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Reactive oxygen species mediate doxorubicin induced p53-independent apoptosis
W P Tsang1, Sophia P Y Chau, S K Kong
1Department of Biochemistry, The Chinese University of Hong Kong, Shatin, Hong Kong, PR China.
Abstract:
Doxorubicin (DOX) is a common anticancer drug. The mechanisms of DOX induced apoptosis and the involvement of reactive oxygen species (ROS) in apoptotic signaling were investigated in p53-null human osteosarcoma Saos-2 cells. Accumulation of pre-G1 phase cells and induction of DNA laddering, which are the hallmarks of apoptosis, were detected in cells at 48 h upon DOX treatment. Furthermore, DOX increased the intracellular hydrogen peroxide and superoxide levels, followed by mitochondrial membrane depolarization, cytochrome c release, caspase-3 activation, prior to DNA laddering in Saos-2 cells. In addition, DOX treatment also upregulated Bax and downregulated Bcl-2 levels in the cells. The role of ROS in DOX induced cell death was confirmed by the suppression effect of catalase on DOX induced ROS formation, mitochondrial cytochrome c release, procaspase-3 cleavage, and apoptosis in Saos-2 cells. The catalase treatment however only suppressed DOX induced Bax upregulation but had no effect on Bcl-2 downregulation. Results from the present study suggested that ROS might act as the signal molecules for DOX induced cell death and the process is still functional even in the absence of p53.
Insights
Reactive oxygen species (ROS) mediate doxorubicin-induced apoptosis in p53-null osteosarcoma cells. Catalase suppressed ROS, but not Bcl-2 changes, confirming ROS
Area of Science:
- Cell Biology
- Molecular Oncology
- Biochemistry
Background:
- Doxorubicin (DOX) is a widely used chemotherapy agent.
- Apoptosis, or programmed cell death, is a key mechanism in cancer therapy.
- The role of reactive oxygen species (ROS) in DOX-induced apoptosis is not fully understood, especially in p53-deficient cells.
Purpose of the Study:
- To investigate the mechanisms of DOX-induced apoptosis in human osteosarcoma Saos-2 cells lacking functional p53.
- To elucidate the specific involvement of ROS in the apoptotic signaling pathway triggered by DOX.
Main Methods:
- Saos-2 cells were treated with Doxorubicin (DOX).
- Apoptosis was assessed by quantifying pre-G1 phase cell accumulation and DNA laddering.
- Intracellular ROS levels (hydrogen peroxide and superoxide) were measured.
- Mitochondrial function, caspase-3 activation, and Bcl-2/Bax protein expression were analyzed.
- The effect of catalase, an ROS scavenger, on DOX-induced effects was evaluated.
Main Results:
- DOX treatment induced apoptosis, evidenced by pre-G1 cell accumulation and DNA laddering.
- DOX increased intracellular ROS, leading to mitochondrial membrane depolarization, cytochrome c release, and caspase-3 activation.
- DOX upregulated Bax and downregulated Bcl-2 expression.
- Catalase treatment reduced DOX-induced ROS, cytochrome c release, caspase-3 activation, and apoptosis, and suppressed Bax upregulation but not Bcl-2 downregulation.
Conclusions:
- Reactive oxygen species (ROS) act as critical signaling molecules in doxorubicin-induced apoptosis in p53-null osteosarcoma cells.
- The apoptotic pathway initiated by DOX remains functional even in the absence of p53.
- ROS are involved in the modulation of Bcl-2 family proteins (Bax and Bcl-2) during DOX-induced cell death.