microRNA-150 Regulates Mobilization and Migration of Bone Marrow-Derived Mononuclear Cells by Targeting Cxcr4 Nobuko Tano, Ha Won Kim, Muhammad Ashraf* Department of Pathology and Lab Medicine, University of Cincinnati, Cincinnati, Ohio, United States of America Abstract The interaction between chemokine receptor type 4 (CXCR4) and its ligand, stromal cell-derived factor (SDF)-1, plays an important role in stem cell mobilization and migration in ischemic tissues. MicroRNAs (miRs) are key regulators of stem cell function and are involved in regulation of stem cell survival and differentiation to adopt different cell lineages. In this study, we show that ischemia inhibits the expression of miR-150 in BM-derived mononuclear cells (MNC) and activates its target Cxcr4 gene. Our results show that miR-150/CXCR4 cascade enhances MNC mobilization and migration. By using mouse acute myocardial infarction (MI) model, we found that MNCs in peripheral blood (PB) were increased significantly at day 5 after AMI as compared to control group and the number of CXCR4 positive MNCs both in bone marrow (BM) and PB was also markedly increased after MI. Analysis by microarray-based miRNA profiling and real-time PCR revealed that the expression of miR-150 which targets Cxcr4 gene as predicted was significantly downregulated in BM-MNCs after MI. Abrogation of miR-150 markedly increased CXCR4 protein expression suggesting its target gene. To show that miR-150 regulates MNC mobilization, knockdown of miR-150 in BM-MNCs by specific antisense inhibitor resulted in their higher migration ability in vitro as compared to scramble-transfected MNCs. Furthermore, in vivo BM transplantation of MNCs lacking miR-150 expression by lentiviral vector into the irradiated wild type mice resulted in the increased number of MNCs in PB after AMI as compared to control. In conclusion, this study demonstrates that ischemia mobilizes BM stem cells via miR-150/CXCR4 dependent mechanism and miR-150 may be a novel therapeutic target for stem cell migration to the ischemic tissue for neovascularization and repair. Citation: Tano N, Kim HW, Ashraf M (2011) microRNA-150 Regulates Mobilization and Migration of Bone Marrow-Derived Mononuclear Cells by Targeting Cxcr4. PLoS ONE 6(10): e23114. doi:10.1371/journal.pone.0023114 Editor: Piero Anversa, Brigham and Women’s Hospital, United States of America Received June 22, 2011; Accepted July 6, 2011; Published October 19, 2011 Copyright: 2011 Tano et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: Funding was provided by National Institutes of Health grants R37-HL074272; HL-080686; HL087246 (M.A) and HL087288; HL089535; HL106190-01 (K.H.H). The funder had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: Ashrafm@ucmail.uc.edu Introduction Ischemic heart disease is a leading cause of death worldwide. As a result of an insufficient blood supply of the heart muscle by coronary occlusion, loss of viable cardiomyocytes and the reduction of cardiac output can be induced during AMI phase. BM-derived mononuclear cells including endothelial progenitor cells (EPCs) play an important role in the maintenance of vascular integrity [1,2]. Since MNCs/EPCs are able to differentiate into mature endothelial cells and promote repair of damaged endothelium, they are attractive target for the repair of ischemic tissue [3–5]. MNC/EPC number and function are closely associated with coronary endothelial function, and reduced levels of circulating MNCs/EPCs have been shown to be independent predictors of atherosclerotic disease progression [6]. Therefore, sufficient MNC/EPC numbers, as well as the capacity to differentiate into mature endothelial cells are considered to be essential for myocardial functional recovery and infarct size reduction. Interaction between stromal cell-derived factor-1a (SDF-1a, or CXCL12a) and its receptor CXC chemokine receptor 4 (CXCR4, or fusin/CD184) plays a key role in mobilization of vascular stem/ progenitor cells [7]. As one of the strategies to rescue cardiac dysfunctions after AMI, the modification of CXCR4 expression in BM-derived stem cells has been investigated by using various BM- derived stem cells [8]. For instance, hypoxic preconditioning of cardiac stem/progenitor cells (cardiosphere-derived, Lin2 c-kit+ progenitor cells) upregulates CXCR4 expression and increases the recruitment of these cells into the ischemic myocardium, thereby reducing the infarct size and improving the cardiac function after MI [9]. In addition, intravenous delivery of mesenchymal stem cells (MSCs) overexpressing CXCR4 improves cardiac function and remodeling after MI, suggesting CXCR4 as an important therapeutic target for the treatment of cardiovascular diseases [10]. MicroRNAs (miRs) play an important role in the posttranscrip- tional regulation of target mRNA in a range of biological processes, including maintenance of stemness and modulation of mobilization, proliferation and differentiation. miRNAs are short (19–23 nucleotides), noncoding small regulatory RNAs that are loaded into the RNA-induced silencing complex, recognize the 39- untranslated region (UTR) of target genes, and thereby regulate their expression by translational repression or mRNA degradation. [11,12] Donahue and colleagues have previously profiled miR expression in response to Plerixafor (AMD3100, hematopoietic mobilizing agent) and granulocyte colony-stimulating factor (G- PLoS ONE | www.plosone.org 1 October 2011 | Volume 6 | Issue 10 | e23114