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Cytotoxic Efficacy of Photodynamic Therapy in Osteosarcoma Cells In Vitro
Published on: March 18, 2014
[p33(ING1b) enhances chemosensitivity of osteosarcoma cell U2OS to etoposide]
Jin-Jun Zhu1, Wei-Ming Liao, Fo-Bao Li
1Institute of Orthopedic Oncology, Department of Orthopedic Surgery, The First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, PR China.
Background & 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.
Conclusion:
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.
Insights
The ING1 tumor suppressor enhances osteosarcoma cell sensitivity to etoposide chemotherapy. Overexpressing p33(ING1b) boosts apoptosis and increases p53, p21(WAF1), and Bax protein levels, suggesting a p53-dependent mechanism.
Area of Science:
- Oncology
- Molecular Biology
- Cancer Genetics
Background:
- ING1 is a novel tumor suppressor gene with functions similar to p53.
- ING1 plays a role in cell cycle arrest, DNA repair, apoptosis, and chemosensitivity.
Purpose of the Study:
- To investigate the impact of p33(ING1b) on osteosarcoma chemosensitivity.
- To elucidate the underlying molecular mechanisms of p33(ING1b)-mediated chemosensitivity.
Main Methods:
- Overexpression of p33(ING1b) in U2OS osteosarcoma cells via transient transfection.
- Treatment with etoposide and assessment of cell growth inhibition and apoptosis.
- Analysis of p53, p21(WAF1), MDM2, and Bax protein expression using Western blot.
Main Results:
- p33(ING1b) overexpression significantly increased cell growth inhibition and etoposide-induced apoptosis.
- Ectopic p33(ING1b) enhanced the protein expression of p53, p21(WAF1), and Bax.
- MDM2 protein levels showed no significant alteration.
Conclusions:
- p33(ING1b) up-regulates p53 protein expression.
- p33(ING1b) cooperates with p53 to enhance p21(WAF1) and Bax expression.
- These effects contribute to increased etoposide-induced apoptosis through p53-dependent pathways.
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