Bioinformatics and experimental insights into the relationship between ferroptosis and fluoride-induced neurotoxicity.

Introduction

Fluorine is a widely distributed element in the form of fluoride in nature and plays an important role in human health (Ayub et al., 2024). Appropriate fluoride intake supports growth and development, including bone health maintenance and dental caries prevention. However, fluoride concentrations in groundwater can indeed reach extremely high levels, occasionally as high as 42 mg/L (Singh et al., 2018). Residents in these areas are exposed to high levels of fluoride through drinking water or dietary intake, and excessive intake can cause fluorosis, a systemic condition that damages multiple tissues and organs (Wang et al., 2024, Du et al., 2024a). As a global endemic disease, fluorosis is prevalent in more than 20 countries, endangering the health of about 200 million people worldwide (Rasool et al., 2018). Elucidating the mechanism of fluorosis is therefore critical for the prevention and control of fluoride-related health damage.

Fluoride can penetrate the blood-brain barrier (BBB) and subsequently enter brain tissue, leading to neurological damage characterized by symptoms such as memory loss, dizziness, and tremors (Ren et al., 2022). Epidemiological studies have shown that children’s fluoride exposure levels are significantly associated with their diminished non-verbal intellectual abilities, lower intelligence quotient (IQ), and internalizing symptomatology (Adkins et al., 2022, Yu et al., 2021). Furthermore, fluoride can penetrate the placental barrier and BBB during embryonic development, adversely affecting fetal neurodevelopment (Hu and Wu, 1988). Maternal fluoride exposure levels during pregnancy have been associated with lower IQ and an increased risk of attention deficit hyperactivity disorder in childhood (Green et al., 2019, Bashash et al., 2018). Animal experiments have corroborated these findings, showing that fluoride-treated rats have impaired spatial learning and memory, as well as anxiety- and depression-like behaviors (Du et al., 2024a, Zhou et al., 2021). Despite substantial evidence of fluoride-induced neurological impairment, the underlying molecular mechanisms, particularly those involving regulated cell death pathways, remain incompletely understood.

Neuronal death is a key contributor to fluoride-related neurological impairment, involving processes such as autophagy, apoptosis, and necroptosis (Lai et al., 2020, Niu et al., 2018, Wei et al., 2025). Emerging research suggests that ferroptosis, a novel iron-dependent form of regulated cell death, also plays a pivotal role in neurological impairment caused by fluoride (Zhao et al., 2024, Xu et al., 2025). Ferroptosis is characterized by intracellular iron overload, lipid peroxidation, and shrunken or even absent mitochondrial cristae (Zeng et al., 2023). Through Fenton reaction, excessive Fe²- drives the overproduction of reactive oxygen species (ROS), which subsequently oxidize polyunsaturated fatty acids (PUFAs) to generate cytotoxic lipid peroxides, including 4-hydroxynonenal and malondialdehyde (MDA), ultimately triggering ferroptosis (Chen et al., 2021). Essential for ferroptosis control, the system Xc- antiporter is composed of the SLC7A11 (xCT) light chain and the SLC3A2 heavy chain (Jiang et al., 2021). Specifically, SLC7A11 enables the synthesis of glutathione (GSH) and glutathione peroxidase 4 (GPX4) by mediating cystine import into cells. As a pivotal negative regulator of ferroptosis, GPX4 can reduce lipoperoxides in a GSH-dependent manner, thereby mitigating lipoperoxide accumulation and protecting cells from oxidative damage (Ye et al., 2022). Downregulation of SLC7A11 impairs GSH production, leading to lipoperoxide accumulation and ferroptosis (Koppula et al., 2021). Notably, p53 has been shown to promote ferroptosis by inhibiting SLC7A11 (Li et al., 2023a), and elevated p53 expression has been observed in the medial prefrontal cortex of rats and in SH-SY5Y cells treated with fluoride (Javanbakht et al., 2025, Xiang et al., 2024). Therefore, investigating the role of p53-mediated ferroptosis in fluoride-induced neurological impairment could provide critical insights into the underlying mechanisms.

Based on this background, we conducted bioinformatics analysis to identify differentially expressed genes (DEGs) and key signaling pathways associated with fluoride-induced neurological impairment using a transcriptomic dataset (GSE195920) derived from U87-MG human glioblastoma cells (non-neuronal origin), thereby providing preliminary mechanistic clues. We then validated these findings using an in vivo model (rats treated with fluoride from the embryonic period to adulthood) and in vitro models of SH-SY5Y cells. This study seeks to elucidate how fluoride induces neurological impairment, thereby providing a scientific rationale for early intervention.