Oxidative stress and reactive oxygen species in otorhinolaryngological diseases: insights from pathophysiology to targeted antioxidant therapies

ABSTRACT

Oxidative stress, characterized by an imbalance between excessive reactive oxygen species (ROS) production and impaired antioxidant defenses, is closely linked to the pathogenesis of various otorhinolaryngological disorders. Mitochondria, as the primary site of cellular energy production, play a crucial role in modulating oxidative stress. Mitochondrial dysfunction exacerbates ROS generation, leading to cellular damage and inflammatory responses. In otorhinolaryngological diseases, oxidative stress is strongly associated with conditions such as hearing loss, allergic rhinitis, and chronic sinusitis, where oxidative damage and tissue inflammation are key pathological features. Recent studies have highlighted the potential of antioxidant therapies to mitigate oxidative stress and restore homeostasis, offering promising avenues for alleviating symptoms in these diseases. However, despite the encouraging results from early-stage research, the clinical efficacy of antioxidant interventions remains to be fully established. This review provides an overview of the role of oxidative stress in otorhinolaryngological diseases and evaluates the therapeutic potential of antioxidant strategies.

Introduction

Reactive Oxygen Species (ROS) are highly reactive molecules that include free radicals such as the superoxide anion (${\mathrm{O}}_2^{-}$ )and hydroxyl radical (·OH), as well as non-radical oxidants like hydrogen peroxide (${\mathrm{H}}_2{\mathrm{O}}_2$) [1]. These species primarily arise as metabolic byproducts, particularly during mitochondrial oxidative phosphorylation. Under normal physiological conditions, ROS levels are maintained at low concentrations; however, they can increase significantly under stress conditions such as inflammation, radiation, and pollution[2]. ROS have been recognized as crucial signaling molecules since the 1970s[3]. This ROS-generating process is prevalent in neutrophils, epithelial cells, macrophages, and endothelial cells, contributing to immune responses and signal transduction [4]. Under normal physiological conditions, ROS levels are kept low and play essential roles in cellular signaling, growth, differentiation, and apoptosis. However, under stress conditions like inflammation, radiation, and pollution, ROS levels can increase significantly, leading to oxidative stress, which damages DNA, proteins, and lipids[5]. This oxidative stress is increasingly recognized as a key factor in the pathogenesis of otorhinolaryngological diseases, including hearing loss, chronic rhinosinusitis, allergic rhinitis, and obstructive sleep apnea[6–9].

Emerging evidence underscores the pivotal role of oxidative stress in the pathogenesis of various diseases. Initial theories posited that oxidative stress causes pathological changes through direct oxidative damage. However, antioxidant interventions, such as systemic vitamin C and vitamin E trials, have shown limited success in alleviating cardiovascular diseases[10]. Furthermore, the clinical associations between vitamin E and asthma have been inconsistent[11]. These findings suggest that the mechanisms underlying oxidative stress are not yet fully understood. Recent research has revealed that oxidative stress plays a more complex role than previously thought. Using antioxidants to simply inhibit ROS production has shown only modest effectiveness in slowing disease progression, and in some cases, may exacerbate conditions[12]. Effective ROS signaling requires precise regulation to maintain a balance between oxidation and antioxidation, ensuring that beneficial physiological ROS signaling pathways are preserved while preventing harmful pathways from becoming overactive. This complexity likely contributes to the limited efficacy of antioxidant therapies [13].

Given the critical link between ROS signaling and otorhinolaryngological diseases, the regulation of ROS production and clearance has become a key research focus. However, a comprehensive review on the relationship between mitochondrial dysfunction, oxidative stress, and otorhinolaryngological diseases is currently lacking. This review aims to explore the intracellular sources of ROS, elucidate the role of oxidative stress in otorhinolaryngological diseases, and systematically summarize the connections between mitochondrial dysfunction, oxidative stress, and these conditions. Additionally, it will evaluate the effectiveness of ROS-related therapies in otorhinolaryngological disorders and assess the prospects and feasibility of targeted antioxidant treatments.

Major sources of ROS

As research advances, increasing evidence highlights the distinct roles of various ROS in different pathological processes. Identifying the specific ‘footprint’ of ROS – its type, source, and mechanism in particular diseases – is essential for understanding pathophysiology and developing targeted therapies. Key ROS, such as ${\mathrm{O}}_2^{-}$, serve as precursors to other ROS, driving inflammatory responses. ${\mathrm{H}}_2{\mathrm{O}}_2$, a stable ROS capable of diffusing across membranes, modulates signal transduction but can also induce oxidative damage in cells. The ·OH, produced through the Fenton reaction, is highly reactive and directly causes lipid peroxidation and DNA damage. Mitochondria are central to cellular energy production and are also major sources of ROS. ROS are primarily generated in the electron transport chain of the mitochondrial inner membrane, where some electrons leak to ${\mathrm{O}}_2$, forming the ${\mathrm{O}}_2^{-}$·, a key ROS species. There are eight identified sites within mitochondria capable of producing ${\mathrm{O}}_2^{-}$, although the precise contributions of these sites to intracellular ROS levels remain unclear. Theoretically, ${\mathrm{O}}_2^{-}$ produced in the intermembrane space can more readily diffuse through the outer mitochondrial membrane, while ${\mathrm{O}}_2^{-}$in the matrix must traverse both inner and outer membranes to escape the mitochondrion[3]. Research indicates that complexes I and III of the respiratory chain are significant sources of mitochondrial ${\mathrm{O}}_2^{-}$·[14]. Mitochondria not only serve as a primary source of superoxide anions but also generate hydrogen peroxide through enzymatic reactions. Other enzyme systems, such as peroxidases and xanthine oxidase, also contribute to hydrogen peroxide production.

Another important ${\mathrm{O}}_2^{-}$· source is NADPH oxidase (NOX) complexes, which use NADPH as an electron donor to convert oxygen molecules into superoxide. The NOX family comprises NOX1-5, DUOX1, and DUOX2. Initially identified in phagocytes, NOX enzymes are now known to be present in various cell types including immune cells, smooth muscle cells, fibroblasts, and epithelial cells[15, 16]. NOX1, NOX2, and NOX3 are primarily located on the cell membrane and require binding with the stabilizing subunit p22phox for activation, which involves the recruitment of cytosolic active subunits[17]. NOX5, DUOX1, and DUOX2 are activated at the cell membrane but do not require cytosolic subunits; instead, they are activated through $\mathrm{Ca}{}^{2+}$ binding. DUOX1 and DUOX2 also need DUOXA1 and DUOXA2 for maturation, whereas NOX5 is activated solely by $\mathrm{Ca}{}^{2+}$. In contrast, NOX4 does not require cytosolic subunits for activation and is expressed in various organelles, including the nucleus, mitochondria, and endoplasmic reticulum[18]. Unlike other NOX subunits that primarily generate ${\mathrm{O}}_2^{-}$·, NOX4 is unique in directly producing a significant amount of ${\mathrm{H}}_2{\mathrm{O}}_2$ through enzymatic reactions, without requiring extensive conversion of superoxide anion. The activation of NOX enzymes is regulated by external stimuli such as cytokines (e.g. TNF-α, IL-1β), growth factors (e.g. PDGF), and pathogens, playing a crucial role in cellular signaling and functional regulation depending on their subcellular localization and tissue distribution (Figure 1).

Figure 1.

Major sources of ROS: NADPH oxidases (NOXs) are membrane-bound enzymes that play a crucial role in generating ROS. NOX1-3, located at the plasma membrane, produce ${\mathrm{O}}_2^{-}$ in the cytosol. Their activation involves the stabilization of the p22phox subunit and the recruitment of cytosolic active subunits. NOX5 and DUOX1/2, which bind $\mathrm{Ca}{}^{2+}$, also generate superoxide in the cytosol. NOX4 is located on the plasma membrane and the mitochondrial inner membrane, where it produces ${\mathrm{O}}_2^{-}$ and ${\mathrm{H}}_2{\mathrm{O}}_2$ in the cytoplasm and mitochondrial matrix. The respiratory chain complexes I and III, present in the mitochondrial inner membrane, produce O2− and ${\mathrm{H}}_2{\mathrm{O}}_2$ in the intermembrane space. Xanthine oxidase, located in the cytoplasm, generates O2− and ${\mathrm{H}}_2{\mathrm{O}}_2$. Peroxisomes produce small amounts of O2− and ${\mathrm{H}}_2{\mathrm{O}}_2$ during the oxidation of fatty acids and other substrates.

Scavenging system for ROS

The body maintains redox balance through endogenous antioxidant systems, which include both enzymatic and non-enzymatic components. The enzymatic systems consist of superoxide Dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx). The SOD family includes three main isoenzymes: SOD1 (Cu/Zn-SOD, located in the cytoplasm), SOD2 (Mn-SOD, located in the mitochondrial matrix), and SOD3 (EC-SOD, located in the extracellular matrix). All three isoenzymes convert superoxide anion (${\mathrm{O}}_2^{-}$) into hydrogen peroxide (${\mathrm{H}}_2{\mathrm{O}}_2$) and oxygen (${\mathrm{O}}_2$). SOD1 expression is regulated by specificity protein 1 (Sp1) and requires the copper chaperone CCS1 for activation [19]. SOD2 activity is modulated by acetylation, with deacetylation by Sirt3 enhancing its ROS-scavenging capability[20]. SOD3 binds metal ions in the extracellular matrix, thereby protecting matrix proteins from oxidative damage and reducing chronic inflammation[21]. CAT is primarily found in peroxisomes but can also be expressed in mitochondria at high hydrogen peroxide concentrations, where it neutralizes oxidative stress from respiration and protects cells. The GPx family reduces hydrogen peroxide and organic peroxides to water and alcohols at selenium-containing active sites while oxidizing reduced glutathione (GSH) to oxidized glutathione (GSSG), thus maintaining the intracellular GSH/GSSG balance and bolstering antioxidant capacity[22].

Non-enzymatic antioxidant systems also play a crucial role in protecting cells from oxidative damage. Glutathione maintains antioxidant balance by reducing hydrogen peroxide and organic peroxides; vitamin C scavenges free radicals in bodily fluids; vitamin E shields cell membranes from lipid peroxidation; carotenoids capture free radicals, thereby reducing oxidative damage; selenium, as a component of antioxidant enzymes like GPx, enhances peroxide clearance; and phenolic compounds neutralize free radicals, further protecting cells. These mechanisms collectively provide a vital biological foundation for the prevention and treatment of oxidative stress-related diseases (Figure 2).

Figure 2.

Scavenging system for ROS: Superoxide (${\mathrm{O}}_2^{-}$) is converted into hydrogen peroxide (${\mathrm{H}}_2{\mathrm{O}}_2$) in the cytosol and mitochondrial matrix by superoxide dismutases (SOD1 and SOD2). Hydrogen peroxide is subsequently transformed into water (H₂O) by catalase (CAT) in both locations. Additionally, hydrogen peroxide is detoxified by glutathione peroxidases (GPx1 and GPx2 in the cytosol, GPx1 and GPx4 in mitochondria) and peroxiredoxins (PRX1 and PRX2 in the cytosol, PRX3 and PRX5 in mitochondria). Glutathione peroxidases convert reduced glutathione (GSH) into oxidized glutathione (GSSG), which is then regenerated by glutathione reductase (GR) using NADPH. In the cytosol, thioredoxin reductase (TRR1) employs NADPH to regenerate reduced thioredoxin (Trx1), thereby maintaining the activity of PRX1 and PRX2. In mitochondria, mitochondrial thioredoxin reductase (TRR2) uses NADPH to regenerate reduced thioredoxin (Trx2), sustaining the activity of PRX3 and PRX5

Methods for identifying specific ROS and oxidative damage

Accurate detection techniques are essential for analyzing ROS footprints and understanding their pathophysiological mechanisms. Fluorescent probes are commonly used for ROS detection due to their high sensitivity and ability to monitor dynamic changes in real time. For example, MitoSOX Red detects mitochondrial-derived ${\mathrm{O}}_2^{-}$·, revealing dynamic ROS generation in hair cells during noise-induced hearing loss[23]. Similarly, DHE and DCF-DA probes are employed to monitor ROS production due to upregulated NOX activity in nasal-related diseases[7, 24] (Summary in Table 1). Molecular biomarker detection is another important method, using lipid peroxidation products such as malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE) to indirectly quantify ROS levels. Elevated MDA in OSA patients reflects ROS activity, though this method only captures cumulative effects. Enzyme activity assays, which measure SOD, GPx, and CAT activity, are also used to indirectly assess ROS levels[25].

Table 1.

Summary of fluorescent probes for ROS detection and their significance.

ROS Type	Main Fluorescent Probes	Detection Target	Significance
Superoxide (${\mathrm{O}}_2^{-}$·)	- DHE (Dihydroethidium) - MitoSOX Red	- DHE: Measures total intracellular ${\mathrm{O}}_2^{-}$·levels - MitoSOX: Mitochondria-specific ${\mathrm{O}}_2^{-}$ levels	Primary ROS detection from NOX and mitochondria
Hydrogen Peroxide (${\mathrm{H}}_2{\mathrm{O}}_2$)	- Amplex Red - Peroxyfluor series - HyPer (genetically encoded)	- Amplex Red: Quantifies ${\mathrm{H}}_2{\mathrm{O}}_2$ in cells/tissues - HyPer: Monitors real-time ${\mathrm{H}}_2{\mathrm{O}}_2$ dynamics	${\mathrm{H}}_2{\mathrm{O}}_2$ signaling and oxidative stress
Hydroxyl Radical (·OH)	- HPF (Hydroxyphenyl fluorescein) - APF (Aminophenyl fluorescein)	- HPF: Measures total ·OH levels - APF: Specifically detects ·OH, avoiding interference from RNS	Tracks reactive ROS damaging DNA, lipids, proteins
Mitochondrial ROS (Mito-ROS)	- MitoSOX Red - MitoTracker Red CM-H2XRos	- MitoSOX: Detects mitochondrial ${\mathrm{O}}_2^{-}$·levels - MitoTracker Red CM-H2XRos: Detects overall mitochondrial ROS	Mitochondrial ROS, oxidative stress, dysfunction
Global ROS Detection	- DCFH-DA (Dichlorofluorescein Diacetate) - Carboxy-H2DCFDA - CellROX (Deep Red, Green, etc.)	- DCFH-DA: Measures total ROS levels -Carboxy-H2DCFDA: Enhanced stability for reliable quantification - CellROX: General oxidative stress indicator for live cells	Summary of oxidative stress contributions

Recent advancements in detection methods for ROS and oxidative damage have been driven by technological progress. While traditional methods like fluorescence probes and biomarker analysis are widely used, they are limited by specificity and sensitivity, making it difficult to fully elucidate the molecular and spatial distribution of oxidative damage. Innovative approaches have introduced new tools for studying oxidative stress. For example, mass spectrometry (MS) combined with high-resolution proteomics enables precise identification of oxidative protein modifications (e.g. carbonylation, nitration) and changes in abundance[26]. Single-cell analysis techniques help capture cellular heterogeneity, revealing how specific cells, such as inner ear hair cells, respond to ROS[27]. High-resolution imaging technologies, such as confocal and super-resolution microscopy, coupled with specific probes, allow subcellular visualization of ROS distribution, enabling the observation of ROS dynamics[28], such as mitochondrial ROS generation in hair cells[29]. These advanced technologies not only enhance our understanding of oxidative stress in otorhinolaryngological diseases but also lay the groundwork for developing targeted therapeutic strategies (Figure 3).

Figure 3.

ROS Detection Workflow: The diagram highlights the workflow of reactive oxygen species (ROS) detection, starting with advanced detection technologies such as flow cytometry, confocal microscopy, mass spectrometry, and fluorescent probes. Data analysis identifies molecular biomarkers critical for understanding oxidative stress. These findings support potential clinical applications, including diagnosis, therapeutic optimization, and efficacy monitoring (Created with BioRender.com).

Oxidative stress and otorhinolaryngological diseases

Oxidative stress induces cell damage by altering protein structure and function. These changes often involve oxidative modifications, such as side-chain modifications or cross-linking, resulting in protein aggregation, degradation, or loss of activity. Such damage commonly affects mitochondrial, membrane, and cytoskeletal proteins, and has been closely linked to the pathological processes of various otorhinolaryngological diseases.

Sensorineural hearing loss

Sensorineural Hearing Loss (SNHL) is a prevalent auditory disorder resulting from partial or complete dysfunction of the auditory pathways from the ear to the auditory cortex. Its causes include genetic factors, aging, ototoxic drug use, and noise exposure. Research highlights the significant roles of mitochondrial dysfunction and oxidative stress in the pathogenesis of SNHL. Sensory hair cells, the stria vascularis, and spiral ganglion neurons – structures with high metabolic demands – are particularly susceptible to mitochondrial dysfunction. In sensorineural hearing loss, mitochondrial ROS induce oxidative stress, impairing hair cell proteins, particularly those in mitochondrial respiratory complexes I and III. These proteins may undergo oxidative modifications, leading to reduced electron transfer efficiency and disrupted cellular energy metabolism. Furthermore, lipid peroxidation products, such as 4-HNE, exacerbate membrane protein dysfunction, accelerating hair cell apoptosis[30, 31]. Hereditary hearing losses are often linked to mitochondrial gene mutations, such as the A1555G mutation in the mitochondrial 12S rRNA gene, which is associated with maternally inherited deafness[32]. Age-related hearing loss (ARHL) primarily affects high-frequency hearing and is characterized by progressive mitochondrial dysfunction, increased mtDNA mutations, and elevated ${\mathrm{O}}_2^{-}$· production with aging[33]. The enzyme SOD1, crucial for oxidative stress protection, is closely related to ARHL. Its deficiency reduces antioxidant capacity, making cochlear hair cells more vulnerable to damage[34]. ROS-induced DNA damage is associated with decreased cellular energy and mitochondrial dysfunction; excessive ROS can activate the AMPK signaling pathway, accelerating cell apoptosis and worsening ARHL. NADPH oxidase (NOX) also plays a significant role in ARHL; NOX3 knockout reduces ${\mathrm{O}}_2^{-}$· production and inhibits outer hair cell apoptosis, thereby improving hearing. In contrast, p22phox-deficient mice exhibit reduced NOX activity, decreased upregulation of excitatory pathway genes in cochlear tissue, and diminished excitotoxic damage to the cochlea, leading to improved ARHL [35, 36].

Noise-Induced Hearing Loss (NIHL) results from prolonged exposure to high-intensity noise, leading to damage of inner ear hair cells. Excessive ROS generation is a primary mechanism in NIHL. Noise exposure activates the caspase pathway, leading to mitochondrial release of cytochrome c and initiation of apoptosis[37]. Additionally, Excessive mitochondrial ${\mathrm{O}}_2^{-}$· production in cochlear outer hair cells triggers lipid peroxidation via the AMPK signaling pathway, contributing to outer hair cell death and hearing impairment. In rat models of NIHL, decreased mtDNA content, gene expression, and ATP levels are observed, along with impaired TFAM expression and mtDNA binding function; Mitochondrial dysfunction in sensory hair cells has been shown to cause excessive production of ROS, especially ${\mathrm{O}}_2^{-}$·, leading to protein oxidation and lipid peroxidation. Antioxidant treatment markedly reduces levels of 4-HNE and MDA in hair cells, indicating that lipid peroxidation is a central mechanism of cochlear damage[30]. Noise exposure also increases mitochondrial calcium levels and metabolic demands in hair cells, further enhancing ROS production[38]. NADPH oxidase plays a critical role in NIHL, with upregulation of NOX1 and DUOX2 associated with cochlear damage, while downregulation of NOX3 may represent an endogenous protective mechanism. Inhibiting NADPH oxidase to reduce ${\mathrm{O}}_2^{-}$· production is considered a novel therapeutic approach for NIHL. In a NIHL mouse model, NOX4 affects the synaptic ribbons of the auditory nerve, leading to delayed development and early degeneration. This may represent a novel therapeutic target for oxidative stress-induced SNHL [39]. SOD2 converts toxic superoxide anions into hydrogen peroxide, mitigating oxidative damage. Mice lacking SOD2 show reduced survival of inner and outer hair cells under noise damage, particularly at 4 kHz, highlighting the role of oxidative stress in cochlear damage and elevated hearing thresholds [40].

Ototoxicity refers to hearing loss caused by certain drugs, such as cisplatin, which induces damage and apoptosis of inner ear hair cells. Cisplatin induces excessive ${\mathrm{O}}_2^{-}$· accumulation in mitochondria, causing mitochondrial dysfunction and leading to hair cell damage and apoptosis[41]. Inhibiting ROS production has been shown to improve cisplatin-induced hearing loss (CIHL). For instance, inhibiting protein arginine methyltransferase 6 (PRMT6) reduces ROS generation, enhances mitochondrial function, and reverses cisplatin-induced caspase-3 expression and cytochrome c translocation, thus inhibiting inner ear hair cell apoptosis [42]. Cisplatin induces the loss of the antioxidant enzyme PRDX1, leading to mitochondrial ${\mathrm{O}}_2^{-}$· accumulation, lipid peroxidation, and impaired mitochondrial autophagy, ultimately damaging spiral ganglion neurons[29]. Additionally, cisplatin increases mRNA expression of NOX family subtypes, such as NOX1 and NOX4; targeting these subtypes can effectively reduce ${\mathrm{O}}_2^{-}$·and ${\mathrm{H}}_2{\mathrm{O}}_2$ production, thereby alleviating cisplatin-induced ototoxicity, and NOX3 knockout can also alleviate cisplatin-induced apoptosis[43]. Aminoglycoside antibiotics can cause mtROS accumulation and loss of membrane potential, leading to DNA damage and caspase-3 activation, with upregulation of NOX2 expression and its ${\mathrm{O}}_2^{-}$· production being significant contributors to neomycin-induced ototoxicity [44, 45].

In summary, oxidative stress and mitochondrial dysfunction are central to various forms of SNHL. In-depth research into these mechanisms holds the potential to develop new therapeutic strategies to slow or reverse the progression of SNHL and improve patients’ quality of life.

Chronic rhinosinusitis

Chronic Rhinosinusitis (CRS) is a prevalent inflammatory condition affecting the mucosa of the nasal cavity and sinuses, characterized by persistent nasal congestion, hyposmia, and facial pain. CRS is divided into two types based on the presence of nasal polyps: CRS with nasal polyps (CRSwNP) and CRS without nasal polyps (CRSsNP). The pathogenesis of CRS is multifaceted, with recent research emphasizing the critical roles of oxidative stress and mitochondrial dysfunction. Studies have demonstrated that colonization by various bacteria and their products, including Staphylococcus aureus exotoxins (e.g. SEA, SEC1-C3, SED) and Staphylococcus aureus enterotoxin B (SEB), can contribute to the development of CRS. These exotoxins induce significant inflammatory responses by activating T cells, thereby mediating the disease process. A study conducted in China observed significantly elevated levels of SEB and SEA in the nasal tissues of patients with CRSwNP, suggesting that Staphylococcus aureus may play a key role in this condition [46]. SEB regulates p47phox to induce excessive ${\mathrm{O}}_2^{-}$· production in nasal epithelial cells, causing endoplasmic reticulum stress and promoting CRSwNP development [47]. SEB exposure altered the expression of mitochondrial fusion and fission proteins, leading to mitochondrial dysfunction and morphological damage[48].

In CRSwNP, analyses of oxidative stress-related genes have revealed significant upregulation of nitric oxide synthase (iNOS) and heme oxygenase-1 (HO-1), while lactoperoxidase (LPO), myeloperoxidase (MPO), and superoxide dismutase 3 (SOD3) are downregulated. These biomarkers may serve as endogenous predictors for CRSwNP[49]. Loss of epithelial-specific Nrf2 exacerbates oxidative stress, leading to eosinophilic inflammation and epithelial barrier dysfunction, positioning Nrf2 as a potential therapeutic target[50]. Elevated expression of p67phox has been observed in eosinophils and neutrophils within nasal polyp tissues[51]. Additionally, increased expression of DUOX1 and DUOX2 at the apical surface of nasal epithelial cells correlates with heightened levels of cytokines such as Eotaxin, MIG, TNF-α, and IL-8 in both CRSwNP and CRSsNP[52]. In vitro studies have demonstrated elevated levels of NOX1, HO-1, and SOD2 in nasal epithelial cells and macrophages derived from nasal polyps, correlating positively with inflammatory cytokines like IL-5 and IL-6[53]. Immunofluorescence staining has shown that DUOX2-derived ${\mathrm{H}}_2{\mathrm{O}}_2$ triggers the translocation of high mobility group box 1 (HMGB1) from the nucleus to the extracellular space on epithelial cells, amplifying the inflammatory response and upregulating IL-8[54].

Epithelial – mesenchymal transition (EMT) is a critical mechanism in the pathogenesis of CRS. CRSwNP is characterized by goblet cell hyperplasia, extracellular matrix protein deposition, edema, and increased vascularity, while CRSsNP primarily shows epithelial detachment, collagen fiber deposition, and submucosal fibrosis. Oxidative stress regulates TGF-β1 activation and expression by interfering with latency-associated peptide (LAP), which normally binds to and activates TGF-β1. Activated TGF-β1 subsequently impairs mitochondrial function and increases mtROS production. Research indicates that oxidative stress promotes TGF-β1 production in macrophages via the ROS/p38 MAPK/Egr-1 pathway, thereby inducing EMT and contributing to nasal polyp formation [55]. The activation and expression of TGF-β1 are closely associated with disease persistence in CRS, with ROS playing a significant role, particularly under the influence of external factors such as smoking [56]. Studies using DAB staining have shown elevated levels of ${\mathrm{H}}_2{\mathrm{O}}_2$ in the nasal mucosa of CRS patients, with a more pronounced increase in those exposed to secondhand smoke, indicating that smoking significantly enhances ${\mathrm{H}}_2{\mathrm{O}}_2$ production [57].

In summary, oxidative stress is crucial in the development and progression of CRS. Further research into these mechanisms may improve our understanding of the disease's pathological processes and uncover new therapeutic targets for CRS.

Allergic rhinitis

Allergic rhinitis (AR) is an IgE-mediated Type I hypersensitivity reaction to allergens, resulting in inflammation primarily in the nasal mucosa. Although not life-threatening, AR significantly impacts quality of life. Recent research has highlighted the role of oxidative stress in the pathogenesis of AR. Elevated markers of oxidative stress and reduced antioxidant defenses have been observed in AR patients, underscoring the importance of oxidative stress in the disease. Histamine, a key mediator in AR, not only promotes ${\mathrm{O}}_2^{-}$· production but also increases the levels of ERK and p38 proteins[58]. A study involving 226 AR patients found significantly higher serum levels of total antioxidant status (TAS) and total oxidant status (TOS) compared to controls, indicating increased oxidative stress in AR patients. Additionally, the duration of AR is associated with increased antioxidant activity, which may reflect an adaptive response to sustained oxidative stress[59]. Research on grass pollen-induced AR demonstrated that allergen exposure reduced the activity of mitochondrial respiratory chain complex I and impaired mitochondrial coupling within 6 h, suggesting a detrimental effect on mitochondrial function[60]. Pollen-derived NADPH oxidase has been shown to independently induce ROS production in mucosal cells, regardless of adaptive immune responses[61].

The nasal epithelial barrier serves as the primary defense against pathogens and allergens, playing a crucial role in the pathogenesis of AR. Recent studies suggest that ROS contribute to AR by promoting inflammation through epithelial cells or disrupting the epithelial barrier. In nasal epithelial cells, ${\mathrm{O}}_2^{-}$· induces oxidative stress, impairing the function of tight junction proteins (Occludin and Claudin). This dysfunction, likely due to oxidative modifications, compromises the intercellular barrier, increasing permeability and facilitating excessive inflammatory factor leakage[62, 63]. Immunofluorescence analysis reveals a significant reduction in these proteins in the nasal epithelium of allergic rhinitis patients, likely resulting from ROS-induced degradation. Oxidative stress also inhibits tyrosine phosphatases through signaling pathways such as p38 MAPK, p65 NF-kB, and Akt, thereby reducing the expression of tight junction proteins[64]. Studies have shown that in allergic mice, abnormalities in the AMPK/Drp1-mediated mitochondrial fission pathway lead to mitochondrial ${\mathrm{O}}_2^{-}$·, reduced mitochondrial membrane potential, elevated Drp1 expression, increased apoptotic markers, and Th1/Th2 imbalance. These abnormal indicators are associated with exacerbated allergic rhinitis symptoms[65].

NADPH oxidase, expressed in various cell types including epithelial cells, smooth muscle cells, fibroblasts, and immune cells, plays a significant role in AR. NOX1 and NOX4 are predominantly expressed in the nasal mucosal epithelium, with differential expression noted between AR patients and normal controls[66]. Recently, NOX2 has also been identified in nasal mucosal epithelial tissues. Previous studies have shown that Th2 cytokines induce the generation of ROS through DUOX2, promoting the extracellular transport of HMGB1. This process regulates inflammation-related markers, such as IL-5, IL-13, and eosinophil infiltration, playing a key role in the pathogenesis of AR[7]. NOX1 and NOX4 are implicated in processes such as proliferation, differentiation, apoptosis, and angiogenesis and are associated with respiratory diseases like asthma. Whole-genome microarray analysis revealed upregulated NOX4 mRNA in airway smooth muscle, increasing oxidative stress, ${\mathrm{H}}_2{\mathrm{O}}_2$ production, and DNA damage, thereby promoting muscle hypercontraction and contributing to asthma pathogenesis[67, 68]. Excessive ${\mathrm{H}}_2{\mathrm{O}}_2$ production mediated by NOX4 leads to mitochondrial dysfunction, increased mitochondrial ${\mathrm{O}}_2^{-}$·, and activation of mitochondrial-dependent apoptotic pathways and NLRP3/IL-1β-mediated inflammation, resulting in airway inflammation[69]. While the exact roles of these NADPH oxidase subunits in AR require further clarification, their mechanisms may mirror those in asthma, indicating a need for additional research.

Intermittent hypoxia and obstructive sleep apnea

Obstructive Sleep Apnea (OSA) is a disorder characterized by partial or complete obstruction of the upper airway during sleep. This repeated airway blockage leads to reduced blood oxygen levels, fragmented sleep, and a range of complications, including cardiovascular dysfunction and cognitive impairments. During this process, patients experience intermittent hypoxia (IH), which causes fluctuations in blood oxygen levels and reoxygenation. This cycle results in the generation of ROS and systemic oxidative stress. Intermittent hypoxia-induced ROS in endothelial cells of OSA patients alters protein function, potentially through nitration. Studies suggest that oxidative stress may impair endothelial nitric oxide synthase (eNOS) activity by generating nitrotyrosine (3-NT) from peroxynitrite ($\mathrm{ONO}{\mathrm{O}}^{-}$), resulting in reduced vasodilation[70]. This process is closely linked to systemic inflammation and oxidative damage. Studies have found that ROS induce early endothelial dysfunction in OSA by upregulating both leukocyte-specific adhesion molecules (e.g. L-selectin and integrins) and endothelial-specific adhesion molecules (e.g. E-selectin, P-selectin, ICAM-1, and VCAM-1), thereby heightening the risk of cardiovascular diseases [71]. OSA is closely linked to obesity, which represents a chronic inflammatory state and is highly associated with oxidative stress and an increased prevalence of cardiovascular diseases [72]. Studies have shown that SOD activity levels are lower in OSA patients, particularly those with mild to moderate OSA[73]. Recent research highlights the importance of redox status in regulating the activity of suprachiasmatic nucleus neurons and clock gene transcription. Different reactive oxygen species produced by microglia and astrocytes can disrupt circadian clock function, leading to disturbances in circadian rhythms[74].

Sleep disruptions, which are common in OSA patients, may exacerbate oxidative stress, complicating the relationship between oxidative stress and sleep regulation. For instance, one study found that mitochondrial ${\mathrm{O}}_2^{-}$· in the brain of fruitfly Drosophila regulates sleep-inducing neurons by modulating the NADPH/NADP + ratio in the Shaker K Vβ subunit, linking energy metabolism, oxidative stress, and sleep, with potential implications for developing sleep-regulatory drugs[75]. Chronic sleep deprivation has been shown to significantly impact mitochondrial function, including disruption of oxidative phosphorylation, uncoupling of ATP production, reduction in cytochrome c oxidase activity, and decreased mitochondrial membrane potential[76]. Mitochondria are not only a primary source of ROS but also play a crucial role during the reoxygenation phase in OSA patients. Research indicates that neurocognitive impairments associated with OSA are primarily driven by neuroinflammation and oxidative stress resulting from CIH. Mitochondrial ${\mathrm{O}}_2^{-}$· regulates mitophagy by activating the NLRP3 inflammasome. NLRP3 deficiency enhances Parkin-mediated mitophagy, reducing neuronal damage, microglial activation, and oxidative stress (e.g. MDA and SOD levels), while promoting the clearance of damaged mitochondria[77].

In OSA patients, hypoxia-inducible factor-1α (HIF-1α) plays a significant role in the adaptive response to hypoxic conditions and is heavily involved in oxidative stress generation. HIF-1α upregulates enzymes involved in mitochondrial respiration, resulting in increased levels of TNF-α, inflammatory cytokines (IL-2, IL-4, IL-6), lipid peroxidation, and circulating cell-free DNA in OSAS patients [71]. Furthermore, HIF-1α is closely associated with elevated levels of ${\mathrm{O}}_2^{-}$·in OSA patients, which can also be generated by NADPH oxidase[78]. Research has shown that intermittent hypoxia can induce high airway reactivity in rats, with increased expression of the p47phox protein in laryngeal tissues compared to controls, indicating that NADPH oxidase-derived ${\mathrm{O}}_2^{-}$· play a critical role in laryngeal airway hyperreactivity [79].

Oxidative stress not only contributes to the progression of OSA but is also linked to its complications. Exposure to IH significantly increases ${\mathrm{O}}_2^{-}$· production in the aorta, activating HIF-1α and promoting the expression of VEGF, MMP2, and MMP9, thereby accelerating the progression of aortic dissection (AD). In vitro experiments further confirm that ${\mathrm{O}}_2^{-}$·-induced HIF-1α expression plays a crucial role in the induction of MMPs, regulated through the PI3 K/AKT/FRAP pathway[80]. Lipid peroxidation products, such as MDA, are elevated in the serum of OSA patients, contributing to an increased incidence of complications. The study found that in an OSA-related rat model, differentially expressed in chondrocytes (DEC1) regulates oxidative stress in pulmonary arterial smooth muscle cells (PASMCs) by modulating ${\mathrm{O}}_2^{-}$·production and pro-inflammatory cytokine expression. This leads to increased levels of MDA, decreased SOD activity, and reduced GSH-Px expression in PASMCs, ultimately influencing their proliferation and apoptosis[25]. These findings suggest that regulating oxidative stress and mitochondrial function, and inhibiting oxidative stress processes, may offer new therapeutic strategies for OSA.

Oxidative stress in otorhinolaryngology: insights from transgenic models

Research into oxidative stress in Otorhinolaryngology diseases has significantly advanced through the use of transgenic animal models, which help elucidate its impact on these conditions (Summary in Table 2). One notable example is the mitochondrial DNA polymerase γ (POLG) mutant mouse model. POLG mutations lead to a loss of mtDNA proofreading ability, resulting in the accumulation of mtDNA mutations over time. This accumulation causes mitochondrial dysfunction and impaired energy metabolism, ultimately leading to cochlear cell apoptosis and early-onset ARHL. Studies have shown that these POLG mutant mice develop hearing loss at 9–10 months of age, highlighting the critical connection between mtDNA mutations and hearing deterioration[8, 81]. In addition, the mitochondrial methyltransferase TFB1M overexpression mouse model demonstrates that hypermethylation of mitochondrial 12S rRNA leads to ROS-dependent activation of AMP-activated protein kinase (AMPK). This activation contributes to the loss of spiral ganglion neurons (SGNs), inner hair cells (IHCs), and outer hair cells (OHCs), underscoring the significant role of ROS in cochlear cell damage[82].

Table 2.

Overview of transgenic animal models of oxidative stress in otorhinolaryngological diseases.

Gene	Animal model	Phenotype	Author
NOX3 knockout Mitochondrial polymerase γ mtTFB1 Fus1 knockout IDH2 knockout P66shc knockout Mn-SOD knockout Cu/Zn SOD knockout OSBPL2 knockout Nrf2 knockout p22phox knockout SOD3 knockout NOX2 knockout SOD2 knockout NRF2 knockout	mouse ARHL、 CIHL、NIHL mouse ARHL mouse ARHL mouse ARHL mouse ARHL mouse NIHL mouse NIHL mouse ARHL zebrafish mouse Rhinosinusitis mouse Rhinosinusitis mouse Allergic inflammation mouse Allergic inflammation mouse CIH mouse CIH mouse CIH	Reduce oxidative damage to outer hair cells, inner hair cells, supporting cells, spiral ganglion cells, and cochlear basal membrane cells Low to mid-frequency hearing loss appearing at 9–10 months of age Loss of spiral ganglion neurons (SGNs), inner hair cell (IHC) synapses, and outer hair cells (OHCs) Severe stria vascularis atrophy, SGN degeneration, and reduced IHC afferent synapses. Spiral ganglion neurons and hair cell loss Reduced oxidative stress and ischemia markers, enhanced auditory trauma tolerance Slightly increase oxidative stress within the cochlea, but not enough to accelerate age-related cochlear damage during physiological aging Promote cochlear hair cell degeneration Elevated ROS and mitochondrial damage in zebrafish inner ear Goblet cell hyperplasia, increased tissue eosinophils, and significantly elevated mucosal IL-13 levels increase susceptibility to sinusitis Leads to excessive increase of epithelial-derived inflammatory cytokines, including IL-33 and TSLP Reducing mite-induced goblet cell hyperplasia and mucus production Worsening OVA-induced airway remodeling and Th2 cell trafficking Reduces atrial Cx40 and Cx43 loss from intermittent hypoxia Increased mtROS, worsened lung remodeling, and heightened oxidative stress Exacerbated intermittent hypoxia-induced neuroinflammation, leading to neuronal apoptosis	Mohri H et al[35]. Someya S et al[81]. Zhao J et al[82]. Tan W. J. T et al[83]. White K et al[84]. Fetoni A.R et al[85]. Kinoshita M et al[86]. McFadden S.L et al[87]. Wang H et al[88]. London NR Jr al[89]. Yao et al[90]. Nagaraj C et al[91]. Kwon M.J et al[92]. Nair D et al[94]. Song J.Q et al[95]. Qiu X et al[96].

Similarly, studies involving Fus1 knockout mice reveal severe cochlear damage associated with oxidative stress. Fus1-deficient mice exhibit degeneration and atrophy of the stria vascularis, SGN degeneration, a reduction in IHC afferent synapses, and chronic cochlear inflammation. These findings highlight Fus1's critical role in cochlear health, emphasizing its function in regulating ROS production and mitochondrial function[83]. The Isocitrate dehydrogenase (IDH2) knockout model further underscores the protective role of antioxidants against oxidative stress. IDH2 is crucial for maintaining mitochondrial integrity by providing NADPH to combat oxidative damage. IDH2 knockout mice exhibit severe cochlear cell loss and hearing impairment, illustrating the importance of IDH2 in protecting against oxidative stress[84]. Additionally, genetic models targeting p66shc and NOX3 reinforce the involvement of ROS in cochlear health. p66shc-deficient mice show increased resistance to noise-induced cochlear damage, while NOX3 knockout mice exhibit reduced ROS levels in the inner ear, which helps mitigate damage from noise exposure and cisplatin treatment[35, 85].

SOD2 homozygous knockout mice exhibit lethality shortly after birth due to severe cardiovascular disease, which arises from mitochondrial dysfunction and elevated oxidative stres. While partial deficiency of SOD2 increases oxidative stress, its impact on hearing during normal aging is relatively modest[86]. In contrast, SOD1 deficiency results in more pronounced hearing loss. Mice lacking SOD1 demonstrate significantly greater hearing impairment and cochlear hair cell degeneration compared to wild-type mice across various ages [87]. Moreover, Oxysterol-binding protein-like 2 (OSBPL2) has been identified as a novel gene associated with autosomal dominant nonsyndromic hearing loss. Although OSBPL2 is primarily known for its role in cholesterol homeostasis, recent studies have shown that OSBPL2 knockout in zebrafish leads to increased reactive oxygen species (ROS) and mitochondrial damage in the inner ear. This finding highlights a significant link between OSBPL2 and oxidative stress in auditory cells[88].

In nasal diseases, oxidative stress plays a crucial role. The transcription factor Nrf2 (Nuclear factor erythroid 2-related factor 2) regulates the expression of antioxidant genes and is critical for managing oxidative stress. In experimental sinusitis models, Nrf2 deficiency leads to goblet cell hyperplasia, increased eosinophil counts, and elevated IL-13 levels, all of which enhance susceptibility to sinusitis[89]. Furthermore, Nrf2 knockout results in excessive increases in IL-33 and TSLP, further exacerbating sinusitis symptoms[89]. Despite the growing recognition of oxidative stress in nasal diseases, the use of transgenic mouse models in this research area remains limited. However, proteomic studies suggest a shared molecular basis between upper and lower airway diseases, indicating potential commonalities in their pathogenesis[90]. Similarly, oxidative stress is implicated in the pathogenesis of allergic asthma. For example, knockout of p22phox reduces goblet cell hyperplasia, mucus production, and airway hyperreactivity induced by house dust mites[91]. Additionally, SOD3 knockout worsens asthma symptoms, while SOD3 supplementation significantly alleviates these symptoms[92]. The voltage-gated proton channel Hv1 is also associated with oxidative stress and allergic inflammation; its deficiency exacerbates oxidative stress levels and aggravates asthma symptoms[93].

Oxidative stress induced by CIH is a significant feature of OSA. NADPH oxidase plays a prominent role in this context. For instance, NOX2 gene knockout alleviates CIH-induced elevations in arterial blood pressure and improves atrial remodeling associated with intermittent hypoxia [94]. Furthermore, SOD2 knockout mice exposed to CIH display increased mitochondrial ROS and aggravated pulmonary remodeling, activating the NLRP3 inflammasome and leading to elevated IL-1β levels and increased caspase-1 activity[95]. NRF2 gene knockout in CIH-induced mouse models exacerbates the inflammatory response in neural cells, resulting in cognitive impairments[96]. Despite the limited use of transgenic models in otorhinolaryngological research, these findings strongly support their potential for advancing treatment strategies for these diseases.

Research progress on ROS as a clinical therapeutic target for otorhinolaryngology

Oxidative stress is a key contributor to the pathogenesis of otorhinolaryngology diseases, involving damage from both one-electron oxidants (e.g. ${\mathrm{O}}_2^{-}$· and ·OH) and two-electron oxidants (e.g. ${\mathrm{H}}_2{\mathrm{O}}_2$). Due to their distinct reactivity, antioxidant design and application require precise targeting strategies. One-electron oxidants are highly reactive, rapidly initiating chain free radical reactions that damage lipid membranes, proteins, and DNA. In contrast, two-electron oxidants primarily oxidize critical cysteine residues, disrupting protein structure and function. Additionally, different oxidants leave distinct ‘footprints’ in disease progression, highlighting the need to tailor antioxidant therapies based on oxidant type, source, and related pathological mechanisms. Antioxidants, including vitamin C, vitamin E, and N-acetylcysteine (NAC), have been explored as primary interventions to counteract oxidative stress. Preliminary clinical trials have demonstrated their potential efficacy in mitigating symptoms related to oxidative stress (see Table 3).

Table 3.

Overview of antioxidant therapy in otorhinolaryngological diseases.

Antioxidant therapy	Disease	Study design	Antioxidant Effects	References
Global antioxidants Vitamin C	ARHL AR	Clinical research	Vitamin C intake, improvement in mid-frequency hearing (2000 and 3000 Hz) Dietary Vitamin C intake, no association with AR	Kang JW et al[97]. García-García C et al[98].
Vitamin E	Otitis Media AR OSA	Clinical research	No effect on the occurrence of chronic suppurative otitis media with or without cholesteatoma. Supplementation with vitamin E (400 IU/d) has no significant effect on perennial AR Vitamin E (800 mg/day) significantly alleviates daily nasal symptoms in patients with seasonal allergic rhinitis. Improves endothelial function, reduces oxidative stress, and enhances sleep parameters	Boesoirie SF et al[100]. Montaño Velázquez BB et al[101]. Shahar E et al[102]. Boppana TK et al[103].
NAC	CIHL SSNHL NIHL AR OSA	Clinical research	Alleviates cisplatin-induced hearing loss Alleviates hearing loss Secondary outcomes and post-hoc analyses indicate that NAC treatment is superior to placebo Reduce nasal symptoms caused by ragweed allergy Improves sleep disorders by attenuating oxidative stress	Orgel E et al[104]. Bai X et al[105]. Kopke R et al[106]. C.J. Lane et al[107]. Wu K et al [109].
Natural Extracts Targeted Antioxidants Resveratrol	NIHL ARHL CRSwNP AR OSA	Animal study (Noise-exposed mouse model) Animal study (age-related mouse model) Animal study (OVA + SEB - induced mouse model) Animal study (OVA - induced mouse model) Animal study (Chronic intermittent hypoxia mouse model)	Improvement of cochlear synaptic loss Significantly reduces the expression of RIPK3, RIPK1, and MLKL during cochlear aging Reducing nasal mucosa eosinophil infiltration and the degree of subepithelial fibrosis Inhibiting the TXNIP oxidative stress pathway to exert anti-allergic and anti-inflammatory effects Mitigating OSA-associated cardiac damage by targeting Nrf2	Yamaguchi T et al[113]. Yang Z et al[114]. Kim S.W et al[115]. Zhang W et al[116]. Sun Z.M et al[117].
Curcumin	ARHL AR OSA	Animal study (age-related mouse model) Clinical research Human study (nasal fibroblasts) Animal study (Chronic intermittent hypoxia mouse model)	Activate the Nrf2 signaling pathway to prevent mitochondrial dysfunction in hair cells Reduced nasal airflow resistance alleviates nasal symptoms (sneezing and runny nose) and nasal congestion. Activating the Nrf2/HO-1 pathway inhibits oxidative stress in nasal fibroblasts induced by particulate matter. Inhibiting AQP4 and p38 MAPK pathways alleviates brain injury caused by chronic intermittent hypoxia	Li N et al[119]. Wu S et al[120]. Kim J.S et al[121]. Wang B et al[122].
Quercetin	NIHL Otitis Media AR CRS	Animal study (Noise-exposed guinea pig model) Animal study (NTHi induced mouse model) Animal study (OVA - induced mouse model) Animal study (TDI - induced mouse model) Animal and Human study (HSNE and MNSE)	Prevent hair cell loss in the cochlea of guinea pigs Inhibit IKKβ phosphorylation and p38 MAPK to block CXCR4 activation. Improving the imbalance between Th1/Th2 cells and Treg/Th17 cells. Inhibiting nasal rubbing and sneezing in allergic rhinitis (AR) rats Increasing chloride ion transport and ciliary beat frequency in nasal epithelial cells	Hirose, Y et al[123]. Ma Y.K et al[124]. Ke X et al[125]. Kashiwabara M et al[126]. Zhang S et al[127].
Sulforaphane	CIHL AR CRS OSA	Animal study (cisplatin -induced rat model Human study (HSNECs) Human study (SEC) Animal study (Chronic intermittent hypoxia mouse model)	Reduced cisplatin-induced loss of outer hair cells and restored ciliary morphology Restored ZO-1 levels reduced by HDM after activating Nrf2 Suppress apoptosis of sinonasal epithelial cells induced by levofloxacin Activate the Nrf2 pathway to enhance SOD activity and reduce MDA levels to improve cognitive dysfunction Nrf2 functions to reduce cardiac fibrosis and inflammation induced by intermittent hypoxia	Wang J et al[128]. London N.R.Jr et al[129]. Kohanski M.A et al[130]. Qiu X et al. Li X et al. Wang J et al[96, 131].
Catechins	CIHL AR	Animal study (Aminoglycoside-induced Zebrafish model) cisplatin -induced Rat model Animal study (OVA - induced mouse model)	Reduces oxidative stress and apoptosis, significantly protecting inner ear hair cells Reduces oxidative stress and apoptosis markers while preserving the antitumor effects of cisplatin Reduces serum IgE levels, COX-2, IL-1β, IL-4, and IL-6 mRNA and protein expression in nasal mucosa Restores the balance between Th2 and Th1 cell types	Zong Y et al[132]. Borse V et al[133]. Fu M et al. Pan Z et al[134, 135].

Vitamin C and vitamin E

Vitamin C, a water-soluble antioxidant, neutralizes free radicals, reduces oxidative stress, and supports immune function and tissue repair. Research involving individuals aged 50–80 has shown that vitamin C intake is associated with improvements in mid-frequency hearing (2000 and 3000 Hz) [97]. However, a study with 6,293 adults aged 20–49 found no significant effects of vitamin C on asthma and allergic rhinitis, although it was linked to a reduced risk of asthma under high-inflammatory conditions[98]. Intravenous vitamin C supplementation has also demonstrated some beneficial effects in alleviating allergy symptoms[99]. Vitamin E, a fat-soluble antioxidant, mitigates damage from ROS and free radicals. While evidence suggests limited effects of vitamin E on chronic suppurative otitis media and perennial allergic rhinitis[100], supplementation with vitamin E (400 IU/d) has no significant effect on perennial AR[101]. However, high doses (800 mg/day) have been found to significantly reduce nasal and ocular symptoms in patients with seasonal allergic rhinitis [102]. A systematic review and meta-analysis of six studies (five randomized controlled trials and one case – control study) indicated that vitamin C and vitamin E might improve endothelial function, reduce oxidative stress, and enhance sleep parameters[103]. Despite the potential benefits of vitamin C and vitamin E in managing otorhinolaryngological disease symptoms, research findings remain inconsistent. Therefore, further large-scale, long-term clinical trials are necessary to confirm their efficacy and safety, providing a more robust scientific foundation for their use in treating otorhinolaryngological diseases.

N-acetylcysteine (NAC)

NAC, a precursor to glutathione, reduces oxidative stress and has shown potential therapeutic benefits in various otorhinolaryngological diseases. Preliminary studies indicate that intravenous NAC may lower the risk of CIHL in pediatric patients undergoing cisplatin therapy [104]. Additionally, oral NAC combined with dexamethasone has demonstrated superior efficacy in treating sudden sensorineural hearing loss (SSNHL) compared to dexamethasone alone, with significant improvement in hearing, particularly at the 8000 Hz frequency[105]. However, a meta-analysis involving 1,197 participants found no significant overall effect of NAC on sensorineural hearing loss, with benefits observed only in SSNHL cases. NAC showed no preventive effect against CIHL or NIHL. Clinical trials investigating NIHL in military personnel indicated that while secondary outcomes and post-hoc analyses suggested NAC's superiority over placebo, the study design lacked the rigor needed to definitively confirm its efficacy. This underscores the necessity for larger, more robustly designed trials to more accurately assess the protective effects of NAC against hearing loss[106]. In nasal diseases, NAC has seen limited application, with most studies conducted in animal models. Early clinical trials suggested that topical NAC reduced allergic responses to ragweed extract[107]. In animal studies, NAC was shown to alleviate allergen-induced nasal symptoms in allergic rhinitis by reducing oxidative stress in the nasal mucosa[108]. NAC also holds potential in addressing complications of OSA, such as hypertension, stroke, and cardiovascular diseases. Small randomized trials suggest some therapeutic effects of NAC in OSA, including reduced sympathetic overactivity induced by CIH[109]. Despite these promising results, NAC’s antioxidant and anti-inflammatory properties warrant further investigation. Larger, well-designed, multicenter clinical trials are needed to conclusively establish its efficacy and safety in the treatment of otorhinolaryngological diseases.

In otorhinolaryngological diseases, both one-electron oxidants and two-electron oxidants play distinct roles in disease progression, necessitating targeted antioxidant strategies. One-electron oxidants, due to their high reactivity, induce lipid peroxidation, protein oxidation, and DNA damage, contributing to sensory hair cell apoptosis and mitochondrial dysfunction. In contrast, two-electron oxidants oxidize cysteine residues, activating inflammatory pathways such as NF-κB and exacerbating chronic inflammation. Targeted therapies against specific oxidants have shown promise. NAC enhances endogenous antioxidant defenses by replenishing glutathione (GSH), thereby reducing oxidative damage from ${\mathrm{O}}_2^{-}$· and related free radicals, and improving hair cell survival and auditory function [104, 110]. For two-electron oxidants like ${\mathrm{H}}_2{\mathrm{O}}_2$, vitamin C directly scavenges ${\mathrm{H}}_2{\mathrm{O}}_2$ and upregulates antioxidant enzymes such as glutathione peroxidase (GPx) and catalase (CAT), effectively reducing oxidative stress, inflammation, and epithelial barrier dysfunction in AR[98]. Additionally, vitamin E, a lipid-soluble antioxidant, protects cell membranes by interrupting lipid peroxidation chain reactions [111]. These targeted antioxidant strategies offer a promising approach for precision therapy in otorhinolaryngological diseases.

Natural extracts as targeted antioxidants

In the research and treatment of otorhinolaryngological diseases, natural extract-derived antioxidants have gained increasing attention due to their diverse bioactive properties and comparatively lower side effect profiles. These compounds are emerging as promising candidates for the management of otorhinolaryngological conditions. This article reviews several natural antioxidants with widespread application in the otorhinolaryngological field, highlighting their mechanisms of action (see Table 1).

Resveratrol, a polyphenol found in grape skins, blueberries, and peanuts, is celebrated for its powerful antioxidant and anti-inflammatory properties. Several studies have evaluated the antioxidant properties of resveratrol in the context of otorhinolaryngological diseases. For noise-induced hearing loss, resveratrol has demonstrated protective effects. One study found that resveratrol effectively prevents NIHL[112]. Further animal studies have revealed that resveratrol treatment prevents noise-induced cochlear synapse loss and sensorineural hearing loss[113]. Additionally, resveratrol has shown beneficial effects against drug-induced hearing loss and age-related hearing loss[114]. Regarding nasal diseases, preliminary studies have shown that resveratrol significantly reduces eosinophil infiltration and subepithelial fibrosis in the CRSwNP mouse model, with effects comparable to those of budesonide, suggesting its potential in preventing CRSwNP[115]. In animal models of AR, resveratrol lowers oxidative stress markers such as malondialdehyde (MDA) and exerts anti-allergic and anti-inflammatory effects by inhibiting the TXNIP pathway[116]. Furthermore, resveratrol offers myocardial protection against CIH-induced damage by targeting Nrf2 and inhibiting the activation of the NLRP3 inflammasome[117].

Curcumin, a bioactive compound extracted from turmeric, is widely recognized for its antioxidant, anti-inflammatory, and anticancer effects. In the context of hearing loss, clinical research has demonstrated that daily supplementation with curcumin significantly improves hearing in patients with chronic kidney disease-induced hearing loss[118]. Additionally, curcumin helps prevent mitochondrial dysfunction in cochlear hair cells by activating the Nrf2 signaling pathway, thereby mitigating NIHL[119]. Curcumin also shows promise for the treatment of nasal diseases. A randomized, double-blind trial with 241 patients with AR found that oral curcumin supplementation reduced nasal airflow resistance, alleviated nasal symptoms, and inhibited the production of inflammatory cytokines such as IL-4, IL-8, and TNF-α[120]. Moreover, Curcumin can inhibit oxidative stress in human nasal fibroblasts exposed to urban particulate matter by activating the Nrf2/HO-1 pathway, suggesting its potential as a treatment for particulate matter-induced nasal diseases[121]. Additionally, curcumin may protect against brain damage caused by chronic intermittent hypoxia by inhibiting the AQP4 and p38 MAPK pathways[122].

Quercetin, a flavonoid prevalent in fruits and vegetables, is acclaimed for its strong antioxidant and anti-inflammatory capabilities. Research has highlighted several beneficial effects of quercetin in otorhinolaryngological diseases. It has been shown to prevent hair cell loss in the lateral line of zebrafish and the cochlea of guinea pigs [123]. Additionally, quercetin directly inhibits the phosphorylation of IKKβ and suppresses p38 MAPK, thereby blocking CXCR4 activation and reducing inflammation in a mouse model of otitis media[124]. In an AR mouse model, oral quercetin reduced serum levels of IgE, IgG1, and histamine, corrected Th1/Th2 cell imbalances, and alleviated AR symptoms[125]. Furthermore, Oral administration of quercetin for 5 and 7 days effectively reduces nasal rubbing and sneezing in AR rats, suggesting its potential as a supplementary treatment for managing AR[126]. Moreover, research indicates that quercetin can enhance chloride ion transport and ciliary beating frequency, which points to its potential in improving mucociliary clearance in patients with CRS[127]. These findings underscore quercetin's promise as a therapeutic agent for various otorhinolaryngological conditions, although additional research is required to confirm its clinical efficacy.

Sulforaphane, an isothiocyanate present in cruciferous vegetables, is well-regarded for its potent antioxidant and anti-inflammatory properties. Research on tubby mice has demonstrated that sulforaphane improves cochlear cell loss and alleviates cochlear degeneration. Additionally, sulforaphane mitigates cisplatin-induced loss of outer hair cells, restores ciliary morphology, and improves hearing loss[128]. It has also been shown to restore epithelial zonula occludens-1 (ZO-1) levels under house dust mite stimulation by activating Nrf2, thereby enhancing nasal epithelial barrier function[129]. A clinical trial further revealed that sulforaphane significantly reduces Th2 cytokines such as IL-4, IL-5, and IL-13 in nasal mucus of AR patients. Another study found that sulforaphane pre-treatment inhibits apoptosis induced by levofloxacin, particularly in sinus epithelial cells from sinus surgery patients, highlighting its antioxidant properties[130]. In the context of OSA, sulforaphane activates the Nrf2 pathway, increases SOD activity, and reduces MDA levels, thereby decreasing cardiac fibrosis and oxidative damage caused by IH[96, 131].

Catechins, primarily found in tea, are renowned for their potent antioxidant and anti-inflammatory properties, with epigallocatechin gallate (EGCG) being the most active component. In a zebrafish model, EGCG has been shown to significantly protect inner ear hair cells from aminoglycoside-induced ototoxic damage by reducing oxidative stress[132]. Additionally, EGCG mitigates cisplatin-induced hearing loss by lowering oxidative stress and apoptosis markers in cochlear cells without impacting the antitumor efficacy of cisplatin[133]. In nasal diseases, catechins exhibit substantial antioxidant effects. EGCG treatment in AR mice significantly reduces sneezing and nasal rubbing, lowers serum levels of IgE and histamine, and decreases IL-1β, IL-4, and IL-6 levels in nasal lavage fluid[134]. Furthermore, catechins can dose-dependently alleviate allergic symptoms such as sneezing and nasal rubbing in AR mice, reduce serum IL-5 and IL-13 levels via the NF-κB/TSLP pathway, and restore the balance of Th2/Th1 cells[135].

Natural extracts have demonstrated dual antioxidant effects in otorhinolaryngological diseases, targeting both one-electron and two-electron oxidants through a combination of scavenging oxidants and modulating the endogenous antioxidant system. For example, quercetin not only inhibits NADPH oxidase activity to reduce ${\mathrm{O}}_2^{-}$· production but also alleviates oxidative stress-induced inflammation by regulating ROS-related NF-κB signaling, exhibiting both antioxidant and anti-inflammatory properties [111]. Similarly, resveratrol directly scavenges ${\mathrm{H}}_2{\mathrm{O}}_2$, reducing ROS levels in nasal epithelial cells, while regulating mitochondrial antioxidant enzymes like SOD2 to reduce MDA levels, thus mitigating oxidative stress and inflammation [116, 136]. These dual mechanisms enable natural antioxidants to target both one-electron and two-electron oxidants, providing a multi-layered protective effect and laying a solid theoretical foundation for precision antioxidant therapies in otorhinolaryngological diseases.

Discussion

In recent years, innovative strategies targeting ROS regulation have shown significant potential in the research and treatment of otorhinolaryngological diseases. Research primarily focuses on precise modulation of ROS production sources, such as the NOX family and mitochondria, as well as enhancing antioxidant enzyme systems. The NOX family is central to ROS generation. NOX2, predominantly expressed in immune cells, produces ${\mathrm{O}}_2^{-}$· that activate the NF-κB signaling pathway, inducing pro-inflammatory cytokines like IL-6 and TNF-α, exacerbating acute inflammation, and damaging the mucosal barrier [17]. Inhibiting NOX2 with small molecule inhibitors or RNA interference has shown notable anti-inflammatory effects in animal models, indicating therapeutic potential for acute inflammatory diseases[137]. NOX4 is a major driver of chronic inflammation and fibrosis, continuously generating ${\mathrm{H}}_2{\mathrm{O}}_2$ to activate the TGF-β1 signaling pathway, promoting fibrosis and epithelial cell proliferation [137]. Targeted inhibition of NOX4 with GKT137831 effectively reduces inflammation in allergic rhinitis, offering potential for chronic disease treatment[138]. NOX3, specifically expressed in the inner ear, is the primary source of cochlear ROS. The ${\mathrm{O}}_2^{-}$· generated by NOX3 induces apoptosis of outer hair cells in noise-induced hearing loss and drug-induced ototoxicity [35]. Inhibiting NOX3 significantly reduces ROS levels, protecting hair cells from oxidative stress and providing new therapeutic directions for hearing disorders[139]. DUOX2, highly expressed in nasal epithelial cells, generates ${\mathrm{H}}_2{\mathrm{O}}_2$ to maintain mucosal antimicrobial defense under normal conditions. However, in allergic rhinitis, DUOX2 overactivation elevates ${\mathrm{H}}_2{\mathrm{O}}_2$ levels, triggering Th2-type inflammation and worsening local inflammation. Gene interventions or small molecule inhibitors targeting DUOX2 effectively alleviate these pathological processes [7].

Mitochondria are another critical ROS source, producing ${\mathrm{O}}_2^{-}$· and ${\mathrm{H}}_2{\mathrm{O}}_2$ through the electron transport chain (ETC). Mitochondrial dysfunction leads to excessive ROS accumulation, inducing cytochrome c release, activating apoptotic pathways, and amplifying inflammation via NF-κB and NLRP3 pathways. Mitochondria-targeted antioxidants, such as MitoQ and MitoTEMPO, clear mitochondrial ROS, reduce cellular damage, and protect hair cells from oxidative stress [140]. Enhancing mitophagy to eliminate damaged mitochondria, such as activating the PINK1/Parkin pathway, also shows therapeutic potential in mitigating noise-induced inner ear damage [141].

Beyond inhibiting ROS production, boosting antioxidant enzyme systems is equally crucial. Activating the Nrf2 signaling pathway upregulates SOD, CAT, and GPx expression, effectively reducing ROS accumulation and inflammation [142]. Gene therapy approaches, such as using CRISPR/Cas9 to enhance SOD2 activity in chronic rhinosinusitis models, not only clear mitochondrial ROS but also restore epithelial redox balance[95, 143].

Future antioxidant therapies for otorhinolaryngological diseases will focus on precise ROS regulation, enhancement of antioxidant enzyme systems, and personalized, multi-target strategies. Selective inhibition of specific ROS sources, such as NOX isoforms (NOX2, NOX3, NOX4), combined with nanoparticle or liposomal delivery systems, may improve drug targeting and tissue penetration. Mitochondrial ROS regulation can be optimized using mitochondria-targeted antioxidants and promoting mitophagy. Enhancing antioxidant enzyme function by activating the Nrf2 pathway to upregulate SOD, CAT, and GPx, or employing CRISPR/Cas9 to boost enzyme activity, enables efficient ROS clearance and redox balance restoration. Multi-target therapies combining NOX inhibitors, antioxidants, and anti-inflammatory agents may simultaneously reduce ROS levels, inflammation, and tissue damage.

Developing real-time ROS detection technologies is equally crucial. Advanced imaging techniques, such as fluorescence microscopy and high-resolution mass spectrometry, alongside biomarkers like MDA and 4-HNE, allow dynamic ROS monitoring and identification of disease-specific characteristics. Research should further investigate the balance between ROS signaling and oxidative stress, focusing on their transition from physiological signaling molecules to drivers of oxidative damage. Understanding ROS localization within organelles like mitochondria and the endoplasmic reticulum can reveal tissue-specific mechanisms. Translating these findings into clinical applications is essential. ROS signatures could serve as biomarkers for early diagnosis and treatment optimization, with NOX-derived ROS levels aiding in classifying chronic sinusitis subtypes and guiding therapies. Dynamic ROS monitoring tools could also assess antioxidant therapy efficacy. By integrating precise detection, targeted interventions, and dynamic monitoring, antioxidant therapies hold great potential to advance precision medicine for otorhinolaryngological diseases.

Author contributions

Shi L was involved in the conception of the study, revision of the paper writing. Meng L wrote the first draft of the manuscript. Liu S, Luo J, Tu Y, Li T, Yu J were involved in the literature search, as well as in reviewing and editing the manuscript.

Funding Statement

This work was supported by National Natural Science Foundation of China: [grant Number 82371117].

Disclosure statement

No potential conflict of interest was reported by the author(s).