Ruthenium red, inhibitor of mitochondrial Ca2+ uniporter, inhibits curcumin-induced apoptosis via the prevention of intracellular Ca2+ depletion and cytochrome c release

doi:10.1016/S0006-291X(03)00479-0

Biochemical and Biophysical Research Communications

Volume 303, Issue 4, 18 April 2003, Pages 1073-1079

https://doi.org/10.1016/S0006-291X(03)00479-0 Get rights and content

Abstract

Curcumin, a natural, biologically active compound extracted from rhizomes of Curcuma species, has been shown to possess potent anti-inflammatory, anti-tumor, and anti-oxidative properties. The mechanism by which curcumin initiates apoptosis remains poorly understood. In the present report we investigated the effect of curcumin on the activation of the apoptotic pathway in human leukemia U937 cells. Curcumin induces apoptosis in U937 cells via a mechanism that appears to involve down-regulation of the anti-apoptotic Bcl-xL, and IAP proteins, release of cytochrome c, and activation of caspase 3. Ruthenium red, an inhibitor of mitochondrial uniporter, specifically inhibits curcumin-induced apoptosis in U937 cells. Cotreatment with ruthenium red markedly prevented the activation of caspase 3, cytochrome c release, and cell death, suggesting a role for intracellular Ca²⁺ in this process. Curcumin induced a marked depletion of [Ca²⁺]_i in Caki cells bathed with both Ca²⁺-containing and -free solutions. Thapsigargin (TG), cyclopiazonic acid (CPA), and dantolene (DAN) had no effect. Ruthenium red, an inhibitor of mitochondrial uniporter, only attenuated the curcumin-induced [Ca²⁺]_i depletion in a dose-dependent manner. These data indicate that curcumin acts as a stimulator of intracellular Ca²⁺ uptake into mitochondria via uniporter pathway and may involve in the execution of apoptosis.

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Materials and methods

Cells and materials. Human leukemia U937 and Caki cells were obtained from the American Type Culture Collection (ATCC: Rockville, MD, USA). The culture medium used throughout these experiments was Dulbecco’s modified Eagle’s medium, containing 10% fetal calf serum (FCS), 20 mM Hepes buffer, and 100 μg/ml gentamicin. Curcumin was directly added to cell cultures at the indicated concentrations. Cells were plated on poly-d-lysin-coated glass coverslip (25 mm in diameter) at a density of 2×10⁵ cells

Curcumin induces apoptosis in U937 cells

In order to determine the nature of apoptosis induced by curcumin, U937 cells were treated for 24 h with various concentrations of curcumin. In order to quantify the degree of apoptosis, we analyzed the amount of sub-G1 DNA by flow cytometry of fixed nuclei (Fig. 1A). U937 cells were exposed to various concentrations of curcumin for 24 h. As shown in Fig. 1B, curcumin treatment in U937 cells resulted in a markedly increased accumulation of sub-G1 phase in a dose-dependent manner (Fig. 1B). Recent

Discussion

Our results show that curcumin induces apoptosis in U937 cells. At the apoptosis-inducing concentrations, curcumin also stimulates proteolytic activities of caspase 3 and release of cytochrome c. Ruthenium red, an inhibitor of mitochondrial uniporter, specifically inhibits curcumin-induced apoptosis in U937 cells. Cotreatment with ruthenium red markedly prevented activation of caspase 3, cytochrome c release, and cell death, suggesting a role for intracellular Ca²⁺ in this process. Thus, our

Acknowledgements

This work was supported by Grant No. R13-2002-028-01002-0 from the MRC Program of the Korea Science and Engineering Foundation and partially by a grant from Korea Research Foundation (KRF-2002-015-DS0002).

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View full text

Ruthenium red, inhibitor of mitochondrial Ca2+ uniporter, inhibits curcumin-induced apoptosis via the prevention of intracellular Ca2+ depletion and cytochrome c release

Abstract

Section snippets

Materials and methods

Curcumin induces apoptosis in U937 cells

Discussion

Acknowledgements

Cell Calcium

Exp. Neurol.

Neuroscience

Cell

Phamacology of Curcuma longa

Planta Med.

Inhibitory effect of curcumin, chlorogenic acid, caffeic acid, and ferulic acid on tumor promotion in mouse skin by 12-O-tetradecanoylphorbol-13-acetate

Cancer Res.

Inhibitory effects of curcumin on tumorigenesis in mice

J. Cell Biochem. Suppl.

Antiproliferative effect of curcumin (diferuloylmethane) against human breast tumor cell lines

Anticancer Drugs

Inhibitory effects of dietary curcumin on forestomach, duodenal, and colon carcinogenesis in mice

Cancer Res.

Chemopreventive drug development: perspectives and progress

Cancer Epidemiol. Biomarkers Prev.

Ruthenium red, inhibitor of mitochondrial Ca²⁺ uniporter, inhibits curcumin-induced apoptosis via the prevention of intracellular Ca²⁺ depletion and cytochrome c release