Flavonoids activated caspases for apoptosis in human glioblastoma T98G and U87MG cells but not in human normal astrocytes.
BACKGROUND
Human glioblastoma is a deadly brain cancer that continues to defy all current therapeutic strategies. The authors induced apoptosis in human glioblastoma T98G and U87MG cells after treatment with apigenin, (-)-epigallocatechin, (-)-epigallocatechin-3-gallate (EGCG), and genistein, which did not induce apoptosis in human normal astrocytes.


METHODS
Induction of apoptosis was examined using Wright staining and ApopTag assay. Production of reactive oxygen species (ROS) and increase in intracellular free Ca(2+) were measured by fluorescent probes. Analysis of mRNA and Western blotting indicated increases in expression and activities of the stress kinases and cysteine proteases for apoptosis. JC-1 showed changes in mitochondrial membrane potential (DeltaPsi(m)), and use of specific inhibitors confirmed activation of kinases and proteases in apoptosis.


RESULTS
Treatment of glioblastoma cells with apigenin, (-)-epigallocatechin, EGCG, or genistein triggered ROS production that induced apoptosis with phosphorylation of p38 mitogen-activated protein kinase (MAPK) and activation of the redox-sensitive c-Jun N-terminal kinase 1 pathway. Pretreatment of cells with ascorbic acid attenuated ROS production and p38 MAPK phosphorylation. Increases in intracellular free Ca2+ and activation of caspase-4 indicated involvement of endoplasmic reticulum stress in apoptosis. Other events in apoptosis included overexpression of Bax, loss of DeltaPsi(m), mitochondrial release of cytochrome c and Smac into the cytosol, down-regulation of baculoviral inhibitor-of-apoptosis repeat-containing proteins, and activation of calpain, caspase-9, and caspase-3. (-)-Epigallocatechin and EGCG also induced caspase-8 activity. Apigenin, (-)-epigallocatechin, EGCG, and genistein did not induce apoptosis in human normal astrocytes.


CONCLUSIONS
Results strongly suggest that flavonoids are potential therapeutic agents for induction of apoptosis in human glioblastoma cells.
Das
Arabinda
A
Department of Neurosciences, Medical University of South Carolina, Charleston, SC 29209, USA. swapan.ray@uscmed.sc.edu
Banik
Naren L
NL
Ray
Swapan K
SK
D017209:Apoptosis; D001253:Astrocytes / Q000201:enzymology* / Q000378:metabolism; D051023:BH3 Interacting Domain Death Agonist Protein / Q000378:metabolism; D001932:Brain Neoplasms / Q000201:enzymology*; D053181:Caspase 8 / Q000378:metabolism; D020169:Caspases / Q000378:metabolism*; D002392:Catechin / Q000031:analogs & derivatives / Q000494:pharmacology; D002460:Cell Line; D045744:Cell Line, Tumor; D004789:Enzyme Activation; D005419:Flavonoids / Q000494:pharmacology*; D005909:Glioblastoma / Q000201:enzymology*; D006801:Humans; D053078:Membrane Potential, Mitochondrial; D048055:Mitogen-Activated Protein Kinase 8 / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism
D051023:BH3 Interacting Domain Death Agonist Protein; D005419:Flavonoids; D017382:Reactive Oxygen Species; D002392:Catechin; C045651:epigallocatechin gallate; D048055:Mitogen-Activated Protein Kinase 8; D053181:Caspase 8; D020169:Caspases; C057580:gallocatechol