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Human ovarian tissue vitrification/warming has minor effect on the expression of apoptosis-related genes.

ABSTRACT: BACKGROUND:In this study, we evaluated the incidence of apoptosis at the ultrastructural levels and expression of some apoptosis-related genes in vitrified human ovarian tissue just after warming. METHODS:Human ovarian tissue biopsies from 23 women after caesarean section were transported to the laboratory within 2 hours, and then they were cut into small pieces. Some pieces were vitrified and warmed and the other samples were considered as control. Apoptosis was assessed by a transmission electron microscope and also by molecular analysis of pro-apoptotic (Fas, FasL, Bax, p53, caspase8, and caspase3) and antiapoptotic (Bcl-2 and BIRC5) genem RNA levels using real-time RT-PCR before and after vitrification. RESULTS:No sign of apoptosis was shown ultrastructurally in vitrified samples. The level of FasL, Bcl-2, Bax, p53, and caspase3 mRNA and Bax:Bcl-2 ratio were similar in non-vitrified and vitrified groups; however, the expression of Fas and caspase8 genes was higher and BIRC5 was lower in vitrified samples compared to non-vitrified group (P<0.05). CONCLUSION:The fine structure of human vitrified ovarian tissue was well preserved; moreover, vitrification was shown to affect the expression of some apoptosis-related genes. However, additional study is needed to confirm this observation.

SUBMITTER: Abdollahi M

PROVIDER: S-EPMC3882920 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Human ovarian tissue vitrification/warming has minor effect on the expression of apoptosis-related genes.

Abdollahi Maasoume M Salehnia Mojdeh M Salehpour Saghar S Ghorbanmehr Nassim N

Iranian biomedical journal 20130101 4

<h4>Background</h4>In this study, we evaluated the incidence of apoptosis at the ultrastructural levels and expression of some apoptosis-related genes in vitrified human ovarian tissue just after warming.<h4>Methods</h4>Human ovarian tissue biopsies from 23 women after caesarean section were transported to the laboratory within 2 hours, and then they were cut into small pieces. Some pieces were vitrified and warmed and the other samples were considered as control. Apoptosis was assessed by a tra ...[more]

PMID: 23999713

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Project description:BackgroundAfter ovarian tissue transplantation, ischemia-reperfusion injury and free radicals cause follicle depletion and apoptosis. Therefore, the use of antioxidants to reduce the production of free radicals is an important method to address the consequences of ischemia-reperfusion injury. Resveratrol is a natural active polyphenol compound with anti-inflammatory, antitumor, strong antioxidant and anti-free radical properties. The aim of this study was to investigate whether resveratrol could improve the effect of autologous ovarian transplantation after cryopreserve-thawn mouse ovarian tissue.MethodsWhole-ovary vitrification and autotransplantation models were used to investigate the effects of resveratrol. Six-week-old female mice from the Institute of Cancer Research (ICR) were subjected to vitrification. All ovaries were preserved in liquid nitrogen for 1 week before being thawed. After thawing, ovarian tissues were autotransplanted in the bilateral kidney capsules. Mice (n = 72) were randomly divided into four groups to determine the optimal concentration of resveratrol (experiment I). Treatments were given as follows: saline, 5 mg/kg resveratrol, 15 mg/kg resveratrol and 45 mg/kg resveratrol, which were administered orally for one week. Grafted ovaries were collected for analysis on days 3, 7, and 21 after transplantation. Ovarian follicle morphology was assessed by hematoxylin and eosin staining. Serum FSH and E2 levels were measured to estimate the transplanted ovarian reserve and endocrine function. Other mice were randomly divided into two groups-saline and 45 mg/kg resveratrol to further evaluate the effect of resveratrol and explore the mechanisms underlying this effect (experiment II). Ovarian follicle apoptosis was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assays. Immunohistochemistry, qRT-PCR and western blotting (MDA, SOD, NF-κB, IL-6 and SIRT1) were used to explore the mechanisms of resveratrol. Moreover, oocytes derived from autotransplanted ovaries at 21 days were cultured and fertilized in vitro.ResultsThe proportions of morphologically normal (G1) follicles at 3, 7 and 21 days were significantly higher in the 45 mg/kg resveratrol group than in the saline group. The TUNEL-stained follicles (%) at 7 days were significantly decreased in the 45 mg/kg resveratrol group compared with the saline group. Western blot analysis revealed that SOD2 and SIRT1 levels were significantly higher in the 45 mg/kg resveratrol group than in the saline group at day 7 and that MDA and NF-κB levels were lower in the saline group on day 3. Likewise, IL-6 was lower in the saline group on day 7. These results are basically consistent with the qRT-PCR results. In addition, the mean number of retrieved oocytes and fertilization and cleavage were significantly increased in the 45 mg/kg resveratrol group compared with the saline group.ConclusionsAdministration of resveratrol could improve the quality of cryopreserved mouse ovarian tissue after transplantation and the embryo outcome, through anti-inflammatory and antioxidative mechanisms.

Project description:BACKGROUND: The effect of hormonal estrus induction on maternal effect (MATER - maternal antigen that embryo requires, ZAR-1 - zygote arrest 1, and BMP15 - bone morphogenetic protein 15) and apoptosis-related genes expression (BCL-2 and BAX) in porcine cumulus-oocyte complexes (COCs) and selected follicular parameters was investigated in this study. METHODS: Gilts were divided into three groups: (I) with natural estrus; (II) stimulated with PMSG/hCG; and (III) with PMSG/hCG?+?PGF2alpha. Analysis of maternal effect and apoptosis-related transcripts expression in COCs, and progesterone synthesis pathway genes expression (P450scc and 3betaHSD) in granulosa cells was performed by qPCR. BMP15 protein expression in follicular fluid (FF) was analyzed by western blot. Oocyte nuclear maturation was assessed by aceto-orcein staining. Progesterone (P4) and estradiol (E2) concentrations in FF and serum were measured by ELISA. Data were analyzed with the one-way ANOVA and Bonferroni post-test or Kruskal-Wallis test and Dunns post-test. RESULTS: The highest expression of MATER, ZAR-1, and BMP15 genes was found in COCs recovered from gilts treated with PMSG/hCG when compared to PMSG/hCG?+?PGF2alpha-stimulated or non-stimulated gilts. Hormonal treatment did not affect the BMP15 protein expression in FF, but increased the expression of genes participating in P4 synthesis in granulosa cells. The higher percentage of immature oocytes was found in PMSG/hCG-treated when compared to the non-stimulated gilts. The expression of BCL-2 and BAX mRNA, and BCL-2/BAX mRNA ratio was significantly higher in COCs derived from PMSG/hCG-treated when compared to PMSG/hCG + PGF2alpha-treated or non-stimulated subjects. The level of P4 in serum was similar in animals from all experimental groups, while its concentration in FF was greater in gilts subjected to PMSG/hCG treatment than in PMSG/hCG + PGF2alpha-stimulated and non-stimulated gilts. The concentration of E2 did not differ in the serum or FF between the control group and the hormonally stimulated groups. CONCLUSIONS: Hormonal induction of estrus affected maternal effect gene transcripts levels in COCs and and oocyte nuclear maturation. The inclusion of PGF2alpha into the stimulation protocol enabled maintaining of physiological concentration of P4 in FF. Additionally, both hormonal treatments seem to be beneficial for apoptosis prevention through increasing BCL-2/BAX transcript ratio.

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Human ovarian tissue vitrification/warming has minor effect on the expression of apoptosis-related genes.

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Human ovarian tissue vitrification/warming has minor effect on the expression of apoptosis-related genes.

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