In experiments using influenza virus, autologous B-LCL were infected overnight, whereafter the B-LCL were irradiated and washed selleck kinase inhibitor extensively. After 4–6 days of culture, the allo-specific proliferation of responder T cells was analyzed by flow cytometry. For measurement of suppression on IL-2 production CFSE-labeled D1.50 was cocultured with the indicated M1-specific T-cell

clone at a 1:1 ratio together with autologous B-LCL in the presence of 50 IU/mL IL-2. The Treg clone was prestimulated with 5 μg/mL cognate peptide. After 24 h D1.50 was stimulated with 5 μg/mL cognate peptide, and 1 h later 10 μg/mL Brefeldin A (Sigma-Aldrich) was added. After overnight incubation, the cells were fixed, permeabilized, and stained for CD4 and intracellular IL-2 as described earlier 39. The percentage of IL-2-producing cells was analyzed by flow cytometry. We would like to thank Klara Broadway for technical assistance. The authors declare no conflict of interest. This study was financially supported by a grant from the Netherlands

Organization for Scientific Research (Zon/Mw 917.56.311 to S.H.v.d.B.). Conflict of interest: The authors declare no financial or commercial conflict of interest. “
“Stress activates the hypothalamic-pituitary-adrenocortical axis to promote the release of corticosterone (CORT), which consequently suppresses pathogenic stimulation of the immune system. Paradoxically, however, stress often promotes autoimmunity through yet unknown mechanisms. Opaganib mw Here we investigated how chronic variable stress (CVS), and the associated alterations in CORT levels, affect the susceptibility to experimental autoimmune encephalomyelitis (EAE) in female and male C57BL/6 mice. Under baseline (nonstressed) conditions, females exhibited substantially higher CORT levels and an attenuated EAE with less mortality

than males. However, CVS induced a significantly worsened EAE in females, which Florfenicol was prevented if CORT signaling was blocked. In addition, females under CVS conditions showed a shift toward proinflammatory Th1/Th17 versus Th2 responses and a decreased proportion of CD4+CD25+ Treg cells. This demonstrates that whereas C57BL/6 female mice generally exhibit higher CORT levels and an attenuated form of EAE than males, they become less responsive to the immunosuppressive effects of CORT under chronic stress and thereby prone to a higher risk of destructive autoimmunity. It has been well established that stress may substantially affect the homeostatic regulation of the immune system [1-3]. In most animal models studied thus far, stressful triggers such as fear, maternal deprivation, social threat, or physiological challenge have been shown to induce immunosuppression associated with increased susceptibility to allergies and infectious diseases [1, 4, 5]. These effects are mediated by the hypothalamic-pituitary-adrenal (HPA) axis, a complex network linking the nervous, endocrine and immune systems [6, 7].