Journal
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
Volume 23, Issue 12, Pages -Publisher
MDPI
DOI: 10.3390/ijms23126527
Keywords
co-culture; secretome; azidonorleucine; click chemistry; BONCAT; mesenchymal stromal cells
Funding
- Korea Health Industry Development Institute [HI14C3484]
- National Research Foundation of Korea [2017M3C9A5031595, 2017M3A9F9030559]
- KIST intramural program
- National Research Foundation of Korea [2017M3A9F9030559, 2017M3C9A5031595] Funding Source: Korea Institute of Science & Technology Information (KISTI), National Science & Technology Information Service (NTIS)
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Co-culture system is advantageous in mimicking in vivo niche of cells. This study presents a strategy to analyze the secretome of a specific cell type under co-culture condition and applies it to profile the secretome of mesenchymal stem cells.
Co-culture system, in which two or more distinct cell types are cultured together, is advantageous in that it can mimic the environment of the in vivo niche of the cells. In this study, we presented a strategy to analyze the secretome of a specific cell type under the co-culture condition in serum-supplemented media. For the cell-specific secretome analysis, we expressed the mouse mutant methionyl-tRNA synthetase for the incorporation of the non-canonical amino acid, azidonorleucine into the newly synthesized proteins in cells of which the secretome is targeted. The azidonorleucine-tagged secretome could be enriched, based on click chemistry, and distinguished from any other contaminating proteins, either from the cell culture media or the other cells co-cultured with the cells of interest. In order to have more reliable true-positive identifications of cell-specific secretory bodies, we established criteria to exclude any identified human peptide matched to bovine proteins. As a result, we identified a maximum of 719 secreted proteins in the secretome analysis under this co-culture condition. Last, we applied this platform to profile the secretome of mesenchymal stem cells and predicted its therapeutic potential on osteoarthritis based on secretome analysis.
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