Mechanism of extracellular ATP- and adenosine-induced apoptosis of cultured pulmonary artery endothelial cells

Abstract

Apoptosis may be important in the exacerbation of endothelial cell injury or limitation of endothelial cell proliferation. We have found that extracellular ATP (exATP) and adenosine cause endothelial apoptosis and that the development of apoptosis is linked to intracellular metabolism of adenosine [Dawicki, D. D., D. Chatterjee, J. Wyche, and S. Rounds. Am. J. Physiol. 273 ( Lung Cell Mol. Physiol. 17): L485–L494, 1997]. In the present study, we investigated the mechanism of this effect. We found that exATP, adenosine, and the S-adenosyl-l-homocysteine (SAH) hydrolase inhibitor MDL-28842 caused apoptosis and decreased the ratio of S-adenosyl-l-methionine to SAH compared with untreated control cells. Using release of soluble [3H]thymidine as a measure of DNA fragmentation, we found that the effect of adenosine on soluble DNA release was potentiated by coincubation with homocysteine. These results suggest that the mechanism of exATP- and adenosine-induced endothelial cell apoptosis involves inhibition of SAH hydrolase. exATP-induced apoptosis was enhanced by an inhibitor of adenosine deaminase, whereas exogenous adenosine-induced apoptosis was partially inhibited by an adenosine deaminase inhibitor. These results suggest that adenosine deaminase may also be involved in the mechanism of adenosine-induced endothelial cell apoptosis. Adenosine and MDL-28842 caused intracellular acidosis as assessed with the fluorescent probe 2′,7′-bis(2-carboxyethyl)-5(6)-carboxyfluorescein. The cell-permeant base chloroquine prevented adenosine-induced acidosis but not apoptosis. Thus, although intracellular acidosis is associated with adenosine-induced apoptosis, it is not necessary for this effect. We speculate that exATP- and adenosine-induced endothelial cell apoptosis may be due to an inhibition of methyltransferase(s) activity. Purine-induced endothelial cell apoptosis may be important in limiting endothelial cell proliferation after vascular injury.

Role of elevated S-adenosylhomocysteine in rat hepatocyte apoptosis: Protection by betaine

“…These naïve hepatocytes were then exposed in vitro to agents that are known to increase intracellular SAH levels by exploiting the unique characteristic of the enzyme, SAH hydrolase, which catalyzes the reverse hydrolysis of SAH to adenosine and homocysteine. Incubations with SAHhydrolase inhibitor, or under conditions of excess adenosine and homocysteine either added alone or in combination have all been shown to generate increased intracellular SAH [24][25][26]32]. But in our study, we did not use homocysteine as an agent to achieve increased intracellular SAH levels but instead focused on adenosine.…”

“…More recent studies using various cell types that implicate increased adenosine toxicity via increased SAH levels have employed comparable adenosine concentrations as used by us [24,[26][27][28][29]42]. The mechanism of action of adenosine appears to be via its ability to act as a substrate for as well as an inhibitor of SAH hydrolase thereby elevating intracellular SAH.…”

“…In recent years there have been reports of increased intracellular SAH levels inducing apoptosis in many different cell types [23][24][25][26][27][28][29]. Therefore, we put forth the hypothesis that increased intracellular SAH generation may be responsible for hepatocyte apoptosis seen after ethanol administration.…”

“…We accomplished this undertaking by exploiting the unique characteristic of the enzyme SAH hydrolase-the only enzyme involved in SAH catabolism. Since SAH hydrolase catalyzes the reversible hydrolysis to adenosine and homocysteine with equilibrium of the reaction favoring the synthesis of SAH [30,31], altering the levels of its products (adenosine and/or homocysteine) has been shown to play a critical role in the control of tissue levels of SAH [24][25][26]32]. Therefore in this study, hepatocytes were exposed to varying concentrations of adenosine in order to promote the generation of SAH in order to increase intracellular SAH levels.…”

Sustained Adenosine Exposure Causes Lung Endothelial Barrier Dysfunction via Nucleoside Transporter–Mediated Signaling

“…Adenosine has been shown to cause EC apoptosis (41). However, exposure of confluent endothelial monolayer to (A1D) for 40 hours did not significantly increase EC apoptosis (Figure 4e).…”