http://hdl.handle.net/2144/2856 2013-05-20T08:31:35Z http://hdl.handle.net/2144/3309 Increased Frequency and Compromised Function of T Regulatory Cells in Systemic Sclerosis (SSc) Is Related to a Diminished CD69 and TGFβ Expression Radstake, Timothy R. D. J.; van Bon, Lenny; Broen, Jasper; Wenink, Mark; Santegoets, Kim; Deng, Yanhui; Hussaini, Anila; Simms, Robert; Cruikshank, William W.; Lafyatis, Robert BACKGROUND. Regulatory T cells (Tregs) are essential in the control of tolerance. Evidence implicates Tregs in human autoimmune conditions. Here we investigated their role in systemic sclerosis (SSc). METHODS/PRINCIPAL FINDINGS. Patients were subdivided as having limited cutaneous SSc (lcSSc, n=20) or diffuse cutaneous SSc (dcSSc, n=48). Further subdivision was made between early dcSSc (n=24) and late dcSSc (n=24) based upon the duration of disease. 26 controls were studied for comparison. CD3+ cells were isolated using FACS and subsequently studied for the expression of CD4, CD8, CD25, FoxP3, CD127, CD62L, GITR, CD69 using flow cytometry. T cell suppression assays were performed using sorted CD4CD25highCD127- and CD4CD25lowCD127high and CD3+ cells. Suppressive function was correlated with CD69 surface expression and TGFβ secretion/expression. The frequency of CD4+CD25+ and CD25highFoxP3highCD127neg T cells was highly increased in all SSc subgroups. Although the expression of CD25 and GITR was comparable between groups, expression of CD62L and CD69 was dramatically lower in SSc patients, which correlated with a diminished suppressive function. Co-incubation of Tregs from healthy donors with plasma from SSc patients fully abrogated suppressive activity. Activation of Tregs from healthy donors or SSc patients with PHA significantly up regulated CD69 expression that could be inhibited by SSc plasma. CONCLUSIONS/SIGNIFICANCE. These results indicate that soluble factors in SSc plasma inhibit Treg function specifically that is associated with altered Treg CD69 and TGFβ expression. These data suggest that a defective Treg function may underlie the immune dysfunction in systemic sclerosis. 2009-06-22T00:00:00Z http://hdl.handle.net/2144/3308 The Pronounced Th17 Profile in Systemic Sclerosis (SSc) Together with Intracellular Expression of TGFβ and IFNϒ Distinguishes SSc Phenotypes Radstake, Timothy R. D. J.; van Bon, Lenny; Broen, Jasper; Hussiani, Anila; Hesselstrand, Roger; Wuttge, Dirk M.; Deng, Yanhui; Simms, Robbert; Lubberts, Erik; Lafyatis, Robert BACKGROUND. Systemic sclerosis (SSc) is an autoimmune disease where controversy on Th1/Th2 balance dominates. We investigated whether the recently discovered Th17 pattern was present in SSc. METHODOLOGY AND PRINCIPAL FINDINGS. Patients were subdivided as having limited cutaneous SSc (lcSSc, n=12) or diffuse cutaneous SSc (dcSSc, n=24). A further arbitrary subdivision was made between early dcSSc (n=11) and late dcSSc (n=13) based upon the duration of disease. As a comparator group 14 healthy controls were studied. CD3+ cells were isolated using FACS and subsequently studied for the expression of CD4, CD8, CD25, CD45Ro, CD45Ra, IL-23, GITR, CD69 and intracellular expression of IL-17, TGFβ and IFNϒ using flow cytometry. Levels of IL-17, IL-6, IL-1a and IL-23 were measured using Bioplex assays. SSc patients had more and more activated CD4+ cells. In addition, CD4, CD45Ro and CD45Ra cells from all SSc patients highly expressed the IL23R, which was associated with a higher IL-17 expression as well. In contrast, IFNϒ and TGFβ were selectively up regulated in SSc subsets. In line with these observation, circulating levels of IL-17 inducing cytokines IL-6, IL-23 and IL-1a were increased in all or subsets of SSc patients. CONCLUSION AND SIGNIFICANCE. The combination of IL-17, IFNϒ and TGFβ levels in CD45Ro and CD45Ra cells from SSc patients is useful to distinguish between lSSc, ldSSc or edSSc. Blocking Th17 inducing cytokines such as IL-6 and IL-23 may provide a useful tool to intervene in the progression of SSc. 2009-06-17T00:00:00Z