Sulforaphane attenuates oxidative stress, senescence, and ferroptosis induced by cigarette smoke extract in vitro and in vivo via upregulating the expression of SIRT1

Amin Reihani, Ebrahim Mohammadi, Fereshteh Talebpour Amiri, Mohammad Seyedabadi, Fatemeh Shaki

Abstract


Background and purpose: Cigarette smoking induces lung toxicity by triggering oxidative stress, leading to apoptosis, ferroptosis, and senescence. Sulforaphane (SFN), a potent antioxidant, activates the SIRT1 pathway, enhancing cellular stress resistance and survival. This study aimed to evaluate the protective effects of SFN against cigarette smoke extract (CSE)-induced damage in human airway epithelial cells (BEAS-2B) and in mouse lungs, focusing on its role in upregulating SIRT1 expression.

Experimental approach: BEAS-2B cells were treated with CSE and SFN, and cell viability was assessed using the MTT assay. Cellular senescence was assessed using the SA-β-gal assay and the expression of genes associated with senescence (p16 and p21). The expression levels of SIRT1, senescence-associated secretory phenotype (SASP) cytokines (IL-1β, IL-6, IL-8, TNF-α), GPX4, and SLC7A11 were quantified using qRT-PCR. Additionally, ROS production, GSH and MDA levels, and iron content were measured. An emphysema mouse model was induced by intraperitoneal administration of CSE (7.2 mg/kg) alone or in combination with SFN (10.2 mg/kg) over 28 days, and subsequent histopathological changes were evaluated.

Findings/Results: Our findings revealed that SFN co-treatment effectively mitigated CSE-induced cytotoxicity, senescence, and SASP cytokine secretion, as well as the pronounced emphysematous changes in lung tissues. Furthermore, SFN reversed CSE-induced downregulation of SIRT1 and upregulation of NF-κB. Notably, SFN also inhibited CSE-induced ferroptosis by increasing GPX4 and SLC7A11 expression while reducing iron and MDA levels.

Conclusion and implications: The findings of the present study demonstrated that sulforaphane offers protective effects against CSE-induced toxicity by mitigating oxidative stress, ferroptosis, and cellular senescence.

 

 


Keywords


Cellular senescence; Cigarette smoke extract; Ferroptosis; SIRT1; Sulforaphane.

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References


Smith CJ, Hansch C. The relative toxicity of compounds in mainstream cigarette smoke condensate. Food Chem Toxicol. 2000;38(7):637-46.DOI: 10.1016/s0278-6915(00)00051-x.

Patel RR, Ryu JH, Vassallo R. Cigarette smoking and diffuse lung disease. Drugs. 2008;68(11):1511-1527.DOI: 10.2165/00003495-200868110-00004.

Zeng XL, Yang XN, Liu XJ. Resveratrol attenuates cigarette smoke extract induced cellular senescence in human airway epithelial cells by regulating the miR-34a/SIRT1/NF-κB pathway.Medicine. 2022;101(46):e31944,1-8.DOI:10.1097/md.0000000000031944.

Yang SR, Wright J, Bauter M, Seweryniak K, Kode A, Rahman I. Sirtuin regulates cigarette smoke-induced proinflammatory mediator release via RelA/p65 NF-κB in macrophages in vitro and in rat lungs in vivo:implications for chronic inflammation and aging. Am J Physiol Lung Cell Mol Physiol.2007;292(2):L567-L576.DOI: 10.1152/ajplung.00308.2006.

Satrialdi, Pratiwi C, Khaeranny RN, Mudhakir D. The development of mitochondria-targeted quercetin for rescuing Sertoli cells from oxidative stress. Res Pharm Sci. 2025;20(1):109-120.DOI: 10.4103/RPS.RPS_226_23.

Ma N, Deng TT, Wang Q, Luo ZL, Zhu CF, Qiu JF, et al. Erythromycin regulates cigarette smoke-induced proinflammatory mediator release through sirtuin 1-nuclear factor κB axis in macrophages and mice lungs. Pathobiology. 2019;86(5-6):237-247.DOI: 10.1159/000500628.

Barnes PJ. Senescence in COPD and its comorbidities. Annu Rev Physiol.

;79:517-639.DOI: 10.1146/annurev-physiol-022516-034314.

Zhang Y, Huang W, Zheng Z, Wang W, Yuan Y, Hong Q, et al. Cigarette smoke-inactivated SIRT1 promotes autophagy-dependent senescence of alveolar epithelial type 2 cells to induce pulmonary fibrosis. Free Radic Biol Med. 2021;166:116-127.DOI: 10.1016/j.freeradbiomed.2021.02.013.

Easter M, Bollenbecker S, Barnes JW, Krick S. Targeting aging pathways in chronic obstructive pulmonary disease. Int J Mol Sci. 2020;21(18):6924,1-17.DOI: 10.3390/ijms21186924.

Campisi J. Aging, cellular senescence, and cancer. Annu Rev Physiol. 2013;75:685-705.DOI: 10.1146/annurev-physiol-030212-183653.

González-Gualda E, Baker AG, Fruk L, Muñoz-Espín D. A guide to assessing cellular senescence in vitro and in vivo. FEBS. 2021;288(1):56-80.DOI: 10.1111/febs.15570.

van Deursen JM. The role of senescent cells in ageing. Nature. 2014;509(7501):439-446.DOI: 10.1038/nature13193.

Du Y, Ding Y, Shi T, He W, Mei Z, Feng X, et al. Suppression of circXPO1 attenuates cigarette smoke-induced inflammation and cellular senescence of alveolar epithelial cells in chronic obstructive pulmonary disease. Int Immunopharmacol. 2022;111:109086.DOI: 10.1016/j.intimp.2022.109086.

Xia H, Wu Y, Zhao J, Cheng C, Lin J, Yang Y, et al. N6-Methyladenosine-modified circSAV1 triggers ferroptosis in COPD through recruiting YTHDF1 to facilitate the translation of IREB2. Cell Death Differ. 2023;30(5):1293-1304.DOI: 10.1038/s41418-023-01138-9.

Feng L, Sun J, Xia L, Shi Q, Hou Y, Zhang L, et al. Ferroptosis mechanism and Alzheimer's disease. Neural Regen Res. 2024;19(8):1741-1750.DOI: 10.4103/1673-5374.389362.

Chen X, Li J, Kang R, Klionsky DJ, Tang D. Ferroptosis:machinery and regulation. Autophagy. 2021;17(9):2054-2081.DOI: 10.1080/15548627.2020.1810918.

Riegman M, Sagie L, Galed C, Levin T, Steinberg N, Dixon SJ, et al. Ferroptosis occurs through an osmotic mechanism and propagates independently of cell rupture. Nat Cell Biol. 2020;22(9):1042-1048.DOI: 10.1038/s41556-020-0565-1.

Chen J, Deng X, Xie H, Wang C, Huang J, Lian N. Circular RNA_0025843 alleviated cigarette smoke extract induced bronchoalveolar epithelial cells ferroptosis. Int J Chron Obstruct Pulmon Dis. 2024;19:363-374.DOI: 10.2147/copd.s444402.

Haigis MC, Sinclair DA. Mammalian sirtuins:biological insights and disease relevance. Annu Rev Pathol. 2010;5(1):253-295.DOI: 10.1146/annurev.pathol.4.110807.092250.

Hall JA, Dominy JE, Lee Y, Puigserver P. The sirtuin family's role in aging and age-associated pathologies. J Clin Invest. 2013;123(3):973-979.DOI: 10.1172/JCI64094.

Yao H, Sundar IK, Ahmad T, Lerner C, Gerloff J, Friedman AE, et al. SIRT1 protects against cigarette smoke-induced lung oxidative stress via a FOXO3-dependent mechanism. Am J Physiol Lung Cell Mol. 2014;306(9):L816-L28.DOI: 10.1152/ajplung.00323.2013.

Ebrahim NA, Elnagar MR, El-Gamal R, Habotta OA, Albadawi EA, Albadrani M, et al. Melatonin mitigates doxorubicin induced chemo brain in a rat model in a NRF2/p53-SIRT1 dependent pathway. Heliyon. 2024;10(19):e38081,1-15.DOI:10.1016/j.heliyon.2024.e38081.

Guarente L. Sirtuins, aging, and medicine. N Engl J Med. 2011;364(23):2235-2244.DOI: 10.1056/NEJMra1100831.

Biswas S, W Hwang J, A Kirkham P, Rahman I. Pharmacological and dietary antioxidant therapies for chronic obstructive pulmonary disease. Curr Med Chem. 2013;20(12):1496-1530.DOI: 10.2174/0929867311320120004.

Danilov CA, Chandrasekaran K, Racz J, Soane L, Zielke C, Fiskum G. Sulforaphane protects astrocytes against oxidative stress and delayed death caused by oxygen and glucose deprivation. Glia. 2009;57(6):645-656.DOI: 10.1002/glia.20793.

Juge N, Mithen RF, Traka M. Molecular basis for chemoprevention by sulforaphane:a comprehensive review. Cell. Mol Life Sci. 2007;64(9):1105-1127.DOI: 10.1007/s00018-007-6484-5.

Ping Z, Liu W, Kang Z, Cai J, Wang Q, Cheng N, et al. Sulforaphane protects brains against hypoxic-ischemic injury through induction of Nrf2-dependent phase 2 enzyme. Brain Res. 2010;1343:178-185.DOI: 10.1016/j.brainres.2010.04.036.

Zhang Y, Cao J, Chen Y, Chen P, Peng H, Cai S, et al. Intraperitoneal injection of cigarette smoke extract induced emphysema, and injury of cardiac and skeletal muscles in BALB/C mice. Exp Lung Res. 2013;39(1):18-31.DOI: 10.3109/01902148.2012.745910.

Reihani A, Shaki F, Azari A. Zinc oxide nanoparticles decrease acrylamide cytotoxicity and oxidative stress in HepG2 cells. Curr Res Nutr Food Sci. 2025;55(3):481-492.DOI: 10.1108/NFS-07-2023-0147.

Zhao Z, Xu Z, Chang J, He L, Zhang Z, Song X, et al. Sodium pyruvate exerts protective effects against cigarette smoke extract-induced ferroptosis in alveolar and bronchial epithelial cells through the GPX4/Nrf2 axis. J Inflamm. 2023;20(1):28,1-11.DOI: 10.1186/s12950-023-00347-w.

Uchil PD, Nagarajan A, Kumar P. Assay for β-galactosidase in extracts of mammalian cells. Cold Spring Harb Protoc. 2017;2017(10):837-841.DOI: 10.1101/pdb.prot095778.

Jiao Z, Chang J, Li J, Nie D, Cui H, Guo D. Sulforaphane increases Nrf2 expression and protects alveolar epithelial cells against injury caused by cigarette smoke extract. Mol Med Rep. 2017;16(2):1241-1247. DOI: 10.3892/mmr.2017.6700.

Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods. 2001;25(4):402-408.DOI:10.1006/meth.2001.1262.

Jalili C, Korani M, Pazhouhi M, Ghanbari A, Zhaleh M, Davoudi S, et al. Protective effect of gallic acid on nicotine-induced testicular toxicity in mice. Res Pharm Sci. 2021;16(4):414-424.DOI: 10.4103/1735-5362.319579.

Chiaradia E, Sansone A, Ferreri C, Tancini B, Latella R Tognoloni A, et al. phospholipid fatty acid remodeling and carbonylated protein increase in extracellular vesicles released by airway epithelial cells exposed to cigarette smoke extract. Eur J Cell Biol. 2023;102(1):151285,1-11.DOI: 10.1016/j.ejcb.2022.151285.

Guo P, Pi H, Xu S, Zhang L, Li Y, Li M, et al. Melatonin improves mitochondrial function by promoting MT1/SIRT1/PGC-1 alpha-dependent mitochondrial biogenesis in cadmium-induced hepatotoxicity in vitro. Toxicol Sci. 2014;142(1):182-195.DOI: 10.1093/toxsci/kfu164.

Bendary MM, Ali MAM, Abdel Halim AS, Boufahja F, Chaudhary AA, Elkelish A, et al. Investigating Sulforaphane's anti-virulence and anti-quorum sensing properties against Pseudomonas aeruginosa. Front Pharmacol. 2024;15:1406653,1-21.DOI:10.3389/fphar.2024.1406653.

Yasuda S, Horinaka M, Sakai T. Sulforaphane enhances apoptosis induced by Lactobacillus pentosus strain S-PT84 via the TNFα pathway in human colon cancer cells. Oncol Lett. 2019;18(4):4253-4261.DOI: 10.3892/ol.2019.10739.

Rajaei N, Rahgouy G, Panahi N, Razzaghi-Asl N. Bioinformatic analysis of highly consumed phytochemicals as P-gp binders to overcome drug-resistance. Res Pharm Sci. 2023;18(5): 505-516.DOI: 10.4103/1735-5362.383706.

Zanichelli F, Capasso S, Cipollaro M, Pagnotta E, Cartenì M, Casale F, et al. Dose-dependent effects of R-sulforaphane isothiocyanate on the biology of human mesenchymal stem cells, at dietary amounts, it promotes cell proliferation and reduces senescence and apoptosis, while at anti-cancer drug doses, it has a cytotoxic effect. Age (Dordrecht, Netherlands). 2012;34(2):281-293.DOI: 10.1007/s11357-011-9231-7.

Jackson SJ, Singletary KW. Sulforaphane inhibits human MCF-7 mammary cancer cell mitotic progression and tubulin polymerization. J Nutr. 2004;134(9):2229-2236.DOI: 10.1093/jn/134.9.2229.

Wu H, Ma H, Wang L, Zhang H, Lu L, Xiao T, et al. Regulation of lung epithelial cell senescence in smoking-induced COPD/emphysema by microR-125a-5p via Sp1 mediation of SIRT1/HIF-1a. Int J Biol Sci. 2022;18(2):661-674.DOI: 10.7150/ijbs.65861.

Karakuyu NF, Özseven A, Akın SE, Çamaş HE, Özmen Ö, Cengiz Ç. L-carnitine protects the lung from radiation-induced damage in rats via the AMPK/SIRT1/TGF-1ß pathway. Naunyn Schmiedebergs Arch Pharmacol. 2024;397(10):8043-8051.DOI: 10.1007/s00210-024-03157-w.

Yeung F, Hoberg JE, Ramsey CS, Keller MD, Jones DR, Frye RA, et al. Modulation of NF-kappaB-dependent transcription and cell survival by the SIRT1 deacetylase. EMBO J. 2004;23(12):2369-23480.DOI: 10.1038/sj.emboj.7600244.

Deng M, Tong R, Bian Y, Hou G. Astaxanthin attenuates cigarette smoking-induced oxidative stress and inflammation in a sirtuin 1-dependent manner. Biomed Pharmacother. 2023;159:114230,1-14.DOI: 10.1016/j.biopha.2023.114230.

Chen W, Sun Z, Wang XJ, Jiang T, Huang Z, Fang D, et al. Direct interaction between Nrf2 and p21(Cip1/WAF1) upregulates the Nrf2-mediated antioxidant response. Mol Cell. 2009;34(6):663-673.DOI: 10.1016/j.molcel.2009.04.029.

Kubo E, Chhunchha B, Singh P, Sasaki H, Singh DP. Sulforaphane reactivates cellular antioxidant defense by inducing Nrf2/ARE/Prdx6 activity during aging and oxidative stress. Sci Rep. 2017;7(1):14130,1-17.DOI: 10.1038/s41598-017-14520-8.

Yu Z, He W, Shi W. Sulforaphane (Sul) reduces renal interstitial fibrosis (RIF) by controlling the inflammation and TGF-β/Smad signaling pathway. Appl Biol Chem. 2024;67(1):8,1-11.DOI: 10.1186/s13765-024-00858-x.

Pogorzelska A, Świtalska M, Wietrzyk J, Mazur M, Milczarek M, Medyńska K, et al. Antitumor and antimetastatic effects of dietary sulforaphane in a triple-negative breast cancer models. Sci Rep. 2024;14(1):16016,1-11.DOI: 10.1038/s41598-024-65455-w.

Yan M, Xu S, Wang H, Dong S, Mo C. Ferroptosis in chronic obstructive pulmonary disease:from cellular mechanisms to therapeutic applications. Chin Med J. 2024;137(10):1237-1239.DOI: 10.1097/cm9.0000000000003079.

Terzi EM, Sviderskiy VO, Alvarez SW, Whiten GC, Possemato R. Iron-sulfur cluster deficiency can be sensed by IRP2 and regulates iron homeostasis and sensitivity to ferroptosis independent of IRP1 and FBXL5. Sci Adv. 2021;7(22):1-11.DOI: 10.1126/sciadv.abg4302.


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