Macrophage-Associated Mesenchymal Stem Cells Assume an Activated, Migratory, Pro-Inflammatory Phenotype with Increased IL-6 and CXCL10 Secretion Kevin Anton 1 , Debabrata Banerjee1,2, John Glod1,3* 1 Department of Pharmacology, The Cancer Institute of New Jersey, Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, New Brunswick, New Jersey, United States of America, 2 Department of Medicine, The Cancer Institute of New Jersey, Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, New Brunswick, New Jersey, United States of America, 3 Department of Pediatrics, The Cancer Institute of New Jersey, Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, New Brunswick, New Jersey, United State of America Abstract Mesenchymal stem cells (MSCs) exhibit tropism for sites of tissue injury and tumors. However, the influence of the microenvironment on MSC phenotype and localization remains incompletely characterized. In this study, we begin to define a macrophage-induced MSC phenotype. These MSCs secrete interleukin-6 (IL-6), CCL5, and interferon gamma-induced protein-10 (CXCL10) and exhibit increased mobility in response to multiple soluble factors produced by macrophages including IL-8, CCL2, and CCL5. The pro-migratory phenotype is dependent on activation of a c-Jun N-terminal kinase (JNK) pathway. This work begins to identify the influence of macrophages on MSC biology. These interactions are likely to play an important role in the tissue inflammatory response and may provide insight into the migratory potential of MSCs in inflammation and tissue injury. Citation: Anton K, Banerjee D, Glod J (2012) Macrophage-Associated Mesenchymal Stem Cells Assume an Activated, Migratory, Pro-Inflammatory Phenotype with Increased IL-6 and CXCL10 Secretion. PLoS ONE 7(4): e35036. doi:10.1371/journal.pone.0035036 Editor: Dimas Tadeu Covas, University of Sao Paulo – USP, Brazil Received November 4, 2011; Accepted March 8, 2012; Published April 4, 2012 Copyright: 2012 Anton 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: This work was supported by a grant from the New Jersey Commission on Cancer Research (09-1959-CCR-E0. The funders 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: glodjo@umdnj.edu Introduction It is well established that mesenchymal stem cells (MSCs) exhibit tropism for sites of tissue injury and the tumor microenvironment [1]. While the migratory ability of MSCs is well documented, the precise molecular mechanisms responsible for MSC homing to specific in vivo targets remain incompletely characterized. A better understanding of the MSC migratory process may identify therapeutic targets for the treatment of neoplastic and inflamma- tory disease and facilitate novel uses of MSCs such as targeted drug-delivery and gene therapy [2–4]. The inflammatory response plays a major role in forming the microenvironment of both injuries and tumors and the complex interplay between cellular components within this milieu influ- ences the pathophysiology of these conditions. The microenviron- ment of tumor types such as breast cancer and high-grade gliomas is characterized by a dense population of macrophages [5–9]. Tumor-associated macrophages (TAMs) promote tumor progres- sion by stimulating angiogenesis, inducing tumor cell invasion and metastasis, and conferring chemoresistant properties to tumor cells [10,11]. Macrophages also play a pivotal role in normal wound healing and tissue repair [12,13]. We tested the hypothesis that macrophages stimulate MSC localization to tumors and sites of injury and influence their molecular phenotype within the microenvironment. Our data show that macrophages, through the release of soluble factors, stimulate MSC motility and alter their cytokine secretion profile. Macrophages promote MSC migration through the production of soluble factors that activate the c-Jun NH2-terminal kinase (JNK) signaling pathway. A single cytokine or chemokine does not elicit the maximum MSC response. Our data suggest that the interaction between macrophages and MSCs impacts the inflammatory microenvironment. Results Macrophage-associated MSCs Up-regulate the Expression of Pro-inflammatory Cytokines Macrophages and MSCs are highly plastic cells that undergo changes in phenotype based upon local environmental cues. Both cell types are integral components of inflammation and are likely to interact during processes such as wound healing and solid tumor growth. In order to better define the impact of a macrophage-rich microenvironment on MSC phenotype we first identified changes in mRNA expression in response to culture in macrophage conditioned medium. MSCs exhibit changes in gene expression indicative of a pro-inflammatory phenotype, with increased expression of CCL7, IL-8, CCL20, CXCL6, CCL2 and CCL5 mRNA (Figure 1). We then determined whether increases in mRNA expression were accompanied by changes in cytokine secretion for five of the up-regulated genes in addition to IL-6, a cytokine that is highly expressed by pro- inflammatory MSC1 cells [14]. MSCs activated by macrophages had increased secretion of IL-6, CXCL10, and CCL5. To PLoS ONE | www.plosone.org 1 April 2012 | Volume 7 | Issue 4 | e35036