Investigation of the Effects of Bilberry Extract on Methotrexate-Induced Hepatotoxicity

Öz

Background: Methotrexate (Mtx) is widely used in the treatment of cancer and autoimmune diseases. But it can cause toxicity in the liver, which is responsible for the metabolism of many chemotherapeutics such as Mtx. Bilberry (Bb) fruit has antioxidant and anti-inflammatory effects thanks to the phenolic compounds it contains. This study aims to investigate the effects of oral Bb extract on Mtx-induced hepatotoxicity. Materials and Methods: For the study, 21 Spraque Dawley adult rats were divided into three groups equal in number. One group was given a single dose of 30 mg/kg intraperitoneal (ip.) Mtx on the first day of the experi-ment (MtxG). The other group was given a single dose of 30 mg/kg ip. Mtx and 200 mg/kg Bb extract by oral gavage once a day for five days (Mtx+BbG). No procedure was applied to the control group (CG). While some of the liver tissues taken at the end of the experiment were evaluated and scored histologically, the other part was evaluated biochemically. Results: Malondialdehyde (MDA) and caspase 3 levels were higher in liver tissues of MtxG than CG. Although it is not significant according to CG, there was an increase in 8-hydroxy-2'-deoxyguanosine (8-OHdG) and oxidative stress index (OSI), but a decrease in superoxide dismutase (SOD), catalase (CAT) and total antioxidant capacity (TOS) in MtxG. In Mtx+BbG, MDA levels decreased compared to MtxG. In this group, a nonsignificant statististical decrease was also observed in caspase 3, OSI and 8-OHdG levels compared to MtxG. In addition, while the SOD level in this group was lower than in CG; CAT and total antioxidant status (TAS) levels were higher than in MtxG, although not statistically significant. In MtxG, there was an increase in total damage score, vacuoliza-tion/degeneration/pyknosis in hepatocytes, Kupffer cells and sinusoidal vasodilation/congestion compared to the control. In Mtx+BbG, there was a decrease in all these histopathological findings and the total damage score. Conclusions: Orally administered Bb extract may reduce Mtx-induced oxidative stress, apoptosis, and the hepato-toxicity caused by them in rat liver.

Anahtar Kelimeler

• Liver
• methotrexate
• oxidative stress
• bilberry

Yaban Mersini Ekstresinin Metotreksat Kaynaklı Hepatotoksisite Üzerine Etkilerinin Araştırılması

Öz

Amaç: Metotreksat (Mtx), kanser ve otoimmün hastalıkların tedavisinde yaygın olarak kullanılmaktadır. Ancak Mtx, kendisi gibi birçok kemoterapötiğin metabolizmasından sorumlu olan karaciğerde toksisiteye neden olabilir. Yaban mersini (Ym) meyvesi, yapısında yoğun olarak bulunan fenolik bileşikler sayesinde antioksidan ve antienflamatuvar etkiye sahiptir. Bu çalışma, Ym ekstresinin, Mtx kaynaklı hepatotoksisiteye olan etkilerini araştırmayı amaçlamakta-dır. Materyal ve metod: Çalışma için 21 adet Spraque Dawley ırkı erişkin sıçan, sayıca eşit üç gruba bölündü. Bir gruba, deneyin ilk günü tek doz 30 mg/kg intraperitoneal (ip.) Mtx verildi (MtxG). Diğer gruba tek doz 30 mg/kg ip. Mtx ve beş gün boyunca günde bir kez oral gavajla 200 mg/kg Ym ekstresi verildi (Mtx+YmG). Kontrol grubuna ise herhangi bir işlem yapılmadı (KG). Deney sonunda alınan karaciğer dokularının bir kısmı histolojik olarak değer-lendirilerek skorlanırken diğer kısmı da biyokimyasal olarak değerlendirildi. Bulgular: MtxG’nin karaciğer dokularında malondialdehid (MDA) ve kaspaz 3 düzeyleri KG’ye göre yüksekti. KG’ye göre anlamlı olmamakla birlikte MtxG’de, 8-hidroksi-2′-deoksiguanozin (8-OHdG) ve oksidatif stres indeksi (OSİ)’nde artış; süperoksit dismutaz (SOD), katalaz (CAT) ve total antioksidan kapasite (TOS)’de ise azalma mevcut-tu. Mtx+YmG’de MDA düzeyleri MtxG’ye göre azaldı. Bu grupta, kaspaz 3, OSI ve 8-OHdG düzeylerinde de MtxG'ye göre istatistiksel olarak anlamlı olmayan belirgin bir azalma görüldü. Ayrıca bu gruptaki SOD seviyesi KG’ye göre düşükken CAT ve total antioksidan kapasite (TAS) seviyeleri ise MtxG’ye göre istatistiksel anlamlı olmasa da yüksekti. MtxG’de; total hasar skorunda, hepatositlerdeki vakuolizasyon/dejenerasyon/piknoziste, Kupffer hücrelerinde ve vazodilatasyon/konjesyonda kontrole göre artış mevcuttu. Mtx+YmG’de ise tüm bu histopatolojik bulgularda ve total hasar skorunda azalma mevcuttu. Sonuç: Oral yolla uygulanan Ym ekstresi, sıçan karaciğerindeki Mtx kaynaklı oksidatif stresi, apoptozu ve bunların neden olduğu hepatotoksisiteyi azaltabilir.

Anahtar Kelimeler

• Karaciğer
• metotreksat
• oksidatif stress
• yaban mersini

Kaynakça

1. 1. Mudd TW, Guddati AK. Management of hepatotoxicity of chemotherapy and targeted agents. Am J Cancer Res. 2021 Jul 1511(7):3461-3474.
2. 2. Işeri S, Ercan F, Gedik N, Yüksel M, Alican I. Simvastatin attenuates cisplatin-induced kidney and liver damage in rats. Toxicology. 2007 Feb;230(2-3):256-264.
3. 3. Gedikli S, Şengül E. Ratlarda Siklofosfamid ile İndüklenen Hepatotoksisite Üzerine Kuersetinin Etkileri. Dicle Tıp Dergi-si. 2019;46(1):41-50.
4. 4. Özgöçmen M, Aşcı H, Doğan HK, İlhan İ, Pekgöz Ş, Mustafa-oğlu A. A study on Wistar Albino rats: investigating protecti-ve role of ramelteon on liver damage caused by methot-rexate. Drug Chem Toxicol. 2022;45(6):2678–2685.
5. 5. Hamed KM, Dighriri IM, Baomar AF, Alharthy BT, Alenazi FE, Alali GH et al. Overview of Methotrexate Toxicity: A Comp-rehensive Literature Review. Cureus. 2022 Sep 23;14(9):e29518.
6. 6. Koca I, Ucar M, Tutoglu A, Berhoglu A, Boyaci A. Henoch-Schönlein Purpura induced by Low-Dose Methotrexate. Har-ran Üniversitesi Tıp Fakültesi Dergisi. 2014;11(3):316-320.
7. 7. Ezhilarasan D. Hepatotoxic potentials of methotrexate: Understanding the possible toxicological molecular mecha-nisms. Toxicology. 2021 Jun 30;458:152840.
8. 8. Abdul-Hamid M, Abdel-Reheim ES, Hegazy W, Allam A, Othman SI, Alwaele MA. Effect of gervital in attenuating he-patotoxicity caused by methotrexate or azathioprine in adult albino rats. Environ Sci Pollut Res Int. 2022 Jul;29(31):46788-46801

9. 9. Shahani S, Mehraban N, Talebpour Amiri F, Abedi SM, Noaparast Z, Mohammadinia S. Melissa Officinalis L. aque-ous extract pretreatment decreases methotrexate-induced hepatotoxicity at lower dose and increases99mTc-phytate li-ver uptake, as a probe of liver toxicity assessment, in rats. Ann Nucl Med. 2023 Mar;37(3):166-175.
10. 10. Drishya S, Dhanisha SS, Guruvayoorappan C. Antioxidant-rich fraction of Amomum subulatum fruits mitigates experi-mental methotrexate-induced oxidative stress by regulating TNF-α, IL-1β, and IL-6 proinflammatory cytokines. J Food Bi-ochem. 2022 Apr;46(4):e13855.
11. 11. Pires TCSP, Caleja C, Santos-Buelga C, Barros L, Ferreira ICFR. Vaccinium myrtillus L. Fruits as a Novel Source of Phenolic Compounds with Health Benefits and Industrial Applications - A Review. Curr Pharm Des. 2020;26(16):1917-1928.
12. 12. Ștefănescu R, Laczkó-Zöld E, Ősz BE, Vari CE. An Updated Systematic Review of Vaccinium myrtillus Leaves: Phytoc-hemistry and Pharmacology. Pharmaceutics. 2022 Dec 21;15(1):16.
13. 13. Sharma A, Lee HJ. Anti-Inflammatory Activity of Bilberry (Vaccinium myrtillus L.). Curr Issues Mol Biol. 2022 Sep 30;44(10):4570-4583.
14. 14. Popović D, Đukić D, Katić V, Jović Z, Jović M, Lalić J. Antioxi-dant and proapoptotic effects of anthocyanins from bilberry extract in rats exposed to hepatotoxic effects of carbon tet-rachloride. Life Sci. 2016 Jul 15;157:168-177.
15. 15. Haga S, YiMin, Yamaki H, Jin S, Sogon T, Morita N. Extracts of bilberry (Vaccinium myrtillus L.) fruits improve liver steatosis and injury in mice by preventing lipid accumulation and cell death. Biosci Biotechnol Biochem. 2019 Nov;83(11):2110-2120.
16. 16. Pandir D, Kara O. Chemopreventive effect of bilberry (Vac-cinium myrtillus) against cisplatin-induced oxidative stress and DNA damage as shown by the comet assay in peripheral blood of rats. Biologia. 2014;69:811–816
17. 17. Demir S, Turan I, Demir F, Ayazoglu Demir E, Aliyazicioglu Y. Cytotoxic Effect of Laurocerasus officinalis Extract on Human Cancer Cell Lines. Marmara Pharmaceutical Journal. 2017;21(1):121-126.
18. 18. Tusat M, Mentese A, Demir S, Alver A, Imamoglu M. Medical ozone therapy reduces oxidative stress and testicular dama-ge in an experimental model of testicular torsion in rats. Int Braz J Urol. 2017;43(6):1160-1166.
19. 19. Pekgöz S, Asci H, Erzurumlu Y, Savran M, Ilhan I, Hasseyid N. Nebivolol alleviates liver damage caused by methotrexate via AKT1/Hif1α/eNOS signaling. Drug Chem Toxicol. 2022 Sep;45(5):2153-2159.
20. 20. Abdel-Daim MM, Khalifa HA, Abushouk AI, Dkhil MA, Al-Quraishy SA. Diosmin Attenuates Methotrexate-Induced Hepatic, Renal, and Cardiac Injury: A Biochemical and Histo-pathological Study in Mice. Oxid Med Cell Longev. 2017;2017:3281670.
21. 21. Samdanci ET, Huz M, Ozhan O, Tanbek K, Pamukcu E, Akatli AN. Cytoprotective effects of molsidomine against methot-rexate-induced hepatotoxicity: an experimental rat study. Drug Des Devel Ther. 2018 Dec 20;13:13-21.
22. 22. Al-Khawalde AAA, Abukhalil MH, Jghef MM, Alfwuaires MA, Alaryani FS, Aladaileh SH. Punicalagin Protects against the Development of Methotrexate-Induced Hepatotoxicity in Mice via Activating Nrf2 Signaling and Decreasing Oxidative Stress, Inflammation, and Cell Death. Int J Mol Sci. 2022 Oct 15;23(20):12334.
23. 23. Aslankoç R, Demirci D, İnan Ü, Yıldız M, Öztürk A, Çetin M. Oksidatif stres durumunda antioksidan enzimlerin rolü - Sü-peroksit dismutaz (SOD), katalaz (CAT) ve glutatyon peroksi-daz (GPX). Süleyman Demirel Üniversitesi Tıp Fakültesi Der-gisi. 2019; 26(3): 362-369.
24. 24. Pizzino G, Irrera N, Cucinotta M, Pallio G, Mannino F, Arcora-ci V. Oxidative Stress: Harms and Benefits for Human Health. Oxid Med Cell Longev. 2017;2017:8416763.
25. 25. Ghoneum M, El-Gerbed MSA. Human placental extract ameliorates methotrexate-induced hepatotoxicity in rats via regulating antioxidative and anti-inflammatory responses. Cancer Chemother Pharmacol. 2021 Dec;88(6):961-971.
26. 26. Türk S, Kazak F, Peker P, Kısaçam M.A. Rutin Attenuates Methotrexate-Induced Hepatic Oxidative Stress in Rats. Ata-türk Üniversitesi Veteriner Bilimleri Dergisi. 2021;16(3):291-297.
27. 27. Kızıl HE, Caglayan C, Darendelioğlu E, Ayna A, Gür C, Kande-mir FM, Küçükler S. Morin ameliorates methotrexate-induced hepatotoxicity via targeting Nrf2/HO-1 and Bax/Bcl2/Caspase-3 signaling pathways. Mol Biol Rep. 2023 Apr;50(4):3479-3488.
28. 28. Özgöçmen M, Yesilot S. The role of resveratrol in hepata-toxicity caused by methotrexate. Mehmet Akif Ersoy Üni-versitesi Veteriner Fakültesi Dergisi. 2021;6(2):57-63.
29. 29. Obeng E. Apoptosis (programmed cell death) and its signals - A review. Braz J Biol. 2021;81(4):1133-1143.
30. 30. Gürel A, Kaya K. Bromelain has Antioxidant Effect on Met-hotrexate Hepatotoxicity and Nephrotoxicity . Van Sağlık Bi-limleri Dergisi. 2022;15(1):37-42.
31. 31. Parthasarathy M, Prince SE. Andrographis paniculata (Burm.f.) Nees Alleviates Methotrexate-Induced Hepato-toxicity in Wistar Albino Rats. Life (Basel). 2023 May 12;13(5):1173.
32. 32. Ozlem K, Birkan Y, Mustafa K, Emin K. Protective effect of Vaccinium myrtillus on ischemia- reperfusion injury in rat ovary. Taiwan J Obstet Gynecol. 2018 Dec;57(6):836-841.
33. 33. Luo H, Lv XD, Wang GE, Li YF, Kurihara H, He RR. Anti-inflammatory effects of anthocyanins-rich extract from bil-berry (Vaccinium myrtillus L.) on croton oil-induced ear edema and Propionibacterium acnes plus LPS-induced liver damage in mice. Int J Food Sci Nutr. 2014;65(5):594-601.
34. 34. Bao L, Abe K, Tsang P, Xu JK, Yao XS, Liu HW. Bilberry extract protect restraint stress-induced liver damage through atte-nuating mitochondrial dysfunction. Fitoterapia. 2010 Dec;81(8):1094-101.
35. 35. Ozdemir A, Mercantepe T, Erdivanli B, Sen A, Mercantepe F, Tumkaya L. Neuroprotective effects of Vaccinium myrtillus on damage-related brain injury. J Chem Neuroanat. 2023 Jan;127:102193.
36. 36. Wang Y, Zhao L, Lu F, Yang X, Deng Q, Ji B. Retinoprotective Effects of Bilberry Anthocyanins via Antioxidant, Anti-Inflammatory, and Anti-Apoptotic Mechanisms in a Visible Light-Induced Retinal Degeneration Model in Pigmented Rabbits. Molecules. 2015 Dec 14;20(12):22395-410.