Pifithrin-alpha decreases the radioprotective efficacy of a Podophyllum hexandrum Himalayan mayapple fraction REC-2006 in HepG2 cells.
Inhibition of the tumour suppressor p53 by PFT (pifithrin-alpha) promotes p53-mediated apoptosis and protects against doxorubicin-induced apoptosis. The present study was carried out to evaluate the effect of PFT on the radioprotective potential of Podophyllum hexandrum fraction (REC-2006) in HepG2 (p53++) cell line. REC-2006 (10-5 microg/ml) treatment at 2 h before irradiation (10 Gy) rendered 80+/-3% protection in HepG2 cells, whereas PFT debilitated the radioprotective potential of REC-2006. REC-2006 increased the expression of Hsp70 (heat-shock protein 70), HSF1 (heat-shock factor 1) and Bcl-2 in irradiated HepG2 cells, whereas PFT when treated with REC-2006 decreased the expression of Hsp70, HSF1 and Bcl-2 in HepG2 cells. REC-2006 facilitated post-irradiation DNA repair by pausing cell-cycle progression at G1- and G2-phase, whereas no such cell-cycle arrest was observed in irradiated HepG2 cells pretreated with PFT in irradiated HepG2 cells. No change was observed in Mdm2 (murine double minute 2) and Ras-GAP (Ras-GTPase-activating protein) expression with or without PFT treatment. Decrease in the expression of caspase 3 and Bax was observed in HepG2 cells when REC-2006 treatment was given 2 h before irradiation; however, PFT treatment increased the expression of Bax leading to apoptosis. It can be concluded that p53 expression plays a major role in the REC-2006-mediated protection against acute irradiation in HepG2 cells. PFT treatment reduced the radioprotective efficacy of REC-2006 by inhibiting the expression of HSF1 and Hsp70 and thereby the expression of Bcl-2, by up-regulating the cell-cycle-regulatory proteins and therefore reducing the span of time for DNA repair and also by inducing Bax-mediated apoptosis. PFT did not, however, show any effect on p53 regulating protein (Mdm2) and pro-survival protein (Ras-GAP).
Singh
Pankaj Kumar
PK
Division of Radiation Biology and Radioprotection, Institute of Nuclear Medicine and Allied Sciences (INMAS), Delhi-110 054, India.
Kumar
Raj
R
Sharma
Ashok
A
Arora
Rajesh
R
Jain
Swatantra Kumar
SK
Sharma
Rakesh Kumar
RK
D017209:Apoptosis / Q000187:drug effects; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D052160:Benzothiazoles / Q000494:pharmacology*; D018797:Cell Cycle Proteins / Q000096:biosynthesis; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004273:DNA, Neoplasm / Q000528:radiation effects; D004268:DNA-Binding Proteins / Q000096:biosynthesis; D004347:Drug Interactions; D018840:HSP70 Heat-Shock Proteins / Q000096:biosynthesis; D000076249:Heat Shock Transcription Factors; D006801:Humans; D010936:Plant Extracts / Q000494:pharmacology*; D011035:Podophyllum / Q000737:chemistry*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis; D051736:Proto-Oncogene Proteins c-mdm2 / Q000096:biosynthesis; D011837:Radiation-Protective Agents / Q000494:pharmacology*; D014050:Toluene / Q000031:analogs & derivatives* / Q000494:pharmacology; D014157:Transcription Factors / Q000096:biosynthesis; D016159:Tumor Suppressor Protein p53 / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D020703:ras GTPase-Activating Proteins / Q000096:biosynthesis
D051017:Apoptosis Regulatory Proteins; D052160:Benzothiazoles; D018797:Cell Cycle Proteins; D004273:DNA, Neoplasm; D004268:DNA-Binding Proteins; C000619706:HSF1 protein, human; D018840:HSP70 Heat-Shock Proteins; D000076249:Heat Shock Transcription Factors; D010936:Plant Extracts; D019253:Proto-Oncogene Proteins c-bcl-2; D011837:Radiation-Protective Agents; C495901:TP53 protein, human; D014157:Transcription Factors; D016159:Tumor Suppressor Protein p53; D020703:ras GTPase-Activating Proteins; D014050:Toluene; C121565:pifithrin; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2