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J Biol Chem, Vol. 273, Issue 21, 12858-12862, May 22, 1998
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From the Oxidants are important human toxicants. Increased
intracellular free Ca2+ may be critical for oxidant
toxicity, but this mechanism remains controversial. Furthermore,
oxidants damage the endoplasmic reticulum (ER) and release ER
Ca2+, but the role of the ER in oxidant toxicity and
Ca2+ regulation during toxicity is also unclear.
tert-Butylhydroperoxide (TBHP), a prototypical organic
oxidant, causes oxidative stress and an increase in intracellular free
Ca2+. Therefore, we addressed the mechanism of
oxidant-induced cell death and investigated the role of ER stress
proteins in Ca2+ regulation and cytoprotection after
treating renal epithelial cells with TBHP. Prior ER stress induces
expression of the ER stress proteins Grp78, Grp94, and calreticulin and
rendered cells resistant to cell death caused by a subsequent TBHP
challenge. Expressing antisense RNA targeted to grp78
prevents grp78 induction sensitized cells to TBHP and
disrupted their ability to develop cellular tolerance. In addition,
overexpressing calreticulin, another ER chaperone and
Ca2+-binding protein, also protected cells against TBHP.
Interestingly, neither prior ER stress nor calreticulin expression
prevented lipid peroxidation, but both blocked the rise in
intracellular free Ca2+ after TBHP treatment. Loading cells
with EGTA, even after peroxidation had already occurred, also prevented
TBHP-induced cell death, indicating that buffering intracellular
Ca2+ prevents cell killing. Thus, Ca2+ plays an
important role in TBHP-induced cell death in these cells, and the ER is
an important regulator of cellular Ca2+ homeostasis during
oxidative stress. Given the importance of oxidants in human disease, it
would appear that the role of ER stress proteins in protection from
oxidant damage warrants further consideration.
Adirondack Biomedical Research Institute,
Lake Placid, New York 12946 and the ¶ Division of Toxicology,
Leiden Amsterdam Center for Drug Research, Leiden University,
2300 RA, Leiden, The Netherlands
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