Potential Therapeutic Effect of Bee Pollen and Metformin Combination on Testosterone and Estradiol Levels, Apoptotic Markers and Total Antioxidant Capacity in A Rat Model of Polycystic Ovary Syndrome

Document Type : Original Article


1 Student Research Committee, Kermanshah University of Medical Sciences, Kermanshah, Iran

2 Fertility and Infertility Research Center, Health Technology Institute, Kermanshah University of Medical Sciences, Kermanshah, Iran


Background: Polycystic ovary syndrome (PCOS) is associated with metabolic disorder as well as infertility. Many
traditional remedies have been reported to show estrogenic and antioxidant potential. Bee pollen is a natural compound,
reported as one such remedy. The present study aimed to investigate the effects of BP extract and metformin
(MET) on estradiol (E2) and testosterone (T) levels, apoptotic markers, and total antioxidant capacity (TAC) inarat
model of PCOS.

Materials and Methods: In this experimental study, 54 female Wistar (n=6/group) rats received 2 mg of estradiol
valerate (EV) intramuscularly and 6 additional rats were considered the control without EV injection. The rats were
treated with BP (50, 100, and 200 mg/kg), MET (300 mg/kg) and BP+MET (50 BP+300 MET, 100 BP+300 MET,
and 200 BP+300 MET mg/kg). Serum levels of E2 and T were assessed by ELISA method. TAC of serum was also
determined. The expressions of Bcl-2, Bax and Caspase-3 (Cas-3), and Sirt-1 genes were evaluated by real-time polymerase
chain reaction (PCR). Data were statistically analyzed using one-way ANOVA.

Results: In the untreated PCOS group E2 and T levels (p <0.01), and Bcl-2 (P=0.007) expression were increased, but TAC
(P=0.002) and expression of Bax (P=0.001), Cas-3 and Sirt1 (p <0.01) were decreased significantly. The levels of E2 and T,
as well as the expressions of Bcl-2 were decreased in all treated groups compared to the untreated PCOS group (p <0.01). On
the other hand, TAC and expression of Bax, Cas-3 and Sirt1 were increased in the BP- and MET-treated groups (p <0.05).

Conclusion: BP and MET synergistically improved serum E2, T and TAC levels, and expression of apoptotic genes.


1. Azizi M, Elyasi F. Psychosomatic aspects of polycystic ovarian syndrome: a review. Iran J Psychiatry Behav Sci. 2017; 11(2): e6595.
2. Ahmadi A, Nasiri Nejad N, Parivar K. Effect of aqueous extract of the aerial part of the Ruta graveolens on the spermatogenesis ofimmature Balb/c mice. RJMS. 2007; 14(56): 13-20.
3. Dou L, Zheng Y, Li L, Gui X, Chen Y, Yu M, et al. The effect of cinnamon on polycystic ovary syndrome in a mouse model. Reprod Biol Endocrinol. 2018; 16(1): 99.
4. Febri RR, Wiweko B, Hanifah N, Sholihah A, Iffanolida PA, Mutia K, et al. BAX/BCL-2 expression in granulosa cells was not altered in patients with polycystic ovary syndrome. J Phys Conf Ser. 2018; 1073(3).
5. Pourmatroud E, Mohammadjafari R, Roozitalab M. Comparison of metformin and simvastatin administration in women with polycystic ovary syndrome before intra-cytoplasmic sperm injection cycle: a prospective, randomized, clinical trial study. Iran Red Crescent Med J. 2015; 17(12): e20082.
6. Komosinska-Vassev K, Olczyk P, Kaźmierczak J, Mencner L, Olczyk K. Bee pollen: chemical composition and therapeutic application. Evid Based Complement Alternat Med. 2015; 2015: 297425.
7. Medeiros KCP, Figueiredo CAV, Figueredo TB, Freire KRL, Santos FAR, Alcântara-Neves NM, et al. Anti-allergic effect of bee pollen phenolic extract and myricetin in ovalbumin-sensitized mice. J Ethnopharmacol. 2008; 119(1): 41-46.
8. Xie Y, Wan B, Li W. Effect of bee pollen on maternal nutrition and fetal growth. Hua Xi Yi Ke Da Xue Xue Bao. 1994; 25(4): 434-437.
9. Klippel KF, Hiltl DM, Schipp B. A multicentric, placebo-controlled, double-blind clinical trial of β-sitosterol (phytosterol) for the treatment of benign prostatic hyperplasia. German BPH-Phyto Study group. Br J Urol. 1997; 80(3): 427-432.
10. Karimi Jashni H, Kargar Jahromi H, Bagheri Z. The effect of palm pollen extract on polycystic ovary syndrome (POS) in rats. Int J Med Res Health Sci. 2016; 5(5): 317-321.
11. Šarić A, Balog T, Sobočanec S, Kušić B, Šverko V, Rusak G, et al. Antioxidant effects of flavonoid from Croatian Cystus incanus L. rich bee pollen. Food Chem Toxicol. 2009; 47(3): 547-554.
12. Sousa C, Andrade P, Valentão P. Relationships of Echium plantagineum L. bee pollen, dietary flavonoids and their colonic metabolites with cytochrome P450 enzymes and oxidative stress. RSC Advances. 2016; 6(8): 6084-6092.
13. Mirabolghasemi G, Kamyab Z. Changes of the uterine tissue in rats with polycystic ovary syndrome induced by estradiol valerate. Int J Fertil Steril. 2017; 11(1): 47-55.
14. Brawer JR, Munoz M, Farookhi R. Development of the polycystic ovarian condition (PCO) in the estradiol valerate-treated rat. Biol Reprod. 1986; 35(3): 647-655.
15. Tao X, Cai L, Chen L, Ge S, Deng X. Effects of metformin and exenatide on insulin resistance and AMPKα-SIRT1 molecular pathway in PCOS rats. J Ovarian Res. 2019; 12(1): 86.
16. Abdel-gawad E. Potential impact of bee pollen administration during pregnancy in rats. J Am Sci. 2010; 6: 44-53.
17. Wellington D, Mikaelian I, Singer L. Comparison of ketamine-xylazine and ketamine-dexmedetomidine anesthesia and intraperitoneal tolerance in rats. J Am Assoc Lab Anim Sci. 2013; 52(4): 481-487.
18. Ghanbari E, Nejati V, Khazaei M. Antioxidant and protective effects of Royal jelly on histopathological changes in testis of diabetic rats. Int J Reprod Biomed. 2016; 14(8): 519-526.
19. Denisow B, Denisow-Pietrzyk M. Biological and therapeutic properties of bee pollen: a review. J Sci Food Agric. 2016; 96(13): 4303-4309.
20. Desai BN, Maharjan RH, Nampoothiri LP. Aloe barbadensis Mill. formulation restores lipid profile to normal in a letrozole-induced polycystic ovarian syndrome rat model. Pharmacognosy Res. 2012; 4(2): 109-115.
21. Amsterdam A, Gold RS, Hosokawa K, Yoshida Y, Sasson R, Jung Y, et al. Crosstalk among multiple signaling pathways controlling ovarian cell death. Trends Endocrinol Metab. 1999; 10(7): 255-262.
22. Salvetti NR, Panzani CG, Gimeno EJ, Neme LG, Alfaro NS, Ortega HH. An imbalance between apoptosis and proliferation contributes to follicular persistence in polycystic ovaries in rats. Reprod Biol Endocrinol. 2009; 7: 68.
23. Chen J, Shen S, Tan Y, Xia D, Xia Y, Cao Y, et al. The correlation of aromatase activity and obesity in women with or without polycystic ovary syndrome. J Ovarian Res. 2015; 8: 11.
24. Mokhtari M, Ebrahimpoor MR, Harfsheno S. The effects of alcoholic extract of Marrubum vulgare on hormonal parameters in female rat model of polycystic ovarian syndrome. Medical Sciences. 2014; 24(2): 74-80.
25. Malini T, Vanithakumari G. Antifertility effects of beta-sitosterol in male albino rats. J Ethnopharmacol. 1991; 35(2): 149-153.
26. Goodarzi MO, Dumesic DA, Chazenbalk G, Azziz R. Polycystic ovary syndrome: etiology, pathogenesis and diagnosis. Nat Rev Endocrinol. 2011; 7(4): 219-231.
27. Rzepecka-Stojko A, Stojko J, Kurek-Górecka A, Górecki M, Kabała-Dzik A, Kubina R, et al. Polyphenols from bee pollen: structure, absorption, metabolism and biological activity. Molecules. 2015; 20(12): 21732-21749.
28. Kimura Y, Ito H, Ohnishi R, Hatano T. Inhibitory effects of polyphenols on human cytochrome P450 3A4 and 2C9 activity. Food Chem Toxicol . 2010; 48(1): 429-435.
29. Gilman CI, Leusch FDL, Breckenridge WC, MacLatchy DL. Effects of a phytosterol mixture on male fish plasma lipoprotein fractions and testis P450scc activity. Gen Comp Endocrinol. 2003; 130(2): 172-184.
30. Pinto B, Caciagli F, Riccio E, Reali D, Šarić A, Balog T, et al. Antiestrogenic and antigenotoxic activity of bee pollen from Cystus incanus and Salix alba as evaluated by the yeast estrogen screen and the micronucleus assay in human lymphocytes. Eur J Med Chem. 2010; 45(9): 4122-4128.
31. Wang F, Shing M, Huen Y, Tsang SY, Xue H. Neuroactive flavonoids interacting with GABAA receptor complex. Curr Drug Targets CNS Neurol Disord. 2005; 4(5): 575-585.
32. Kitagawa S, Nabekura T, Takahashi T, Nakamura Y, Sakamoto H, Tano H, et al. Structure-activity relationships of the inhibitory effects of flavonoids on P-glycoprotein-mediated transport in KB-C2 cells. Biol Pharm Bull. 2005; 28(12): 2274-2278.
33. Monsefi M, Ghasemi A, Alaee S, Aliabadi E. Effects of Anethum graveolens L. (dill) on oocyte and fertility of adult female rats. J Reprod Infertil. 2015; 16(1): 10-17.
34. Murri M, Luque-Ramírez M, Insenser M, Ojeda-Ojeda M, Escobar-Morreale HF. Circulating markers of oxidative stress and polycystic ovary syndrome (PCOS): a systematic review and meta-analysis. Hum Reprod Update. 2013; 19(3): 268-288.
35. Kohzadi M, Khazaei MR, Choobsaz F, Khazaei M. Relationship between serum levels of anti-mullerian hormone, adiponectin and oxidative stress markers in patients with polycystic ovary syndrome. Int J Fertl Steril. 2020; 14(1): 27-33.
36. Ahmadi M, Rostamzadeh A, Fathi F, Mohammadi M, Rezaie MJ. The effect of Melatonin on histological changes of ovary in induced polycystic ovary syndrome model in mice. Middle East Fertil Soc J. 2017; 22(4): 255-259.
37. Das M, Djahanbakhch O, Hacihanefioglu B, Saridogan E, Ikram M, Ghali L, et al. Granulosa cell survival and proliferation are altered in polycystic ovary syndrome. J Clin Endocrinol Metab. 2008; 93(3): 881-887.
38. Isobe N, Yoshimura Y. Deficient proliferation and apoptosis in the granulosa and theca interna cells of the bovine cystic follicle. J Reprod Dev. 2007; 53(5): 1119-1124.
39. Tao X, Zhang X, Ge SQ, Zhang EH, Zhang B. Expression of SIRT1 in the ovaries of rats with polycystic ovary syndrome before and after therapeutic intervention with exenatide. Int J Clin Exp Pathol. 2015; 8(7): 8276-8283.
40. Lashen H. Role of metformin in the management of polycystic ovary syndrome. Ther Adv Endocrinol Metab. 2010; 1(3): 117-128.