ImmunoMethods最新文献

Most tumor-host interactions occur through the cytokines released by tumor cells and/or tumor-infiltrating leukocytes. Local exogenous administration of cytokines (especially by gene transfer) may result in tumor inhibition. However, cytokine gene transfer as an approach to curing cancer is not without many biases, which are discussed here. The methodology focuses on the detection of cytokines in situations in which few cells are available or ex vivo tissue must be examined, as occurs in studies of tumor-host interactions and when such interactions are modulated by immunological interventions.

Prostaglandin receptors in many tissues have been described and the receptor for prostaglandin E₂ (PGE-R) has been studied extensively. In most tissues this receptor has a high affinity (K_d = nM); however, the capacity varies widely in different tissues. Recent successful cloning of one PGE-R should soon elucidate the structure of the receptor. In many tissues, PGE-R are coupled to adenyl cyclase to stimulate cAMP formation; however, PGE-R that inhibit cAMP, as well as PGE-R that are coupled to phosphoinositide metabolism, have been described. Despite this complexity, several schemes of classifying PGE-R have been proposed. Our studies in a murine mammary tumor system provide evidence for a functional role of the PGE-R in tumor metastasis. Using several PGE-R antagonists, we have shown that pharmacologic inhibition of [³H]PGE₂ binding and PGE₂-mediated cAMP elevation are associated with enhanced mammary tumor cell metastasis. These receptor antagonists also inhibit natural killer cell-mediated conjugation with and lysis of susceptible target cells. In addition, tumor cell adhesion to laminin is inhibited. Thus, we have proposed that the PGE-R may interact directly or indirectly with adhesive cell functions critical to the metastatic process.

The data presented in this work illustrate that seminal plasma components inhibit the function of both NK cell and T lymphocytes and that the E series prostaglandins are responsible for a major proportion of this suppression. Fractions rich in PGE and 19-OH PGE suppressed cell function, extraction of the PGEs removed most of the immunosuppressive activity, and the immunosuppressive properties of purified E series prostaglandin closely resembled those of whole seminal plasma. To obtain further insight into the immunosuppressive properties of the E series prostaglandins and their relationship to molecular structure, we have also compared the immunosuppressive properties in vitro of PGE₁, PGE₂, and 19-OH PGE and a number of analogues, including the 5-trans, 11β, 15R, and 8-iso variants. The immunosuppressive effects were evaluated in three immunological systems known to be modulated by prostaglandins: mitogen-induced lymphocyte proliferation, IL2 and transferrin receptor expression, and NK cell-mediated cytotoxicity. These studies revealed that PGE₁ and PGE₂ exerted a greater immunosuppressive effect than 19-OH PGE. Furthermore, the studies with analogues indicated that subtle changes in the structure of the PGE molecule resulted in the substantial loss of immunosuppressive activity. We also show that seminal prostaglandins can induce apoptosis in thymocytes and compared the abilities of PGEs and their analogues to induce this phenomenon. Our data go some way in explaining the unusually high concentration of prostaglandins in seminal plasma. We discuss the relevance of our data to understanding the etiology of AIDS and other sexually transmitted diseases, infertility, and malignancies of the urogenital tract.

The role of mast cell activation and the generation of mediator release are important factors in determining the reactivity of lung tissue during allergic reactions and asthma. In this article, the advantages of our procedure for isolating mast cells from lung tissue over other methods are discussed. Our studies have demonstrated the importance of cross talk between mast cells and other cell types that also release mediators of bronchoconstriction during activation, thus amplifying the response. In addition, it has been found that inositol phospholipid turnover in response to various mast cell mediators is enhanced in hyperresponsive lung tissue. The consequences to the contractility of the tissue of such alterations in the cellular signaling system in lung tissue are discussed.

Prostaglandins are potent mediators of inflammatory and immune responses in human skin and are important effector molecules in the cellular responses to cytokines, especially interleukin-1. In vitro studies have demonstrated that prostaglandins are synthesized and released from cellular components of the skin immune system, including keratinocytes, antigen-presenting cells such as Langerhans cells, monocytes/macrophages, and dendritic cells, T cells, mast cells, and the microvascular endothelial cells, as well as from dermal fibroblasts. Interleukin-1 is a prime initiator of cutaneous inflammatory response to nonspecific exogenous stimuli such as ultraviolet irradiation, mechanical trauma, and contact allergens. Interleukin-1 induces release of prostaglandin E₂ in a time- and concentration-dependent manner from human epidermal keratinocytes, dermal fibroblasts, and microvascular endothelial cells. Furthermore, preincubation with Interleukin-1 "primes" skin cells and potentiates prostaglandin release induced by classical pro-inflammatory agonists such as histamine and bradykinin. These prostaglandins down-regulate cellular responses to and synthesis of cytokines by stimulating a transient increase in intracellular cyclic AMP levels. Comparative pharmacological studies of prostaglandin synthesis induced by cytokines and classical inflammatory molecules such as histamine have established the existence of multiple rate-limiting reactions. Recently, a second isoform of the prostaglandin endoperoxide synthase (EC 1.14.99.1) has been identified and shown to be a glucocorticoid-sensitive transcriptionally regulated form of the enzyme. The existence of different isoforms of this enzyme allows reinterpretation of the results of earlier pharmacological experiments and also reemphasizes the importance of prostaglandins as autocrine and paracrine regulators of inflammatory and immune cellular responses.

The widely held view that prostaglandin E₂ (PGE₂) only down-regulates immune function has been challenged by recent data from this and other laboratories. In the case of B lymphocytes, PGE₂ has potent regulatory effects on activation and immunoglobulin class switching. Furthermore, the maturation stage at which a B cell encounters PGE₂ can determine whether the lymphocyte continues along its differentiative path or is terminated by apoptosis. Our studies have combined the use of murine B lymphomas and normal B lymphocytes to unravel the mechanisms by which PGE₂ affects B cells. Here we describe the strategies and approaches that have led to a new understanding of PGE₂ regulation of immunity. These models may be extended to the study of other prostanoids and to human B lymphocytes.

Eicosanoids, both prostaglandins and leukotrienes, have been implicated as mediators in a number of physiological processes. Various tissues have been found to produce different types and quantities of eicosanoids. Tissues also differ in their eicosanoid profiles when the eicosanoids are produced under different conditions. The total amount of prostaglandins formed in response to cellular stimuli depends upon the release of arachidonic acid and its metabolism by PGH synthase (cyclooxygenase). Currently available assays for PGH synthase activity are too expensive, cumbersome, and insensitive to be used in screening a large number of samples for enzyme activity. The most sensitive assays available for prostaglandin detection are radioimmunoassays for specific prostaglandins. We discuss in this article the development of radioimmunoassays using monoclonal antibodies, both specific and pan-specific, for recognition of the prostaglandins.

Endogenous mediators, such as the arachidonic acid metabolite PGE₂, can affect cytokine production by T cells and other cell types. Further, the effects of PGE₂ on cytokine production differ for the Th1 and Th2 T-cell subsets. Production of the genetically and functionally related cytokines TNF and lymphotoxin by Th1 T-cell clones stimulated with immobilized anti-CD3 or Con A was inhibited by PGE₂. Production of TNF by the Th2 T-cell clone D1OG.4 also was inhibited by PGE₂; however, the extent of inhibition was less than that observed for several Th1 clones. The partial inhibitory effect of PGE₂ on TNF production by D1OG.4 cells was observed by assessing bioactivity using the WEHI-164 fibrosarcoma cell line, and the results were confirmed by Northern blot analyses which showed that the accumulation of TNF mRNA also was partially reduced. These results were consistently observed even though the D10G.4 cells produced, in general, less TNF than the Th1 clones studied. Dose-dependent increases in intracellular cyclic AMP levels, which were maximal by 3 min, were observed when the Th1 T-cell clone F1.28 was stimulated with PGE₂. The description of prostaglandin receptor subtypes on many different cells types and the high-affinity binding of PGE₂ to T cells suggest that receptor subtypes for PGE₂ also may be present on T-cell subsets. Further, different intracellular signaling mechanisms are initiated after the interaction of PGE₂ with each receptor subtype. The inhibitory effect of PGE₂ on TNF/LT production by F1.28 cells was observed only with PGE₂ and PGE₁, which bind to the EP1, EP2, and EP3 receptors. In contrast, no effect was evident with the EP1- and EP2-selective agonists 17-phenyl-PGE₂ and 11-deoxy-PGE₁, respectively. These data suggest that more than one intracellular signaling pathway may mediate the inhibitory effect of PGE₂ on TNF production.