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Actin-sequestering protein, thymosin beta-4, induces paclitaxel resistance through ROS/HIF-1alpha stabilization in
1Department of Bioscience and Biotechnology, Sejong University, Seoul, Republic of Korea.
Aims:
We investigated whether actin-sequestering protein, thymosin beta-4 (TB4)-induced reactive oxygen species (ROS) affect the stabilization of hypoxia-inducible transcription factor (HIF)-1alpha and paclitaxel-resistance induction.
Main Methods:
HeLa human cervical tumor cells were used. The percentage of cell survival was determined by MTT assay. ROS production, cell cycle and hypodiploid cell formation were assessed by flow cytometry analysis. HIF-1alpha stabilization and molecular changes were analyzed by western blotting or RT-PCR. NF-kappaB activation was assessed by EMSA and western blotting.
Key Findings:
TB4 protein (TB4P) significantly increased intracellular ROS level and HIF-1alpha. The increased level of HIF-1alpha by TB4P was reduced by the treatment with N-acetylcysteine (NAC), a well-known ROS scavenger. TB4P-induced ROS production was confirmed by the activation of nuclear factor kappa B. TB4P-induced Erk phosphorylation was attenuated by the treatment with NAC. In addition, tumor cell death was decreased by TB4 gene overexpression and TB4P treatment. NAC treatment attenuated tumor cell density increased by TB4P. Tumor cell death by paclitaxel was also increased by NAC treatment or the transfection with HIF-1alpha-siRNA. Paclitaxel-induced B16F10 mouse melanoma regression was physiologically inhibited in TB4-transgenic mice compared to wildtype mice.
Significance:
These findings demonstrate that TB4-induced ROS and ROS-mediated HIF-1alpha stabilization could play a role in tumor cell resistance to anticancer agents like paclitaxel. It suggests that soluble TB4 could be a novel endogenous regulator to control intracellular ROS production in tumor cells.
Insights
Thymosin beta-4 (TB4) increases reactive oxygen species (ROS), stabilizing hypoxia-inducible factor-1alpha (HIF-1alpha) and promoting paclitaxel resistance in tumor cells. Reducing ROS with N-acetylcysteine (NAC) reverses this resistance.
Area of Science:
- Cell Biology
- Molecular Oncology
- Biochemistry
Background:
- Actin-sequestering protein thymosin beta-4 (TB4) plays a role in cell proliferation and survival.
- Reactive oxygen species (ROS) are implicated in various cellular processes, including cancer progression and drug resistance.
- Hypoxia-inducible factor-1alpha (HIF-1alpha) is a key regulator of cellular response to hypoxia and is often overexpressed in tumors, contributing to therapeutic resistance.
Purpose of the Study:
- To investigate the role of TB4-induced ROS in HIF-1alpha stabilization.
- To determine if TB4-induced ROS contribute to paclitaxel resistance in tumor cells.
- To explore TB4 as a potential regulator of intracellular ROS and its impact on cancer therapy.
Main Methods:
- HeLa human cervical tumor cells were utilized for in vitro experiments.
- Cell survival was assessed using MTT assays.
- ROS production, cell cycle, and apoptosis were analyzed via flow cytometry.
- HIF-1alpha stabilization and gene expression were evaluated by western blotting and RT-PCR.
- NF-kappaB activation was measured using EMSA and western blotting.
Main Results:
- TB4 protein significantly elevated intracellular ROS levels and HIF-1alpha stabilization.
- N-acetylcysteine (NAC), a ROS scavenger, reversed TB4-induced HIF-1alpha stabilization and reduced ROS production.
- TB4 overexpression and treatment decreased tumor cell death and enhanced resistance to paclitaxel.
- Paclitaxel efficacy was improved by NAC treatment or HIF-1alpha knockdown.
- TB4-transgenic mice exhibited inhibited paclitaxel-induced melanoma regression.
Conclusions:
- TB4-induced ROS and subsequent HIF-1alpha stabilization contribute to tumor cell resistance against paclitaxel.
- TB4 acts as an endogenous regulator of intracellular ROS in tumor cells.
- Targeting TB4-mediated ROS pathways may offer novel strategies to overcome chemoresistance.
