مطالعه اثر عصاره فلفل کوهی (Vitagnus castus) بر بلوغ و لقاح آزمایشگاهی اووسیت موش‌های سوری مبتلاشده به سندروم تخمدان پلی‌کیستیک

نوع مقاله: علمی پژوهشی

نویسندگان

1 دانشجوی فیزیولوژی دام، دانشکده کشاورزی و منابع طبیعی اهر، دانشگاه تبریز، تبریز، ایران.

2 استادیار گروه علوم دامی دانشکده کشاورزی و منابع طبیعی اهر، دانشگاه تبریز، تبریز، ایران.

3 دانشیار گروه علوم آناتومی، دانشکده دامپزشکی، دانشگاه ارومیه، ارومیه، ایران.

4 استادیار گروه علوم طیور، دانشکدة کشاورزی، دانشگاه تربیت مدرس، تهران، ایران.

10.30495/jvcp.2020.1881036.1245

چکیده

عدم کاهش کلاژن دیواره فولیکولی منجر به تجمع فولیکول­ها و بروز سندرم تخمدان پلی­کیستیک و کاهش کیفیت اووسیت­ها میگردد. هدف ازمطالعه حاضر بررسی اثر عصاره گیاه فلفل­کوهی (Vitagnus castus) بر بلوغ و توان باروری آزمایشگاهی اووسیتها در موش­های سوری مبتلاشده به سندرم تخمدان پلی­کیستیک (ovarian syndrome; PCOpolycystic) بود. بدین منظور تعداد 32 سر موش سوری ماده NMRI (Naval Medical Research Institute) ماده نابالغ 25 روزه با میانگین وزنی 25 گرم تهیه­شده از دانشگاه تبریز، به­طور تصادفیبه 4 گروه آزمایشی به شرح زیر تقسیم شدند: گروه شاهد بدون دریافت عصاره، گروه کنترل PCO بدون دریافت عصاره،  گروه PCO دریافت­کننده365 میلی­گرم عصاره فلفل­کوهی و گروه PCO دریافت­کننده730میلی­گرم عصاره فلفل‌کوهی. برای تهیه اووسیت، موش­های آزمایشی توسط تزریق درون صفاتی هورمون PMSG ­(pregnant mare serum gonadotropin) تیمار شدند و بعد از آسان­کشی،از فولیکول­های تخمدانی تخمک­گیری به­عمل آمده و برای ارزیابی بلوغ و لقاح آزمایشگاهی مورد استفاده قرار گرفتند.نتایج نشان داد که مقادیر هورمون­های تستوسترون و استروژن در گروه­های PCO تیمارشده با عصاره فلفل­کوهی نسبت به گروه ­PCOدریافت­کننده استرادیول ­والرات به طور معنی­داری پایین­تر بود (05/0>p < /em>).همچنین تعداد اووسیت­های نابالغی که به مرحله متافازII رسیدند در گروهPCOدریافت­کننده730 میلی­گرم عصاره فلفل­کوهی نسبت به گروه PCO دریافت­کننده استرادیول­والرات به طور معنی­داری بالاتربود (05/0>p < /em>). اما درصد اووسیت­های لقاح یافته و نیز بلاستوسیست­های تولیدی بین گروه­های آزمایشی تفاوت معنی­داری با هم نداشتند (05/0<p < /em>). از طرف دیگر درصد رویان­های هچ­شده در گروه  PCO­ دریافت کننده365 میلی­گرم عصاره فلفل­کوهی به طور معنی­داری بالاتر از گروه ­PCOدریافت­کننده استرادیول­والرات بود (05/0>p < /em>). یافته­های پژوهش حاضر نشان داد که عصاره گیاه فلفل کوهی باعث تکامل بیشتر اووسیت­ها جهت لقاح و نیز افزایش تکامل رویان­های حاصل از لقاح آزمایشگاهی در حیوانات درگیربا سندرم تخمدان پلی­کیستیک می­شود.

کلیدواژه‌ها


عنوان مقاله [English]

Therapeutic Effects of Vitex (Vitagnus castus) Extract on In vitro Maturation and In vitro Fertilization Oocytes of Mice as Model of Polycystic Ovary Syndrome

نویسندگان [English]

  • tohid ghorbani 1
  • amir karimi 2
  • gholamreza najafi 3
  • maghsud besharti 2
  • M. Sharafi 4
1 MSc Student, Department of Animal Science, Faculty of Ahar Agriculture and Natural Resources, University of Tabriz, Tabriz, Iran.
2 Assistant Professor, Department of Animal Science, Faculty of Ahar Agriculture and Natural Resources, University of Tabriz, Tabriz, Iran.
3 Associated Professor, Department of Anatomy, Faculty of Veterinary Medicine, Urmia University, Urmia, Iran.
4 Assistant Professor, Department of Poultry Sciences, Faculty of Agriculture, Tarbiat Modarres University, Tehran, Iran.
چکیده [English]

Introduction: Nowadays, to get rid of the side effects of chemical drugs, the use of medicinal plants is taken into consideration. Polycystic Ovary Syndrome (PCOs) is an important reproductive and endocrine disorder in women in the reproductive age. Also, Similar to PCOs, some disorders have been identified in other animals such as sheep, rat and mouse. PCOs is known as one of the most important endocrine disorders and a common cause of anovulation and infertility. There are some studies about the effects of vitex (Vitagnus castus) on different aspects of PCOs animals such as endocrinology (hormonal profiles), ovarian histology and changes in estrus cycle, but there is no study about the effects of vitex on embryo qualities or efficiency of embryo production. The aim of this study was evaluation of vitex extract on in vitro maturation (IVM) and in vitro fertilization (IVF) of oocytes in mice exposured to polycystic ovary syndrome.
Material and Methods: A total of 32 adult virgin NMRI female mice were kept in a temperature-controlled room under a 12 hours on and 12 hours off light cycle. Animals were randomly and equally allocated into four experimental groups included: 1) Control group (con) which did not receive any injection or other manipulations, 2) polycystic ovary syndrome group (PCO) received an IM injection of 4 mg/kg estradiol valerate dissolved
in 0.4 ml of sesame oil) (Aburaihan Co., Iran), to induce PCOs, and groups 3 and 4 were induced PCO that orally received vitex extract at 365 mg/kg/day and 730 mg/kg/day dosage, respectively for 30 days. Each eight of them were housed in to a cage with unlimited food and water access. Thirty days after the estradiol valerate injection, animals were IM administered by 7.5 IU of PMSG. Subsequent of 48 hours, the mice in experimental groups were anesthetized with ketamine and xylazine and then sacrificed, after that their ovaries were removed and separated from other tissues. In order to provide oocytes, GV oocytes were isolated from ovaries and cultured in TEM199. After insemination of MII oocytes resulted from incubation of GV oocytes, the investigation of fertilized oocytes (zygotes), blastocysts and hatched embryos were carried out by inverted microspore. Then the ovaries were fixed at least for a day in formaldehyde 10%. Prepared tissues according to standard protocols were used for counting of different type of ovarian follicles. Tissue staining was carried out with hematoxylin and eosin. All types of follicles were counted in each ovary.
Results and Discussion: Numerous experimental models for PCOs have been developed using estradiol valerate. Estradiol valerate can produce a PCOs model in mouse with interrupting in metabolic and physiologic processes. In this study, morphological examination of the ovaries showed that exposure of young adult female mice to a single dose of exogenous estradiol valerate can inhibit ovulation and consequently induces PCOs. The amounts of serum testosterone and estradiol substantially decreased in treated groups in comparison with PCO mice (p < 0.05). Increase of testosterone is one of the most important symptoms in PCOs, but use of vitex extract decrease testosterone concentration whether in groups consuming 365 or 730 mg/kg/day that is because of less number of follicular cysts in these groups versus PCO group (p < 0.05). In this regard it is observed total number of follicles with less than 70 µm and more than 200 µm significantly was higher than PCO group (p < 0.05). These results are expected because polycystic ovary is a disorder of follicle development. In PCOs, follicles arrest in immature stages and therefore dominant follicle selection will not occur. This may be due to lack of positive estrogen feedback to the hypothalamus and pituitary axis. As the result of this lacking, LH surge will not occur and therefore ovulation will be disrupted and finally follicular cysts remain on the ovaries.
The number of matured oocytes (MII) in PCO group + 730 mg/kg/day was significantly higher than PCO group (p < 0.05). Also there were no significant differences in percentage of fertilized oocytes (zygotes) and Two-cell embryos between experimental groups (p>0.05). On the other hand, the percentage of produced blastocyst and hatched embryos in group PCO + 365 mg/kg/day was higher than PCO group (p < 0.05). With administration of vitex extract more ovarian follicles ovulated and their oocytes were higher in quality (matured oocytes) that finally, produced more blastocysts and hatched embryos.
Conclusion: The result of present study showed that PCOs can be induced in adult mice with a single dose injection of Estradiol valerate. Also it seems vitex extract consumption can induce more oocytes for fertilization and consequently, production of embryos in animals with PCO syndrome.
Conflict of interest: None declared.
Keywords: PCOS, Vitex, In Vitro Maturation, In Vitro Fertilization.

کلیدواژه‌ها [English]

  • PCOs
  • Vitex
  • In vitro maturation
  • in vitro fertilization
  • Altonen, J., Laitinen, M.P., Voujolainen, K., Jaatinen, R., Horelli-­Kuitunen, N. and Seppä, L. (1999). Human growth differentiation factor 9 (GDF-9) and its novel homolog GDF-9B are expressed in oocytes during early folliculogenesis. The Journal of Clinical Endocrinology and Metabolism, 84(8): 2744-2750.
  • Borrione, P., Di Luigi, L., Maffulli, N. and Pigozzi, F. (2008). Herbal supplements: cause for concern? Journal of Sport Science and Medicine, 7(4): 562-564.
  • Brock, B., Smidt, K., Ovesen, P., Schmitz, O. and Rungby, J. (2005). Is metformin therapy for Polycystic Ovary Syndrome safe during pregnancy? Basic & Clinical Pharmacology and Toxicology, 96(6): 410-412.
  • Daghigh Kia, H., Sadeghi Sadegh Abad, F., Ebrahimi, M. and Samadian, F. (2017). Comparative effect of different concentrations of hydro-ethanolic extract of chamomile on freeze-thawn semen quality of rams. Veterinary Clinical Pathology, 11(41): 13-23. [In Persian]
  • Daniele, C., Thopson, C.J., Pittler, M.H. and Ernest, E. (2005). Vitex agnus-­castus a systemathic review of adverse event. Drug Safety, 28(4): 319-322.
  • Dumesic, D.A., Meldrum, D.R., Katz-Jaffe, M.G., Krisher, R.L. and Schoolcraft, W.B. (2015). Oocyte environment: follicular fluid and cumulus cells are critical for oocyte health. Fertility and Sterility, 103(2): 303-316.
  • Dumesic, D.A, Padmanabhan, V. and Abbott, D.H. (2008). Polycystic ovary syndrome and oocyte developmental competence. Obstetrics and Gynecology, 63(1): 39-48.
  • Franks, S. (2009). Do Animal Models of Polycystic Ovary Syndrome Help to Understand Its Pathogenesis and Management? Yes, but Their Limitations should be Recognized. Endocrinology, 150(9): 3983-3985.
  • Gonzalez, F., Thusu, K., Abdel-Rahman, E., Prabhala, A., Tomani, M. and Dandona, P. (1999). Elevated serum levels of tumor necrosis factor alpha in normal-weight women with polycystic ovary syndrome. Metabolism, 48(4): 437-441.
  • Henmi, H., Endo, T., Nagasawa, K., Hayashi, T., Chida, M. and Akutagawa, N. (2001). Lysyl oxidase and MMP-2 expression in dehydroepiandrosterone induced polycystic ovary in rats. Biology of Reproduction, 64(1): 157-162.
  • Hossain Rashidi, B. and Nemati, M. (2017). Effects of Vitex agnus-castus extract on the secretory function of pituitary-gonadal axis and pregnancy rate in patients with premature ovarian aging (POA). Journal of Herbal Medicine, 10: 24-30.
  • Handa, R.J., Pak, T.R., Kudwa, A.E., Lund, T.D. and Hinds, L. (2008) An alternate pathway for androgen regulation of brain function: activation of estrogen receptor beta by the metabolite of dihydrotestosterone, 5alpha-androstane-3beta,17 beta-diol. Hormones and Behavior, 53(5): ­741-752.
  • Jelodar, G.A. and Askari, K. (2012). Effect of Vitex agnus castus fruits hydroalcoholic extract on sex hormones in rat with induced polycystic ovary syndrome (PCOS). Journal of Physiology and Pharmacology, 16(1): 62-69. [In Persian]
  • Jelodar, G.A. and Askari, K. (2017). Effect of hydroalcoholic extract of Vitex agnus-castus fruit on fertility and estrous cycle in letrozole-­induced polycystic ovary syndrome in rat. Razi Journal of Medical Sciences, 24(156): 42-48. [In Persian]
  • Marx, T.L. and Mehta, A.E. (2003). Polycystic ovary syndrome: pathogenesis and treatment over the short and long term. Cleveland Clinic Journal of Medicine, 70(1): 31-45.
  • Mohammadzadeh, H., Delashoub, M. and Khakpour, M. (2018). Effect of vitamin E in prevention of lipopolysaccharide induced fetal injuries in the rat. Veterinary Clinical Pathology, 11(44): 367-377. [In Persian]
  • Mostahsan, Z. and Mortazavi, P. (2019). The effects of Tilia (Tilia plathyphyllos) extract on gene expression of caspase­3 and caspase­9 in canine mammary gland cancer cell line (CF41.Mg). Veterinary Clinical Pathology, 13(49): 2-14. [In Persian]
  • Nabiuni, M., Mohammadi, S., Kayedpoor, P. and Karimzadeh, L., (2015). The effect of curcumin on the estradiol valerate-induced polycystic ovary in rats. Kashan University of Medical Sciences Journal (FEYZ), 18(6): 515-523. [In Persian]
  • Nasri, S., Oryan, S.h., HaeriRohani, A., Amin, G.H. and Taghizadeh, M. (2005). The effects of Vitexagnuscastus L. extract and interaction with bromocriptine on luteinizing hormone and testosterone in male mice. Medical Journal of Hormozgan University, 9(2): 113-118. [In Persian]
  • Noorafshan, A., Ahmadi, M., Mesbah, S. and Karbalay-Doust, S. (2013). Stereological study of the effect of letrozole and estradiol valerate treatment on the ovary of rats. Clinical and Experimental Reproductive Medicine, 40(3): 115-121.
  • Palep-Singh, M., Picton, H.M., Yates, Z.R., Barth, J.H. and Balen, A.H. (2008). Plasma homocysteine concentrations and the single nucleotide polymorphisms in the methionine synthase gene (MTR 2756A>G): Associations with the polycystic ovary syndrome an observational study. The European Journal of Obstetrics & Gynecology and Reproductive Biology, 138(2): 180-186.
  • Rani, A. and Sharma, A. (2013). The genus Vitex: A review. Pharmacognosy Reviews, 7(14): 188–198.
  • Robinson, J.E., Forsdike, R.A. and Taylor, J.A. (1999). In utero exposure of female lambs to testosterone reduces the sensitivity of the GnRH neuronal network to inhibition by progesterone. Endocrinology, 140(12): 5797-5805.
  • Shafiee, M.N., Malik, D.A., Yunos, R.I.M., Atiomo, W., Omar, M.H., Ghani, N.A.A. et al. (2015). The effect of Metformin on endometrial tumor-regulatory genes and systemic metabolic parameters in polycystic ovarian syndrome–a proof-of-concept study. Gynecological Endocrinology, 31(4): 286-290.
  • Shamsi, M., Nejati, V. and Najafi, G. (2015). Therapeutic Effects of Licorice Extract on In vitro Maturation and In vitro Fertilization in Mice Model of Polycystic Ovary Syndrome. Journal of Mazandaran University of Medical Science, 25(132): 113-121. [In Persian]
  • Stadtmauer, L.A., Toma, S.K., Riehl, R.M. and Talbert, L.M. (2001). Metformin treatment of patients with polycystic ovary syndrome undergoing in vitro fertilization improves outcomes and is associated with modulation of the insulin-like growth factor. Fertility and Sterility, 75(3): 505-509.
  • Tsilchorozidou, T., Overton, C. and Conway, G.S. (2004). The pathophysiology of polycystic ovary syndrome. Clinical Endocrinology, 60(1): 1-17.
  • van Houten, E.L. and Visser, J.A. (2014). Mouse models to study polycystic ovary syndrome: a possible link between metabolism and ovarian function? Reproductive Biology, 14(1): 32-43.