139 PERTURBATION OF THE DYNAMICS OF DNA METHYLATION IN THE PATERNAL GENOME FOLLOWING IN VITRO FERTILIZATION IN CATTLE: THE SECRETS OF HEAT STRESS EFFECTS

2014 ◽  
Vol 26 (1) ◽  
pp. 183
Author(s):  
M. B. Rahman ◽  
T. Rijsselaere ◽  
A. Van Soom
Keyword(s):  
Dna Methylation ◽  
Heat Stress ◽  
In Vitro Fertilization ◽  
Chromatin Condensation ◽  
Histone Variants ◽  
Dna Demethylation ◽  
Paternal Genome ◽  
Vitro Fertilization ◽  
Fourier Harmonic

Ejaculates collected from breeding bulls during the summer season under tropical or subtropical conditions are often reported to have a lower fertilization potential. We recently reported that sperm cells at post-meiotic stages of development were more susceptible to heat stress (scrotal insulation technique). In post-meiotic stages of sperm cell development, extensive incorporation of histone variants and hyperacetylation confirms unstable chromatin. The unstable forms of chromatin are more vulnerable to heat stress and affect sperm DNA-protamine condensation. By using a special stain and nuclear morphometric assessment, Chromomycin A3 and Fourier harmonic analysis, respectively, we observed protamine-deficient spermatozoa with altered chromatin condensation in the heat-stressed ejaculates. In a recent study, we investigated the patterns of global DNA methylation in pronuclear development and fertilization potential of such altered chromatin condensed spermatozoa. To this end, 1239 in vitro-matured oocytes were fertilized with spermatozoa of 3 groups (i.e. standard IVF control, non-heat-stressed control, and heat-stressed spermatozoa). Data were analysed by means of ANOVA. The results showed that heat-stressed spermatozoa with altered chromatin condensation perturb the dynamics of DNA methylation reprogramming in the paternal pronucleus (i.e. disordered the active demethylation followed by a de novo methylation pattern during one cell stage). In addition, there was a tendency for a decrease in the size of both paternal and maternal pronuclei developed after fertilization with heat-stressed spermatozoa in comparison with standard IVF control spermatozoa, leading to lower fertilization rates (54 v. 82%). Future studies should focus on the mechanisms of perturbed DNA demethylation in the paternal genome after in vitro fertilization of oocytes with spermatozoa having altered chromatin condensation.

Altered chromatin condensation of heat-stressed spermatozoa perturbs the dynamics of DNA methylation reprogramming in the paternal genome after in vitro fertilisation in cattle

2014 ◽  
Vol 26 (8) ◽  
pp. 1107 ◽  
Author(s):  
Mohammad Bozlur Rahman ◽  
Md. Mostofa Kamal ◽  
Tom Rijsselaere ◽  
Leen Vandaele ◽  
Mohammed Shamsuddin ◽  
...  
Keyword(s):  
Dna Methylation ◽  
De Novo ◽  
Chromatin Condensation ◽  
Dna Demethylation ◽  
In Vitro Fertilisation ◽  
Paternal Genome ◽  
Time Points ◽  
Fertilisation Rate ◽  
De Novo Methylation

Shortly after penetration of the oocyte, sperm DNA is actively demethylated, which is required for totipotent zygotic development. Aberrant DNA methylation is thought to be associated with altered chromatin condensation of spermatozoa. The objectives of this study were to investigate the dynamics of DNA methylation reprogramming in the paternal pronucleus and subsequent fertilisation potential of heat-stressed bull spermatozoa having altered chromatin condensation. Hence, bovine zygotes (n = 1239) were collected at three different time points (12, 18 and 24 h post insemination, hpi), and stained with an antibody against 5-methylcytosine. Fluorescence intensities of paternal and maternal pronuclei were measured by ImageJ. DNA methylation patterns in paternal pronuclei derived from heat-stressed spermatozoa did not differ between time points (P > 0.05), whereas control zygotes clearly showed demethylation and de novo methylation at 18 and 24 hpi, respectively. Moreover, heat-stressed spermatozoa showed a highly reduced (P < 0.01) fertilisation rate compared with non-heat-stressed or normal control spermatozoa (53.7% vs 70.2% or 81.5%, respectively). Our data show that the normal pattern of active DNA demethylation followed by de novo methylation in the paternal pronucleus is perturbed when oocytes are fertilised with heat-stressed spermatozoa, which may be responsible for decreased fertilisation potential.

scholarly journals DNA METHYLATION DIFFERENCES IN LIVER OF MICE CONCEIVED BY IN VITRO FERTILIZATION

2020 ◽  
Vol 114 (3) ◽  
pp. e153-e154
Author(s):  
Saúl Lira-Albarrán ◽  
Xiaowei Liu ◽  
Paolo Rinaudo
Keyword(s):  
Dna Methylation ◽  
In Vitro Fertilization ◽  
Vitro Fertilization

scholarly journals SUN-715 IIM May Influence Matured Oocytes’ DNA Methylation of PCOS Patients

2020 ◽  
Vol 4 (Supplement_1) ◽  
Author(s):  
Congru Li ◽  
Yang Yu
Keyword(s):  
Dna Methylation ◽  
In Vitro Fertilization ◽  
Embryonic Development ◽  
Embryo Transfer ◽  
Cpg Island ◽  
Reproductive Technologies ◽  
Childbearing Age ◽  
Vitro Fertilization ◽  
The Impact

Abstract Polycystic ovary syndrome (PCOS) is the most common endocrine disorder in women of childbearing age and is the main cause of anovulatory infertility. To increase the number of oocytes obtained, controlled ovarian stimulation (COS) has become a routine choice for in vitro fertilization-embryo transfer (IVF-ET), which is one of the common assisted reproductive technologies for PCOS patients. However, for these patients, there is a high risk of ovarian hyperstimulation syndrome (OHSS). Obtaining in vitro maturation (IVM) of immature oocytes, and then in vitro fertilization and embryo transfer of mature oocytes provides a possible way for people to solve the above problems. Since the IVM technology will expose oocytes to in vitro conditions for a longer period of time, theoretically increasing the risk of the oocytes being affected by the culture environment, further research and explorations are needed for study in gene programming, epigenetics, etc. Therefore, to explore the impact of IVM operation on embryonic development is of great significance for further clarifying assisted reproductive safety and improving IVM operation conditions. Here we focused on DNA methylation reprogramming process which was essential for embryonic development. We tested the DNA methylation of sperm, IVM oocytes and IVM generated early stage embryos including pronucleus, 4cell, 8cell, morula, inner cell mass, trophoectoderm (TE) as well as six-week embryos by Nimble Gen Human DNA Methylation 3x729K CpG Island Plus RefSeq Promoter Array and compared the data with our published genome-wide DNA methylomes of human gametes and early embryos generated from in vivo maturation oocytes. We showed that IVM embryos show abnormal DNA methylation reprogramming pattern. By analyzing the abnormally reprogrammed promoters, we further found that IVM may affect the functions of demethylation related genes. Oocytes from IVM manipulation were tested with higher DNA methylation levels, and their abnormal methylated promoters mainly enriched in immune and metabolism pathways. Furthermore, we investigated the DNA methylation of TE, which was directly related with implantation process and revealed the abnormal methylated promoters were related with metabolism pathway too. Our data support that IVM may influence the DNA methylome of oocytes, which in turn affects the methylome of their embryos. However, due to the limited number of samples and the inability of the chip to cover all CpG sites, the results of this study require further research and validation.

50 DNA METHYLATION PROFILES OF UPSTREAM ELEMENTS OF Oct-3/4 GENE IN IN VITRO FERTILIZATION (IVF) AND SOMATIC CELL NUCLEAR-TRANSFERRED (SCNT) EMBRYOS

2007 ◽  
Vol 19 (1) ◽  
pp. 143
Author(s):  
M. Kawasumi ◽  
Y. Unno ◽  
M. Nishiwaki ◽  
K. Matsumoto ◽  
M. Anzai ◽  
...  
Keyword(s):  
Dna Methylation ◽  
In Vitro Fertilization ◽  
Somatic Cell ◽  
Methylation Level ◽  
Nuclear Transfer ◽  
Upstream Region ◽  
Sequencing Analysis ◽  
Vitro Fertilization ◽  
Cpg Sites

Cloning by adult somatic cell nuclear transfer (SCNT) has proven to be successful for the production of clones from many species (Keith 2004 Cytogenet. Genome Res. 105, 285). However, somatic cloning is currently highly inefficient. One of the reasons for this is that SCNT is believed to be associated with epigenetic errors including abnormal DNA methylation of the reconstructed embryo. The Oct-3/4 gene, a member of the POU transcription factor family, is expressed throughout the pre-implantation embryo. Abnormal expression of the Oct-3/4 gene in the nuclear-transferred embryo is either directly or indirectly caused by nuclear transfer and is suggested to be indicative of a general failure to reset the genetic program (Boiani et al. 2002 Genes Dev. 16, 1209). In this study, we investigated the DNA methylation profiles of the Oct-3/4 gene in the genome of SCNT embryos, using bisulfite sequencing analysis. Then, we observed the detailed subcellular localization of Oct-3/4 proteins in SCNT embryos using immunocytochemical (ICC) analysis. Nuclear transfer of cumulus cell nuclei was carried out as previously described (Wakayama et al. 1998 Nature 394, 369). After nuclear transfer, embryos were subsequently cultured in KSOM media to the morula and blastocyst stages. We compared the methylation profiles of 3 transcriptional control elements (distal enhancer, DE; proximal enhancer, PE; and promoter) of the upstream region of the Oct-3/4 gene with the genome of in vitro fertilization (IVF) and SCNT embryos. The methylation rate of CpG sites in the DE and promoter regions of both IVF and SCNT embryos was low at both the morula and the blastocyst stages. What&apos;s interesting is that there was a significant difference in the methylation level on CpG sites in the PE element between IVF and SCNT embryos. At the morula stage, the methylation level on CpG sites in the PE element was very low in the IVF embryo and moderately high in the SCNT embryo (0.9&percnt; and 26.3&percnt;). Conversely, at the blastocyst stage, CpG sites in the PE element showed high methylation in the IVF embryo and low methylation in the SCNT embryo (55.2&percnt; and 10.5&percnt;). CpG sites in the PE element were lightly methylated (3.0&percnt;) in the inner cell mass (ICM) of the IVF embryo. This means that the main portion of methylation in the IVF blastocyst embryo occurred at the trophectoderm (TE). On the other hand, in ICM of the SCNT embryo, the methylation level of each embryonic cell was almost the same in the whole blastocyst embryo (9.8&percnt; and 10.5&percnt;). As a result, it is highly possible that the CpG sites in the PE element of ICM were methylated as in the TE. ICC analysis revealed that some SCNT embryos showed aberrant Oct-3/4 expression in the TE. These results indicate that the methylation of CpG sites in the Oct-3/4 PE element may be related to expression of Oct-3/4 in the mouse IVF and SCNT embryos. These differences in methylation level between IVF and SCNT embryos were reflected as abnormal expressions of Oct-3/4 on SCNT embryos. This study was supported by the 21st COE Program of MEST. M.K. is a JSPS Research Fellow and supported by Grant-in Aid for Scientific Research (No. 1751132) of JSPS.

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