mRNA expression analysis of a variety of apoptosis-related genes, including the novel gene of the BCL2-family, BCL2L12, in HL-60 leukemia cells after treatment with carboplatin and doxorubicin.
Apoptosis is a type of programmed cell death involved in many crucial biological processes. It represents the basic mechanism for the action of chemotherapeutic agents, such as doxorubicin and carboplatin. Both are able to cause cell death through the induction of apoptosis in the human leukemic cell line HL-60. We investigated the possible alterations in the expression of apoptosis-related genes, including the novel BCL2L12 gene, which was recently cloned in our group. The kinetics of apoptosis induction and cell toxicity was investigated by DNA laddering and by the MTT method, respectively. Total RNA was extracted and cDNA was prepared by reverse transcription. BCL2 , BAX , FAS , caspase-9, caspase-3 and BCL2L12 were amplified by PCR. Overexpression of FAS , BCL2L12 and caspase-3 was observed after treatment of HL-60 cells for 3 or 6 h with carboplatin, while their expression was decreased after a 12-h treatment, demonstrating that these genes may take part in the early stages of apoptosis. Overexpression of the same genes was also observed after 6 h of treatment with doxorubicin (concomitantly with DNA laddering). In the case of carboplatin-induced apoptosis we detected down-regulation of BAX , BCL2 and caspase-9, whereas in the case of doxorubicin, BAX and BCL2 remained at control levels and caspase-9 was increased.
Floros
Kostas V
KV
National Center for Scientific Research 'Demokritos', GR-15310 Athens, Greece.
Thomadaki
Hellinida
H
Katsaros
Nikos
N
Talieri
Maroulio
M
Scorilas
Andreas
A
D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000235:genetics*; D001483:Base Sequence; D016190:Carboplatin / Q000494:pharmacology*; D017931:DNA Primers; D004317:Doxorubicin / Q000494:pharmacology*; D019254:Genes, bcl-2; D018922:HL-60 Cells; D006801:Humans; D007938:Leukemia / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D000970:Antineoplastic Agents; D017931:DNA Primers; D012333:RNA, Messenger; D004317:Doxorubicin; D016190:Carboplatin