Regulation of intracellular calcium levels and urokinase activity in MDA MB 231 cells by quercetin.
BACKGROUND
The common plant bioflavonoid, quercetin, is cytotoxic to various tumor cell lines, particularly breast cancer, by affecting the protein-kinase-C-dependent signal pathways and by cell cycle regulation. However, its role in breast cancer metastasis has not been studied so far. Increased uPA activity is evident in highly metastatic breast cancer, which is calcium dependent.


METHODS
MDA MB 231 cells were treated with various concentrations of quercetin (15-45 microg/ml). The cytotoxic effect of quercetin was assessed by MTT assay and DNA fragmentation analysis. Intracellular calcium levels were measured using Fura-2, a specific Ca2+ fluorescence indicator. Calcium uptake and release in cells treated with quercetin were measured using radioactive 45Ca2+. Urokinase enzyme activity was assayed by a casein zymogram.


RESULTS
Quercetin elicited dose- and time-dependent cytotoxicity as evidenced by the MTT assay. The maximum effect was observed at 48 h with a quercetin concentration of (45 microg/ml). DNA agarose gel electrophoresis showed dose-dependent DNA fragmentation on quercetin treatment. Quercetin caused significant depletion of cytosolic calcium levels and decreased calcium uptake from the intracellular stores. Casein zymogram showed a marked reduction of urokinase activity as evident by clear lysis bands on a dark background on treatment with quercetin.


CONCLUSIONS
Quercetin was found to exhibit cytotoxicity in the highly invasive breast cancer cell line MDA MB 231 in a dose- and time-dependent manner. Quercetin inhibited calcium dependent urokinase activity and hence may prove to be an effective antimetastatic agent.
Devipriya
S
S
Department of Biochemistry, University of Madras, Guindy Campus, Chennai, India.
Vani
G
G
Ramamurthy
N
N
Shyamaladevi
C S
CS
D000230:Adenocarcinoma; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D001943:Breast Neoplasms; D002118:Calcium / Q000378:metabolism*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D053938:DNA Fragmentation; D004305:Dose-Response Relationship, Drug; D004587:Electrophoresis, Agar Gel; D005260:Female; D006801:Humans; D011794:Quercetin / Q000494:pharmacology*; D014568:Urokinase-Type Plasminogen Activator / Q000037:antagonists & inhibitors*
D000972:Antineoplastic Agents, Phytogenic; D011794:Quercetin; D014568:Urokinase-Type Plasminogen Activator; D002118:Calcium