A2A receptor dependent and A2A receptor independent effects of extracellular adenosine on murine thymocytes in conditions of adenosine deaminase deficiency (original) (raw)
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Blood, 1997
(2) the A2a, but not the A1 or A3, receptors are the major absence of adenosine deaminase activity (ADA) activity results in lymphocyte depletion and in severe combined im-expressed and functionally coupled adenosine receptors in mouse peripheral T and B lymphocytes, and the adenosine-munodeficiency (ADA SCID), which is currently explained by direct cell death-causing effects of intracellular products of induced cAMP accumulation in lymphocytes correlates with the expression of A2a receptors; (3) the specific agonist of adenosine metabolism. We explored the alternative mechanisms of peripheral T-cell depletion as due to inhibition of A2a receptor, CGS21680, induces increases in [cAMP]i in lymphocytes, whereas the specific antagonist of A2a recep-T-cell expansion by extracellular adenosine-mediated signaling through purinergic receptors. The strong inhibition of tor, CSC, inhibits the effects of Ado and CGS21680; and (4) the increases in [cAMP]i mimic the adenosine-induced inhi-the T-cell receptor (TCR)-triggered proliferation and of upregulation of interleukin-2 receptor a chain (CD25) mole-bition of TCR-triggered CD25 upregulation and splenocyte proliferation. These studies suggest the possible role of cules, but not the direct lymphotoxicity, were observed at low concentrations of extracellular adenosine. These effects adenosine receptors in the regulation of lymphocyte expansion and point to the downregulation of A2a purinergic re-of extracellular adenosine (Ado) are likely to be mediated by A2a receptor-mediated signaling rather than by intracellular ceptors on T cells as a potentially attractive pharmacologic target. toxicity of adenosine catabolites, because (1) poorly metabolized adenosine analogs cause the accumulation of cAMP ᭧ tively low concentrations of adenosine rather then the direct,
Biochemical Pharmacology, 2003
Adenosine deaminase (ADA) catalyzes the conversion of adenosine and deoxyadenosine to inosine and deoxyinosine, respectively. ADA-deficient individuals suffer from severe combined immunodeficiency and are unable to produce significant numbers of mature T or B lymphocytes. This occurs as a consequence of the accumulation of ADA substrates or their metabolites. dATP is a candidate toxic metabolite because its concentration in RBCs of ADA-deficient patients correlates with the severity of disease. Murine fetal thymic organ culture (FTOC) under ADA-deficient conditions can be used as a model system to investigate the biochemical mechanism responsible for the inhibition of thymopoiesis. In ADA-deficient FTOCs initiated at day 15 of gestation, thymocyte development was arrested at the CD4 À CD8 À CD44 lo CD25 þ to CD4 À CD8 À CD44 lo CD25 À transition. Apoptosis appeared to be involved because the cultures could be rescued by the pan-caspase inhibitor zVADfmk, a Bcl-2 transgene, or deletion of apoptotic protease activating factor-1. As in ADAdeficient patients, dATP was also elevated in ADA-deficient FTOCs. dATP levels were normalized and thymocyte development was rescued in cultures treated with an inhibitor of adenosine kinase, the enzyme that phosphorylates deoxyadenosine to dAMP. zVADfmk also prevented the accumulation of dATP in ADA-deficient FTOCs, suggesting that deoxyadenosine was derived from thymocytes undergoing apoptosis as a consequence of failing the b selection checkpoint. In contrast, dATP levels remained elevated in ADA-deficient FTOCs with fetal thymuses from Bcl-2 transgenic mice. These data suggest that thymocyte apoptosis as a consequence of failing developmental checkpoints involves one or more caspases that are not regulated by Bcl-2.
Journal of Clinical Investigation, 2000
Murine fetal thymic organ culture was used to investigate the mechanism by which adenosine deaminase (ADA) deficiency causes T-cell immunodeficiency. C57BL/6 fetal thymuses treated with the specific ADA inhibitor 2′-deoxycoformycin exhibited features of the human disease, including accumulation of dATP and inhibition of S-adenosylhomocysteine hydrolase enzyme activity. Although T-cell receptor (TCR) Vβ gene rearrangements and pre-TCR-α expression were normal in ADA-deficient cultures, the production of αβ TCR + thymocytes was inhibited by 95%, and differentiation was blocked beginning at the time of β selection. In contrast, the production of γδ TCR + thymocytes was unaffected. Similar results were obtained using fetal thymuses from ADA gene-targeted mice. Differentiation and proliferation were preserved by the introduction of a bcl-2 transgene or disruption of the gene encoding apoptotic protease activating factor-1. The pan-caspase inhibitor carbobenzoxy-Val-Ala-Asp-fluoromethyl ketone also significantly lessened the effects of ADA deficiency and prevented the accumulation of dATP. Thus, ADA substrates accumulate and disrupt thymocyte development in ADA deficiency. These substrates derive from thymocytes that undergo apoptosis as a consequence of failing to pass developmental checkpoints, such as β selection.
Memory of Extracellular Adenosine A2A Purinergic Receptor-mediated Signaling in Murine T Cells
Journal of Biological Chemistry, 1997
Accumulation of extracellular and intracellular adenosine (Ado) under hypoxic conditions or in the absence of adenosine deaminase results in lymphocyte depletion and in severe combined immunodeficiency, which are currently explained by direct intracellular lymphotoxicity of Ado metabolites. In support of the alternative, "signaling" mechanism, we show that extracellular Ado
Journal of Clinical Investigation, 2002
Thymocyte development past the CD4-CD8stage is markedly inhibited in adenosine deaminase-deficient (ADA-deficient) murine fetal thymic organ cultures (FTOCs) due to the accumulation of ADA substrates derived from thymocytes failing developmental checkpoints. Such cultures can be rescued by overexpression of Bcl-2, suggesting that apoptosis is an important component of the mechanism by which ADA deficiency impairs thymocyte development. Consistent with this conclusion, ADA-deficient FTOCs were partially rescued by a rearranged T cell receptor β transgene that permits virtually all thymocytes to pass the β-selection checkpoint. ADA-deficient cultures were also rescued by the adenosine kinase inhibitor 5′-amino-5′-deoxyadenosine (5′A5′dAdo), indicating that the metabolite responsible for the inhibition of thymocyte development is not adenosine or deoxyadenosine, but a phosphorylated derivative of an ADA substrate. Correction of ADA-deficient FTOCs by 5′A5′dAdo correlated with reduced accumulation of dATP, implicating this compound as the toxic metabolite. In ADA-inhibited FTOCs rescued with a Bcl-2 transgene, however, dATP levels were superelevated, suggesting that cells failing positive and negative selection continued to contribute to the accumulation of ADA substrates. Our data are consistent with dATP-induced mitochondrial cytochrome c release followed by apoptosis as the mechanism by which ADA deficiency leads to reduced thymic T cell production.
The Journal of Immunology
The expression of surface adenosine deaminase (ADA) and CD26 in activated human T cells was studied by flow cytometry. PBLs and CD3+ or CD4+ cells, when subjected to a variety of stimuli (anti-CD3 Abs plus IL-2 or phorbol esters), presented two structurally different cell populations, which differed in size and cellular complexity (populations B1 and B2). In PBLs triggered by an anti-CD3 mAb there was no significant increase of expression of either surface ADA or CD26 in cells of population B1, whose structure is similar to that of nonstimulated cells. In contrast, there was a significant increase in the percentage of expression of ADA and CD26 in the population B2, which corresponds to structurally more complex and larger cells. In the case of activation via TCR-CD3 but in the presence of IL-2 or via phorbol esters, the increase was found in cells from both populations, but B2 cells always showed a higher percentage of expression than B1 cells. The results of increased expression o...
Purinergic Signalling, 2005
We have previously reported that ATPgS, a slowly hydrolyzed analog of ATP, inhibits the activation of human CD4 + T lymphocytes by anti-CD3 and anti-CD28 mAb. In this report we have partially characterized the signaling mechanisms involved in this immunosuppressive effect. ATPgS had no inhibitory effect on CD4 + T-cell activation induced by PMA and anti-CD28, indicating that it acts proximally to the TCR. It had no effect on the calcium rise induced by CD3/CD28 stimulation, but inhibited the phosphorylation of three kinases, ERK2, p38 MAPK and PKB, that play a key role in the activation of T cells. The receptor involved in these actions remains unidentified.