thioinosine and 3-7-dimethyl-1-propargylxanthine

Previously, we have shown that hydrogen peroxide (H2O2) and glucose deprivation (GD) induced ATP loss and cell death in astrocytes. Here, we reported that adenosine and related purine nucleos(t)ides recovered cellular ATP level and completely prevented the cell death in rat primary astrocytes co-treated with H2O2 and glucose deprivation. Time- and concentration-dependently, H2O2 induced cell death and ATP loss in glucose-deprived astrocytes. Adenosine or ATP prevented both astrocytic death and ATP loss caused by H2O2/GD in dose-dependent manner. Further, inhibition of adenosine deamination or transport with erythro-9-(-hydroxy-3-nonyl)adenosine or S-(4-nitrobenzyl)-6-thioinosine largely attenuated the protective effect of adenosine. Other purine nucleos(t)ides such as inosine, guanosine, ADP, AMP, ITP and GTP also showed similar protective effects. Adenosine or ATP also blocked the mitochondrial dysfunction and glutathione (GSH) depletion in H2O2-treated glucose-deprived astrocytes. The present results suggest that adenosine and related purine nucleos(t)ides may protect astrocytes from H2O2 and glucose deprivation induced the potentiated death by restoration of cellular ATP level.

Topics: Adenine; Adenosine; Analysis of Variance; Animals; Animals, Newborn; Astrocytes; Benzimidazoles; Carbocyanines; Cell Death; Cells, Cultured; Dose-Response Relationship, Drug; Drug Interactions; Glucose; Hydrogen Peroxide; In Vitro Techniques; L-Lactate Dehydrogenase; Membrane Potentials; Mitochondria; Purinergic P1 Receptor Agonists; Purinergic P1 Receptor Antagonists; Purines; Rats; Rats, Sprague-Dawley; Theobromine; Thioinosine; Time Factors; Xanthines

The regulatory actions of adenosine on ion channel function are mediated by four distinct membrane receptors. The concentration of adenosine in the vicinity of these receptors is controlled, in part, by inwardly directed nucleoside transport. The purpose of this study was to characterize the effects of adenosine on ion channels in A549 cells and the role of nucleoside transporters in this regulation. Ion replacement and pharmacological studies showed that adenosine and an inhibitor of human equilibrative nucleoside transporter (hENT)-1, nitrobenzylthioinosine, activated K(+) channels, most likely Ca(2+)-dependent intermediate-conductance K(+) (I(K)) channels. A(1) but not A(2) receptor antagonists blocked the effects of adenosine. RT-PCR studies showed that A549 cells expressed mRNA for I(K)-1 channels as well as A(1), A(2A), and A(2B) but not A(3) receptors. Similarly, mRNA for equilibrative (hENT1 and hENT2) but not concentrative (hCNT1, hCNT2, and hCNT3) nucleoside transporters was detected, a result confirmed in functional uptake studies. These studies showed that adenosine controls the function of K(+) channels in A549 cells and that hENTs play a crucial role in this process.

Topics: 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; Adenosine; Affinity Labels; Amiloride; Autocrine Communication; Cell Line; Cell Polarity; Clotrimazole; Diuretics; Epithelial Cells; Equilibrative Nucleoside Transporter 1; Equilibrative-Nucleoside Transporter 2; Growth Inhibitors; Humans; Membrane Transport Proteins; Patch-Clamp Techniques; Potassium; Potassium Channels; Quinazolines; Receptors, Purinergic P1; Respiratory Mucosa; Theobromine; Thioinosine; Triazoles; Uridine; Xanthines

We have investigated the hypothesis that adenosine, a purine nucleoside produced within hypoxic regions of solid tumors, may interfere with the recognition of tumor cells by cytolytic effector cells of the immune system. We measured the adhesion of murine spleen-derived anti-CD3-activated killer (AK) lymphocytes to syngeneic MCA-38 colon adenocarcinoma cells in a model system. Adenosine, in the presence of the adenosine deaminase inhibitor coformycin to prevent the breakdown of adenosine, inhibited adhesion by up to 60%. The inhibitory effect of adenosine was exerted on the AK cells and not on the MCA-38 targets. The response to adenosine was generated at the cell surface, since the inhibition of adhesion was not abrogated by S-(4-nitrobenzyl)-6-thioinosine or dipyridamole, which block adenosine uptake. The inhibition of adhesion due to adenosine was not blocked by either the A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine or the A2 receptor antagonist 3,7-dimethyl-1-propargylxanthine. This suggested that a non-A1, A2 receptor might be involved. The relative order of potencies of adenosine and common analogues was: 5'-N-ethylcarboxamidoadenosine = adenosine = (R)-phenylisopropyladenosine > N6-cyclopentyladenosine > 2-chloro-N6-cyclopentyladenosine = 2-p-(2- carboxyethyl)phenethylamino-5'-N-ethyl-carboxamidoadenosine. This agonist potency profile was again inconsistent with either the A1 or the A2 receptor subtype but indicated that the recently described A3 receptor subtype might be responsible for the inhibition of adhesion. Consistent with this suggestion, aminophenylethyladenosine, an adenosine analogue that binds with high affinity to A3 receptors, inhibited the adhesion of AK cells to MCA-38 tumor cells with high potency (50% inhibitory concentration approximately 1 nM). Adenosine, therefore, interferes with the AK cell recognition of colorectal tumor targets by acting through an A3 receptor on the effector cells. We suggest that this mechanism of immunosuppression, secondary to tissue hypoxia, may be important in the resistance of colorectal and other solid cancers to immunotherapy.

Topics: Adenocarcinoma; Adenosine; Animals; Antibodies; CD3 Complex; Cell Adhesion; Colonic Neoplasms; Killer Cells, Natural; Mice; Theobromine; Thioinosine; Tumor Cells, Cultured; Xanthines