D vitamini eksikliği ve 1,25(OH)2D3 uygulamasının etanole bağlı karaciğer hasarında oksidatif stres ve Nrf2-antioksidan sinyal sistemi üzerine etkisi

Year 2021, Volume: 6 Issue: 3, 127 - 133, 09.12.2021

İlknur Bingül , Fatih Aydın , Canan Kucukgergin , Asiye Işın Doğan Ekici , Semra Dogru Abbasoglu , Mujdat Uysal

Abstract

Amaç: D vitamini eksikliğinin (VDE) kronik karaciğer hastalıklarında karaciğer hasarını arttırdığı ileri
sürülmektedir. Bununla birlikte, VDE ile alkole bağlı karaciğer hasarı arasındaki ilişkiye ait bilgiler sınırlıdır.
Bu nedenle, bu çalışmada VDE'nin etanol (EtOH) ile indüklenen karaciğer hasarı üzerindeki etkisi araştırıldı.
Ayrıca, EtOH uygulanan sıçanlarda 1,25(OH)2D3'ün karaciğeri koruyucu potansiyelinde Nrf2-antioksidan sinyal
yolunun rolü de araştırıldı.
Yöntemler: Erkek Wistar sıçanlar 12 hafta süreyle VDE diyet ile beslendi. Son 8 hafta içme suyunda artan
konsantrasyonlarda EtOH (% 5-20) verildi. Ayrıca, bir grup sıçana bu süreçte 11,25(OH)2D3 (5μg/kg; haftada iki
kez; i.p.) uygulandı. Karaciğerde trigliserit (TG) ve hidroksiprolin düzeyleri, inflamasyon göstergeleri, lipid
peroksitler, protein karboniller, Nrf2, süperoksit dismutaz (SOD) ve glutatyon peroksidaz (GSH-Px)’ın mRNA
ekspresyonları, SOD ve GSH-Px aktiviteleri, glutatyon düzeyleri ve histopatolojik incelemeler yapıldı.
Bulgular: EtOH uygulaması steatoz ve fibroza, karaciğerde TG, lipit peroksit, protein karbonil ve hidroksiprolin
düzeyleri ile inflamasyon göstergelerinde artışa neden oldu. VDE, EtOH’a bağlı karaciğer hasarını, steatoz ve
fibrozu yoğunlaştırmadı. Ancak, reaktif oksijen türleri ve lipit peroksit düzeylerinde VDE+EtOH grubunda
ılımlı artış bulundu. Nrf2-SOD-GSH-Px’in gen ekspresyonları, enzim aktiviteleri ve glutatyon düzeyleri de
VDE+EtOH grubunda EtOH grubundan daha yüksekti. Ayrıca, 1,25(OH)2D3, EtOH uygulanan sıçanlarda
karaciğerde SOD ve GSH-Px’ın mRNA ekspresyonlarını ve aktivitelerini arttırdı.
Sonuç: Bulgularımız VDE diyetin EtOH ile indüklenen karaciğer hasarı üzerinde ek bir etki oluşturmadığını
gösterdi. Ayrıca EtOH’a bağlı karaciğer hasarında 11,25(OH)2D3'ün koruyucu etkisinde Nrf2-antioksidan sinyal
yolunun aktivasyonunun rol oynayabileceği saptandı. 

Keywords

D vitamini eksikliği, 1,25(OH)2D3, Etanol, Karaciğer hasarı, Oksidatif stres, Nrf2

Supporting Institution

İstanbul Üniversitesi Bilimsel Araştırma Projeleri Koordinasyon Birimi

Project Number

TSA-2018-30443

Thanks

Bu çalışma İstanbul Üniversitesi Araştırma Fonu tarafından desteklenmiştir

The effect of vitamin D deficiency and 1,25(OH)2D3 treatment on oxidative stress and Nrf2-antioxidant signaling in ethanol-induced hepatotoxicity

Abstract

Aim: Vitamin D deficiency (VDD) is suggested to enhance hepatotoxicity in chronic liver diseases. However,
there is limited knowledge about the association between VDD and alcoholic liver damage. Therefore, the effect
of VDD on ethanol (EtOH)-induced hepatotoxicity was investigated in this study. Moreover, the role of the
Nrf2-antioxidant signaling pathway in the hepatoprotective potential of 1,25(OH)2D3 was also searched in
EtOH-treated rats.
Methods: Male Wistar rats were fed on VDD-diet for 12 weeks. EtOH (5-20%) was applied in drinking water in
increasing concentrations for the last 8 weeks. In addition, one group of rats were injected with 1,25(OH)2D3
(5μg/kg; twice a week; i.p.) during this period. Hepatic triglyceride and hydroxyproline levels, inflammation
markers, lipid peroxides, protein carbonyls, mRNA expressions of Nrf2, superoxide dismutase (SOD), and
glutathione peroxidase (GSH-Px), SOD and GSH-Px activities, glutathione levels and histopathology were
examined.
Results: EtOH application caused steatosis and fibrosis, elevated hepatic TG, lipid peroxide, protein carbonyls
and hydroxyproline levels and inflammation markers. VDD did not aggravate EtOH-induced liver damage,
steatosis and inflammation, but reactive oxygen species and lipid peroxide levels were slightly increased in
VDD+EtOH group. Gene expressions of Nrf2-SOD-GSH-Px, enzyme activities and glutathione levels were also
higher in VDD+EtOH group than EtOH group. Additionally, 1,25(OH)2D3 elevated mRNA expressions and
activities of SOD and GSH-Px in EtOH-treated rats.
Conclusion: Our results indicate that VDD diet did not cause an additive effect on EtOH-induced hepatotoxicity.
Moreover, it was detected that the activation of Nrf2-antioxidant signaling pathway may play a role in the
protective effect of 1,25(OH)2D3 against EtOH-induced hepatotoxicity

Keywords

Vitamin D deficiency, 1,25(OH)2D3, Ethanol, Liver damage, Oxidative stress, Nrf2

Project Number

TSA-2018-30443

References

• 1. Kennel KA, Drake MT, Hurley DL. Vitamin D deficiency in adults: when to test and how to treat. Mayo Clin Proc. 2010; 85: 752-757
• 2. Wang H, Chen W, Li D, Yin X, Zhang X, Olsen N, et al. Vitamin D and chronic diseases. Aging Dis. 2017; 8: 346-353.
• 3. Uberti F, Morsanuto V, Molinari C. Vitamin D in oxidative stress and diseases. ‘A Critical Evaluation of Vitamin D-Basic Overview. Ed: Sivakumar Joghi Thatha Gowder, IntechOpen, Chapter 2, p.47-73. 2017.
• 4. Berridge MJ. Vitamin D signalling in health and disease. Biochem Biophys Res Commun. 2015; 460: 53-71.
• 5. Iruzubieta P, Terán Á, Crespo J, Fábrega E. Vitamin D deficiency in chronic liver disease. World J Hepatol. 2014; 6: 901-915.
• 6. Elangovan H, Chahal S, Gunton JE. Vitamin D in liver disease: Current evidence and potential directions. Biochim Biophys Acta 2017; 1863: 907-916.
• 7. Sakaguchi S, Takahashi S, Sasaki T, Kumagai T, Nagata K. Progression of alcoholic and non-alcoholic steatohepatitis: Common metabolic aspects of innate immune system and oxidative stress. Drug Metab Pharmacokinet. 2011; 26: 30-46.
• 8. Zhu CG, Liu YX, Wang H, Wang BP, Qu HQ, Wang BL, et al. Active form of vitamin D ameliorates non-alcoholic fatty liver disease by alleviating oxidative stress in a high-fat diet rat model. Endocr J. 2017; 64: 663-673.
• 9. Maia-Ceciliano TC, Dutra RR, Aguila MB, Mandarim-De-Lacerda CA. The deficiency and the supplementation of vitamin D and liver: Lessons of chronic fructose-rich diet in mice. J Steroid Biochem Mol Biol. 2019; 192: 105399.
• 10. Karatayli E, Stokes CS, Lammert F. Vitamin D in preclinical models of fatty liver disease. Anticancer Res. 2020; 40: 527-534.
• 11. Bingul I, Aydın AF, Küçükgergin C, Dogan-Ekici AI, Dogru-Abbasoglu S, Uysal MM. The effect of 1,25-dihydroxyvitamin D3 on liver damage, oxidative stress and advanced glycation end products in experimental nonalcoholic- and alcoholic- fatty liver disease. Turk J Med Sci. 2021 (doi: 10.3906/sag-2007-289).
• 12. Kong M, Zhu L, Bai L, Zhang X, Chen Y, Liu S, et al. Vitamin D deficiency promotes nonalcoholic steatohepatitis through impaired enterohepatic circulation in animal model. Am J Physiol Gastrointest Liver Physiol. 2014; 307: G883-G893.
• 13. Su D, Nie Y, Zhu A, Chen Z, Wu P, Zhang L, et al. Vitamin D signaling through induction of paneth cell defensins maintains gut microbiota and improves metabolic disorders and hepatic steatosis in animal models. Front Physiol. 2016; 7: 498.
• 14. Liu XJ, Wang BW, Zhang C, Xia MZ, Chen YH, Hu CQ, et al. Vitamin D deficiency attenuates high-fat diet-induced hyperinsulinemia and hepatic lipid accumulation in male mice. Endocrinology 2015; 156 :2103-2113.
• 15. Lee K. Sex-specific relationships between alcohol consumption and vitamin D levels: The Korea National Health and Nutrition Examination Survey 2009. Nutr Res Pract. 2012; 6: 86-90. 16. Anthy R, Canivet CM, Patouraux S, Ferrari-Panaia P, Saint-Paul MC, et al. Severe vitamin D deficiency may be an additional cofactor for the occurrence of alcoholic steatohepatitis. Alcohol Clin Exp Res. 2015; 39: 1027-1033.
• 17. Tardelli VS, do Lago MPP, da Silveira DX, Fidalgo TM. Vitamin D and alcohol: A review of the current literature. Psychiat Res. 2017; 248: 83-86.
• 18. Zhang H, Xue L, Li B, Zhang Z, Tao S. Vitamin D protects against alcohol-induced liver cell injury within an NRF2-ALDH2 feedback loop. Mol Nutr Food Res. 2019; 63: e1801014.
• 19. Yuan F, Xu Y, You K, Zhang J, Yang F, Li YX. Calcitriol alleviates ethanol-induced hepatotoxicity via AMPK/mTOR-mediated autophagy. Arch Biochem Biophys. 2021; 697: 108694.
• 20. Trepo E, Ouziel R, Pradat P, Momozawa Y, Quertinmont E, Gervy C, et al. Marked 25-hydroxyvitamin D deficiency is associated with poor prognosis in patients with alcoholic liver disease. J Hepatol. 2013; 59: 344-350.
• 21. Hu CQ, Bo QL, Chu LL, Hu YD, Fu L, Wang GX, et al. Vitamin D deficiency aggravates hepatic oxidative stress and inflammation during chronic alcohol-induced liver injury in mice. Oxid Med Cell Longev. 2020; 2020: 5715893.
• 22. Mokhtari Z, Hekmatdoost A, Nourian M. Antioxidant efficacy of vitamin D. J Parathyroid Dis. 2017; 5: 11-16.
• 23. Nakai, H. Fujii, K.Kono, Goto S, Kitazawa R, Kitazawa S, et al. Vitamin D activates the Nrf2-Keap1 antioxidant pathway and ameliorates nephropathy in diabetic rats, Am J Hypertens. 2014; 27: 586-595.
• 24. Saad El-Din S, El-Din S, Rashed L, Medhat E, Aboulhoda BE, Badawy D , et al. Active form of vitamin D analogue mitigates neurodegenerative changes in Alzheimer's disease in rats by targeting Keap1/Nrf2 and MAPK-38p/ERK signaling pathways. Steroids 2020; 156: 108586.
• 25. Folch J, Lees M, Stanley GHS. A simple method for the isolation and purification of total lipids from animal tissues. J Biol Chem. 1957; 226: 497-500.
• 26. Rachmilewitz D, Stamler JS, Karmeli F, Mullins ME, Singel DJ, Loscalzo J, et al. Peroxynitrite-induced rat colitis - a new model of colonic inflammation. Gastroenterology 1993; 105: 1681-1688.
• 27. Wang H, Joseph JA. Quantifying cellular oxidative stress by dichlorofluorescein assay using microplate reader. Free Radic Biol Med. 1999; 27: 612-616.
• 28. Buege JA, Aust SD. Microsomal lipid peroxidation. Methods Enzymol. 1978; 52: 302–310.
• 29. Reznick AZ, Packer L. Oxidative damage to proteins: spectrophotometric method for carbonyl assay. Methods Enzymol 1994; 233: 357–363.
• 30. Benzie IFF, Strain JJ. The ferric reducing ability of plasma (FRAP) as a measure of ‘antioxidant power’’: the FRAP assay. Anal Biochem. 1996; 239: 70-76.
• 31. Beutler E, Duron O, Kelly BM. Improved method for the determination of blood glutathione. J Lab Clin Med. 1963; 61: 882-888.
• 32. Mylorie AA, Collins H, Umbles C, Kyle J. Erythrocyte superoxide dismutase activity and other parameters of copper status in rats ingesting lead acetate. Toxicol Appl Pharmacol. 1986; 82: 512-520.
• 33. Lawrence RA, Burk RF. Glutathione peroxidase activity in selenium deficient rat liver. Biochem Biophys Res Commun. 1976; 71: 952-958.
• 34. Smith PK, Krohn RI, Hermanson GT, Mallia AK, Gartner FH, Provenzano MD, et al. Measurement of protein using bicinchoninic acid. Anal Biochem. 1985; 150: 76-85.
• 35. Bingül I, Aydın AF, Başaran-Küçükgergin C, Doğan-Ekici I, Çoban J, Doğru-Abbasoğlu S, et al. High-fat diet plus carbon-tetrachloride-induced liver fibrosis is alleviated by betaine treatment in rats. Int Immunopharmacol. 2016; 39: 199-207.
• 36. Kawaratani H, Tsujimoto T, Douhara A, Takaya H, Moriya K, Namisaki T, et al. The effect of inflammatory cytokines in alcoholic liver disease. Mediators Inflamm. 2013; 2013: 495156.
• 37. Wu D, Cederbaum AI. Oxidative stress and alcoholic liver disease. Semin Liver Dis. 2009; 29: 141-154.
• 38. You M, Arteel GE. Effect of ethanol lipid metabolism. J Hepatol. 2019; 70: 237-248.
• 39. Balkan J, Kanbağlı Ö, Aykaç-Toker G, Uysal M. Taurine treatment reduces hepatic lipids and oxidative stress in chronically ethanol-treated rats. Biol Pharmaceut Bull. 2002; 25: 1231-1233.
• 40. Prathibha P, Rejitha S, Harikrishnan R, Das SS, Abhilash PA, Indira M. Additive effect of alpha-tocopherol and ascorbic acid in combating ethanol-induced hepatic fibrosis. Redox Rep. 2013; 18: 36-46.
• 41. Liu J, Wang X, Liu R, Liu Y, Zhang T, Fu H, et al. Oleanolic acid co-administration alleviates ethanol-induced hepatic injury via Nrf-2 and ethanol-metabolizing modulating in rats. Chem Biol Interact. 2014; 221: 88-98.
• 42. Xiong ZE, Dong WG, Wang BY, Tong QY, Li ZY. Curcumin attenuates chronic ethanol-induced liver injury by inhibition of oxidative stress via mitogen-activated protein kinase/nuclear factor E2-related factor 2 pathway in mice. Pharmacogn Mag. 2015; 11: 707-715. 43. Song M, Chen T, Prough RA, Cave MC, McClain CJ. Chronic alcohol consumption causes liver injury in high-fructose-fed male mice through enhanced hepatic inflammatory response. Alcohol Clin Exp Res. 2016; 40: 518-528.
• 44. Nagappan A, Kim JH, Jung DY, Jung MH. Cryptotanshinone from the Salvia miltiorrhiza Bunge attenuates ethanol-induced liver injury by activation of AMPK/SIRT1 and Nrf2 signaling pathways. Int J Mol Sci. 2020; 21: 265.
• 45. Mallya SM, Corrado KR, Saria EA, Yuan FNF, Tran HQ, Saucier K, et al. Modeling vitamin D deficiency and moderate deficiency in adult mice via dietary cholecalciferol restriction. Endocrin Res. 2016; 41: 290-299.
• 46. Shaw P, Chattopadhyay A. Nrf2-ARE signaling in cellular protection: Mechanism of action and the regulatory mechanisms. J Cell Physiol. 2019; 2019:1-12.
• 47. Dai Y, Zhang J, Xiang J, Li Y, Wu D, Xu J. Calcitriol inhibits ROS-NLRP3-IL-1β signaling axis via activation of Nrf2-antioxidant signaling in hyperosmotic stress stimulated human corneal epithelial cells. Redox Biol. 2019; 21: 101093.
• 48. Teixeira TM, da Costa DC, Resende AC, Soulage CO, Bezerra FF, Daleprane JB. Activation of Nrf2-antioxidant signaling by 1,25-dihydroxycholecalciferol prevents leptin-induced oxidative stress and inflammation in human endothelial cells. J Nutr. 2017; 147: 506-513.