More importantly, accumulating evidence has pointed to the critical role of stem cell niche in the regulation of stem cell functions. The balance between proliferation-inhibiting and proliferation-promoting signals provided by the specialized niche microenvironment is the key to homeostatic regulation of stem cell maintenance versus tissue regeneration [12]. Therefore, any changes in the niche components will lead to alterations of stem cell functions. A decline in signaling from the niche that is essential to the maintenance of stem cells can lead to aging and even loss of stem cell number and function [74,75], thus contributing to aging or degenerative diseases. On the other hand, deregulation or alteration of the niche by dominant proliferation-promoting signals may lead to overgrowth of resident stem cells, thus contributing to hyper-proliferative diseases, benign and malignant tumors [12]. Keloid, a fibro-proliferative benign tumor of skin, is characterized by chronic inflammation, an increased infiltration of inflammatory cells, an enriched milieu of cytokines and growth factors, and an abundant accumulation of ECM [1,2,57], thus providing a unique inflammatory niche for the resident stem cells. Using keloid as a niche-related disease model we explore the critical niche components and their role in the regulation of stem cell functions. IL-6, an inflammatory cytokine, plays a critical role in the pathogenesis of most fibrotic diseases [18] and chronic inflamma- tion-associated tumorigenesis [23,24]. Consistent with previous reports [5,6], we have demonstrated that IL-6 is significantly up- regulated in keloid scars versus their normal skin counterparts, and KPCs constitutively express a higher level of IL-6 and IL-6R compared to SKPs. Moreover, our results show that IL-6 can significantly stimulate the expression of both Oct-4 and telome- rase, two important stem cell function genes, in cultured KPCs and SKPs, and incubation with specific IL-6 neutralizing antibodies suppressed the constitutive expression of telomerase in KPCs. These findings support our hypothesis that the locally enriched IL-6 in keloid scars might constitute a major component of the inflammatory niche where KPCs reside and play a critical role in the regulation of KPC functions. Since an animal model for keloid scar does not currently exist, we proposed to generate keloid tumor using dermal derived precursor cells. To recapitulate the ‘‘pathological’’ niche of keloid, we utilized a newly introduced carrier Extracel-HPTM Hydrogel conjugated with IL-6 for the in vivo transplantation of KPCs [61,62]. We have demonstrated that IL-6 stimulates the expression of Oct-4 and telomerase in vivo and significantly promotes the growth of KPCs and SKPs-generated transplants. More impor- tantly, abundant keloid-like collagen fibrils were accumulated in transplants of KPCs in the presence of IL-6 that resembled the characteristic histological features of keloid scars. Using the current KPCs-generated transplant as a keloid model we attempted to treat with IL-6 neutralizing antibody using standard intra-lesional injection approach and showed significant reduction in keloid size. Furthermore, pre-treatment of KPCs with IL-6 neutralizing antibody significantly suppressed the in vivo growth of KPCs-generated transplant, suggesting a potential anti-scarring preventive and therapeutic effect. Taken together, these important findings further support the notion that IL-6, as a major component of the inflammatory niche in keloid scars, may play a critical role in the acquirement of benign tumor-like stem cell phenotypes, therefore the benign tumor growth. IL-17, another recently identified inflammatory cytokine, exclusively produced by activated T cells and whose receptor is widely expressed in many cell types [42], plays a pivotal role in several inflammatory diseases [38–41]. IL-6, in concert with TGF- b, induces TH17 cell differentiation from na¨ıve T cells by Niche-Driven Overgrowth of KPC orchestrating a series of ‘‘downstream’’ cytokine-dependent signaling pathways [67–69]. More importantly, IL-6 has been reported to function both upstream and downstream of IL-17, constituting a positive feedback loop that promotes autoimmune and allergic diseases [50,51]. In this study, we have demonstrated for the first time to our knowledge that IL-17 is up-regulated in keloid tissues, and KPCs express relatively higher IL-17R than normal skin-derived precursor cells. Furthermore, we demonstrat- ed that blocking IL-6 abolished IL-17-induced up-regulation of hTERT and Oct-4 expression; however, blocking IL-17 failed to attenuate IL-6-induced up-regulation of hTERT and Oct-4 expression in KPCs. These findings support our hypothesis that elevated IL-6 and TGF-b in keloids promote differentiation of TH17 cells, which in turn secretes more IL-17 to further enhance the release of IL-6 from keloid stem cells or other stromal cells. The IL-17-triggered positive-feedback loop via paracrine IL-6 induction constitutes the ‘‘pathological’’ niche or altered inflam- matory niche that drives KPCs toward proliferation, and thus tumor-like keloid growth (Figure S6). However, it is important to emphasize that despite the tumor-like growth behavior, KPCs were distinct from cancer cells or cancer stem-like cells (25) in that KPCs displayed a normal karyotype and an absence of cytogenetic abnormalities. Since inflammation plays a critical role in tumorigenesis (23, 24), longer in vivo experimental observation, up to 2 months in our study, was carried out to determine the long-term effects of inflammatory niche components, specifically the IL-17/IL-6 axis, on the acquired properties of KPCs, including their replicative/proliferative capabilities, and cellular transformation. Due to the limitation of the biological activity of the hydrogel-released growth factors, we were not able to prolong the study beyond the above time point. Further studies will be needed to determine the effect of chronic inflammation on potential tumorigenesis. In addition to providing proliferation and differentiation signals, stem cell niche also provides homing signals to recruit bone marrow MSCs to sites of injury and inflammation [10,12], whereby a variety of inflammatory mediators, including hypoxia, reactive oxygen species (ROS), inflammatory chemokines and cytokines, may trigger the migration of BMMSCs [76]. In response to the unique tumor microenvironment of chronic inflammation [77,78], BMMSCs may acquire the cancer-associ- ated fibroblast (CAF) phenotype and subsequently promote tumor growth and metastasis via the production of pro-angiogenic and tumor stimulating paracrine factors [79,80]. Using a dual- chamber cell-migration assay, keloid-derived fibroblasts have been shown to induce human BMMSCs chemoattraction toward keloid cells, and develop myofibroblastic changes characterized by abundant rough endoplasmic reticulum, secretion of collagen-like fibers as well as actin-type microfilament bundles [7]. These findings suggest that the unique keloid microenvironment or niche may preferentially recruit bone marrow MSCs and sustain their interaction with resident fibroblasts, thus contributing to keloid formation by producing abundant extracellular matrix at the scar site. The interaction between the keloid niche and keloid derived stem cells, KPCs, has been demonstrated here in our study, which is sustained by the IL-17/IL-6 feedback loop. It remains to be determined whether IL-17/IL-6 axis contributes to the recruit- ment of BMMSCs to keloid lesion and whether KPCs represent a distinct population of resident MSCs or derived from BMMSCs. In addition, a recent study reported that methylprednisolone is capable of inhibiting IL-17 production in lymphocytes and lymph node cells [81]. Further studies are needed to determine whether the anti-scarring effect of the intralesional steroid therapy in keloid scar correlates with its role on the regulation of the IL-17/IL-6. PLoS ONE | www.plosone.org 13 November 2009 | Volume 4 | Issue 11 | e7798