Cytoplasmic p21 induced by p65 prevents doxorubicin-induced cell death in pancreatic carcinoma cell line.
BACKGROUND
Studies have shown the existence of p21 induction in a p53-dependent and -independent pathway. Our previous study indicates that DOX-induced p65 is able to bind the p21 promoter to activate its transactivation in the cells.


METHODS
Over-expression and knock-down experiments were performed in Human Pancreatic Carcinoma (PANC1) cells. Cell cycle and cell death related proteins were assessed by Western Blotting. Cytotoxicity assay was checked by CCK-8 kit. Cell growth was analyzed by flow cytometers.


RESULTS
Here we showed that over-expression of p65 decreased the cytotoxic effect of DOX on PANC1 cells, correlating with increased induction of cytoplasmic p21. We observed that pro-caspase-3 physically associated with cytoplasmic p21, which may be contribution to prevent p21 translocation into the nucleus. Our data also suggested that no clear elevation of nuclear p21 by p65 provides a survival advantage by progression cell cycle after treatment of DOX. Likewise, down-regulation of p65 expression enhanced the cytotoxic effect of DOX, due to a significant decrease of mRNA levels of anti-apoptotic genes, such as the cellular inhibitor of apoptosis-1 (c-IAP1), and the long isoform of B cell leukemia/lymphoma-2 (Bcl-2), leading to efficient induction of caspase-3 cleavage in the cells. More, we present evidence that over-expression of p53 or p53/p65 in the PANC1 cells were more sensitive to DOX treatment, correlated with activation of caspase-3 and clear elevation of nuclear p21 level. Our previous data suggested that expression of p21 increases Gefitinib-induced cell death by blocking the cell cycle at the G1 and G2 phases. The present findings here reinforced this idea by showing p21's ability of potentiality of DOX-induced cell death correlated with its inhibition of cell cycle progression after over-expression of p53 or p53/p65.


CONCLUSIONS
Our data suggested p65 could increase p53-mediated cell death in response to DOX in PANC1 cells. Thus, it is worth noting that in p53 null or defective tumors, targeting in down-regulation of p65 may well be useful, leading to the potentiality of chemotherapeutic drugs.
Tongji University School of Medicine, Shanghai, China.
YingQi
Y
Zhou
Gang
G
Li
Yuan
Y
Ji
Chen
C
Liu
JingPing
J
Zhu
YanJun
Y
Lu
D000903:Antibiotics, Antineoplastic / Q000633:toxicity*; D015153:Blotting, Western; D016923:Cell Death; D045744:Cell Line, Tumor; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000096:biosynthesis* / Q000235:genetics; D004317:Doxorubicin / Q000633:toxicity*; D006801:Humans; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D051996:Transcription Factor RelA / Q000378:metabolism*; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
D000903:Antibiotics, Antineoplastic; D050759:Cyclin-Dependent Kinase Inhibitor p21; D012333:RNA, Messenger; D051996:Transcription Factor RelA; D016159:Tumor Suppressor Protein p53; D004317:Doxorubicin