Affiliation:
1. From the Respiratory Division, Hôpital Cantonal Universitaire de Genève (Switzerland).
Abstract
Abstract
We have investigated the effects of reactive O
2
metabolites generated by the hypoxanthine–xanthine oxidase (HX-XO) system on intracellular Ca
2+
and its relation with protein synthesis in human umbilical vein endothelial cells (HUVECs). Spectrofluorometry with fura 2 showed that the oxidative stress induced a rapid transient rise in cytosolic [Ca
2+
], followed by a sustained elevation above the baseline value. In the presence of La
3+
, which blocks Ca
2+
influx from the extracellular medium, a transient [Ca
2+
] increase was still observed, but the sustained rise was suppressed. The HX-XO–related [Ca
2+
] changes were completely prevented by pretreatment with thapsigargin, which depletes intracellular Ca
2+
stores. Hence, the effects of HX-XO on Ca
2+
homeostasis were due to mobilization of Ca
2+
from the intracellular stores with subsequent influx of extracellular Ca
2+
. HX-XO mobilized more of sequestered Ca
2+
than did thrombin, a receptor agonist that depletes only a part of the intracellular Ca
2+
stores (the hormone-sensitive stores). To determine the relevance of the HX-XO–related depletion of Ca
2+
stores for cell function, we investigated the role of Ca
2+
mobilization in the regulation of protein synthesis. Overall protein synthesis in HUVECs was markedly reduced by thapsigargin, which depletes both hormone-sensitive and -insensitive stores, but was not substantially affected by thrombin. Manipulation of the refilling of the Ca
2+
stores via the availability of extracellular Ca
2+
significantly influenced the thapsigargin-related and the HX-XO–related inhibition of overall protein synthesis. A corresponding effect of extracellular [Ca
2+
] was seen in polyribosome distribution profiles, which reflected an inhibition of translation initiation in both treatments. Thus, depletion of Ca
2+
stores appeared to be involved in the inhibition of protein synthesis at the initiation level by both thapsigargin and HX-XO. These results indicate that (1) the cytosolic [Ca
2+
] changes induced by HX-XO result from mobilization of Ca
2+
from intracellular stores and subsequent influx of extracellular Ca
2+
, (2) the HX-XO–related mobilization of sequestered Ca
2+
includes hormone-insensitive pools, and (3) the depletion of hormone-insensitive Ca
2+
stores appears to be in part responsible for the inhibition of protein synthesis by HX-XO.
Publisher
Ovid Technologies (Wolters Kluwer Health)
Subject
Cardiology and Cardiovascular Medicine,Physiology
Cited by
57 articles.
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