Article pubs.acs.org/jpr
Proteomic Analysis of Microvesicles Derived from Human Mesenchymal Stem Cells Han-Soo Kim,†,‡,◆ Do-Young Choi,§,◆ So Jeong Yun,⊥,◆ Seong-Mi Choi,¶ Jeong Won Kang,§ Jin Woo Jung,§ Daehee Hwang,*,⊥,#,∇ Kwang Pyo Kim,*,§ and Dong-Wook Kim*,‡,¶,∥ †
Department of Laboratory Medicine, ‡Cell Therapy Center, ¶Severance Biomedical Science Institute, and ∥Department of Physiology, Yonsei University College of Medicine, 50 Yonsei-ro, Seodaemoon-gu, Seoul 120-752, Republic of Korea § Department of Molecular Biotechnology, WCU Program, Konkuk University, Seoul, 143-701, Republic of Korea ⊥ School of Interdisciplinary Bioscience and Bioengineering, #Department of Chemical Engineering, and ∇Integrative Bioscience and Biotechnology, POSTECH, Pohang, 790-784, Republic of Korea S Supporting Information *
ABSTRACT: Mesenchymal stem cells (MSCs) have emerged as a promising means for treating degenerative or incurable diseases. Recent studies have shown that microvesicles (MVs) from MSCs (MSC-MVs) contribute to recovery of damaged tissues in animal disease models. Here, we profiled the MSC-MV proteome to investigate their therapeutic effects. LC−MS/MS analysis of MSCMVs identified 730 MV proteins. The MSC-MV proteome included five positive and two variable known markers of MSCs, but no negative marker, as well as 43 surface receptors and signaling molecules controlling self-renewal and differentiation of MSCs. Functional enrichment analysis showed that cellular processes represented by the MSC-MV proteins include cell proliferation, adhesion, migration, and morphogenesis. Integration of MSC’s self-renewal and differentiationrelated genes and the proteome of MSC-conditioned media (MSC-CM) with the MSC-MV proteome revealed potential MV protein candidates that can be associated with the therapeutic effects of MSC-MVs: (1) surface receptors (PDGFRB, EGFR, and PLAUR); (2) signaling molecules (RRAS/NRAS, MAPK1, GNA13/GNG12, CDC42, and VAV2); (3) cell adhesion (FN1, EZR, IQGAP1, CD47, integrins, and LGALS1/LGALS3); and (4) MSC-associated antigens (CD9, CD63, CD81, CD109, CD151, CD248, and CD276). Therefore, the MSC-MV proteome provides a comprehensive basis for understanding the potential of MSC-MVs to affect tissue repair and regeneration. KEYWORDS: mesenchymal stem cells, microvesicle, proteomics, self-renewal, tissue regeneration
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INTRODUCTION Mesenchymal stem cells (MSCs) are a heterogeneous population of stem cells that have the capabilities to self-renew and also differentiate into distinct mesodermal lineages1 and other embryonic lineages, such as endodermal and ectodermal cells.2−4 In addition, MSCs possess immunomodulatory or anti-inflammatory properties and trophic effects.5,6 Compared to embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs), and other tissue-specific stem cells, MSCs have greater availability, fewer ethical concerns, and low immunogenicity with immunomodulatory capacity, thus serving as ideal candidates for clinical applications in regenerative medicine. MSCs have been isolated from the brain, liver, lung, fetal blood, umbilical cord blood, kidneys, adipose tissue, and placentas.7 This broad distribution throughout the body suggests that MSCs play critical roles in tissue homeostasis.8 Numerous studies have demonstrated the ability of MSCs to differentiate into mesenchymal lineage cells, such as bone, cartilage, tendon, and fat in vitro.2 This has led to preclinical and clinical studies © 2011 American Chemical Society
that have further confirmed their therapeutic potential in bone and cartilage repairs.9 Moreover, clinical studies of MSC-based cellular therapy have shown therapeutic promise for a broad spectrum of human diseases including chronic graft-versus-host diseases (GvHD) upon allogeneic bone marrow transplantation or hematopoietic stem cell transplantation, myocardial infarction, systemic lupus erythematosus, liver cirrhosis, diabetes, acute kidney injury, and various neurodegenerative diseases.10−16 However, the mechanisms underlying the regenerative potential of MSCs are still elusive. Furthermore, the efficacy of MSC-based cell therapy has been questioned because the therapeutic effects have often been observed in the absence of MSC engraftment to target tissues other than connective tissues and organs. Frequently, the level of improvement of diseased tissues showed no correlation with that of observed cellular engraftments and differentiation of MSCs to tissue cells, Received: July 19, 2011 Published: December 8, 2011 839
dx.doi.org/10.1021/pr200682z | J. Proteome Res. 2012, 11, 839−849
Journal of Proteome Research
Article
Isolation and Characterization of MSCs from Human Bone Marrow Samples
suggesting that they may play indirect roles in the tissue regeneration. In a murine hindlimb ischemia model, Kinnaird et al.17 further demonstrated that the injection of MSCconditioned media (CM) alone was sufficient to mediate the improved function of hindlimbs. Thus, it has been postulated that the therapeutic effects exerted by MSCs can be mediated by trophic factors secreted by MSCs. Recent studies have suggested that some of the regenerative effects are mediated by microvesicles (MVs) derived from MSCs.18−20 Plasma membrane-derived MVs with a diameter of
