Subject(s)
Antibodies, Anti-Idiotypic/immunology , Mesenchymal Stem Cell Transplantation , Mesenchymal Stem Cells/metabolism , Animals , Cross Reactions , Dogs , Flow Cytometry , Humans , Intervertebral Disc Degeneration/pathology , Intervertebral Disc Degeneration/therapy , Low Back Pain/pathology , Low Back Pain/therapy , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/immunologyABSTRACT
Recruitment of mesenchymal stem cells (MSC) following cardiac injury, such as myocardial infarction, plays a critical role in tissue repair and may contribute to myocardial recovery. However, the mechanisms that regulate migration of MSC to the site of tissue damage remain elusive. Here, we demonstrate in vitro that activated platelets substantially inhibit recruitment of MSC toward apoptotic cardiac myocytes and fibroblasts. The alarmin high mobility group box 1 (HMGB1) was released by platelets upon activation and mediated inhibition of the cell death-dependent migratory response through Toll-like receptor (TLR)-4 expressed on the MSC. Migration of MSC to apoptotic cardiac myocytes and fibroblasts was driven by hepatocyte growth factor (HGF), and platelet activation was followed by HMGB1/TLR-4-dependent down-regulation of HGF receptor MET on MSC, thereby impairing HGF-driven MSC recruitment. We identify a novel mechanism by which platelets, upon activation, interfere with MSC recruitment to apoptotic cardiac cells, a process that may be of particular relevance for myocardial repair and regeneration.
Subject(s)
Apoptosis/physiology , Blood Platelets/metabolism , Cell Movement/physiology , Down-Regulation/physiology , Fibroblasts/metabolism , HMGB1 Protein/metabolism , Mesenchymal Stem Cells/metabolism , Myocytes, Cardiac/metabolism , Platelet Activation/physiology , Proto-Oncogene Proteins c-met/biosynthesis , Toll-Like Receptor 4/metabolism , Blood Platelets/cytology , Fibroblasts/cytology , HMGB1 Protein/genetics , Hepatocyte Growth Factor/genetics , Hepatocyte Growth Factor/metabolism , Humans , Mesenchymal Stem Cells/cytology , Myocardium/cytology , Myocardium/metabolism , Myocytes, Cardiac/cytology , Proto-Oncogene Proteins c-met/genetics , Regeneration/physiology , Toll-Like Receptor 4/geneticsABSTRACT
Several strategies have been developed to facilitate the prospective isolation of bone marrow-derived mesenchymal stem/stromal cells (BM-MSCs) based on the selective expression or absence of surface markers. Recently, we described the monoclonal antibodies W3D5 and W5C5, which selectively react with BM-MSCs, but not with hematopoietic cells. Both antibodies showed an identical reactivity pattern, indicating that they may recognize the same molecule. To identify the cognate antigen, cultured MSCs were sorted for cells expressing either very high levels of W5C5/W3D5 antigen or for cells which were negative for this antigen. Further processing of these cells for microarray analysis revealed a 20-fold enrichment of the type 1 integral membrane protein Sushi domain containing 2 (SUSD2) in the in W5C5(+) subset. To confirm the identity of the W5C5/W3D5 antigen to SUSD2, HEK293 cells were transfected with the full-length coding sequence of human SUSD2 followed by reactivity analysis of W5C5 and W3D5 antibodies with the transfected line. Flow cytometric analysis showed that both antibodies selectively recognized HEK293/huSUSD2 cells, but not the parental cell line. In line with this, SUSD2 siRNA treatment of SUSD2(+) WERI-RB-1 retinoblastoma cells reduced the expression levels of W3D5 and W5C5 antigens to ~39% and 37%, respectively. Finally, FACSorting and colony assays revealed that only SUSD2(+), but not SUSD2(-) BM cells give rise to colony-forming units-fibroblasts and are able to differentiate into osteoblasts, adipocytes, and chondrocytes. In conclusion, we identified SUSD2 as a novel and specific marker for the prospective isolation of BM-MSCs.