The human body contains between 50 and 120 mg of copper (Cu), with approximately 70% of it localized in muscle and bone tissues (Olivares and Uauy, 1996; Turnlund et al., 1998). In bone tissue, the physiological concentration of Cu is typically 5–6 mg/kg dry weight. Copper is essential for regulating collagen production, bone and cartilage matrix metabolism, and remodeling processes. It also enhances bone density and strength, promoting normal bone development and maintaining bone and cartilage homeostasis (Wang et al., 2016; Baino et al., 2018; Antoniadis et al., 2019; Ciosek et al., 2023). This paper will explore Cu’s regulatory effects on various cellular components within bone and joint tissues.

A follow-up study has suggested that copper deficiency might be an overlooked diagnostic factor, as it can significantly affect patient health by altering various metabolic indicators (Fujikawa and Haruta, 2023). Copper deficiency has widespread consequences on cellular and systemic metabolism due to its impact on the function of several essential enzymes, potentially leading to pathological conditions such as Menkes disease (Guthrie et al., 2020). Within the context of bone and cartilage, copper deficiency primarily results in detrimental effects due to the impaired function of copper-dependent enzymes (Sarazin et al., 2000), direct damage to bone and chondrocytes (Alshenibr et al., 2017; Liu et al., 2022), and decreased anti-inflammatoryand antioxidant defenses (Gaffney-Stomberg, 2019; Pantic et al., 2023).

Copper deficiency adversely affects the function of copper-dependent enzymes, including lysyl oxidase (LOX) (Vallet and Ricard-Blum, 2019), ceruloplasmin (CP) (Yiannikourides and Latunde-Dada, 2019), and superoxide dismutase (SOD) (Sun et al., 2024). The diminished activity of LOX severely disrupts collagen cross-linking, undermining the stability of the cartilage extracellular matrix and reducing bone strength (Sarazin et al., 2000; Alshenibr et al., 2017; Liu et al., 2022). Additionally, decreased SOD activity, a key enzyme in combating oxidative stress, results in a diminished capacity to clear reactive oxygen species (ROS) in bone and cartilage, leading to oxidative damage and subsequent deterioration of these tissues (Pantic et al., 2023).Furthermore, dietary copper deficiency exacerbates oxidative stress by reducing NRF2/HO-1 activity (Pan et al., 2024). From an osteogenic perspective, copper deficiency weakens bone integrity by depleting intracellular glutathione (GSH) (Bernhardt et al., 2021)and directly inhibiting osteoblast activity, thereby impairing bone remodeling (Pantic et al., 2023), while having minimal impact on osteoclasts (de Romaña et al., 2011; Tokuda et al., 2013). Collectively, these processes increase the susceptibility of bone and articular cartilage, heightening the risk of developing OA (Medeiros, 2016).

Moreover, copper deficiency influences iron metabolism by decreasing CP activity (Broderius et al., 2010; Yiannikourides and Latunde-Dada, 2019), leading to reduced extracellular iron excretion and intracellular iron overload (Miyajima, 2015; Zheng et al., 2018; Villadsen et al., 2023). This iron accumulation disrupts lipid metabolism (Ben-Hamouda et al., 2017; Raia et al., 2023) and heightens the risk of ferroptosis, a form of iron-induced cell death, particularly in chondrocytes. As ferroptosis contributes to OA development, exploring the interplay between copper deficiency, CP, iron metabolism, and cell death could offer a novel therapeutic approach to treating OA.

Research indicates that copper concentrations in both serum and joint fluid are markedly elevated in patients with OA compared to normal levels (Yazar et al., 2005; Yang et al., 2023), with degenerated cartilage showing higher copper content than healthy cartilage (Li et al., 2023c). These findings suggest a strong association between excessive copper and the development of OA, implicating it in cartilage damage (Guan et al., 2023; Li S. et al., 2023). In vitro studies of chondrocytes cultured with excess copper reveal abnormal cytoskeletal morphology, including altered microfilaments, microtubules, and fibers, as well as the inactivation of integrin-linked kinases (Zou et al., 2018), suggesting an aberrant state of chondrocytes (Terpstra et al., 2003). Elevated copper levels are also linked to the promotion of inflammatory states, which may exacerbate arthritis pathogenesis (Wang W. et al., 2023).Excess copper disrupts cellular function by competitively inhibiting the binding of cofactors to enzyme proteins, leading to redox imbalances (Guerra et al., 2023). In bone and cartilage, elevated copper levels damage osteoblasts, osteoclasts, and chondrocytes (León et al., 2014; Schamel et al., 2017; Bernhardt et al., 2021). Additionally, data have been compiled on the copper concentrations that induce damage to chondrocytes, osteoblasts, osteoclasts, macrophages, and bone marrow stromal cells (BMSCs) (Figure 1)

In osteoblasts, osteoclasts, and bone marrow stromal cells (BMSCs), excessive copper stabilizes the conformation of hypoxia-inducible factor 1-alpha (HIF-1α), leading to tissue hypoxia (Stelling et al., 2019). This condition downregulates the expression of Runt-related transcription factor 2 (Runx2) and decreases the levels of collagen type I, osteocalcin (OCN), insulin-like growth factor I (IGF-I), and bone morphogenetic protein 2 (BMP-2) (Qi et al., 2021). These molecular changes collectively impair the proliferative activity, osteogenic function, and osteogenic differentiation potential of osteoblasts. Additionally, excessive copper directly inhibits osteoclast function (Zofková et al., 2013; Bernhardt et al., 2021), thereby disrupting the balance between osteogenesis and osteoclast activity (Li et al., 2014). In chondrocytes, elevated copper levels suppress antioxidant enzymes, particularly superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), significantly reducing the cells’ antioxidant defense capacity. This suppression may facilitate the development of OA by promoting oxidative stress and related damage. Moreover, excessive copper induces a senescent phenotype in chondrocytes by impairing extracellular matrix synthesis (Massie et al., 1990).

Metallothioneins (MT-1 and MT-2), components of the metallothionein family, are stably expressed across various tissues. The expression of MT1/2 can be upregulated in response to stimuli such as copper excess, oxidative stress, and inflammation (Wake and Mercer, 1985), enabling the sequestration of excess copper ions for detoxification (Freedman and Peisach, 1989). The regulation of MTs may involve upstream factors such as hypoxia-inducible factor 2-alpha (HIF-2α), nicotinamide phosphoribosyltransferase (NAMPT), and the zinc transporter ZIP8-MTF1 axis (Park et al., 2023). This represents a cellular negative feedback mechanism against copper overload, establishing a critical link between copper and zinc metabolism (Park et al., 2023). Both metallothioneins (MTs) and glutathione (GSH) have significant potential in mitigating oxidative stress. Therefore, further exploration of copper metabolism and its related pathways may uncover potential therapeutic targets for treating related diseases, offering novel insights for clinical interventions.

Recent studies have identified a novel mode of programmed cell death induced by copper ions, referred to as “cuproptosis,” which may involve apoptosis, cysteine protease activity, or the accumulation of reactive oxygen species (ROS). This copper-mediated cell death, characterized by multiple factors and pathways, differs significantly from other forms of cell death, such as apoptosis and pyroptosis (Tsvetkov et al., 2022). Initially, excess copper binds to mitochondrial lipoic acid-related protein components (e.g., DLAT) (Rowland et al., 2018), leading to the formation of insoluble DLAT aggregates in mitochondria and the loss of FDX1-dependent Fe-S clusters. This process results in the oligomerization of DLAT and reduced Fe-S cluster stability, disrupting the tricarboxylic acid (TCA) cycle and triggering cell death through a protein toxicity-related stress response. Ferredoxin 1 (FDX1), identified as the central regulatory gene in copper-induced cell death, modulates this process by influencing the TCA cycle, specifically through the regulation of DLAT thioctyl acylation (Tsvetkov et al., 2022). Moreover, an excess of Cu⁺ ions can generate a substantial quantity of ROS through a mechanism similar to the Fenton reaction involving hydrogen peroxide (Ma et al., 2019), exacerbating oxidative stress-induced cellular damage. Elevated copper levels have been shown to activate the ROS/HO-1/NQO1 and NRF2/HO-1 pathways while concurrently inhibiting P38-MAPK-related signaling pathways (Fang et al., 2021a). ROS-induced damage to Fe-S cluster proteins, particularly the cyclical transformation between [4Fe-4S] and [2Fe-2S] clusters, is essential for maintaining equilibrium and adapting to changes in the redox environment (Read et al., 2021). However, substantial Cu⁺ reduction by histone H3/H4 in the nucleus further compromises Fe-S cluster proteins through ROS generation via a Fenton-like reaction.Proteins such as Ssq1, Jac1, and Nfs1 are integral to the assembly, maturation, and repair of mitochondrial Fe-S proteins. Copper-mediated superoxide dismutase (SOD) loss results in damage to these Fe-S cluster proteins (Strain et al., 1998). Additionally, copper can directly bind to glutathione (GSH), and excess copper leads to GSH depletion, a reduction in glutathione peroxidase 4 (GPX4) activity, and a subsequent decrease in cellular antioxidant capacity. Furthermore, excessive copper levels may enhance susceptibility to ferroptosis by affecting associated pathways, thereby exacerbating the progression of OA (Nong et al., 2023).In the subsequent sections, a comprehensive analysis will be conducted on how dysregulated copper metabolism influences ferroptosis sensitivity. Additionally, the interconnections between abnormal copper metabolism, cuproptosis, and ferroptosis will be elucidated.

Recent genome-wide CRISPR/Cas9 screenings have identified seven genes upregulated in copper ion-induced cell death: ferredoxin 1 (FDX1), lipoyl transferase 1 (LIPT1), lipoic acid synthetase (LIAS), dihydrolipoic acid dehydrogenase (DLD), dihydrolipoic acid S-acetyltransferase (DLAT), pyruvate dehydrogenase E1 alpha subunit (PDHA1), and pyruvate dehydrogenase E1 beta subunit (PDHB). Simultaneously, three genes—metal-regulated transcription factor 1 (MTF1), glutaminase (GLS), and cyclin-dependent kinase inhibitor 2A (CDKN2A)—were downregulated. The regulatory interplay of these genes with copper metabolism offers insights into their roles in OA (Figure 2) (Chang et al., 2023).

Fron-induced cell death, known as ferroptosis, is an iron-dependent mechanism characterized by the abnormal accumulation of lipid peroxides and their derivatives. Elevated levels of lipid peroxides, such as malondialdehyde (MDA) and the pro-inflammatory molecule 4-hydroxynonenal (4HNE), accumulate on cellular membranes and organelles, leading to membrane damage, rupture, and ultimately cell death (Chen X. et al., 2021). The Fenton reaction accelerates lipid peroxidation chain reactions, and under normal physiological conditions, these lipid peroxidation products are mitigated by the cell’s intrinsic reducing mechanisms, with selenium-dependent glutathione peroxidase 4 (GPX4) playing a critical role (Yang et al., 2020; Liu Y. et al., 2021). Glutathione (GSH) is a critical cofactor for GPX4 in this context. The reduction pathway also involves the SLC3A2 and SLC7A11 membrane transporters, essential for cystine uptake and subsequent GSH synthesis. Notably, Botulinum toxin A (Zeng et al., 2024) and Ruscogenin (Ruan et al., 2024), a bioactive compound from Radix Ophiopogon japonicus, have been shown to inhibit ferroptosis in chondrocytes via the SLC7A11/GPX4 pathway. While the GPX4 system is predominant, it is not the sole pathway; GPX4-independent mechanisms, such as the FSP1/CoQ10, DHODH, and GCH1 (GTP cyclohydrolase 1)/BH4 (tetrahydrobiopterin) pathways, also play significant roles (Bersuker et al., 2019; Gan, 2021; Yang H. et al., 2022). Moreover, elements like transferrin, the transferrin receptor, metal reductase 3, and FTH-1 are integral to the iron REDOX metabolic pathway (Park and Chung, 2019; Fang et al., 2021b; Yang M. et al., 2022; Ye et al., 2022). Mitochondrial dysfunction, due to its high iron content, is central to the pathogenesis of ferroptosis (Yang F. et al., 2022). This dysfunction exacerbates lipid peroxidation through mechanisms such as oxidative respiratory chain abnormalities, cysteine depletion, mitochondrial dihydroorotate dehydrogenase (DHODH) inhibition, and mitochondrial coenzyme Q10 (CoQ10) inhibition. As a prototypical ferroptosis inhibitor, nuclear factor erythroid 2-related factor 2 (NRF2) can upregulate various antioxidant enzymes, including heme oxygenase-1 (HO-1) and GPX4 (Lin Q. et al., 2023).

Ferroptosis is implicated in a wide range of physiological and pathological processes. The well-known tumor suppressor gene P53 can inhibit the transport of glutathione (GSH) precursors by downregulating the expression of SLC7A11 in the System Xc-transporter, thereby inhibiting GPX4 and inducing ferroptosis in cancer cells (Shao et al., 2022). Recent research has shown that P21 can stabilize GPX4 by modulating its recruitment to the linear ubiquitin chain assembly complex (LUBAC) and influencing the level of GPX4 M1-linked ubiquitination. Functionally, the knockdown of P21 enhances the sensitivity of chondrocytes to Erastin-induced ferroptosis, decreases Col2A1 expression, and increases MMP13 expression (Zheng et al., 2024). From an inflammatory perspective, both IL-1β and TNF-α have been shown to induce and enhance the expression of transferrin and its receptor, thereby increasing intracellular iron transport. Consequently, this inflammatory environment creates conditions that favor ferroptosis (Feng et al., 2020). This observation has led to investigations into the role of ferroptosis in the pathogenesis of OA. In the following sections, the relationship between ferroptosis and OA chondrocytes will be examined.

A study examining the concentration of iron in articular fluid among patients with OA compared to healthy individuals revealed a significant elevation of iron levels in the articular fluid of those with OA (Yazar et al., 2005). Further research involving the knockout of the HFE gene, which regulates iron (Simão et al., 2019), and the observed increase in serum ferritin levels (Nugzar et al., 2018) has also established a link between iron dysregulation and OA. Moreover, prior research has demonstrated that chondrocyte ferroptosis contributes to the progression of OA, as evidenced by both in vivo and in vitro experiments. This phenomenon is not solely due to exposure to ferric ammonium citrate (FAC), which induces iron overload and subsequent ferroptosis. Notably, chondrocyte ferroptosis has been observed in a prototypical IL-1β-induced inflammation model of primary mouse chondrocytes. In this model, expression levels of type II collagen, System Xc, and GPX4 protein decreased, while expression of MMP13, ACSL4, and P53 increased (Yao et al., 2021). Recent in vitro experiments have shown that gossypol acetic acid (GAA) induces resistance to ferroptosis in chondrocytes within OA rat models by inhibiting GPX4 methylation (Shang et al., 2024b).

Fer-1, a lipophilic antioxidant, has been effective in mitigating Erastin-induced ferroptosis by clearing lipid peroxides (Dixon et al., 2012). Conversely, research indicates that desferrioxamine (DFO), an iron chelator, inhibits the development of IL-1β-induced chondrocyte models and slows disease progression in OA mouse models (Guo et al., 2022). DFO has been shown to inhibit ferroptosis through iron chelation, similar to the mechanism of Ferrostatin-1 (Fer-1) in mitigating lipid peroxidation. Additionally, both DFO and Fer-1 were found to enhance the expression of GPX4, HO-1, and other antioxidant enzymes by promoting the nuclear translocation of NRF2 in chondrocytes. This action reduces chondrocyte injury in OA mouse models, increases type II collagen expression, and decreases MMP13 levels (Guo et al., 2022). NF-κB, a key regulator of inflammatory processes, plays a significant role in OA pathogenesis by mechanisms involving both inflammation and ferroptosis. Upon activation, NF-κB induces the expression of HIF-2α and exacerbates ferroptotic activity by downregulating glutathione (GSH) levels and directly inhibiting the activity of GPX4. Furthermore, D-mannose has been shown to protect chondrocytes against ferroptosis by downregulating HIF-2α and promoting GPX4 expression (Zhou et al., 2021; 2022).

The transient receptor potential vanilloid 1 (TRPV1) has emerged as a promising therapeutic target in OA management. TRPV1 activation has been found to inhibit the ferroptosis pathway in chondrocytes, thereby decelerating OA progression. In tandem with this, innovative therapeutic strategies have been developed, incorporating TRPV1-targeting nanomaterials, which have demonstrated substantial efficacy in vivo (Li et al., 2024; Lv et al., 2024). A clinical study identified heightened ferroptosis in chondrocytes located in weight-bearing regions of the joint, implicating mechanical stress as a key driver of OA pathology (Wang et al., 2022). Excessive mechanical stress initiates catabolic processes in chondrocytes and disrupts glutathione (GSH) synthesis viacalcium influx through mechanically activated Piezo channels, ultimately impairing glutathione peroxidase 4 (GPX4) activity and promoting ferroptosis. Furthermore, in vitro studies using IL-1β-induced chondrocyte models revealed that moderate mechanical stress activates the NRF2/GPX4/HO-1 axis while inhibiting the NF-κB P65 pathway, thus alleviating chondrocyte inflammation and ferroptosis (Han J. et al., 2024b). This evidence underscores the dualistic effect of mechanical stress on chondrocyte ferroptosis and provides a promising foundation for future research and therapeutic advancements.

Transferrin receptor 1 (TFR1), a transmembrane glycoprotein widely expressed across various tissue cells, plays a pivotal role in binding and facilitating the cellular uptake of ferric ions, which are subsequently reduced to ferrous ions by metal reductase 3. Recent studies have demonstrated that modulating TFR1 expression can influence ferroptosis inOA chondrocytes by regulating iron homeostasis. In a murine model of OA established through DMM surgery, researchers observed a significant increase in the positive rate of TFR1 expression within the model group (Wang W. et al., 2024). Further in vivo experiments using IL-1β-induced primary chondrocytes from mice revealed a dose-dependent increase in TFR1 expression, establishing a positive correlation between TFR1 expression and OA progression.Knockout of the TFR1 gene led to reduced reactive oxygen species (ROS) levels, increased expression of ferroptosis markers GPX4 and SLC7A11 (Wang J. et al., 2024), decreased expression of cartilage catabolic markers MMP3 and MMP13, and upregulation of anabolic markers COL2A1 and SOX9. Mitochondrial dysfunction, which plays a central role in iron metabolism, can trigger both inflammatory changes and ferroptosis in chondrocytes. Mitochondrial damage, along with the release of mitochondrial DNA (mtDNA) into the cytoplasm, can activate inflammatory responses via the cGAS-STING signaling pathway (Dela Cruz and Kang, 2018). The knockout of TFR1 has been shown to restore mitochondrial morphology by inhibiting IL-1β-induced cGAS/STING activity and mitigating mtDNA/cGAS/STING-mediated mitochondrial damage caused by iron overload (Wang W. et al., 2024). These findings establish TFR1 as a critical target for therapeutic intervention in ferroptosis related to OA and enhance the understanding of chondrocyte behavior in OA through the perspective of iron metabolic pathways. Such insights provide valuable directions for future research and clinical strategies.Recent studies have demonstrated that the overexpression of FTH-1, both in vitro and in vivo, enhances chondrocyte resistance to ferroptosis and inhibits cartilage extracellular matrix degradation. This is achieved through the regulation and inhibition of the MAPK pathway in the reversal of the DMM model, as well as the upregulation of SOX9 and Acan. Consequently, these mechanisms collectively contribute to a therapeutic effect on osteoarthritis (Yuan et al., 2024). A study investigating the role of GPR30 in ferroptosis within osteoarthritic (OA) cartilage demonstrates that the activation of GPR30 enhances the resistance of OA chondrocytes to ferroptosis. This effect is mediated through the modulation of YAP1 phosphorylation and the upregulation of FTH-1 expression, as validated under controlled conditions using G1 agonists and G15 inhibitors (Zhao et al., 2024).

Research has demonstrated that elevated levels of free fatty acids are present in patients with OA before the onset of classical symptoms (Bonner et al., 1975). The accumulation of lipid droplets in the cartilage matrix and chondrocytes accelerates cartilage degradation and the development of OA. Abnormal lipid metabolism within chondrocyte cytoplasm exacerbates cellular damage through various mechanisms (Zhou et al., 2021; Park et al., 2022). In the context of ferroptosis in chondrocytes, lipid metabolism is integral to the accumulation of lipid peroxides and subsequent oxidative stress, hallmarks of ferroptosis. Key enzymes such as ACSL4 and LPCAT3 play key roles in the synthesis of polyunsaturated fatty acid-containing phospholipids (PUFA-Pls), which are substrates for lipid peroxidation (LPO) (Klinger et al., 2002). ACSL4 facilitates the conjugation of long-chain polyunsaturated fatty acids (PUFAs), like adrenalic acid (ADA) and arachidonic acid (AA), with Coenzyme A (CoA) to produce ADA-CoA and AA-CoA. These conjugates are then re-esterified by LPCAT3 and incorporated into phospholipids (PLs), forming PUFA-PLs (Xu et al., 2023a). PKCβII enhances ACSL4 activity through phosphorylation, while acetyl-CoA carboxylase promotes PUFA synthesis. This process is driven by non-enzymatic autoxidation mediated by the iron-catalyzed Fenton reaction and enzymatic reactions involving ALOX or cytochrome P450 oxidoreductase, which further propagate lipid peroxidation (LPO). The combined effects of these processes lead to LPO, ultimately resulting in ferroptosis of chondrocytes in OA.

The SLC7A11/GSH/GPX4 pathway is a vital antioxidant mechanism within cells, particularly in the context of ferroptosis (Jiang et al., 2021). Both ferroptosis and cuproptosis can impair this pathway through various mechanisms involving excess copper. Firstly, copper (Cu) can directly bind to cysteine residues on glutathione peroxidase 4 (GPX4), leading to abnormal aggregation of GPX4 (Xue et al., 2023a), loss of its antioxidant activity, and the initiation of GPX4 autophagy-related reactions via the TAX1BP1 receptor. This sequence ultimately results in autophagy-mediated ferroptosis (Xue et al., 2023b). Notably, knocking down autophagy-related genes ATG5 and TAX1BP1 partially upregulates GPX4 and mitigates Cu-induced ferroptosis (Xue et al., 2023a). Similarly, the administration of copper-chelating agents has been shown to reduce sensitivity to ferroptosis, highlighting copper’s role in this process.Copper can also directly bind to glutathione (GSH), the key cofactor for GPX4, thereby depleting GSH levels. This depletion significantly reduces GPX4 activity and enhances susceptibility to ferroptosis (Scheiber et al., 2013). GSH also plays a regulatory role in maintaining intracellular copper homeostasis by modulating ATP7A and ATP7B (Banci et al., 2010), and its depletion may further exacerbate intracellular copper accumulation. However, the inactivation of GPX4 in certain cells has been associated with enhanced antioxidant capacity (Viswanathan et al., 2017), potentially indicating a compensatory mechanism involving alternative antioxidant pathways. This compensatory response, although not yet fully understood, is expected to become a focal point of future research, with significant implications for strategies aimed at mitigating oxidative stress and ferroptosis.

Wilson’s disease, a genetic disorder of aberrant copper metabolism, leads to excessive copper accumulation in the liver and is associated with premature OA due to metabolic disturbances caused by high copper levels (Ye et al., 2017). In patients with Wilson’s disease, elevated copper levels are accompanied by increased selenium and selenium-related transport-associated glycoprotein (SELENOP) concentrations in hepatic cells, alongside reduced activity. This reduction is possibly due to excessive accumulation of APOE, which binds to SELENOP and inhibits its secretion (Jin et al., 2020). Research indicates an antagonistic relationship between copper (Cu) and selenium (Se) in inflammatory and physiological contexts (Baudry et al., 2020). Excess copper also inhibits selenium-related enzymes, such as GPX4 (Schwarz et al., 2020). Targeting selenium metabolism—either by increasing selenium content or enhancing SELENOP expression—may alleviate oxidative stress and ferroptosis induced by copper overload.Research into the interactions between copper (Cu), selenium (Se), and associated proteins is essential for understanding and addressing inflammatory and age-related diseases. MEMO1, which sequesters copper and binds iron (Dolgova et al., 2024), has been shown to enhance GPX4 activity when knocked down in cellular models (Dolgova et al., 2024), highlighting the complex interplay between copper and iron.

SLC7A11 is a vital transporter for cystine uptake, essential for glutathione (GSH) synthesis and cellular antioxidant defense, thereby playing a key role in resisting ferroptosis. Excessive copper accumulation induces mitochondrial dysfunction, leading to increased reactive oxygen species (ROS) levels and DNA damage. This condition results in the direct degradation of SLC7A11 (Lei et al., 2022), diminishing GSH synthesis and reducing glutathione peroxidase 4 (GPX4) activity, which ultimately triggers ferroptosis.

Conversely, copper deficiency also disrupts the SLC7A11/GSH/GPX4 pathway, increasing susceptibility to ferroptosis. Under such conditions, hypoxia-inducible factor 1-alpha (HIF-1α) has been shown to upregulate SLC7A11, a ferroptosis suppressor protein. However, when intracellular copper ion levels decrease, the stabilizing effect of copper on HIF-1α is reduced. This attenuation leads to the downregulation of SLC7A11 and a subsequent reduction in GSH synthesis, thereby increasing cellular vulnerability to ferroptosis (Tan et al., 2021). Furthermore, COMMD10, a protein involved in copper metabolism, interacts with ATP7A, ATP7B to facilitate copper excretion, thereby down-regulating HIF-1α-associated SLC7A11 and CP and enhancing sensitivity to ferroptosis (Yang H. et al., 2022).

The FSP1 pathway offers an antioxidant defense mechanism distinct from the SLC7A11/GSH/GPX4 pathway. FSP1 works by interacting with DHODH to convert CoQ to CoQH2 via NAD(P)H. CoQH2 then neutralizes reactive oxygen species (ROS) at both the inner mitochondrial and plasma membranes (Koppula et al., 2022), thus reducing oxidative stress and preventing ferroptosis. Excess copper exposure can inhibit FSP1 expression, leading to decreased CoQ levels. This reduction impairs cellular antioxidant defenses and the clearance of ROS, thus contributing to ferroptosis (Zhong et al., 2024). This disruption in the FSP1-CoQ10 pathway due to excessive copper introduces a novel mechanism of copper-induced ferroptosis, distinct from the traditional SLC7A11/GSH/GPX4 pathway, and presents a potential target for therapeutic interventions aimed at modulating ferroptosis.

Additionally, excessive copper has been shown to elevate the protein expression levels of ACLS4 and NCOA4 (Zhong et al., 2024). ACLS4 promotes ferroptosis through fatty acid metabolism (Liu Z. et al., 2021; Zhao et al., 2021), while NCOA4 facilitates ferroptosis through ferritinophagy (Li et al., 2020). Moreover, NRF2 is crucial in regulating the expression of ferroptosis-related genes (Dodson et al., 2019; Li et al., 2020) and mitigating oxidative stress. Excess copper may downregulate NRF2 expression by promoting its inhibitor, Keap1, potentially contributing to ferroptosis (Zhong et al., 2024). Further research is needed to validate these mechanisms in osteoarthritic chondrocytes and explore their implications for therapeutic strategies.

The imbalance between ATP7A/B, which are essential for copper excretion and cellular endocrine functions (Dmitriev and Patry, 2024), and CTR1, the main copper uptake transporter (Ross et al., 2024), can significantly impact copper and iron metabolism, thereby influencing ferroptosis sensitivity (Li et al., 2023b). Elesclomol, a small molecule that binds Cu²⁺, targets and degrades ATP7A. This degradation leads to abnormal copper accumulation within the cell, which promotes the production of reactive oxygen species (ROS) through the Fenton reaction. The resulting oxidative stress damages mitochondria and degrades SLC7A11, diminishing GPX4 activity and ultimately triggering ferroptosis (Halliwell and Gutteridge, 1984; Kirshner et al., 2008; Gao et al., 2021).

Ceruloplasmin (CP) plays a pivotal role in copper transport and iron homeostasis by oxidizing Fe²⁺ to Fe³⁺, which facilitates iron binding to transferrin and enhances intracellular iron excretion (Vashchenko and MacGillivray, 2013). CP also helps mitigate iron overload and ROS production via the Fe²⁺-mediated Fenton reaction (Bakhautdin et al., 2014). In OA, severe oxidative stress can reduce CP levels (Kvien, 2010). This reduction can lead to iron overload, increasing the risk and sensitivity to ferroptosis (Mo et al., 2024). Additionally, copper deficiency can reduce CP levels, potentially disrupting iron metabolism and contributing to OA susceptibility (Rolić et al., 2024). Further research is needed to validate these mechanisms in the context of OA and explore potential therapeutic interventions.

Research suggests that P53 plays a pivotal role in regulating copper homeostasis (Xiong et al., 2023) and heightens sensitivity to copper-induced cell death by inhibiting glycolysis (Liu et al., 2019; Tsvetkov et al., 2022). Elevated copper levels activate P53, which subsequently downregulates SLC7A11 (Huang et al., 2023) and upregulates SAT1 and GLS2, ultimately leading to ferroptosis. SAT1 further amplifies the expression of arachidonic acid 15-lipoxygenase (ALOX15), intensifying oxidative stress and lipid peroxidation, thereby increasing cellular vulnerability to ferroptosis (Ou et al., 2016). GLS2, in concert with P53, promotes the conversion of glutamate to α-ketoglutarate (α-KG) and induces the production of reactive oxygen species (ROS). In hepatic cells lacking STP7B, GLS2 levels significantly rise under elevated copper conditions, suggesting that high copper concentrations may exert effects on GLS2 similar to those mediated by P53 (Yin et al., 2022). Thus, P53 may act as a key intermediary linking dysregulated copper/iron metabolism to ferroptosis, making it a potential target in conditions associated with copper overload and resistance to ferroptosis. Moreover, excessive copper directly damages mitochondrial DNA, causing extensive mitochondrial damage. This mitochondrial impairment leads to the release of free iron and the formation of multiple free iron pools, thereby greatly increasing susceptibility to ferroptosis (Ren et al., 2021).