Administration of Ginkgo biloba Extract (EGb761) Alone and in Combination with FK506 Promotes Liver Regeneration in a Rat Model of Partial Hepatectomy

Abstract

Background: Free radical damage is known to occur during liver regeneration. The Ginkgo biloba extract EGb761 has antioxidant properties due to its ability to scavenge free radicals. FK506 has been widely used as an immunosuppressant that stimulates hepatocyte proliferation following partial hepatectomy.Aims: To explore whether EGb761 enhances liver regeneration after hepatectomy in rats, we investigated the effects of EGb761 alone and in combination with FK506 on the liver regenerative process.Study Design: Animal experimentation.Methods: A total of 75 Wistar albino rats weighing 340.08±11.66 g were randomly divided into five experimental groups: sham, control, FK506, EGb761, and FK506 + EGb761. According to the study groups, rats were administered FK506 at a dose of 0.1 mg/kg/day and EGb761 at 25 mg/kg/day three times via the intraperitoneal route. Then, two-thirds hepatectomy was performed according to the Higgins and Anderson technique in all the rats. At postoperative 48 h, 53 surviving rats were sacrificed. Serum and plasma samples were collected for analyzing thymidine kinase and oxidative stress marker levels. The regenerated liver was entirely resected, weighed, and sectioned. The mitotic index was assessed using hematoxylin-eosin staining. The extent of liver regeneration was calculated using the Child’s formula. The data were statistically analyzed using ANOVA, with a significance level of 5% (p<0.05).Results: Rats who received EGb761 showed significantly higher levels of liver regeneration than those who received FK506 or FK506 + EGb761 (p<0.01). Thymidine kinase level and mitotic index were significantly higher in the EGb761 (p<0.005) and FK506 (p<0.05) groups than in the control and sham groups. In addition, the liver regeneration percentage was significantly higher in the EGb761 group than in the FK506 group (p<0.01). Myeloperoxidase and malondialdehyde levels were significantly correlated between the EGb761 and FK506 groups, even at lower levels in the EGb761 group (p<0.001).Conclusion: EGb761, which is an antioxidant, reduces liver damage and stimulates liver regeneration following partial hepatectomy in rats through its anti-inflammatory and antioxidative effects.

Keywords

• EGb761, FK506, hepatectomy, liver, oxidative stress, regeneration, thymidine kinase, mitosis

Kaynakça

1. 1. Miyahara S, Yokomuro K, Takahashi H, Kimura Y. Regeneration and the immune system. I. In vitro and in vivo activation of lymphocytes by liver regeneration and the role of Kupffer cells in stimulation. Eur J Immunol 1983;13:878-83.
2. 2. Guerrieri F, Vendemiale G, Grattagliano I, Cocco T, Pellecchia G, Altomare E. Mitochondrial oxidative alterations following partial hepatectomy. Free Radic Biol Med 1999:26;34-41.
3. 3. Nash K, Shah ZA. Current perspectives on the beneficial role of Ginkgo biloba in neurological and cerebrovascular disorders. Integr Med Insights 2015;10:1-9.
4. 4. Bridi R, Crossetti FP, Steffen VM, Henriques AT. The antioxidant activity of standardized extract of Ginkgo biloba (EGb 761) in rats. Phytother Res 2001;15:449-51.
5. 5. Koch E. Inhibition of platelet activating factor (PAF)-induced aggregation of human thrombocytes by ginkgolides: considerations on possible bleeding complications after oral intake of Ginkgo biloba extracts. Phytomedicine 2005;12:10-6.
6. 6. Yoo DY, Nam Y, Kim W, Yoo KY, Park J, Lee CH, et al. Effects of Ginkgo biloba extract on promotion of neurogenesis in the hippocampal dentate gyrus in C57BL/6 mice. J Vet Med Sci 2011;73:71-6.
7. 7. Backes AN, Tannuri AC, Backes FN, Queiroz AJ, Coelho MC, da Silva EL, et al. Effects of tacrolimus and insulin in a liver regeneration model in growing animals with portal vein stenosis: immunohistochemical and molecular studies. Pediatr Surg Int 2014;30:423-9.
8. 8. Ohtake M, Sakaguchi T, Yoshida K, Muto T. Hepatic branch vagotomy can suppress liver regeneration in partially hepatectomized rats. HPB Surg 1993;6:277-86.

9. 9. Emond J, Capron-Laudereau M, Meriggi F, Bernuau J, Reynes M, Houssin D. Extent of hepatectomy in therat. Evaluation of basal conditions and effect of therapy. Eur Surg Res 1989;21:251-9.
10. 10. Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. Protein measurement with the folin phenol reagent. J Biol Chem 1951;193,265-74.
11. 11. Kagure K, Zhang YQ, Shibata H, Kojima I. Immediate onset of DNA synthesis in remnant rat liver after 90% hepatectomy by an administration of follistatin. J Hepatol 1998;28:977-84.
12. 12. Fishback FC. A morphologic study of regeneration of the liver after partial removal. Arch Pathol 1929;7:956-77.
13. 13. Okhawa H, Ohishi N, Yagi K. Assay for lipid peroxides in animal tissue by Thiobarbituric acid reaction. Anal Biochem 1979;95:351-8.
14. 14. Bradley PP, Priebat DA, Christensen RD, Rothstein G. Measurement of cutaneous inflamation. Estimation of neutrophil content with an enzyme marker. J Invest Dermatol 1982;78:206-9.
15. 15. Beauchamp C, Fridovich I. Superoxide dismutase improved assays and an assay applicable to acrilamide gels. Anal Biochem 1971:44;276-87.
16. 16. Paglia DE, Valentine WN. Studies on the quantitative and qualitative characterization of erythrocyte glutathione peroxidase. J Lab Clin Med 1967;70:158-69.
17. 17. Goldberg DM, Spooner RJ. “Glutathione Reductase,” In: Bermeyer HU, Bergmeyer J, GraBI M, editors. Methods of Enzymatic Analysis, 3rd Edition, Verlag Chemie: Weinheim; 1983:258-65.
18. 18. Wang G, Zhao C, Chen S, Li X, Zhang L, Chang C, et al. A preliminary in vivo study of the effects of OPN on rat liver regeneration induced by partial hepatectomy. Mol Biol Rep 2016;43:1371-82.
19. 19. Batmunkh B, Choijookhuu N, Srisowanna N, Byambatsogt, Synn Oo P, Noor Ali M, et al. Estrogen accelerates cell proliferation through estrogen receptor α during rat liver regeneration after partial hepatectomy. Acta Histochem Cytochem 2017;50:39- 48.
20. 20. Zhou JB, Yang XK, Ye QF, Ming YZ, Xia ZJ. Effect of extract of ginkgo biloba leaves on theprecondition of liver graft in rat liver transplantation. Zhong Nan Da Xue Xue Bao Yi Xue Ban 2007;32:54-8.
21. 21. Yoshikawa T. Food Factors for Health Promotion. In: Wang Y, Ho CT, editors. Metabolism of Flavonoids. Kyoto: Basel Karger; 2009:64-74.
22. 22. Tighilet B, Lacour M. Pharmacological activity of the Ginkgo biloba extract (EGb 761) on equilibrium function recovery in the unilateral vestibular neurectomized cat. J Vestib Res 1995;5:187-200.
23. 23. Sakr MF, Zetti GM, Hassanein TI, Farghali H, Nalesnik MA, Gavaler JS, et al. FK 506 ameliorates the hepatic injury associated with ischemia and reperfusion in rats. Hepatology 1991;13:947-51.
24. 24. Griffiths EJ, Halestrap AP. Protection by cyclosporin a of ischemic/reperfusioninduced damage in isolated rat hearts. J Mol Cell Cardiol 1993;25:1461-9.
25. 25. Zhu Z, Zhou X, He B, Dai T, Zheng C, Yang C, et al. Ginkgo Biloba Extract (EGb 761) Promotes peripheral nerve regeneration and neovascularization after acellular nerve allografts in a rat model. Cell Mol Neurobiol 2015;35:273-82.
26. 26. Field FJ, Mathur SN, LaBrecque DR. Cholesterol metabolism in regenerating liver of the rat. Am J Physiol 1985;249:679-84.
27. 27. O'Brien PJ, Slaughter MR, Polley SR, Kramer K. Advantages of glutamate dehydrogenase as a blood biomarker of acute hepatic injury in rats. Lab Anim 2002;36:313-21.
28. 28. Liu SQ, Yu JP, Chen HL, Luo HS, Chen SM, Yu HG. Therapeutic effects and molecular mechanisms of Ginkgo biloba extract on liver fibrosis in rats. Am J Chin Med 2006;34:99-114.
29. 29. Moreno D, Neri L, Vicente E, Vales A, Aldabe R. Use of thymidine kinase recombinant adenovirus and ganciclovir mediated mouse liver preconditioning for hepatocyte xenotransplantation. Methods Mol Biol 2017;1506:179-92.