Rapid and transient reduction in circulating thyroid hormones following systemic antigen priming: implications for functional collaboration between dendritic cells and thyroid.

The thyroid hormones T(3) (tri-iodothyronine) and T(4) (thyroxine) are disseminated throughout the body via the circulation and are maintained across a range of physiological concentrations under the control of thyroid-stimulating hormone (TSH). T(3) (and T(4) after conversion to T(3)) influences many biological activities, including gene expression and protein synthesis, though little is known about the nature of pituitary-thyroid immune interactions. In the present study we show that serum T(3) and T(4) levels are sharply but transiently reduced during the first 24 h of systemic antigen exposure and that this is followed by suppressed levels of free T(4), after which there is rapid recovery to normal levels. Splenic dendritic cells, depending upon the stage of maturation/activation, were found to be a rich source of TSH, and CD11c(+) cells with dendritic cell morphology were present in the thyroid 1-3 days after antigen exposure. Moreover, antigen priming of hypophysectomized mice that are unable to make pituitary-derived TSH resulted in significant increases in circulating T(4), implying that compensation in the drop in thyroid hormones can be regulated from extrapituitary sources. These findings thus identify a novel set of immune-endocrine interactions that transpire during the early phase of antigen exposure, and they suggest that under appropriate conditions the immune system directly participates in the process of maintaining physiological homeostasis by contributing to the regulatory control of thyroid hormone activity.

CD43 potentiates CD3-induced proliferation of murine intestinal intraepithelial lymphocytes.

The involvement of CD43 in cell proliferation of murine intestinal intraepithelial lymphocytes (IEL) has been studied in in vitro CD3-stimulated cell cultures. In the presence of either IL-2 or IL-15, CD3 stimulation of IEL resulted in low levels of proliferation as measured by thymidine incorporation, whereas no proliferation occurred upon CD3 stimulation in the absence of cytokines. The combination of both cytokines to IEL cultures synergistically enhanced CD3-induced proliferation by approximately threefold that of cultures supplemented with either cytokine alone. Most importantly, however, proliferation of IEL was significantly greater when CD3 stimulation occurred in conjunction with CD43 triggering, indicating that CD43 functions as a coactivational signal for murine IEL. These findings indicate that a spectrum of potential proliferative responses exist among murine IEL depending on the types and combinations of signals received, and that because under normal conditions murine IEL are largely devoid of CD28 expression, a classical T-cell coactivational molecule, the capacity for high-level IEL proliferation may reside with CD43.

The thyrotropin (thyroid-stimulating hormone) receptor is expressed on murine dendritic cells and on a subset of CD45RBhigh lymph node T cells: functional role for thyroid-stimulating hormone during immune activation.

Thyroid-stimulating hormone (TSH), a central neuroendocrine mediator of the hypothalamus-pituitary-thyroid axis, has been shown to affect various aspects of immunological development and function. To gain a better understanding of TSH involvement within the mammalian immune system, the expression and distribution of the TSH receptor (TSHr) has been studied by immunoprecipitation and by flow cytometric analyses. Using highly enriched populations of B cells, T cells, and dendritic cells, trace amounts of TSHr were precipitated from B cells and T cells, whereas high levels of TSHr were precipitated from the dendritic cell fraction. Flow cytometric analyses of TSHr expression on splenic and lymph node T cells revealed a major difference between those tissues in that only 2-3% of splenic T cells were TSHr+, whereas 10-20% of CD4+8- and CD4-8+ lymph node T cells expressed the TSHr, which was exclusively associated with CD45RB(high) cells and was not expressed during or after activation. The TSHr was not present on cells of the immune system during fetal or neonatal life. However, recombinant TSHbeta was found to significantly enhance the phagocytic activity of dendritic cells from adult animals and to selectively augment IL-1beta and IL-12 cytokine responses of dendritic cells following phagocytic activation. These findings identify a novel immune-endocrine bridge associated with professional APCs and naive T cells.

Origins of intestinal intraepithelial lymphocytes: direct evidence for a thymus-derived gamma delta T cell component.

The contribution of T cell precursors from the thymus and the bone marrow to the pool of intestinal intraepithelial lymphocvtes (IELs) has been studied in a system using donor cells from enhanced-green fluorescent protein (EGFP+ ) transgenic mice a doptively transferred into EGFP- recipient mice. Consistent with previous studies, regeneration of gamma delta and alpha beta T cell populations in the intestinal epithelium occurred within 2-3 weeks of bone marrow transfer into irradiatiated EGFP- animals and prior to T cell repopulation of the spleen, of interest, however, although transfer of whole adult EGFP+ thymocytes to non-irradiated EGFP- congenitally-athymic nude mice produced alpha beta T cells in both the spleen and intestine. Gamma delta T cells in significant number were detected only in the intestine of recipient mice. In contrast, transfer of CD3-, CD4-, CD8- immature thymocytes resulted in no detectable T cells in either the intestine or the spleen of nude mice up to twelve weeks post-cell transfer, suggesting that intestinal IELs generated from thymocytes arose from differentiated lineage-committed cells rather than from immature thymocytes. These findings provide direct evidence for both thymus-independent and thymus-dependent sources of intestinal gamma delta T cells, and they suggest that murine IELs consist of diverse groups of T cells with distinct developmental origins.

Differential expression of CD43 isoforms on murine T cells and their relationship to acute intestinal graft versus host disease: studies using enhanced-green fluorescent protein transgenic mice.

Three mAb (R2/60, S7 and 1B11) were used to study the expression of murine CD43 on peripheral T cells and intestinal intraepithelial lymphocytes (IEL) from normal mice, and from mice during acute graft versus host disease (GVHD). In the spleen, essentially all T cells expressed the R2/60 and S7 antigens, whereas the 1B11 antigen was expressed on about half of the CD8(+) cells and approximately 15% of CD4(+) T cells. Interestingly, a significant proportion of resting splenic B cells expressed the 1B11 and R2/60 antigens, but not the S7 antigen. The majority of IEL expressed R2/60 antigen; however, the S7 and 1B11 markers were differentially expressed on CD8alpha, CD8beta, TCRalphabeta and TCRgammadelta cells. Immunoprecipitation and Western blotting analyses identified characteristic 115 and 130 kDa reactive components from IEL lysates with mAb S7 and 1B11 respectively, and reactivity to both molecular entities by mAb R2/60. During acute intestinal GVHD induced by injecting CB6F(1) athymic nude mice with spleen cells from C57BL/6 enhanced-green fluorescent protein transgenic mice, 80-90% of donor T cells in the intestine epithelium expressed all CD43 isoforms; however, the level of expression of the 130 kDa CD43 antigen increased significantly and the level of the 115 kDa antigen decreased on GVHD donor T cells compared to cells at the time of transfer. Using EL4 cells, a similar shift in the expression of CD43 isoforms occurred experimentally following treatment with neuraminidase, suggesting that the type of CD43 isoform expressed on T cells is strongly influenced by conditions which affect membrane charge. The significance of these findings for intestinal immunopathology is discussed.

Neuroendocrine-induced synthesis of bone marrow-derived cytokines with inflammatory immunomodulating properties.

Although cytokines and other soluble regulators of immunity are known to be involved in hematopoiesis, little is known about the signals that induce the synthesis of those mediators locally. Based on recent studies linking the neuroendocrine hormone thyrotropin [thyroid-stimulating hormone (TSH)] to immune cell function in other tissues, we investigated the capacity of TSH to activate cytokine responses from bone marrow cells. These studies reveal that stimulation of the TSH receptor on bone marrow cells-using highly purified or recombinant TSH or by direct stimulation with anti-TSH receptor antibodies-rapidly induces the synthesis of cytokines from bone marrow cells that are classically used in the regulation of inflammatory responses. Of 13 cytokines screened for activity by ELISA or by RNase protection assays for gene expression, IL-6, IFN-beta, TNFalpha, TNFbeta, TGFbeta2, and lymphotoxin-beta responses were reproducibly induced by TSH within 2-3 h of stimulation. Intracellularly, TSH stimulation of bone marrow cells caused rapid increases in cAMP levels and induced the phosphorylation of the Jak2 protein kinase, thereby defining a novel G-protein-coupled receptor/cytokine synthesis pathway. These findings demonstrate that TSH can serve as a primary inductive signal of cytokine production by bone marrow cells.

Cell surface sialic acid and the regulation of immune cell interactions: the neuraminidase effect reconsidered.

It has been known for over a decade that sialidase (neuraminidase) treatment could substantially enhance the capacity of resting B cells to stimulate the proliferation of allogeneic and antigen specific, syngeneic T cells. Thus, cell-surface sialic acid was implicated as a potential modulator of immune cell interaction. However, little progress has been made in either identifying explicit roles for sialic acid in this system or in hypothesizing mechanisms to explain the "neuraminidase effect." Here we show for the first time that cell surface sialic acid on medium incubated B cells blocks access to costimulatory molecules on the B cell surface, and that this is the most likely explanation for the neuraminidase effect. Further, we show that it is likely to be upregulation of ICAM-1 and its subsequent engagement of LFA-1 rather than loss of cell surface sialic acid that in part regulates access to CD86 and other costimulatory molecules. However, we cannot exclude a role for CD86-bound sialic acid on the B cell in modulating binding to T cell CD28. Because sialidase treatment of resting B cells but not resting T cells enables T cell activation, we suggest that sialidase treatment may still be an analogue for an authentic step in B cell activation, and show that for highly activated B cells (activated with polyclonal anti-IgM plus INF-gamma) there is specific loss 2, 6-linked sialic acid. Potential roles for sialic acid in modulating B cell/T cell collaboration are discussed.

Glycosylation of native MHC class Ia molecules is required for recognition by allogeneic cytotoxic T lymphocytes.

The question whether or not target cell N-linked carbohydrate participates in cytotoxic T lymphocyte (CTL) mediated cytolysis has been in contention for well over a decade. Much of the evidence supporting a role for N-linked carbohydrate stems from the treatment of target cells with the glycosylation inhibitor tunicamycin (TM). In this report, we show that treatment of targets with TM, but not other inhibitors of complex carbohydrate formation (swainsonine, 1-deoxynojirimycin, and 1-deoxymannojirimycin), abrogates the lytic response. We conclude that N-linked complex carbohydrates are not absolutely required for the maturation or function of any target cell glycoconjugate directly involved in the lytic process. However, we show that TM treatment of target cells leads to the loss of an MHC class Ia epitope known to be required for T cell triggering. Further, we show that only target cells expressing class Ia molecules with functional glycosylation sequons are subject to the TM effect. From these results we conclude that while N-linked glycosylation of native MHC class Ia molecules is required for antigen presentation, the exact structural motif of the glycan is not important.