Exploring the Role of Ferroptosis in the Pathophysiology and Circadian Regulation of Restless Legs Syndrome
Abstract
1. Introduction
2. Materials and Methods
3. Results
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
Abbreviations
FC | Fold Change |
FDR | False Discovery Rate |
KEGG | Kyoto Encyclopedia of Genes and Genomes |
PBMCs | Peripheral Blood Mononuclear Cells |
PCA | Principal Component Analysis |
PLMS | Periodic Limb Movements during Sleep |
RIN | RNA Integrity Number |
RLS | Restless Legs Syndrome |
RNA | Ribonucleic Acid |
ROS | Reactive Oxygen Species |
SCN | Suprachiasmatic Nucleus |
References
- Koo, B.B. Restless Leg Syndrome Across the Globe: Epidemiology of the Restless Legs Syndrome/Willis-Ekbom Disease. Sleep Med. Clin. 2015, 10, 189–205. [Google Scholar] [CrossRef] [PubMed]
- Ulfberg, J.; Nystrom, B.; Carter, N.; Edling, C. Prevalence of restless legs syndrome among men aged 18 to 64 years: An association with somatic disease and neuropsychiatric symptoms. Mov. Disord. Off. J. Mov. Disord. Soc. 2001, 16, 1159–1163. [Google Scholar] [CrossRef]
- Ulfberg, J.; Nystrom, B.; Carter, N.; Edling, C. Restless Legs Syndrome among working-aged women. Eur. Neurol. 2001, 46, 17–19. [Google Scholar] [CrossRef]
- Allen, R.P.; Picchietti, D.L.; Garcia-Borreguero, D.; Ondo, W.G.; Walters, A.S.; Winkelman, J.W.; Zucconi, M.; Ferri, R.; Trenkwalder, C.; Lee, H.B.; et al. Restless legs syndrome/Willis-Ekbom disease diagnostic criteria: Updated International Restless Legs Syndrome Study Group (IRLSSG) consensus criteria—History, rationale, description, and significance. Sleep Med. 2014, 15, 860–873. [Google Scholar] [CrossRef] [PubMed]
- Mogavero, M.P.; DelRosso, L.M.; Lanza, G.; Lanuzza, B.; Bruni, O.; Strambi, L.F.; Ferri, R. Changes in time structure of periodic leg movements during sleep in restless legs syndrome: Effects of sex and age. Sleep Med. 2024, 115, 137–144. [Google Scholar] [CrossRef]
- Mogavero, M.P.; Lanza, G.; DelRosso, L.M.; Lanuzza, B.; Bruni, O.; Ferini Strambi, L.; Ferri, R. Exploring sex differences in periodic leg movements during sleep across the lifespan of patients with restless legs syndrome. Sleep Med. 2024, 122, 253–257. [Google Scholar] [CrossRef] [PubMed]
- Mogavero, M.P.; Antelmi, E.; Lanza, G.; Marelli, S.; Castelnuovo, A.; Tinazzi, M.; DelRosso, L.M.; Silvestri, R.; Ferri, R.; Ferini Strambi, L. Sex-based disparities in dopamine agonist response in patients with restless legs syndrome. J. Sleep Res. 2024, 34, e14311. [Google Scholar] [CrossRef]
- Manconi, M.; Garcia-Borreguero, D.; Schormair, B.; Videnovic, A.; Berger, K.; Ferri, R.; Dauvilliers, Y. Restless legs syndrome. Nat. Rev. Dis. Primers 2021, 7, 80. [Google Scholar] [CrossRef] [PubMed]
- Trenkwalder, C.; Allen, R.; Hogl, B.; Clemens, S.; Patton, S.; Schormair, B.; Winkelmann, J. Comorbidities, treatment, and pathophysiology in restless legs syndrome. Lancet Neurol. 2018, 17, 994–1005. [Google Scholar] [CrossRef]
- Mogavero, M.P.; Salemi, M.; Lanza, G.; Rinaldi, A.; Marchese, G.; Ravo, M.; Salluzzo, M.G.; Antoci, A.; DelRosso, L.M.; Bruni, O.; et al. Unveiling the pathophysiology of restless legs syndrome through transcriptome analysis. iScience 2024, 27, 109568. [Google Scholar] [CrossRef]
- Mogavero, M.P.; Mezzapesa, D.M.; Savarese, M.; DelRosso, L.M.; Lanza, G.; Ferri, R. Morphological analysis of the brain subcortical gray structures in restless legs syndrome. Sleep Med. 2021, 88, 74–80. [Google Scholar] [CrossRef]
- Rizzo, G.; Plazzi, G. Neuroimaging Applications in Restless Legs Syndrome. Int. Rev. Neurobiol. 2018, 143, 31–64. [Google Scholar] [CrossRef]
- Ferri, R.; Zucconi, M.; Manconi, M.; Bruni, O.; Ferini-Strambi, L.; Vandi, S.; Montagna, P.; Mignot, E.; Plazzi, G. Different periodicity and time structure of leg movements during sleep in narcolepsy/cataplexy and restless legs syndrome. Sleep 2006, 29, 1587–1594. [Google Scholar] [CrossRef]
- Dixon, S.J.; Lemberg, K.M.; Lamprecht, M.R.; Skouta, R.; Zaitsev, E.M.; Gleason, C.E.; Patel, D.N.; Bauer, A.J.; Cantley, A.M.; Yang, W.S.; et al. Ferroptosis: An iron-dependent form of nonapoptotic cell death. Cell 2012, 149, 1060–1072. [Google Scholar] [CrossRef] [PubMed]
- Stockwell, B.R.; Friedmann Angeli, J.P.; Bayir, H.; Bush, A.I.; Conrad, M.; Dixon, S.J.; Fulda, S.; Gascon, S.; Hatzios, S.K.; Kagan, V.E.; et al. Ferroptosis: A Regulated Cell Death Nexus Linking Metabolism, Redox Biology, and Disease. Cell 2017, 171, 273–285. [Google Scholar] [CrossRef] [PubMed]
- Morris, G.; Berk, M.; Carvalho, A.F.; Maes, M.; Walker, A.J.; Puri, B.K. Why should neuroscientists worry about iron? The emerging role of ferroptosis in the pathophysiology of neuroprogressive diseases. Behav. Brain Res. 2018, 341, 154–175. [Google Scholar] [CrossRef] [PubMed]
- Winkelman, J.W.; Berkowski, J.A.; DelRosso, L.M.; Koo, B.B.; Scharf, M.T.; Sharon, D.; Zak, R.S.; Kazmi, U.; Falck-Ytter, Y.; Shelgikar, A.V.; et al. Treatment of restless legs syndrome and periodic limb movement disorder: An American Academy of Sleep Medicine clinical practice guideline. J. Clin. Sleep Med. 2024, 21, 137–152. [Google Scholar] [CrossRef]
- Wang, X.; Wang, M.; Zhi, H.; Li, J.; Guo, D. REV-ERBα inhibitor rescues MPTP/MPP+-induced ferroptosis of dopaminergic neuron through regulating FASN/SCD1 signaling pathway. Heliyon 2024, 10, e40388. [Google Scholar] [CrossRef]
- Clemens, S.; Rye, D.; Hochman, S. Restless legs syndrome: Revisiting the dopamine hypothesis from the spinal cord perspective. Neurology 2006, 67, 125–130. [Google Scholar] [CrossRef]
- Ondo, W.G.; He, Y.; Rajasekaran, S.; Le, W.D. Clinical correlates of 6-hydroxydopamine injections into A11 dopaminergic neurons in rats: A possible model for restless legs syndrome. Mov. Disord. Off. J. Mov. Disord. Soc. 2000, 15, 154–158. [Google Scholar] [CrossRef]
- Pina-Leyva, C.; Lara-Lozano, M.; Rodriguez-Sanchez, M.; Vidal-Cantu, G.C.; Barrientos Zavalza, E.; Jimenez-Estrada, I.; Delgado-Lezama, R.; Rodriguez-Sosa, L.; Granados-Soto, V.; Gonzalez-Barrios, J.A.; et al. Hypothalamic A11 Nuclei Regulate the Circadian Rhythm of Spinal Mechanonociception through Dopamine Receptors and Clock Gene Expression. Life 2022, 12, 1411. [Google Scholar] [CrossRef]
- Schilling, C.; Schredl, M.; Strobl, P.; Deuschle, M. Restless legs syndrome: Evidence for nocturnal hypothalamic-pituitary-adrenal system activation. Mov. Disord. Off. J. Mov. Disord. Soc. 2010, 25, 1047–1052. [Google Scholar] [CrossRef]
- Damiola, F.; Le Minh, N.; Preitner, N.; Kornmann, B.; Fleury-Olela, F.; Schibler, U. Restricted feeding uncouples circadian oscillators in peripheral tissues from the central pacemaker in the suprachiasmatic nucleus. Genes Dev. 2000, 14, 2950–2961. [Google Scholar] [CrossRef] [PubMed]
- Walters, A.S.; LeBrocq, C.; Dhar, A.; Hening, W.; Rosen, R.; Allen, R.P.; Trenkwalder, C. Validation of the International Restless Legs Syndrome Study Group rating scale for restless legs syndrome. Sleep Med. 2003, 4, 121–132. [Google Scholar] [CrossRef]
- Jimenez-Jimenez, F.J.; Alonso-Navarro, H.; Garcia-Martin, E.; Agundez, J.A.G. Inflammatory factors and restless legs syndrome: A systematic review and meta-analysis. Sleep Med. Rev. 2023, 68, 101744. [Google Scholar] [CrossRef] [PubMed]
- Gottlieb, D.J.; Somers, V.K.; Punjabi, N.M.; Winkelman, J.W. Restless legs syndrome and cardiovascular disease: A research roadmap. Sleep Med. 2017, 31, 10–17. [Google Scholar] [CrossRef] [PubMed]
- Earley, C.J.; Jones, B.C.; Ferre, S. Brain-iron deficiency models of restless legs syndrome. Exp. Neurol. 2022, 356, 114158. [Google Scholar] [CrossRef]
- Link, W.; Ferreira, B.I. FOXO Transcription Factors: A Brief Overview. Methods Mol. Biol. 2025, 2871, 1–8. [Google Scholar] [CrossRef]
- Guo, N.; Wang, X.; Xu, M.; Bai, J.; Yu, H.; Le, Z. PI3K/AKT signaling pathway: Molecular mechanisms and therapeutic potential in depression. Pharmacol. Res. 2024, 206, 107300. [Google Scholar] [CrossRef]
- Dixon, S.J.; Olzmann, J.A. The cell biology of ferroptosis. Nat. Rev. Mol. Cell Biol. 2024, 25, 424–442. [Google Scholar] [CrossRef]
- Tang, D.; Chen, X.; Kang, R.; Kroemer, G. Ferroptosis: Molecular mechanisms and health implications. Cell Res. 2021, 31, 107–125. [Google Scholar] [CrossRef] [PubMed]
- Ma, Y.; Jiang, T.; Zhu, X.; Xu, Y.; Wan, K.; Zhang, T.; Xie, M. Efferocytosis in dendritic cells: An overlooked immunoregulatory process. Front. Immunol. 2024, 15, 1415573. [Google Scholar] [CrossRef] [PubMed]
- Walters, A.S.; Paueksakon, P.; Adler, C.H.; Moussouttas, M.; Weinstock, L.B.; Spruyt, K.; Bagai, K. Restless Legs Syndrome Shows Increased Silent Postmortem Cerebral Microvascular Disease with Gliosis. J. Am. Heart Assoc. 2021, 10, e019627. [Google Scholar] [CrossRef]
- Schormair, B.; Zhao, C.; Bell, S.; Didriksen, M.; Nawaz, M.S.; Schandra, N.; Stefani, A.; Hogl, B.; Dauvilliers, Y.; Bachmann, C.G.; et al. Genome-wide meta-analyses of restless legs syndrome yield insights into genetic architecture, disease biology and risk prediction. Nat. Genet. 2024, 56, 1090–1099. [Google Scholar] [CrossRef]
- Para, K.S.; Chow, C.A.; Nalamada, K.; Kakade, V.M.; Chilakamarri, P.; Louis, E.D.; Koo, B.B. Suicidal thought and behavior in individuals with restless legs syndrome. Sleep Med. 2019, 54, 1–7. [Google Scholar] [CrossRef]
- Zhu, Y.; Fujimaki, M.; Rubinsztein, D.C. Autophagy-dependent versus autophagy-independent ferroptosis. Trends Cell Biol. 2025. [Google Scholar] [CrossRef] [PubMed]
- Bass, J.; Takahashi, J.S. Circadian integration of metabolism and energetics. Science 2010, 330, 1349–1354. [Google Scholar] [CrossRef]
- Ying‑Hao, P.; Yu‑Shan, Y.; Song‑Yi, C.; Hua, J.; Peng, Y.; Xiao‑Hu, C. Time of day-dependent alterations of ferroptosis in LPS-induced myocardial injury via Bmal-1/AKT/Nrf2 in rat and H9c2 cell. Heliyon 2024, 10, e37088. [Google Scholar] [CrossRef]
- Xu, S.; Tang, Q.; Du, H.; Xie, J.; He, R.; Wang, R.; Sun, Q. Mechanism of Circadian Regulation in Ferroptosis of the BMAL1/NRF2 Pathway in Renal Ischemia-Reperfusion. Biomedicines 2025, 13, 1375. [Google Scholar] [CrossRef]
- Walters, A.S.; Zee, P.C. Why the worsening at rest and worsening at night criteria for Restless Legs Syndrome are listed separately: Review of the circadian literature on RLS and suggestions for future directions. Front. Neurol. 2023, 14, 1153273. [Google Scholar] [CrossRef]
- Liu, Y.; Wang, Y.; Liu, J.; Kang, R.; Tang, D. The circadian clock protects against ferroptosis-induced sterile inflammation. Biochem. Biophys. Res. Commun. 2020, 525, 620–625. [Google Scholar] [CrossRef] [PubMed]
- Liguori, C.; Holzknecht, E.; Placidi, F.; Izzi, F.; Mercuri, N.B.; Hogl, B.; Stefani, A. Seasonality of restless legs syndrome: Symptom variability in winter and summer times. Sleep Med. 2020, 66, 10–14. [Google Scholar] [CrossRef] [PubMed]
- Lincoln, G. A brief history of circannual time. J. Neuroendocrinol. 2019, 31, e12694. [Google Scholar] [CrossRef]
- Garcia-Borreguero, D.; Aragon, A.G.; Moncada, B.; Romero, S.; Granizo, J.J.; Quintas, S.; Castillo, M. Treatment of Sleep, Motor and Sensory Symptoms with the Orexin Antagonist Suvorexant in Adults with Idiopathic Restless Legs Syndrome: A Randomized Double-Blind Crossover Proof-of-Concept Study. CNS Drugs 2024, 38, 45–54. [Google Scholar] [CrossRef]
- Mogavero, M.P.; Fowowe, M.; Sanni, A.; Goli, M.; Lanza, G.; L’Episcopo, F.; Ferini-Strambi, L.; Mechref, Y.; Ferri, R. Evidence of Involvement of the Calcitonin Gene-Related Peptide in Restless Legs Syndrome. Mov. Disord. Off. J. Mov. Disord. Soc. 2025, 40, 1148–1159. [Google Scholar] [CrossRef]
- Imai, N. Molecular and Cellular Neurobiology of Circadian and Circannual Rhythms in Migraine: A Narrative Review. Int. J. Mol. Sci. 2023, 24, 10092. [Google Scholar] [CrossRef]
- Ryan, S.K.; Ugalde, C.L.; Rolland, A.S.; Skidmore, J.; Devos, D.; Hammond, T.R. Therapeutic inhibition of ferroptosis in neurodegenerative disease. Trends Pharmacol. Sci. 2023, 44, 674–688. [Google Scholar] [CrossRef] [PubMed]
ID | Term Description | Fold Change | Lowest p | Highest p | Up-Regulated | Down-Regulated |
---|---|---|---|---|---|---|
hsa04151 | PI3K-Akt signaling pathway | 1.39 | 3.30 × 10−18 | 2.89 × 10−14 | PIK3CB, PIK3R3, PIK3R1, EGF, ITGA5, LPAR2, FGF22, PDGFA, GNG11, SYNE1, COL1A1, FGF1, MAP2K2, MAP3K5, COL4A6, LAMA2, LAMB1 | SPP1, MMP9, ITGA5, COL1A1, COL4A6, LAMB1, LAMA2, LAMA4, COL4A5, LAMB2, COL4A4, LAMC2, COL4A3, COL4A2 |
hsa04068 | FoxO signaling pathway | 1.95 | 1.12 × 10−15 | 3.34 × 10−8 | MAP3K5, FOXO4, MAP2K2, BAD, PIK3CB, PIK3R3, PIK3R1, GABARAPL1, CALM3, GABARAP, CALM2, CALM1 | MAP3K5, PIK3CB, PIK3R3, PIK3R1, SOX4, IRS1, BAD, GABARAPL1, GABARAP, CALM3, CALM2, CALM1 |
hsa04140 | Autophagy—animal | 1.73 | 2.00 × 10−13 | 1.16 × 10−13 | IRS1, PIK3CB, PIK3R3, PIK3R1, MAP3K5, ATG2B, ATG4C, ATG7, RB1CC1, STX17, GABARAPL1, GABARAP, CALM3, CALM2, CALM1 | IRS1, PIK3CB, PIK3R3, PIK3R1, MAP3K5, ATG2B, ATG4C, ATG7, RB1CC1, STX17, GABARAPL1, GABARAP, CALM3, CALM2, CALM1 |
hsa04728 | Dopaminergic synapse | 1.36 | 3.84 × 10−9 | 1.09 × 10−4 | MAOA, VMAT2, GNAI1, GNG5, GNG3, GNG4, GNAQ, CAMK2B, CAMK2A, GRIN1, GRIA1, GRID1, GABRA2, CALM2, CALM1 | SLC6A3, LEP, CALM1, CALM2, CAMK2A, CAMK2B, GNG5, GNG4, GNG3, GNAI1 |
hsa04920 | Adipocytokine signaling pathway | 1.29 | 3.45 × 10−8 | 1.04 × 10−2 | CRPRA, ACSL1, LEP, POMC, ADIPOR1, STK11, RXRA, SLC2A1, TNFSF4, CAMKK2 | NR1H3, PTPN11, SOCS3, IRS1, RXRA, PRKAA1, PRKAA2, LEP, RXRG, AKT3, MAPK8, STAT1, STAT3 |
hsa00190 | Oxidative phosphorylation | 1.09 | 7.27 × 10−6 | 1.01 × 10−5 | NUDFB8, COX6C, TTC19, ATP6V2, ATP6V1B2, ATP6V1E1, COX7A2, ATP6V1G2 | NUDFB6, NUDFB4, NDUFA1, NUDFB9, NUDFB5, NDUFA2, COX6C, ATP6V1E1, ATP6V1B2 |
hsa04148 | Efferocytosis | 1.32 | 3.10 × 10−5 | 1.30 × 10−3 | AGER, GRK6, ROCK1, ERBB4, MAP3K5, BNIP3L, ITGA5, SYNE1, MAP2K2, GABARAPL1, PIK3CB, PIK3R1, PIK3R3, COL4A6, LAMA2, LAMB1 | METRNL, OXTR, ITGA1, S1PR1, CD47, HVCN1, LRP1, VAV1, TUBB2B, CSF1R, GRK6, BNIP3L, MAP3K5, SYNE1, AGER |
hsa05208 | Chemical carcinogenesis—reactive oxygen species | 1.23 | 1.05 × 10−6 | 1.04 × 10−5 | PIK3CB, MAP3K5, MAP2K2, FGF1, CYBA, NCF1, NCF2, NDUFA2, COX6C, PRDX2, PRDX6, PRDX1, MAPKAPK2, MAPKAPK3, KEAP1, ASMT, SOD2 | PIK3CB, MAP3K5, MAP2K2, FGF1, CYBA, NCFA, NCF2, PRDX2, PRDX1, PRDX6, MAPKAPK2, MAPKAPK3, KEAP1, ASMT, SOD2 |
hsa04710 | Circadian rhythm | 2.89 | 3.91 × 10−16 | 4.62 × 10−3 | DBP, BHLHE41 | RORA, PER1, PER2, PER3, CLOCK |
hsa04070 | Phosphatidylinositol signaling system | 1.35 | 6.61 × 10−8 | 3.73 × 10−2 | PTEN, CALM1, CALM2, PIK3R1, PIK3R3, DGKZ, GRIN1, CALM3, CALML4, CAMK2A, CAMK2B, CAMK2G, MAP2K2, MAP3K5 | PIK3CB, MAP3K5, MAP2K2, GRIN1, CALM1, CALM2, CALM3, CAMK2A, CAMK2B, CAMK2G |
hsa04713 | Circadian entrainment | 1.29 | 4.36 × 10−7 | 4.85 × 10−3 | GRIN1, CALM1, CALM2, CAMK2A, CAMK2B, CAMK2G, MAP2K2, MAP3K5 | GRIN1, CALM1, CALM2, CAMK2A, CAMK2B, CAMK2G |
hsa04216 | Ferroptosis | 1.57 | 2.45 × 10−8 | 4.60 × 10−2 | TP53, SLC1A4, PINK1, SLC7A11, ACSL4 | TP53, SLC1A4, PINK1, SLC7A11, ACSL4 |
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content. |
© 2025 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
Share and Cite
Mogavero, M.P.; Marchese, G.; Ventola, G.M.; Lanza, G.; Bruni, O.; Ferini-Strambi, L.; Ferri, R. Exploring the Role of Ferroptosis in the Pathophysiology and Circadian Regulation of Restless Legs Syndrome. Biomolecules 2025, 15, 1184. https://doi.org/10.3390/biom15081184
Mogavero MP, Marchese G, Ventola GM, Lanza G, Bruni O, Ferini-Strambi L, Ferri R. Exploring the Role of Ferroptosis in the Pathophysiology and Circadian Regulation of Restless Legs Syndrome. Biomolecules. 2025; 15(8):1184. https://doi.org/10.3390/biom15081184
Chicago/Turabian StyleMogavero, Maria Paola, Giovanna Marchese, Giovanna Maria Ventola, Giuseppe Lanza, Oliviero Bruni, Luigi Ferini-Strambi, and Raffaele Ferri. 2025. "Exploring the Role of Ferroptosis in the Pathophysiology and Circadian Regulation of Restless Legs Syndrome" Biomolecules 15, no. 8: 1184. https://doi.org/10.3390/biom15081184
APA StyleMogavero, M. P., Marchese, G., Ventola, G. M., Lanza, G., Bruni, O., Ferini-Strambi, L., & Ferri, R. (2025). Exploring the Role of Ferroptosis in the Pathophysiology and Circadian Regulation of Restless Legs Syndrome. Biomolecules, 15(8), 1184. https://doi.org/10.3390/biom15081184