[p33(ING1b) enhances chemosensitivity of osteosarcoma cell U2OS to etoposide].
OBJECTIVE
As a new tumor suppressor gene, ING1 shared many biological functions with p53, such as cell cycle arrest, DNA repair, apoptosis, and chemosensitivity. The aim of this study was to investigate the effect of p33(ING1b) on chemosensitivity of osteosarcoma cells and its mechanism.


METHODS
p33(ING1b) was overexpressed in osteosarcoma cell line U2OS through transient transfection. After transfection, U2OS cells were treated with etoposide for 24 hours, then cell growth inhibitory rates were detected by trypan blue exclusion assay, and apoptosis was assessed using flow cytometry analysis and fluorescent microscopy. Furthermore, the protein expression of p53, p21(WAF1), MDM2 and Bax were determined by Western blot analysis.


RESULTS
After transient transfection with p33(ING1b) vector for 24 hours, U2OS cells were treated with 20 microg/ml VP-16 for 24 hours. The results showed that the cell growth inhibitory rates strongly increased [(63.1+/-5.1)%], and etoposide- induced apoptosis was increased(62.7%). Ectopic overexpression of p33(ING1b) increased the protein expression of p53 and strongly enhanced the expression of endogenous p21(WAF1) and Bax. Moreover, after transfection and treatment with 20 microg/ml VP-16, the protein expression of p53, p21(WAF1), and Bax strongly increased compared with other groups. The protein expression of MDM2 showed no significant difference.


CONCLUSIONS
These observations suggest that p33(ING1b) up-regulates p53 protein, and cooperate with p53 in stimulating expression of p21(WAF1) and Bax gene, thus to enhance etoposide-induced apoptosis via p53-dependent pathways.
Zhu
Jin-Jun
JJ
Institute of Orthopedic Oncology, Department of Orthopedic Surgery, The First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, PR China.
Liao
Wei-Ming
WM
Li
Fo-Bao
FB
Zhu
Xiao-Feng
XF
Zhou
Jun-Min
JM
Liu
Zong-Chao
ZC
D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis; D001859:Bone Neoplasms / Q000378:metabolism* / Q000473:pathology; D018797:Cell Cycle Proteins; D002455:Cell Division; D045744:Cell Line, Tumor; D004268:DNA-Binding Proteins; D004357:Drug Synergism; D005047:Etoposide / Q000494:pharmacology*; D016147:Genes, Tumor Suppressor; D006801:Humans; D000076023:Inhibitor of Growth Protein 1; D047908:Intracellular Signaling Peptides and Proteins; D009687:Nuclear Proteins; D012516:Osteosarcoma / Q000378:metabolism* / Q000473:pathology; D011506:Proteins / Q000235:genetics / Q000494:pharmacology*; D014162:Transfection; D025521:Tumor Suppressor Proteins
D000972:Antineoplastic Agents, Phytogenic; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C501503:ING1 protein, human; D000076023:Inhibitor of Growth Protein 1; D047908:Intracellular Signaling Peptides and Proteins; D009687:Nuclear Proteins; D011506:Proteins; D025521:Tumor Suppressor Proteins; D005047:Etoposide