ABSTRACT
Objective: Over the last years, vitrification has been widely used for oocyte cryopreservation, in animals and humans; however, it frequently causes minor and major epigenetic modifications. The effect of oocyte vitrification on levels of acetylation of histone H4 at lysine 12 [AcH4K12], and histone acetyltransferase [Hat] expression, was previously assessed; however, little is known about the inhibition of Hat expression during oocyte vitrification. This study evaluated the effect of anacardic acid [AA] as a Hat inhibitor on vitrified mouse oocytes
Materials and Methods: In this experimental study, 248 mouse oocytes at metaphase II [MII] stage were divided in three experimental groups namely, fresh control oocytes [which were not affected by vitrification], frozen/thawed oocytes [vitrified] and frozen/thawed oocytes pre-treated with AA [treatment]. Out of 248 oocytes, 173 oocytes were selected and from them, 84 oocytes were vitrified without AA [vitrified group] and 89 oocytes were pretreated with AA, and then vitrified [treatment group]. Fresh MII mouse oocytes were used as control group. Hat expression and AcH4K12 levels were assessed by using real-time quantitative polymerase chain reaction [PCR] and immunofluoresce staining, respectively. In addition, survival rate was determined in vitrified and treatment oocytes
Results: Hat expression and AcH4K12 modification significantly increased [4.17 +/- 1.27 [P.0.001] and 97.57 +/- 6.30 [P<0.001], respectively] in oocytes that were vitrified, compared to the fresh oocytes. After treatment with AA, the Hat mRNA expression and subsequently H4K12 acetylation levels were significantly reduced [0.12 +/- 0.03 [P.0.001] and 89.79 +/- 3.20 [P.0.05], respectively] in comparison to the vitrified group. However, the survival rate was not significantly different between the vitrified [90.47%] and treatment [91.01%] groups [P>0.05]
Conclusion: The present study suggests that AA reduces vitrification risks caused by epigenetic modifications, but does not affect the quality of vitrification. In fact, AA as a Hat inhibitor was effective in reducing the acetylation levels of H4K12
ABSTRACT
Objective: In vitro maturation technique [IVM] is shown to have an effect on full maturation of immature oocytes and the subsequent embryo development. Embryonic genome activation [EGA] is considered as a crucial and the first process after fertilization. EGA failure leads to embryo arrest and possible implantation failure. This study aimed to determine the role of IVM in EGA-related genes expression in human embryo originated from immature oocytes and recovered from women receiving gonadotrophin treatment for assisted reproduction
Materials and Methods: In this experimental study, germinal vesicle [GV] oocytes were cultured in vitro. After intracytoplasmic sperm injection of the oocytes, fertilization, cleavage and embryo quality score were assessed in vitro and in vivo. After 3-4 days, a single blastomere was biopsied from the embryos and then frozen. Afterwards, the expression of EGA-related genes in embryos was assayed using quantitative reverse transcriptase-polymerase chain reaction [PCR]
Results: The in vitro study showed reduced quality of embryos. No significant difference was found between embryo quality scores for the two groups [P=0.754]. The in vitro group exhibited a relatively reduced expression of the EGA- related genes, when compared to the in vivo group [all of them showed P=0.0001]
Conclusion: Although displaying the normal morphology, the IVM process appeared to have a negative influence on developmental gene expression levels of human preimplanted embryos. Based on our results, the embryo normal morphology cannot be considered as an ideal scale for the successful growth of embryo at implantation and downstream processes
ABSTRACT
Background: oocyte developmental competence is one of the key factors for determining the success rate of assisted reproductive technique
Objective: the aim of the current study was to investigate the effect of L-carnitine [LC] supplementation during in vitro maturation [IVM], on preimplantation embryo development and expression of genes involved in embryo competence derived from oocytes selected with brilliant cresyl blue [BCB] test
Materials and Methods: cumulus-oocyte complexes [COCs] were obtained from NMRI mice ovaries. COCs were stained with BCB and then BCB+ [colored cytoplasm] oocytes cultured in IVM medium supplemented with 0.3 or 0.6 mg/ml LC. COCs untreated with LC were used as control. Fertilization rate and blastocyst development rate were determined after in vitro fertilization. In addition, quantitative reverse transcriptase polymerase chain reaction was used to measure relative genes expression related with development [Ccnb1, Mos, Ces5, and Dppa2] and apoptosis [Bax and Bcl-xL] in oocytes and embryos
Results: oocytes treated with both LC concentrations showed higher blastocyst development rate compared with untreated oocytes [p<0.01]. Moreover, fertilization rate was increased in oocytes treated with 0.6 mg/ml LC [p<0.01]. Treatment of oocytes with both LC concentrations increased [p<0.01] the level of Ccnb1 mRNA in MII oocytes. The two-cell stage embryos and blastocysts derived from LC-treated oocytes [0.6 mg/ml] showed increased the expression levels of Dppa2 and Bcl-xl mRNA, respectively [p<0.01]
Conclusion: the results of the present study show that adding of LC to the IVM medium of BCB+ oocytes can ameliorate reproductive success following in vitro fertilization