUSIONS: These results indicate there is a minimum of two bioactive molecules in CM, one found in the 3-kDa filtrate and one in the 10-50–kDa fraction, and that bioactive molecules in both fractions must be present to ensure RPE survival on Bruch's membrane. Mass spectrometry analysis suggested proteins to test in future studies to identify proteins that may contribute to CM bioactivity. TRANSLATIONAL RELEVANCE: Results of this study are the first steps in development of an adjunct to cell-based therapy to ensure cell transplant survival and functionality in AMD patients. |
format | Online Article Text |
id | pubmed-4771074 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2016 |
publisher | The Association for Research in Vision and Ophthalmology |
record_format | MEDLINE/PubMed |
spelling | pubmed-47710742016-03-01 Two Bioactive Molecular Weight Fractions of a Conditioned Medium Enhance RPE Cell Survival on Age-Related Macular Degeneration and Aged Bruch's Membrane Sugino, Ilene K. Sun, Qian Springer, Carola Cheewatrakoolpong, Noounanong Liu, Tong Li, Hong Zarbin, Marco A. Transl Vis Sci Technol Articles PURPOSE: To characterize molecular weight fractions of bovine corneal endothelial cell conditioned medium (CM) supporting retinal pigment epithelium (RPE) cell survival on aged and age-related macular degeneration (AMD) Bruch's membrane. METHODS: CM was subject to size separation using centrifugal filters. Retentate and filtrate fractions were tested for bioactivity by analyzing RPE survival on submacular Bruch's membrane of aged and AMD donor eyes and behavior on collagen I-coated tissue culture wells. Protein and peptide composition of active fractions was determined by mass spectrometry. RESULTS: Two bioactive fractions, 3-kDa filtrate and a 10-50–kDa fraction, were necessary for RPE survival on aged and AMD Bruch's membrane. The 3-kDa filtrate, but not the 10-50–kDa fraction, supported RPE growth on collagen 1‐coated tissue culture plates. Mass spectrometry of the 10-50–kDa fraction identified 175 extracellular proteins, including growth factors and extracellular matrix molecules. Transforming growth factor (TGF)β-2 was identified as unique to active CM. Peptides representing 29 unique proteins were identified in the 3-KDa filtrate. CONCLUSIONS: These results indicate there is a minimum of two bioactive molecules in CM, one found in the 3-kDa filtrate and one in the 10-50–kDa fraction, and that bioactive molecules in both fractions must be present to ensure RPE survival on Bruch's membrane. Mass spectrometry analysis suggested proteins to test in future studies to identify proteins that may contribute to CM bioactivity. TRANSLATIONAL RELEVANCE: Results of this study are the first steps in development of an adjunct to cell-based therapy to ensure cell transplant survival and functionality in AMD patients. The Association for Research in Vision and Ophthalmology 2016-02-22 /pmc/articles/PMC4771074/ /pubmed/26933521 http://dx.doi.org/10.1167/tvst.5.1.8 Text en http://creativecommons.org/licenses/by-nc-nd/4.0/ This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License. |
spellingShingle | Articles Sugino, Ilene K. Sun, Qian Springer, Carola Cheewatrakoolpong, Noounanong Liu, Tong Li, Hong Zarbin, Marco A. Two Bioactive Molecular Weight Fractions of a Conditioned Medium Enhance RPE Cell Survival on Age-Related Macular Degeneration and Aged Bruch's Membrane |
title | Two Bioactive Molecular Weight Fractions of a Conditioned Medium Enhance RPE Cell Survival on Age-Related Macular Degeneration and Aged Bruch's Membrane |
title_full | Two Bioactive Molecular Weight Fractions of a Conditioned Medium Enhance RPE Cell Survival on Age-Related Macular Degeneration and Aged Bruch's Membrane |
title_fullStr | Two Bioactive Molecular Weight Fractions of a Conditioned Medium Enhance RPE Cell Survival on Age-Related Macular Degeneration and Aged Bruch's Membrane |
title_full_unstemmed | Two Bioactive Molecular Weight Fractions of a Conditioned Medium Enhance RPE Cell Survival on Age-Related Macular Degeneration and Aged Bruch's Membrane |
title_short | Two Bioactive Molecular Weight Fractions of a Conditioned Medium Enhance RPE Cell Survival on Age-Related Macular Degeneration and Aged Bruch's Membrane |
title_sort | two bioactive molecular weight fractions of a conditioned medium enhance rpe cell survival on age-related macular degeneration and aged bruch's membrane |
topic | Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4771074/ https://www.ncbi.nlm.nih.gov/pubmed/26933521 http://dx.doi.org/10.1167/tvst.5.1.8 |
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