Ca2+ influx mediates apoptosis induced by 4-aminopyridine, a K+ channel blocker, in HepG2 human hepatoblastoma cells.
Apoptosis appears to be implicated in the pathogenesis and therapeutic applications of cancer. In this study we investigated the induction of apoptosis by 4-aminopyridine (4-AP), a K(+) channel blocker, and its mechanism in HepG2 human hepatoblastoma cells. 4-AP reduced cell viability and induced DNA fragmentation, a hallmark of apoptosis, in a dose-dependent manner. In addition, 4-AP induced a sustained increase in intracellular Ca(2+) concentration, which was completely inhibited by the extracellular Ca(2+) chelation with EGTA. 4-AP also induced Mn(2+) influx, indicating that the 4-AP-induced increased intracellular Ca(2+) levels were due to activation of Ca(2+) influx pathway. 4-AP also depolarized membrane potential that was measured by using di-O-C(5)(3), a voltage-sensitive fluorescent dye. 4-AP-induced Ca(2+) influx was significantly inhibited not by voltage-operative Ca(2+) channel blockers (nifedipine or verapamil), but by flufenamic acid (FA), a known nonselective cation channel blocker. Quantitative analysis of apoptosis by the flow cytometry revealed that treatment with either FA or BAPTA, an intracellular Ca(2+) chelator, significantly inhibited the 4-AP-induced apoptosis. Taken together, these results suggest that the observed 4-AP-induced apoptosis in the HepG2 cells may result from Ca(2+) influx through the activation of voltage-sensitive Ca(2+)-permeable non-selective cation channels. These results further suggest that membrane potential change by modulation of K(+) channel activity may be involved in the mechanism of apoptosis in human hepatoma cells.
Kim
J A
JA
Department of Physiology, College of Pharmacy, Yeungnam University, Kyongsan, Korea.
Kang
Y S
YS
Jung
M W
MW
Kang
G H
GH
Lee
S H
SH
Lee
Y S
YS
D015761:4-Aminopyridine / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology*; D002118:Calcium / Q000378:metabolism / Q000502:physiology*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects; D002614:Chelating Agents / Q000494:pharmacology; D053938:DNA Fragmentation / Q000187:drug effects; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D005439:Flufenamic Acid / Q000494:pharmacology; D018197:Hepatoblastoma / Q000473:pathology*; D006801:Humans; D008113:Liver Neoplasms / Q000473:pathology*; D008345:Manganese / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009543:Nifedipine / Q000494:pharmacology; D026902:Potassium Channel Blockers; D015221:Potassium Channels / Q000502:physiology; D014407:Tumor Cells, Cultured / Q000187:drug effects; D014700:Verapamil / Q000494:pharmacology
D002121:Calcium Channel Blockers; D002614:Chelating Agents; D026902:Potassium Channel Blockers; D015221:Potassium Channels; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D008345:Manganese; D004533:Egtazic Acid; D005439:Flufenamic Acid; D015761:4-Aminopyridine; D014700:Verapamil; D009543:Nifedipine; D002118:Calcium