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High throughput proteomic analysis of the secretome in an explant model of articular cartilage inflammation
This study employed a targeted high-throughput proteomic approach to identify the major proteins present in the secretome of articular cartilage. Explants from equine metacarpophalangeal joints were incubated alone or with interleukin-1beta (IL-1β, 10 ng/ml), with or without carprofen, a non-steroid...
Autores principales: | , , , , , |
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Formato: | Texto |
Lenguaje: | English |
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Elsevier
2011
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3078332/ https://www.ncbi.nlm.nih.gov/pubmed/21354348 http://dx.doi.org/10.1016/j.jprot.2011.02.017 |
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author | Clutterbuck, Abigail L. Smith, Julia R. Allaway, David Harris, Pat Liddell, Susan Mobasheri, Ali |
author_facet | Clutterbuck, Abigail L. Smith, Julia R. Allaway, David Harris, Pat Liddell, Susan Mobasheri, Ali |
author_sort | Clutterbuck, Abigail L. |
collection | PubMed |
description | This study employed a targeted high-throughput proteomic approach to identify the major proteins present in the secretome of articular cartilage. Explants from equine metacarpophalangeal joints were incubated alone or with interleukin-1beta (IL-1β, 10 ng/ml), with or without carprofen, a non-steroidal anti-inflammatory drug, for six days. After tryptic digestion of culture medium supernatants, resulting peptides were separated by HPLC and detected in a Bruker amaZon ion trap instrument. The five most abundant peptides in each MS scan were fragmented and the fragmentation patterns compared to mammalian entries in the Swiss-Prot database, using the Mascot search engine. Tryptic peptides originating from aggrecan core protein, cartilage oligomeric matrix protein (COMP), fibronectin, fibromodulin, thrombospondin-1 (TSP-1), clusterin (CLU), cartilage intermediate layer protein-1 (CILP-1), chondroadherin (CHAD) and matrix metalloproteinases MMP-1 and MMP-3 were detected. Quantitative western blotting confirmed the presence of CILP-1, CLU, MMP-1, MMP-3 and TSP-1. Treatment with IL-1β increased MMP-1, MMP-3 and TSP-1 and decreased the CLU precursor but did not affect CILP-1 and CLU levels. Many of the proteins identified have well-established extracellular matrix functions and are involved in early repair/stress responses in cartilage. This high throughput approach may be used to study the changes that occur in the early stages of osteoarthritis. |
format | Text |
id | pubmed-3078332 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2011 |
publisher | Elsevier |
record_format | MEDLINE/PubMed |
spelling | pubmed-30783322011-06-28 High throughput proteomic analysis of the secretome in an explant model of articular cartilage inflammation Clutterbuck, Abigail L. Smith, Julia R. Allaway, David Harris, Pat Liddell, Susan Mobasheri, Ali J Proteomics Article This study employed a targeted high-throughput proteomic approach to identify the major proteins present in the secretome of articular cartilage. Explants from equine metacarpophalangeal joints were incubated alone or with interleukin-1beta (IL-1β, 10 ng/ml), with or without carprofen, a non-steroidal anti-inflammatory drug, for six days. After tryptic digestion of culture medium supernatants, resulting peptides were separated by HPLC and detected in a Bruker amaZon ion trap instrument. The five most abundant peptides in each MS scan were fragmented and the fragmentation patterns compared to mammalian entries in the Swiss-Prot database, using the Mascot search engine. Tryptic peptides originating from aggrecan core protein, cartilage oligomeric matrix protein (COMP), fibronectin, fibromodulin, thrombospondin-1 (TSP-1), clusterin (CLU), cartilage intermediate layer protein-1 (CILP-1), chondroadherin (CHAD) and matrix metalloproteinases MMP-1 and MMP-3 were detected. Quantitative western blotting confirmed the presence of CILP-1, CLU, MMP-1, MMP-3 and TSP-1. Treatment with IL-1β increased MMP-1, MMP-3 and TSP-1 and decreased the CLU precursor but did not affect CILP-1 and CLU levels. Many of the proteins identified have well-established extracellular matrix functions and are involved in early repair/stress responses in cartilage. This high throughput approach may be used to study the changes that occur in the early stages of osteoarthritis. Elsevier 2011-05-01 /pmc/articles/PMC3078332/ /pubmed/21354348 http://dx.doi.org/10.1016/j.jprot.2011.02.017 Text en © 2011 Elsevier B.V. https://creativecommons.org/licenses/by/3.0/ Open Access under CC BY 3.0 (https://creativecommons.org/licenses/by/3.0/) license |
spellingShingle | Article Clutterbuck, Abigail L. Smith, Julia R. Allaway, David Harris, Pat Liddell, Susan Mobasheri, Ali High throughput proteomic analysis of the secretome in an explant model of articular cartilage inflammation |
title | High throughput proteomic analysis of the secretome in an explant model of articular cartilage inflammation |
title_full | High throughput proteomic analysis of the secretome in an explant model of articular cartilage inflammation |
title_fullStr | High throughput proteomic analysis of the secretome in an explant model of articular cartilage inflammation |
title_full_unstemmed | High throughput proteomic analysis of the secretome in an explant model of articular cartilage inflammation |
title_short | High throughput proteomic analysis of the secretome in an explant model of articular cartilage inflammation |
title_sort | high throughput proteomic analysis of the secretome in an explant model of articular cartilage inflammation |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3078332/ https://www.ncbi.nlm.nih.gov/pubmed/21354348 http://dx.doi.org/10.1016/j.jprot.2011.02.017 |
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