between stem cells and keloid-derived fibroblasts may contribute to keloid formation by triggering progressive myofibroblastic differentiation and secretion of abundant extracellular matrix [7]. In addition, the fibroproliferative nature of keloids suggests that mesenchymal stem cells may serve as primary source of multipotent cells to rapidly repopulate the wound site in response to exogeneous insults, such as trauma, surgical injury, or infection. Therefore, keloids provide an ideal benign tumor model to study the role of stem cells and their special niche components in promoting and maintaining the proliferative growth of the dermal scar. The fate of the stem cells, undergoing self-renewal or differentiation, is dependent on a specialized microenvironment or niche in which the cells reside [8]. The stem cell niche [9] encompasses all elements immediately surrounding the stem cells, including non-stem cells, the extracellular matrix (ECM), as well as soluble molecules present in the locale [10,11]. Under homeostatic or physiological conditions, the extrinsic niche components or growth factors protect the stem cells from excessive proliferation by providing a balanced proliferation-inhibiting and proliferation- promoting signal [12,13]. Meanwhile, the stem cells must periodically activate to produce specific lineage progenies for the regeneration or repair of tissues [10]. Therefore, the maintenance of a steady state between stem cell quiescence and activity is the hallmark of a functionally normal niche [13], whereas the deregulation of niche signals may lead to the uncontrolled self- renewal and proliferation of the stem cells, thus contributing to the emergence of the so-called niche-generated diseases including pre- cancer and tumorigenesis [11,14]. Interleukin (IL)-6 is produced at the site of acute inflammation [15] and plays a critical role in both cellular and humoral immune responses [16]. IL-6 can trigger the switch from acute to chronic inflammation by enhancing the recruitment of monocytes [17], thereby, serving as a key player in both acute and chronic inflammation [18]. Numerous studies have reported elevated IL-6 levels in several inflammatory diseases, including chronic inflam- matory and fibro-proliferative diseases such as keloids [5,6], rheumatoid arthritis, inflammatory bowel disease (colitis), multiple sclerosis, and pulmonary fibrosis [19,20]. Aside from its established function as an immuno-modulator, IL-6 may contribute to the regulation of stem cell functions via several pathways leading to the inhibition of lymphocyte apoptosis [21] and the maintenance of hematopoietic stem cells [22]. Most recently, IL-6 has been reported as a critical tumor promoter during early colitis- associated tumorigenesis [23,24]. Immune cells, which often infiltrate tumor and pre-neoplastic lesions, are capable of perpetuating a localized inflammatory response via a variety of cytokines and chemokines, and enhancing the growth and survival of premalignant cells. IL-6 has also been demonstrated to trigger malignant features in Notch-3-expressing stem/progenitor cells from human ductal breast carcinoma and normal mammary gland [25] and further up-regulates telomerase activity in human malignancies [26,27]. Since telomerase plays an essential role in the regulation of cell proliferation and senescence [28,29] it has been routinely used as a functional marker to assess stem cell function and tissue homeostasis [30,31]. IL-17, a recently discovered pro-inflammatory cytokine, is secreted by a distinct subtype of activated CD4+ T-cells known as Th17 [32,33]. Most recently, several studies have clearly demonstrated the critical role of TGF-b and IL-6 or other inflammatory cytokines in the differentiation of human Th17 cells [34–37]. The finding of this cytokine has changed our perspectives on chronic inflammatory diseases, including inflammatory bowel disease, rheumatoid arthritis, psoriasis, multiple sclerosis, and Niche-Driven Overgrowth of KPC allergic skin immune responses [38–41]. IL-17 is produced exclusively by activated T cells, however, its receptor is ubiquitously expressed in many cell types, thus making them potential targets [35,42]. IL-17 amplifies the immune response mediated by a variety of cytokines such as IL-6, tumor necrosis factor (TNF)- a and IL-1b, chemokines such as monocyte chemoattachment protein-1 (MCP-1) and macrophage inflamma- tory protein-2 (MIP-2)/IL-8, cell-surface markers such as inter- cellular adhesion molecule-1 (ICAM-1), and pro-inflammatory mediators including prostaglandin E2, nitric oxide, cyclooxygen- ase-2, and C-reactive protein [43–47]. Among these inflammatory cytokines, IL-6 seems to be a major IL-17 signaling target in a variety of cells including macrophages, fibroblasts, osteoblasts, epithelial cells, and chondrocytes [44,45,48–51]. More important- ly, IL-6 not only functions downstream of IL-17 but also acts as a critical upstream target of IL-17, thus forming a paracrine/ autocrine feedback loop that promotes autoimmune and allergic diseases [50,51]. In addition to its immuno-regulatory role, IL-17A promotes the proliferation, migration, and osteogenic differentia- tion of human bone marrow mesenchymal stem cells (BMMSCs) [52]. Based on existing findings we asked whether IL-17 and IL-6 coordinately interact and contribute to the persistent local tissue inflammation in keloid, i.e. sustaining the ‘‘pathological’’ niche, therefore support the proliferation-promoting signal of the benign dermal growth. Previously, stem cells derived from the dermis have been isolated and cultured under either adherent conditions with fetal bovine serum (FBS)-containing medium [53] or neural sphere- forming condition in serum free medium supplemented with epidermal growth factor (EGF) and fibroblast growth factor (FGF)- 2 [54,55,56]. In the current study, using both adherent and sphere-culturing conditions we have isolated and characterized a population of adult stem cells from the dermal layer of keloid scars, named keloid derived precursor cells (KPCs), which are charac- terized by several defined stem cell features including clonogeni- city, self-renewal, expression of distinct embryonic and mesenchy- mal stem cell markers, and multipotent differentiation. We have also defined an essential inflammatory cytokine paracrine loop between IL-17 and IL-6, constituting the ‘‘pathological’’ keloid niche that is capable of elevating the proliferation-promoting signal. Using KPCs transplanted in immunocompromised mice we generate a keloid-like tumor model that is driven by the in vivo inflammatory niche. Understanding how the inflammatory niche regulates tumor-like stem cell growth in keloid is critical in further elucidation of keloid pathogenesis and will open a new avenue for benign tumor stem cell research and ultimately, keloid therapies. Materials and Methods Ethics Statement This study followed the tenets of the Declaration of Helsinki and all keloid tissues were obtained as discarded biological samples from patient donors (aged 20,50 years of age) after a written informed consent was provided. This study was approved by the Institutional Ethics Committee/Institutional Review Board (IRB) of both University of Southern California and Harbor-UCLA/ King Drew Medical Center. Antibodies and reagents All antibodies used in this study were listed in Table S1. Recombinant human interleukin-6 and interleukin-17A were purchased from PeproTech Inc. (Rocky Hills, NJ, USA). All chemical reagents were analytical grade and obtained from Sigma-Aldrich (St. Louis, MO). PLoS ONE | www.plosone.org 2 November 2009 | Volume 4 | Issue 11 | e7798