Inonu University, Turgut Ozal Medical Centre

1. The protective effect of glycyrrhetinic acid and chrysin on ischemia-reperfusion injury in rat ovary Rauf Melekoglu Osman Ciftci Sevil Eraslan Saadet Alan Nese Basak Inonu University, Turgut Ozal Medical Centre

2. GA and CR on ischemia-reperfusion injury in rat ovary • Ovarian torsion refers to twisting of the ovary around its vascular peduncle, the suspensory ligament of the ovary and accounts for nearly 3% of all gynecologic emergencies. • According to the degree of the torsion, venous return diminishes first, and stromal edema and internal hemorrhage occur. • If the arterial blood supply ceases, ischemia and necrosis are inevitable. • Huchon C, Fauconnier A. Adnexal torsion: A literature review. Eur J Obstet Gynecol Reprod Biol 2010; 150: 8–12

3. GA and CR on ischemia-reperfusion injury in rat ovary • Enlarged ovaries due to cysts, stimulation or pregnancy, hyperlaxity of the utero-ovarian or infundibulopelvic ligaments have been considered as risk factors of ovarian torsion. • It is mostly seen at reproductive age, early diagnosis and management are crucial to preserve ovarian function. • Tsafrir Z, Azem F, Hasson J et al. Risk factors, symptoms, and treatment of ovarian torsion in children: The twelve-year experience of one center. J Minim Invasive Gynecol 2012; 19: 29–33.

4. GA and CR on ischemia-reperfusion injury in rat ovary • Recent studies recommend only detorsion and preservation of the twisted ovary even if tissues are cyanotic. • Given that detorsion of the twisted ovary did not increase the risk of pulmonary embolism, adnexectomy is no longer preferred. • Detorsion, however, brings another risk, which is related to reperfusion and neutrophil infiltration, so-called ischemia/reperfusion (I/R) injury. • Excessive reactive oxygen species (ROS) production results in production of toxic substances. • Ergenoglu M, Erbas O, Akdemir A et al. Attenuation of ischemia/reperfusion-induced ovarian damage in rats: Does edaravone offer protection. Eur Surg Res 2013; 51: 21–22

5. GA and CR on ischemia-reperfusion injury in rat ovary • Chrysin (5,7-dihydroxyflavone), which is a natural flavonoid that is present in honey, bee propolis, and many plant extracts is well-known for its various biological activities. • Several studies have indicated antioxidant, antihypertensive, antidiabetogenic, and anxiolytic functions. • Glycyrrhetinic acid, an aglycone saponin extracted from licorice root, is known for its antitumor, antiinflammatory and antihepatitic effects, and has been used worldwide. • Yao Y, Chen L, Xiao J et al. Chrysin protects against focal cerebral ischemia/reperfusion injury in mice through attenuation of oxidative stress and inflammation. Int J Mol Sci. 2014 Nov 13;15(11):20913-26. • Oztanir MN, Ciftci O, Cetin A et al. he beneficial effects of 18β-glycyrrhetinic acid following oxidative and neuronal damage in brain tissue caused by global cerebral ischemia/reperfusion in a C57BL/J6 mouse model.Neurol Sci. 2014 Aug;35(8):1221-8.

6. GA and CR on ischemia-reperfusion injury in rat ovary Objective To evaluate the protective effects of glycyrrhetinic acid and chrysin in experimental ischemia/reperfusion (I/R) injury of rat ovaries. ischemia/reperfusion (I/R) Theanimalsweresacrificedunderetheranesthesia 14 daysafterthetreatmentandtheovariantissueswereimmediatelyremoved and bloodsampleswerecollected Material and Methods ischemia/reperfusion (I/R)+ chrysin (50 mg/kg) Rats were randomly divided into five equal groups (n=10 in each group); ischemia/reperfusion (I/R) + glycyrrhetinicacid (200 mg/kg) Onlychrysin(50 mg/kg) Onlyglycyrrhetinicacid (200 mg/kg) (I/R + G)

7. GA and CR on ischemia-reperfusion injury in rat ovary • Surgical procedures • All surgical interventions were carried out in a proper laboratory environment, under sterile conditions. • After weighing the rats, ketamine (50 mg/kg) and xylazine (10 mg/kg) were injected intramuscularly for anesthesia. • Ischemia/reperfusion (I/R) was performed with vascular clamps by placing clamps approximately 1 cm under the adnexal structure to stimulate ovarian torsion. • After two hour of ovarian torsion, the vascular clamps were opened for allowing reperfusion.

8. GA and CR on ischemia-reperfusion injury in rat ovary Biochemicalassay Malonaldehyde ( MDA) Glutathione (GSH) concentrations Superoxide dismutase ( SOD) Catalase(CAT) Glutathione peroxidase (GSH-Px) activities Hormonal assay Estradiol (E2) , Follicle stimulation hormone (FSH) , Luteinization hormone ( LH) Anti-mullerian hormone (AMH) • Histopathologic examination • Histopathological examination of the tissue damage  hemorrhage, vascular congestion, edema, polymorphonuclear leukocyte (PMNL) infiltration and apoptosis. • Each sample was scored for each criterion using a scale ranging from 0 to 3 • (0, none; 1, mild; 2, moderate; 3, severe).

9. GA and CR on ischemia-reperfusion injury in rat ovary Biochemical results

10. GA and CR on ischemia-reperfusion injury in rat ovary Hormonal assessment

11. GA and CR on ischemia-reperfusion injury in rat ovary Histopathologicscores

12. Effect of endometrioma on ovarian reserve

13. GA and CR on ischemia-reperfusion injury in rat ovary • Conclusion • In conclusion, the treatment of ovarian I/R injury with chrysin and glycyrrhetinicacid were effective in reducing ovarian damage. • The beneficial effects of these agents could be attributed to the antioxidant and vascular effects of CH and G. • The significant improvement in tissue apoptosis scores and hormone levels detected in this study indicate that treatment with CH or G might be an alternate conservative treatment approach for adnexal torsion cases after detorsion surgery. Thank you