Skip to main content

The impact of serum copper on the risk of epilepsy: a mendelian randomization study

Abstract

Background

The relationship between serum copper and epilepsy has been elucidated in observational studies. In this study, we aimed to explore the causal relationship between serum copper and epilepsy using Mendelian randomization (MR) analysis.

Methods

Single nucleotide polymorphisms (SNPs) associated with serum copper were used as instrumental variables for MR analysis to evaluate their causal effects on epilepsy. The main MR results were obtained by using the inverse variance weighting (IVW) method, supplemented by weighted median and MR-Egger regression. In addition, sensitivity analyses such as Cochran's Q test and pleiotropy test were used to assess these SNPs on epilepsy in terms of horizontal pleiotropy and heterogeneity.

Results

The IVW method revealed that the serum copper was associated with an increased risk of generalized epilepsy (OR= 1.07; 95% CI 1.01- 1.14; P = 0.032), and the sensitivity analysis further supports the robustness of the results.

Conclusions

The current study reveals a possible causal role for serum copper in increasing the risk of generalized epilepsy, which provide guidance for identifying potential risk factors for epilepsy.

Background

Epilepsy is a common neurological disorder that affects about 70 million people worldwide, with an incidence of 50.4 to 81.7 per 100,000 per year [1, 2]. The mechanisms of epileptic seizures are not fully understood. Some known causes of epilepsy include genetics, cerebrovascular disease, head injury, neurodegenerative diseases, etc. In recent years, researchers have extensively explored the relationship between trace elements and epilepsy [3]. Identifying this relationship can help discover potential risk factors and provide insight into epilepsy prevention and treatment.

Copper is an essential trace element in the human body, playing an important role in the development and function of the nervous system [4]. Copper is a vital trace element required by many enzymes in the brain. Administration of low doses of copper ion can induce epileptic seizures in animals by inhibiting Mg2+-ATPase and Na+-K+-ATPase, disrupting the Na+-K+ balance and leading to further epileptiform discharges [5, 6]. In addition, copper may produce toxic reactive oxygen species due to its redox activity, causing harm to the brain [7]. Both copper deficiency and excess can seriously affect brain function, leading to neurological diseases [8]. Studies have reported that elevated serum levels of copper may be associated with depression [9], Alzheimer's disease [10, 11], and multiple sclerosis [12]. Observational studies have suggested that elevated serum levels of copper may be associated with an increased risk of epilepsy [13], although another study did not find a correlation between them [14]. It is therefore essential to understand the causal relationship between serum copper and epilepsy.

Mendelian randomization (MR) analysis is a method that uses genetic variation in species as instrumental variables (IVs) to investigate the causality of an exposure on an outcome [15]. MR bears resemblance to randomized controlled studies in that it enables the investigation of causality by circumventing reverse causality and the influences of confounding factors [16]. Consequently, MR has emerged as a widely used epidemiological approach.

In the present study, we used genome-wide association study (GWAS) data on serum copper and epilepsy in an MR approach to further understand the risk factors influencing the development of epilepsy.

Methods

Study design

The MR analysis in this study was based on three key assumptions: (i) the selected genetic variants must be significantly associated with the exposure factor (serum copper); (ii) the selected genetic variants are not associated with other confounding factors; and (iii) the selected genetic variants affect the outcome (epilepsy) only through the exposure factor (serum copper) and do not affect the outcome through other pathways. If these three hypotheses hold, MR analysis could evaluate the causal relationship between the exposure factor and the outcome while avoiding potential confounders [15].

Selection of instrumental variables

For serum copper, we used the large GWAS data from 2603 participants of European ancestry [17]. In this dataset, we used a criterion of P < 1 × 10–5 to screen for genetic variants significantly associated with serum copper and subsequently removed linkage disequilibrium (LD) between single nucleotide polymorphisms (SNPs) by R2 < 0.1. We also used an F-statistic to assess the reliability of the genetic variables, which was generally considered to have stronger instrument strength when F > 10 [18]. R2 was calculated as \({R}^{2}= 2 \times EAF \times (1-EAF) \times {Beta}^{2}\) [18], where Beta represents the estimated effect of genetic variation and EAF represents the effect allele frequency. F was calculated as \(F= {R}^{2}\times (N-2)/(1-{R}^{2})\), where R2 was the cumulative proportion of variance in the phenotype explained by the SNPs included in the exposure factor, and N was the sample size.

Furthermore, we acquired comprehensive GWAS statistical data in association with epilepsy-related subtypes from the International League Against Epilepsy (ILAE). The sample sizes of the datasets are shown in Table 1. More detailed information on epilepsy cases can be found in the original study [19].

Table 1 Characteristics of included GWAS summary-level data of serum copper Levels and epilepsy

Statistical analysis

Three methods of MR were used in this study to analyze the causal effect of serum copper on epilepsy, including the inverse variance weighted (IVW) method, the MR-Egger regression method and the Weighted Median method. Each method uses a different hypothetical model to assess the causal effects and these are then used to check the robustness of the results. The IVW is considered the primary method of analysis as it is the gold standard for MR inference. It is primarily used for basic causal estimation and provides accurate results by calculating a weighted average of the Wald ratio estimates [20]. The MR-Egger method is used to detect sensitivities and it provides calculations after adjusting for pleiotropy [21]. Alternatively, in the median-weighted method, which allows for the presence of a 50% null of SNPs, the estimation of causal effects is then performed [22]. In addition, we also used Cochran's Q statistic to check the heterogeneity of the results in Heterogeneity test and performed a Pleiotropy test to check the polymorphism of results [21].

All analyses were conducted using the R software (version 4.1.3), with the MR analysis performed using the "TwoSampleMR" R package.

Results

In the initial phase, we found that the serum copper was positively associated with the risk of generalized epilepsy (OR= 1.07; 95% CI 1.01–1.14; P = 0.032). In the sensitivity analysis of the significant findings, the Cochran's Q test indicated no heterogeneity between serum copper and generalized epilepsy (Q = 5.789; P = 0.447). Additionally, the polymorphism test demonstrated no evidence of cross-sectional polymorphism (Intercept = -0.006; P = 0.568) in the association between serum copper and generalized epilepsy. The MR estimates of genetically predisposed serum copper and susceptibility to epilepsy are presented in Table 2. The specific sensitivity analysis results are shown in Table 3.

Table 2 Causal effects of serum copper on the risk of epilepsy
Table 3 Evaluation of heterogeneity and pleiotropy using different methods

Scatter plots and funnel plots of significant results are shown in Fig. 1. Forest plots of significant results are shown in Fig. 2.

Fig. 1
figure 1

Scatter plots and funnel plots of MR analysis showed relationships between serum copper and epilepsy risk. a-c Scatter plots of relationships of serum copper with epilepsy (a), generalized epilepsy (b), and focal epilepsy (c). d-f Funnel plots of relationships of serum copper with epilepsy (d), generalized epilepsy (e), and focal epilepsy (f)

Fig. 2
figure 2

Forest plot of MR estimates of the causal effect of serum copper on epilepsy

Finally, detailed information on the SNPs involved in this study is given in Supplementary Tables S1, S2 and S3.

Discussion

In this study, MR analysis showed that the elevated serum copper is associated with an increased risk of generalized epilepsy. However, no causal relationship was observed between serum copper level and focal epilepsy. Furthermore, we revealed no significant heterogeneity or evidence of horizontal pleiotropy in our results.

The copper content in the brain accounts for approximately 9% of the total copper in the human body, ranking the third highest concentration across organs [23]. Disruption of copper metabolism plays a vital role in the pathogenesis of epilepsy. Copper is also involved in the synthesis and release of neurotransmitters [24], regulating neuronal membrane stability and ion channels [25]. Additionally, it may impact the neuronal redox reactions and mitochondrial function. Mitochondria serve as the energy center of neurons, responsible for producing most of the cellular energy. Disturbances in copper metabolism can impair mitochondrial function, leading to disruptions in cellular energy metabolism, thereby affecting neuronal function [26, 27]. These changes may increase neuronal excitability, thereby elevating the risk of epilepsy. Studies have shown that exposure to copper induces oxidative stress and promotes apoptosis of HT22 mouse hippocampal neurons [28]. Furthermore, in recent years, cuproptosis is emerging as a novel mechanism of cell death. It has been demonstrated that copper ions can interact with sulfur transferase proteins in the tricarboxylic acid cycle, promoting abnormal oligomerization of these proteins. Moreover, copper ions can reduce the level of Fe-S cluster proteins, collectively triggering protein toxicity stress response that ultimately leads to cell death [29]. The association between cuproptosis and neurodegenerative disorders such as Alzheimer's disease has been extensively investigated [30, 31]. Wilson's disease, a genetic disorder of copper metabolism, leads to excessive accumulation of copper in the body, resulting in cuproptosis. High levels of copper can be observed in almost all brain regions of patients with Wilson's disease [32]. This condition is currently believed to be associated with seizures in epilepsy [33]. We hypothesize that cuproptosis may be one of the mechanisms underlying epilepsy, and our results also demonstrate that elevated serum copper levels increase the risk of generalized epilepsy.

Similar results have also been found in observational studies. Animal studies have shown that serum copper levels are significantly higher in dogs with controlled and uncontrolled epilepsy compared to normal or untreated dogs [34]. Furthermore, our research findings are consistent with a previous study involving 200 patients with genetic generalized epilepsy, which found a correlation between high serum copper levels and genetic generalized epilepsy [35]. Regarding the potential differences in the roles of copper ions in generalized epilepsy vs in focal epilepsy, it is worth noting that generalized epilepsy is a form of epilepsy that occurs widely throughout the brain, involving extensive neuronal networks [36]. Copper ions may disrupt neuronal function by modulating neuronal excitability and inhibitory pathways. Additionally, copper ions may interact with other ions, altering neuronal membrane potential and leading to abnormal discharges and seizure activity [25]. In focal epilepsy, abnormal discharges and seizures are primarily confined to specific brain regions. These seizures are often associated with specific lesions or injuries, such as brain tumors, infections, or trauma [37, 38]. Compared to generalized epilepsy, the disruption of neuronal networks is more localized in focal epilepsy. Therefore, copper ions may not have sufficient opportunity to enter or accumulate in the specific brain regions that affect seizure activity, resulting in a less significant impact on the occurrence and manifestation of focal epilepsy. However, it is important to note that these results are preliminary, and further research is needed to gain a deeper understanding of the relationship between copper and epilepsy.

This study has some advantages. First, our study was based on data of a large GWAS study provided by the ILAE Consortium, which includes a wide range of subtypes of epilepsy, contributing to a more comprehensive analysis of epilepsy. Second, we performed a sensitivity analysis to ensure that the results are not horizontally pleiotropic or heterogeneous. Finally, our MR analysis is superior to observational studies in that the SNPs were randomly assigned, leading to much less bias due to confounding factors.

However, this study also has certain limitations. First, the results of this study are based on European ancestry and it is unclear whether they can be generalised to other races. Second, the epilepsy dataset provided by the ILAE used in this study contained approximately 14% of cases of non-European descent. Therefore, population stratification is likely to introduce bias here.

Conclusions

To conclude, this MR study offers genetic evidence supporting the association between elevated serum copper levels and an increased risk of generalized epilepsy, specifically within a European population. However, no such relationship was observed for focal epilepsy. These findings provide insights into the identification of potential risk factors of epilepsy. Nevertheless, further large-scale clinical studies are required to validate and expand these findings in the future.

Availability of data and materials

Data and materials related to the current study are openly accessible and available in the IEU Open GWAS Project repository (http://gwas.mrcieu.ac.uk), where they can be retrieved and utilized.

Abbreviations

Beta:

The per-allele effect on cannabis use

EA:

Effect allele

EAF:

Effect allele frequency

F:

F statistic

GWAS:

Genome-wide association study

ILAE:

the International League Against Epilepsy

IVs:

Instrumental variables

IVW:

Inverse variance weighted

MR:

Mendelian randomisation

OA:

Other effect allele

Q_pval:

The P value of Cochran's Q test

R2 :

Percentage of the variation explained by the SNP

SE:

Standard error of Beta

SNPs:

Single nucleotide polymorphisms

References

  1. Fisher RS, Acevedo C, Arzimanoglou A, Bogacz A, Cross JH, Elger CE, et al. ILAE official report: a practical clinical definition of epilepsy. Epilepsia. 2014;55(4):475–82.

  2. Collaborators GBDN. Global, regional, and national burden of neurological disorders, 1990–2016: a systematic analysis for the Global Burden of Disease Study 2016. Lancet Neurol. 2019;18(5):459–80.

    Article  Google Scholar 

  3. Saghazadeh A, Mahmoudi M, Meysamie A, Gharedaghi M, Zamponi GW, Rezaei N. Possible role of trace elements in epilepsy and febrile seizures: a meta-analysis. Nutr Rev. 2015;73(11):760–79.

    Article  PubMed  Google Scholar 

  4. Lutsenko S, Bhattacharjee A, Hubbard AL. Copper handling machinery of the brain. Metallomics. 2010;2(9):596–608.

    Article  CAS  PubMed  Google Scholar 

  5. Donaldson J, St Pierre T, Minnich J, Barbeau A. Seizures in rats associated with divalent cation inhibition of NA + -K + -ATP’ase. Can J Biochem. 1971;49(11):1217–24.

    Article  CAS  PubMed  Google Scholar 

  6. Barbeau A, Donaldson J. Zinc, taurine, and epilepsy. Arch Neurol. 1974;30(1):52–8.

    Article  CAS  PubMed  Google Scholar 

  7. Jomova K, Valko M. Advances in metal-induced oxidative stress and human disease. Toxicology. 2011;283(2–3):65–87.

    Article  CAS  PubMed  Google Scholar 

  8. Scheiber IF, Mercer JF, Dringen R. Metabolism and functions of copper in brain. Prog Neurobiol. 2014;116:33–57.

    Article  CAS  PubMed  Google Scholar 

  9. Ni M, You Y, Chen J, Zhang L. Copper in depressive disorder: A systematic review and meta-analysis of observational studies. Psychiatry Res. 2018;267:506–15.

    Article  CAS  PubMed  Google Scholar 

  10. Ventriglia M, Bucossi S, Panetta V, Squitti R. Copper in Alzheimer’s disease: a meta-analysis of serum, plasma, and cerebrospinal fluid studies. J Alzheimers Dis. 2012;30(4):981–4.

    Article  PubMed  Google Scholar 

  11. Park JH, Lee DW, Park KS. Elevated serum copper and ceruloplasmin levels in Alzheimer’s disease. Asia Pac Psychiatry. 2014;6(1):38–45.

    Article  PubMed  Google Scholar 

  12. De Riccardis L, Buccolieri A, Muci M, Pitotti E, De Robertis F, Trianni G, et al. Copper and ceruloplasmin dyshomeostasis in serum and cerebrospinal fluid of multiple sclerosis subjects. Biochim Biophys Acta Mol Basis Dis. 2018;1864(5 Pt A):1828–38.

  13. Wojciak RW, Mojs E, Stanislawska-Kubiak M, Samborski W. The serum zinc, copper, iron, and chromium concentrations in epileptic children. Epilepsy Res. 2013;104(1–2):40–4.

    Article  CAS  PubMed  Google Scholar 

  14. Tombini M, Squitti R, Cacciapaglia F, Ventriglia M, Assenza G, Benvenga A, et al. Inflammation and iron metabolism in adult patients with epilepsy: does a link exist? Epilepsy Res. 2013;107(3):244–52.

    Article  CAS  PubMed  Google Scholar 

  15. Emdin CA, Khera AV, Kathiresan S. Mendelian Randomization. JAMA. 2017;318(19):1925–6.

    Article  PubMed  Google Scholar 

  16. Hemani G, Zheng J, Elsworth B, Wade KH, Haberland V, Baird D, et al. The MR-Base platform supports systematic causal inference across the human phenome. Elife. 2018;7:e34408.

  17. Evans DM, Zhu G, Dy V, Heath AC, Madden PA, Kemp JP, et al. Genome-wide association study identifies loci affecting blood copper, selenium and zinc. Hum Mol Genet. 2013;22(19):3998–4006.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Burgess S, Thompson SG, Collaboration CCG. Avoiding bias from weak instruments in Mendelian randomization studies. Int J Epidemiol. 2011;40(3):755–64.

    Article  PubMed  Google Scholar 

  19. International League Against Epilepsy Consortium on Complex E. Genome-wide mega-analysis identifies 16 loci and highlights diverse biological mechanisms in the common epilepsies. Nat Commun. 2018;9(1):5269.

  20. Bowden J, Del Greco MF, Minelli C, Davey Smith G, Sheehan N, Thompson J. A framework for the investigation of pleiotropy in two-sample summary data Mendelian randomization. Stat Med. 2017;36(11):1783–802.

    Article  PubMed  PubMed Central  Google Scholar 

  21. Bowden J, Davey Smith G, Burgess S. Mendelian randomization with invalid instruments: effect estimation and bias detection through Egger regression. Int J Epidemiol. 2015;44(2):512–25.

    Article  PubMed  PubMed Central  Google Scholar 

  22. Bowden J, Davey Smith G, Haycock PC, Burgess S. Consistent Estimation in Mendelian Randomization with Some Invalid Instruments Using a Weighted Median Estimator. Genet Epidemiol. 2016;40(4):304–14.

    Article  PubMed  PubMed Central  Google Scholar 

  23. Que EL, Domaille DW, Chang CJ. Metals in neurobiology: probing their chemistry and biology with molecular imaging. Chem Rev. 2008;108(5):1517–49.

    Article  CAS  PubMed  Google Scholar 

  24. D’Ambrosi N, Rossi L. Copper at synapse: Release, binding and modulation of neurotransmission. Neurochem Int. 2015;90:36–45.

    Article  CAS  PubMed  Google Scholar 

  25. Mathie A, Sutton GL, Clarke CE, Veale EL. Zinc and copper: pharmacological probes and endogenous modulators of neuronal excitability. Pharmacol Ther. 2006;111(3):567–83.

    Article  CAS  PubMed  Google Scholar 

  26. Tang D, Chen X, Kroemer G. Cuproptosis: a copper-triggered modality of mitochondrial cell death. Cell Res. 2022;32(5):417–8.

    Article  PubMed  PubMed Central  Google Scholar 

  27. Rossi L, Lombardo MF, Ciriolo MR, Rotilio G. Mitochondrial dysfunction in neurodegenerative diseases associated with copper imbalance. Neurochem Res. 2004;29(3):493–504.

    Article  CAS  PubMed  Google Scholar 

  28. Lu Q, Zhang Y, Zhao C, Zhang H, Pu Y, Yin L. Copper induces oxidative stress and apoptosis of hippocampal neuron via pCREB/BDNF/ and Nrf2/HO-1/NQO1 pathway. J Appl Toxicol. 2022;42(4):694–705.

    Article  CAS  PubMed  Google Scholar 

  29. Chen L, Min J, Wang F. Copper homeostasis and cuproptosis in health and disease. Signal Transduct Target Ther. 2022;7(1):378.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Zhang Y, Zhou Q, Lu L, Su Y, Shi W, Zhang H, et al. Copper Induces Cognitive Impairment in Mice via Modulation of Cuproptosis and CREB Signaling. Nutrients. 2023;15(4):972.

  31. Lai Y, Lin C, Lin X, Wu L, Zhao Y, Lin F. Identification and immunological characterization of cuproptosis-related molecular clusters in Alzheimer’s disease. Front Aging Neurosci. 2022;14: 932676.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Dusek P, Litwin T, Czlonkowska A. Wilson disease and other neurodegenerations with metal accumulations. Neurol Clin. 2015;33(1):175–204.

    Article  PubMed  Google Scholar 

  33. Dening TR, Berrios GE, Walshe JM. Wilson’s disease and epilepsy. Brain. 1988;111(Pt 5):1139–55.

    Article  PubMed  Google Scholar 

  34. Vitale S, Hague DW, Foss K, de Godoy MC, Selmic LE. Comparison of Serum Trace Nutrient Concentrations in Epileptics Compared to Healthy Dogs. Front Vet Sci. 2019;6:467.

    Article  PubMed  PubMed Central  Google Scholar 

  35. Prasad DK, Shaheen U, Satyanarayana U, Surya Prabha T, Jyothy A, Munshi A. Association of serum trace elements and minerals with genetic generalized epilepsy and idiopathic intractable epilepsy. Neurochem Res. 2014;39(12):2370–6.

    Article  CAS  PubMed  Google Scholar 

  36. Blumenfeld H. From molecules to networks: cortical/subcortical interactions in the pathophysiology of idiopathic generalized epilepsy. Epilepsia. 2003;44(Suppl 2):7–15.

    Article  CAS  PubMed  Google Scholar 

  37. Fordington S, Manford M. A review of seizures and epilepsy following traumatic brain injury. J Neurol. 2020;267(10):3105–11.

    Article  PubMed  PubMed Central  Google Scholar 

  38. Nowell M, Miserocchi A, McEvoy AW. Tumors in Epilepsy. Semin Neurol. 2015;35(3):209–17.

    Article  PubMed  Google Scholar 

Download references

Acknowledgements

The authors express sincere gratitude to the ILAE Consortium for generously providing the complete GWAS data, which was instrumental in conducting this study.

Funding

There is no funding for this parper.

Author information

Authors and Affiliations

Authors

Contributions

SC conceptualized and designed the study. SC, WH, and QX conducted the data analysis and drafted the original manuscript. TH and MZ critically reviewed and edited the paper. HX provided supervision throughout the project. All authors have thoroughly reviewed and approved the final version of the manuscript for publication.

Corresponding author

Correspondence to Huiqin Xu.

Ethics declarations

Ethics approval and consent to participate

This study only used publicly available data. No original data were collected. Ethical approval and informed consent for studies included in the analyses were provided in the original publications.

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.

Supplementary Information

Additional file 1:

Supplementary Table S1. Association strength of SNP allele frequencies and effect alleles with serum copper in outcome of epilepsy.

Additional file 2:

Supplementary Table S2. Association strength of SNP allele frequencies and effect alleles with serum copper in outcome of generalized epilepsy.

Additional file 3:

Supplementary Table S3. Association strength of SNP allele frequencies and effect alleles with serum copper in outcome of focal epilepsy.

Rights and permissions

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

Reprints and Permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Chen, S., Huang, W., Xu, Q. et al. The impact of serum copper on the risk of epilepsy: a mendelian randomization study. Acta Epileptologica 5, 15 (2023). https://doi.org/10.1186/s42494-023-00126-3

Download citation

  • Received:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1186/s42494-023-00126-3

Keywords

  • Serum copper
  • Epilepsy
  • Mendelian randomization
  • Genome-wide association studies
  • Single nucleotide polymorphisms