sq-23377 and Colitis

Topics: Animals; Calcium; Cell Culture Techniques; Colitis; Culture Media; Cytokines; Humans; Ionomycin; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Mice, Knockout; T-Lymphocytes, Helper-Inducer

The active form of vitamin D, 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] is synthesized by the 1α-hydroxylase, which is encoded by the Cyp27B1 gene. Using transgenic mice that have replaced the Cyp27B1 gene with the bacterial lacZ reporter gene (β-galactosidase), the inflammatory conditions that induce Cyp27B1 in the immune system were probed. A variety of stimuli including lipopolysaccharide, anti-CD3 or PMA/ionomycin were used to stimulate splenocytes and bone marrow derived macrophage in vitro. Only anti-CD3 stimulation resulted in a low induction of β-galactosidase activity in the spleen, indicating that T cells might be a source of Cyp27B1. In vivo, challenge with lipopolysaccharide, α-galactosylceramide, and Listeria monocytogenes failed to induce β-galactosidase activity outside of the kidneys. During more prolonged and severe inflammation there was staining in both the lungs and the gastrointestinal tract for β-galactosidase. Furthermore, wild-type reconstitution of the hematopoietic cell population in Cyp27B1 KO mice protected the mice from experimental colitis. T cell production of Cyp27B1 activity was shown to be from the CD8+ but not the CD4+ T cell population. CD8+ T cells expressed the reporter gene only after 48 h of stimulation. The data is consistent with a model where CD8+ T cells are activated to produce Cyp27B1 and 1,25(OH)2D3 that serves to turn off the local immune response.

Topics: 25-Hydroxyvitamin D3 1-alpha-Hydroxylase; Animals; beta-Galactosidase; Bone Marrow Cells; CD8-Positive T-Lymphocytes; Colitis; Galactosylceramides; Genes, Reporter; Intestinal Mucosa; Intestines; Ionomycin; Lipopolysaccharides; Listeria monocytogenes; Macrophages; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Promoter Regions, Genetic; Spleen

Induced regulatory T (iTreg) lymphocytes show promise for application in the treatment of allergic, autoimmune and inflammatory disorders. iTreg cells demonstrate advantages over natural Treg (nTreg) cells in terms of increased number of starting population and greater potential to proliferate. Different activation methods to generate iTreg cells result in iTreg cells that are heterogeneous in phenotype and mechanisms of suppression. Therefore it is of interest to explore new techniques to generate iTreg cells and to determine their physiological relevance.. Using phorbol myristate acetate (PMA)/ionomycin and anti-CD3 activation of CD4⁺CD25⁻ cells we generated in vitro functional CD4⁺CD25⁻ iTreg (TregPMA) cells. Functionality of the generated TregPMA cells was tested in vivo in a mouse model of inflammatory bowel disease (IBD) - CD45RB transfer colitis model.. TregPMA cells expressed regulatory markers and proved to ameliorate the disease phenotype in murine CD45RB transfer colitis model. The body weight loss and disease activity scores for TregPMA treated mice were reduced when compared to diseased control group. Histological assessment of colon sections confirmed amelioration of the disease phenotype. Additionally, cytokine analysis showed decreased levels of proinflammatory colonic and plasma IL-6, colonic IL-1 β and higher levels of colonic IL-17 when compared to diseased control group.. This study identifies a new method to generate in vitro iTreg cells (TregPMA cells) which physiological efficacy has been demonstrated in vivo.

Topics: Analysis of Variance; Animals; Body Weight; Calcium Ionophores; CD3 Complex; CD4 Antigens; Colitis; Cytokines; Interleukin-2 Receptor alpha Subunit; Ionomycin; Leukocyte Common Antigens; Lymphocyte Activation; Mice; Mice, Inbred BALB C; T-Lymphocytes, Regulatory; Tetradecanoylphorbol Acetate

The authors have previously reported that homologous immunoglobulin (Ig)G reduces the occurrence of dextran sulfate sodium (DSS)-induced colitis, mainly by suppressing recruitment of immunocompetent cells into colitis lesions. However, the mechanisms of cell recruitment and of its suppression by IgG remain unclear. In addressing these questions, this study focused on the activation of T cells in the presence of macrophages. The authors found that [3H]-thymidine uptake of T cells from DSS-induced colitis mice, but not from normal mice, was significantly enhanced when cultured with DSS-pulsed macrophages. From the profile of cytokine production, it was suggested that T helper 1 (Th1)-type cells become predominant during stimulation. Addition of homologous IgG significantly suppressed T cell proliferation in a dose-dependent manner, while no suppressive effect was observed with heterologous IgG. Mouse IgG F(ab')2, but not Fc, fragments partially mimicked the suppressive effect of whole IgG. These findings provide evidence that Th1-type cells may play an important role in the development of DSS-induced colitis and that homologous IgG exerts its protective action at least in part through the F(ab')2 portion.

Topics: Animals; Antibodies, Monoclonal; CD3 Complex; Cell Division; Colitis; Cytokines; Dextran Sulfate; Disease Models, Animal; Female; Humans; Immunoglobulin G; Ionomycin; Isoantigens; Lymphocyte Activation; Macrophages; Mice; Mice, Inbred BALB C; Mice, Inbred CBA; Mitogens; T-Lymphocytes; Tetradecanoylphorbol Acetate