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Published on: November 11, 2022
Molecular mechanisms regarding potassium bromate‑induced cardiac hypertrophy without apoptosis in H9c2 cells
Shu-Chun Kuo1, Yingxiao Li2, Yung-Ze Cheng3
1Department of Optometry, Chung Hwa University of Medical Technology, Tainan 7170, Taiwan R.O.C.
Insights
Potassium bromate (KBrO3) at low doses induces cardiac hypertrophy in H9c2 cells by increasing reactive oxygen species (ROS) and activating the calcineurin/NFAT pathway, without causing apoptosis. This offers a new tool for studying cardiac hypertrophy.
Area of Science:
- Cardiology
- Cell Biology
- Toxicology
Background:
- Cardiac hypertrophy is a significant factor in cardiac injury.
- Oxidative stress is a known inducer of cardiac hypertrophy and apoptosis.
- Potassium bromate (KBrO3), an oxidizing food additive, prompts investigation into its cellular effects.
Purpose of the Study:
- To investigate the effects of KBrO3 on cardiac hypertrophy in rat-derived H9c2 heart cells.
- To elucidate the underlying mechanisms, including oxidative stress and specific signaling pathways.
- To determine the dose-dependent relationship between KBrO3 exposure and cellular responses, particularly apoptosis.
Main Methods:
- H9c2 cells were treated with varying concentrations of KBrO3.
- Cell size, gene expression (BNP, β-MHC), and apoptosis were assessed.
- Reactive oxygen species (ROS) and superoxide levels were measured using fluorescence assays.
- Protein levels of calcineurin and NFAT were analyzed via Western blot.
- The effects of antioxidants (tiron) and calcineurin inhibitors (cyclosporine A) were evaluated.
Main Results:
- KBrO3 increased H9c2 cell size and promoted gene expression of cardiac hypertrophy biomarkers (BNP, β-MHC) in a dose-dependent manner (<250 µM).
- KBrO3 significantly increased superoxide and ROS levels, which were mitigated by tiron.
- KBrO3 elevated calcineurin and NFAT protein levels, effects reversed by tiron and cyclosporine A.
- Apoptosis was observed only at higher KBrO3 concentrations (>300 µM).
Conclusions:
- Low-dose KBrO3 (<250 µM) induces cardiac hypertrophy in H9c2 cells via ROS generation and activation of the calcineurin/NFAT pathway, without inducing apoptosis.
- KBrO3 serves as a potential tool for inducing cardiac hypertrophy in vitro and in animal models.
- These findings provide a novel perspective on KBrO3's role in cardiac pathophysiology.
Abstract:
Cardiac hypertrophy is commonly involved in cardiac injury. Oxidative stress can induce cardiac hypertrophy with apoptosis. Potassium bromate (KBrO3) has been widely used as a food additive due to its oxidizing properties. In the present study, the rat‑derived heart cell line H9c2 was used to investigate the effect of KBrO3 on cell size. KBrO3 increased cell size at concentrations <250 µM, in a dose‑dependent manner. Additionally, KBrO3 also promoted the gene expression of two biomarkers of cardiac hypertrophy, brain/B‑type natriuretic peptides (BNP) and β‑Myosin Heavy Chain (β‑MHC). However, apoptosis remained unobserved in these cells. Moreover, mediation of free radicals was investigated using a fluorescence assay, and it was observed that superoxide and reactive oxygen species (ROS) levels increased with KBrO3. Effects of KBrO3 were significantly reduced by tiron at concentrations sufficient to produce antioxidant‑like action. Additionally, signals involved in cardiac hypertrophy such as calcineurin and nuclear factor of activated T‑cells (NFAT) were also determined using western blot analysis. KBrO3 increased the protein levels of both these molecules which were decreased by tiron in a dose‑dependent manner. Additionally, cyclosporine A attenuated the cardiac hypertrophy induced by KBrO3 in H9c2 cells at concentrations effective to inhibit calcineurin, in addition to reducing mRNA levels of BNP or β‑MHC. Finally, apoptosis was also identified in H9c2 cells incubated with KBrO3 at concentrations >300 µM. Collectively, these results provided a novel perspective that KBrO3 induces cardiac hypertrophy without apoptosis at a low dose through the generation of ROS, activating the calcineurin/NFAT signaling pathway in H9c2 cells. Therefore, at a dose <250 µM, KBrO3 can be applied as an inducer of cardiac hypertrophy without apoptosis in H9c2 cells. KBrO3 can also be developed as a tool to induce cardiac hypertrophy in animals.
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