Ana Clara Abadía-Molina

Development of chronic colitis is dependent on the cytokine MIF

The cytokine macrophage-migration inhibitory factor (MIF) is secreted by a number of cell types upon induction by lipopolysaccharide (LPS). Because colitis is dependent on interplay between the mucosal immune system and intestinal bacteria, we investigated the role of MIF in experimental colitis. MIF-deficient mice failed to develop disease, but reconstitution of MIF-deficient mice with wild-type innate immune cells restored colitis. In addition, established colitis could be treated with anti-MIF immunoglobulins. Thus, murine colitis is dependent on continuous MIF production by the innate immune system. Because we found increased plasma MIF concentrations in patients with Crohns disease, these data suggested that MIF is a new target for intervention in Crohns disease.

Cutting Edge: The Natural Ligand for Glucocorticoid-Induced TNF Receptor-Related Protein Abrogates Regulatory T Cell Suppression

CD4+25+ regulatory T (Treg) cells maintain immunological self-tolerance through mechanisms that are only in part understood. Previous studies suggest that the glucocorticoid-induced TNFR-related protein (GITR), which is preferentially expressed on the surface of Treg cells, potentially provides a signal that abrogates Treg suppression. In this study, we show that a soluble form of mouse GITR ligand (sGITR-L) induces GITR-dependent NF-κB activation and blocks in vitro suppression mediated by both resting and preactivated polyclonal and Ag-specific Treg cells. Since sGITR-L along with rIL-2 induces proliferation of CD4+25+ cells, it appears that sGITR-L can break the anergic state of Treg cells. Because sGITR-L also up-regulates IL-2 secretion by activated CD4+25 −T cells, these two sGITR-L induced signals synergize to interfere with suppressor activity by CD4+25+ Treg cells.

Follicular Dendritic Cells Are Related to Bone Marrow Stromal Cell Progenitors and to Myofibroblasts

Follicular dendritic cells (FDC) are involved in the presentation of native Ags to B cells during the secondary immune response. Some authors consider FDC to be hemopoietic cells, whereas others believe them to be mesenchymal cells. The low proportion of FDC in the lymphoid follicle, together with technical difficulties in their isolation, make these cells difficult to study. We show that Fibroblast Medium can be used successfully to isolate and maintain FDC lines. In this culture medium, we obtained 18 FDC lines from human tonsils, which proliferated for as long as 18 wk and showed a stable Ag phenotype as detected by flow cytometry and RT-PCR. FDC lines were CD45-negative and expressed Ags associated to FDC (CD21, CD23, CD35, CD40, CD73, BAFF, ICAM-1, and VCAM-1) and Ags specific for FDC (DRC-1, CNA.42, and HJ2). These cell lines were also able to bind B cells and secrete CXCL13, functional activities characteristic of FDC. Nevertheless, the additional expression of STRO-1, together with CD10, CD13, CD29, CD34, CD63, CD73, CD90, ICAM-1, VCAM-1, HLA-DR, alkaline phosphatase, and α-smooth muscle actin (α-SM actin) indicated that FDC are closely related to bone marrow stromal cell progenitors. The expression of α-SM actin also relates FDC with myofibroblasts. Like myofibroblasts, FDC lines expressed stress fibers containing α-SM actin and were able to contract collagen gels under the effect of TGFβ1 and platelet-derived growth factor. These findings suggest that FDC are a specialized form of myofibroblast and derive from bone marrow stromal cell progenitors.

Immune phenotype and cytotoxic activity of lymphocytes from human term decidua against trophoblast

Flow cytometric data were used to compare the phenotype of term decidual lymphocytes and peripheral blood lymphocytes. Unlike peripheral blood lymphocytes, a significant percentage of CD3+, CD4+, CD8+ and CD16+ term decidual lymphocyte populations expressed the CD69 activation marker. The relative proportions of CD38 in CD3+, CD4+ and CD8+ populations were more than twice as large in term decidual lymphocytes as in peripheral blood lymphocytes. As reported for early decidual lymphocytes, the expression of CD38 and CD69 by term decidual lymphocytes suggests that these cells are also regionally activated. However, term decidual lymphocytes showed no spontaneous cytotoxicity against normal trophoblast or its tumoral counterpart, JEG cells. After stimulation with interleukin-2, these lymphocytes became cytotoxic, as did peripheral blood lymphocytes. The relevance of this latter result to the immune control of the physiological and pathological invasion of the decidua by the trophoblast is discussed.

Effect of flavonoids on rat splenocytes, a structure–activity relationship study

Flavonoids are polyphenols frequently consumed in the diet which have been suggested to exert a number of beneficial actions on human health, including intestinal anti-inflammatory activity. Their properties have been studied in numerous cell types, but little is known about their effect on leukocyte biology. We have selected 9 flavonoids (extended to 14 flavonoids plus the related polyphenol resveratrol in some cases) with different structural features to characterize their effects on leukocyte viability, proliferation, and expression of cyclooxygenase 2 (EC 1.14.99.1), inducible nitric oxide synthase (iNOS, EC 1.14.13.39) and proinflammatory cytokines (TNF-alpha, IFN-gamma, IL-2), as well as to elucidate the structural requirements in each case. Quiescent and concanavalin A-stimulated rat splenocytes were used as a model. Flavonoids (50 microM) had a dramatic inhibitory effect on cytokine secretion. Inducible nitric oxide synthase expression was also blocked largely by some flavonoids, especially quercetin, luteolin and apigenin, while cyclooxygenase 2 was downregulated only by apigenin, diosmetin and quercetin. Apigenin, luteolin, genistein and quercetin had substantial cytotoxic/proapoptotic effects, while chrysin, daidzein, hesperetin and kaempferol did not reduce cell viability. In contrast, all flavonoids had powerful antiproliferative effects. However, none of the compounds activated caspase 3 (EC 3.4.22.56), but actually lowered caspase 3 activation and expression in concanavalin A-stimulated cells. The activity of the quercetin metabolite isorhamnetin was generally lower than that of the parent compound. We conclude that flavonoids have powerful effects on lymphocytes with distinct structural requirements that may contribute to their intestinal anti-inflammatory activity. The bioactivity of orally administered flavonoids may be dampened by biotransformation in vivo, particularly in extraintestinal sites.

Decidual Lymphocytes of Human Spontaneous Abortions Induce Apoptosis but Not Necrosis in JEG-3 Extravillous Trophoblast Cells

Abstract The human decidua contains an unusually high proportion of lymphocytes, mainly NK and T cells, which are potentially cytotoxic to the trophoblast when they are stimulated with certain cytokines. Given the high incidence of spontaneous abortion in humans and other species, our working hypothesis is that decidual lymphocytes are involved in immunological mechanisms that attack the trophoblast and induce abortion when any gestational problem arises. To test this hypothesis, flow cytometry was used to compare decidual lymphocyte populations in first-trimester spontaneous abortions and elective terminations of first-trimester pregnancy. We found significantly higher proportions of decidual lymphocytes that expressed activation markers, and of T cells (mainly T helper cells) in spontaneous abortions than in elective terminations of pregnancy. Decidual lymphocytes from spontaneous abortion, like decidual lymphocytes from elective termination of pregnancy and peripheral blood lymphocytes, were however, unable to lyse the JEG-3 extravillous cytotrophoblast cell line in a 51Cr-release assay. Nevertheless, decidual lymphocytes from spontaneous abortion, unlike decidual lymphocytes from elective termination of pregnancy and peripheral blood lymphocytes, induced apoptosis in JEG-3 cells as determined by DNA fragment-release assay. Hematoxylin and eosin staining showed a significantly higher proportion of apoptotic JEG-3 cells when these cells were treated with decidual lymphocytes from spontaneous abortion than when JEG-3 cells were cultured with decidual lymphocytes from elective termination of pregnancy. The ultrastructural signs of apoptosis were confirmed by electron microscopy. These data support the hypothesis that activated decidual lymphocytes participate in human spontaneous abortion by inducing apoptosis but not necrosis of the trophoblast.

The intestinal antiinflammatory agent glycomacropeptide has immunomodulatory actions on rat splenocytes

Bovine glycomacropeptide (GMP) is an immunologically active milk peptide that is a part of the normal human diet. GMP has therapeutic value in preclinical models of intestinal inflammation, and its mechanism may be related to effects on lymphocytes. This study focuses on the actions of GMP on rat splenocytes in vitro and in vivo. Bovine serum albumin and lactoferrin were used for comparative purposes. GMP (0.01-0.1mgmL(-1)) enhanced Concanavalin A (ConA) evoked but not basal splenocyte proliferation. At 1mgmL(-1) GMP lost this effect but augmented basal TNF-alpha secretion and also iNOS and COX2 expression. IFN-gamma, IL-2 and IL-17 were not affected by GMP in quiescent splenocytes, but IL-10 was augmented at all concentrations tested. On the other hand, GMP produced a marked inhibitory effect (70%) on IFN-gamma secretion and to a lower extent (50%) also on TNF-alpha. GMP was shown to block STAT4 but not IkappaB-alpha phosphorylation. The Treg marker Foxp3 was markedly upregulated by GMP. Bovine serum albumin had some effects on splenocyte function which were of lower magnitude and not entirely coincidental, while lactoferrin had a strong antiproliferative effect, as expected, indicating a specific effect of GMP. When administered for 3 days to normal Wistar rats, GMP reproduced the Foxp3 induction effect observed previously in vitro. This was observed in splenocytes but not in thymocytes, and only when administered by the oral rather than the intraperitoneal route. Thus our results support the hypothesis that GMP may limit intestinal inflammation acting at least in part on lymphocytes.

Antigen phenotype of cultured decidual stromal cells of human term decidua

We previously reported that decidual stromal cells (DSC) from early human decidua express antigens associated with hematopoietic cells and develop different immune functions. Here we study the antigenic phenotype of DSC from term decidua and compare it with the phenotype reported for DSC from early decidua. Decidual stromal cells were isolated from human term deciduas and maintained in culture until highly purified DSC cultures were obtained. Most term DSC, like most early DSC, expressed CD10. Term DSC expressed antigens specific for follicular dendritic cells (FDC), such as DRC-1 (CD21L) and HJ2, together with CD21, CD23 and CD80, which are detected on FDC as well. Also like early DSC, term DSC were negative for CD3, CD14, CD15 and CD45. Although early DSC were reported to be HLA-DR-positive and CD86-positive, these antigens were not expressed by term DSC. These discrepant results suggest that two types of cells, or cells at different stages of differentiation (decidualization) were selected during culture of decidual cells from different periods of gestation. This possibility was further supported by the finding that term DSC expressed desmin and prolactin, two markers of decidualization, whereas these molecules have not previously been detected in early DSC.

Phagocytosis by fresh and cultured human decidual stromal cells: opposite effects of interleukin-1α and progesterone

Flow cytometry and transmission electron microscopy have been employed to show that a proportion of fresh and cultured human decidual stromal cells phagocytose latex particles. Phagocytosis of Escherichia coli by cultured decidual stromal cells was, however, very low. Stimulation of cultured decidual stromal cells with interleukin-1 alpha enhanced phagocytosis of both latex particles and E. coli. In contrast, when decidual stromal cells were cultured with progesterone under decidualizing conditions, phagocytic activity was reduced. These results suggest the existence of an immune-endocrine circuit involving decidual stromal cells.

Reduced proportion of decidual DC-SIGN+ cells in human spontaneous abortion

Recent studies showed that some functions of decidual dendritic cells appear to be essential for pregnancy. In humans, decidual dendritic cells are identifiable by their expression of DC-SIGN. We compared the subpopulations of human decidual DC-SIGN+ cells from first-trimester normal pregnancies and spontaneous abortions by flow cytometry. In normal decidua, DC-SIGN+ cells expressed antigens associated with immature myeloid dendritic cells. In samples from spontaneous abortions, we detected decidual DC-SIGN+ cells with an antigen phenotype equivalent to that of DC-SIGN+ cells from normal pregnancies, but at a significantly lower proportion (P < 0.01). Our results support the hypothesis that dendritic cells play a role in normal or pathological human pregnancy outcomes.