Receptors for prostaglandin E(2) that regulate cellular immune responses in the mouse.

Production of prostaglandin E(2) (PGE(2)) is enhanced during inflammation, and this lipid mediator can dramatically modulate immune responses. There are four receptors for PGE(2) (EP1-EP4) with unique patterns of expression and different coupling to intracellular signaling pathways. To identify the EP receptors that regulate cellular immune responses, we used mouse lines in which the genes encoding each of the four EP receptors were disrupted by gene targeting. Using the mixed lymphocyte response (MLR) as a model cellular immune response, we confirmed that PGE(2) has potent antiproliferative effects on wild-type responder cells. The absence of either the EP1 or EP3 receptors did not alter the inhibitory response to PGE(2) in the MLR. In contrast, when responder cells lacked the EP2 receptor, PGE(2) had little effect on proliferation. Modest resistance to PGE(2) was also observed in EP4-/- responder cells. Reconstitution experiments suggest that EP2 receptors primarily inhibit the MLR through direct actions on T cells. Furthermore, PGE(2) modulates macrophage function by activating the EP4 receptor and thereby inhibiting cytokine release. Thus, PGE(2) regulates cellular immune responses through distinct EP receptors on different immune cell populations: EP2 receptors directly inhibit T cell proliferation while EP2 and EP4 receptors regulate antigen presenting cells functions.

Adenosine and inosine increase cutaneous vasopermeability by activating A(3) receptors on mast cells.

Adenosine has potent effects on both the cardiovascular and immune systems. Exposure of tissues to adenosine results in increased vascular permeability and extravasation of serum proteins. The mechanism by which adenosine brings about these physiological changes is poorly defined. Using mice deficient in the A(3) adenosine receptor (A(3)AR), we show that increases in cutaneous vascular permeability observed after treatment with adenosine or its principal metabolite inosine are mediated through the A(3)AR. Adenosine fails to increase vascular permeability in mast cell-deficient mice, suggesting that this tissue response to adenosine is mast cell-dependent. Furthermore, this response is independent of activation of the high-affinity IgE receptor (FcepsilonR1) by antigen, as adenosine is equally effective in mediating these changes in FcepsilonR1 beta-chain-deficient mice. Together these results support a model in which adenosine and inosine induce changes in vascular permeability indirectly by activating mast cells, which in turn release vasoactive substances. The demonstration in vivo that adenosine, acting through a specific receptor, can provoke degranulation of this important tissue-based effector cell, independent of antigen activation of the high-affinity IgE receptor, supports an important role for this nucleoside in modifying the inflammatory response.

Disruption of the A(3) adenosine receptor gene in mice and its effect on stimulated inflammatory cells.

The A(3) adenosine receptor (A3AR) is one of four receptor subtypes for adenosine and is expressed in a broad spectrum of tissues. In order to study the function of A3AR, a mouse line carrying a mutant A(3) allele was generated. Mice homozygous for targeted disruption of the A3AR gene, A3AR(-/-), are fertile and visually and histologically indistinguishable from wild type mice. The lack of a functional receptor in the A3AR(-/-) mice was confirmed by molecular and pharmacological analyses. The absence of A3AR protein expression in the A3AR(-/-) mice was demonstrated by lack of N(6)-(4-amino-3-[(125)I]iodobenzyl)adenosine binding to bone marrow-derived mast cell membranes that were found to express high levels of A3AR in wild type mice. In A3AR(-/-) mice, the density of A(1) and A(2A) adenosine receptor subtypes was the same as in A3AR(+/+) mice as determined by radioligand binding to brain membranes. Additionally, A(2B) receptor transcript expression was not affected by ablation of the A3AR gene. A3AR(-/-) mice have basal heart rates and arterial blood pressures indistinguishable from A3AR(+/+) mice. Functionally, in contrast to wild type mice, adenosine and the A3AR-specific agonist, 2-chloro-N(6)-(3-iodobenzyl)-adenosine-5'-N-methyl-carboxamide (2-Cl-IB-MECA), elicit no potentiation of antigen-dependent degranulation of bone marrow-derived mast cells from A3AR(-/-) mice as measured by hexosaminidase release. Also, the ability of 2Cl-IB-MECA to inhibit lipopolysaccharide-induced tumor necrosis factor-alpha production in vivo was decreased in A3AR(-/-) mice in comparison to A3AR(+/+) mice. The A(2A) adenosine receptor agonist, 2-p-(2-carboxyethyl)phenylamino)-5'-N-ethylcarboxamidoadenosine, produced inhibition of lipopolysaccharide-stimulated tumor necrosis factor-alpha production in both A3AR(-/-) and A3AR(+/+) mice. These results show that the inhibition in vivo can be mediated by multiple subtypes, specifically the A(3) and A(2A) adenosine receptors, and A3AR activation plays an important role in both pro- and anti-inflammatory responses.