Human mesenchymal stem cells secretome investigated by using a label-free proteomic approach

Introduction Mesenchymal stem cells (MSCs) have been shown to have a pivotal role in tissue repair and in local control of inflammation. Although there is evidence from in vivo studies that MSCs are able to reduce inflammatory damage, it is not clear whether their immunomodulatory effects rely on soluble factors or cell-cell contacts. In order to investigate potential activities associated with MSC-secreted proteins, the present study reports the secretome of MSC, isolated from human bone marrow cells, before (unstimulated-MSC) and after (stimulated-MSC) treatment with pro-inflammatory cytokines (TNFs, IL1b and IL6). Methods Secreted proteins were analyzed by a label free quantification based shot-gun proteomic approach using nano-LC-ESI-LTQ Orbitrap Velos Results Two biological replicates (each consisting of stimulated or unstimulated-MSC secretome), were analyzed (five technical replicates of each sample) leading to the identification and quantification of 609 and 608 proteins in the first and second biological replicate, respectively. According to the LFQ statistical analysis (using Perseus , t-test cut-off at 1% permutation-based False Discovery Rate) 231 proteins are differentially expressed in both replicates; in particular: 70 proteins are up-regulated or present only in stimulated MSC-secretome while 161 are down-regulated in stimulated-MSC or present on in unstimulated-MSC secretome. Conclusions The proteomic approach used to identify and quantify the secretome of human MSC cells allowed to pinpoint the main biological processes affected by differential expression of secreted proteins following stimulation of MSC with pro-inflammatory cytokines. This project was founded in part by LINEA B “Piano di Sviluppo Unimi 2014”, Università degli Studi di Milano.