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Introduction Altered phenotypes of circulating monocytes of patients with systemic sclerosis (SSc) have been reported, but the role of these alterations in the pathogenesis of SSc remains unclear. This study was undertaken to identify molecules that are preferentially expressed by SSc monocytes, and to investigate the roles of these molecules in the pathogenic process of SSc. Methods We analyzed circulating CD14+ monocytes isolated from 36 patients with SSc and 32 healthy control subjects. The monocytes' gene expression profiles were assessed by Oligo GEArray® (SABiosciences, Frederic, MA, USA) and semiquantitative or quantitative PCR; their protein expression was evaluated in culture supernatants of unstimulated monocytes by immunoblotting or ELISA, and by immunocytostaining. Monocyte chemoattractant activity of CCL2 was assessed in a TransWell® system (Corning Incorporated, Corning, NY, USA) in the presence or absence of chondroitin sulfate (CS). Results A step-wise approach to profiling gene expression identified that versican and CCL2 were upregulated in SSc monocytes. Subsequent analysis of proteins expressed in monocyte culture supernatants confirmed enhanced production of versican and CCL2 in SSc monocytes compared with control monocytes. CCL2 bound to CS chains of versican and colocalized with versican in the monocytes' Golgi apparatus. Finally, CCL2 had a greater ability to mediate monocyte migration when bound to CS chains, because this binding provided efficient formation of CCL2 gradients and protection from protease attack. Conclusion Circulating monocytes with elevated versican and CCL2 levels may contribute to the fibrotic process in a subset of SSc patients by amplifying a positive feedback loop consisting of versican, CCL2, and the influx of monocytes.

Introduction Behçet's disease (BD) is a multisystem inflammatory disorder, in which a T-helper 1 (Th1)-polarized immune response plays a major role in the pathogenic process. We evaluated the regulatory role of natural killer (NK) cells in Th1-biased immune responses in patients with BD. Methods We studied 47 patients with BD, including 10 with active disease (aBD) and 37 with inactive disease (iBD), and 29 healthy controls. The activation status and cytotoxic activity of NK cells were examined by flow cytometry. The levels of mRNAs for immune modulatory and cytotoxic molecules in NK cells were determined by quantitative PCR. The IL-12 signal strength in NK cells was determined by assessing the phosphorylation state of its downstream component, signal transducer and activator of transduction 4, by immunoblotting. Finally, NK cells' ability to modulate the Th1 response was evaluated by co-culturing NK cells and T cells without cell contact. Results CD69+-activated NK cells were significantly increased in aBD compared with iBD or control samples, although their cytotoxic activities were similar. The iBD NK cells showed downregulated IL-12 receptor β2 mRNA levels compared with aBD or control NK cells. The increased IL-13 expression was detected in a subset of BD patients: most of them had iBD. The IL-13 expression level in iBD patients was significantly higher than the level in controls, but was not statistically different compared with the level in aBD patients. The gene expression profile in iBD patients was consistent with the NK type 2 phenotype, and the shift to NK type 2 was associated with disease remission. NK cells from iBD patients showed impaired IL-12-induced signal transducer and activator of transduction 4 phosphorylation. Finally, iBD, but not control, NK cells suppressed IFNγ expression by aBD-derived CD4+ T cells in vitro. Conclusions NK cells may control disease flare/remission in BD patients via NK type 2-mediated modulation of the Th1 response.

Introduction Microvasculopathy is one of the characteristic features in patients with systemic sclerosis (SSc), but underlying mechanisms still remain uncertain. In this study, we evaluated the potential involvement of monocytic endothelial progenitor cells (EPCs) in pathogenic processes of SSc vasculopathy, by determining their number and contribution to blood vessel formation through angiogenesis and vasculogenesis. Methods Monocytic EPCs were enriched and enumerated using a culture of peripheral blood mononuclear cells and platelets on fibronectin in 23 patients with SSc, 22 patients with rheumatoid arthritis (RA), and 21 healthy controls. To assess the capacity of monocytic EPCs to promote vascular formation and the contribution of vasculogenesis to this process, we used an in vitro co-culture system with human umbilical vein endothelial cells (HUVECs) on Matrigel® and an in vivo murine tumor neovascularization model. Results Monocytic EPCs were significantly increased in SSc patients than in RA patients or healthy controls (P = 0.01 for both comparisons). Monocytic EPCs derived from SSc patients promoted tubular formation in Matrigel® cultures more than those from healthy controls (P = 0.007). Transplantation of monocytic EPCs into immunodeficient mice resulted in promotion of tumor growth and blood vessel formation, and these properties were more prominent in SSc than healthy monocytic EPCs (P = 0.03 for both comparisons). In contrast, incorporation of SSc monocytic EPCs into the tubular structure was less efficient in vitro and in vivo, compared with healthy monocytic EPCs. Conclusions SSc patients have high numbers of aberrant circulating monocytic EPCs that exert enhanced angiogenesis but are impaired in vasculogenesis. However, these cells apparently cannot overcome the anti-angiogenic environment that characterizes SSc-affected tissues.