scholarly journals A new mode of mechano-transduction shakes the oocyte nucleus, thereby fine tunes gene expression modulating the developmental potential

2021 ◽  
Vol 343 (3) ◽  
pp. 223-234
Author(s):  
Maria Almonacid ◽  
Marie-Hélène Verlhac
Keyword(s):  
Gene Expression ◽  
Oocyte Nucleus ◽  
Developmental Potential

The responsiveness of embryonic stem cells to alpha and beta interferons provides the basis of an inducible expression system for analysis of developmental control genes

1993 ◽  
Vol 13 (12) ◽  
pp. 7971-7976
Author(s):  
L M Whyatt ◽  
A Düwel ◽  
A G Smith ◽  
P D Rathjen
Keyword(s):  
Gene Expression ◽  
Inner Cell Mass ◽  
Es Cells ◽  
Expression System ◽  
Cell Mass ◽  
Embryonic Stem ◽  
Expression Vectors ◽  
Developmental Control ◽  
Developmental Potential

Embryonic stem (ES) cells, derived from the inner cell mass of the preimplantation mouse embryo, are used increasingly as an experimental tool for the investigation of early mammalian development. The differentiation of these cells in vitro can be used as an assay for factors that regulate early developmental decisions in the embryo, while the effects of altered gene expression during early embryogenesis can be analyzed in chimeric mice generated from modified ES cells. The experimental versatility of ES cells would be significantly increased by the development of systems which allow precise control of heterologous gene expression. In this paper, we report that ES cells are responsive to alpha and beta interferons (IFNs). This property has been exploited for the development of inducible ES cell expression vectors, using the promoter of the human IFN-inducible gene, 6-16. The properties of these vectors have been analyzed in both transiently and stably transfected ES cells. Expression was minimal or absent in unstimulated ES cells, could be stimulated up to 100-fold by treatment of the cells with IFN, and increased in linear fashion with increasing levels of IFN. High levels of induced expression were maintained for extended periods of time in the continuous presence of the inducing signal or following a 12-h pulse with IFN. Treatment of ES cells with IFN did not affect their growth or differentiation in vitro or compromise their developmental potential. This combination of features makes the 6-16-based expression vectors suitable for the functional analysis of developmental control control genes in ES cells.

scholarly journals In vivo and in vitro maturation of rabbit oocytes differently affects the gene expression profile, mitochondrial distribution, apoptosis and early embryo development

2017 ◽  
Vol 29 (9) ◽  
pp. 1667 ◽  
Author(s):  
M. Arias-Álvarez ◽  
R. M. García-García ◽  
J. López-Tello ◽  
P. G. Rebollar ◽  
A. Gutiérrez-Adán ◽  
...  
Keyword(s):  
Gene Expression ◽  
Rabbit Model ◽  
Developmental Competence ◽  
Gene Markers ◽  
Developmental Potential ◽  
Junction Protein ◽  
Mitochondrial Distribution ◽  
Potential Biomarkers

In vivo-matured cumulus–oocyte complexes are valuable models in which to assess potential biomarkers of rabbit oocyte quality that contribute to enhanced IVM systems. In the present study we compared some gene markers of oocytes and cumulus cells (CCs) from immature, in vivo-matured and IVM oocytes. Moreover, apoptosis in CCs, nuclear maturation, mitochondrial reallocation and the developmental potential of oocytes after IVF were assessed. In relation to cumulus expansion, gene expression of gap junction protein, alpha 1, 43 kDa (Gja1) and prostaglandin-endoperoxide synthase 2 (Ptgs2) was significantly lower in CCs after in vivo maturation than IVM. In addition, there were differences in gene expression after in vivo maturation versus IVM in both oocytes and CCs for genes related to cell cycle regulation and apoptosis (V-Akt murine thymoma viral oncogene homologue 1 (Akt1), tumour protein 53 (Tp53), caspase 3, apoptosis-related cysteine protease (Casp3)), oxidative response (superoxide dismutase 2, mitochondrial (Sod2)) and metabolism (glucose-6-phosphate dehydrogenase (G6pd), glyceraldehyde-3-phosphate dehydrogenase (Gapdh)). In vivo-matured CCs had a lower apoptosis rate than IVM and immature CCs. Meiotic progression, mitochondrial migration to the periphery and developmental competence were higher for in vivo-matured than IVM oocytes. In conclusion, differences in oocyte developmental capacity after IVM or in vivo maturation are accompanied by significant changes in transcript abundance in oocytes and their surrounding CCs, meiotic rate, mitochondrial distribution and apoptotic index. Some of the genes investigated, such as Gja1, could be potential biomarkers for oocyte developmental competence in the rabbit model, helping improve in vitro culture systems in these species.

scholarly journals Developmental and molecular correlates of bovine preimplantation embryos

Reproduction ◽  
2006 ◽  
Vol 131 (5) ◽  
pp. 895-904 ◽  
Author(s):  
Hakan Sagirkaya ◽  
Muge Misirlioglu ◽  
Abdullah Kaya ◽  
Neal L First ◽  
John J Parrish ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methyltransferase ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Blastocyst Stage ◽  
Culture Conditions ◽  
Preimplantation Embryos ◽  
Developmental Potential

Expression of embryonic genes is altered in different culture conditions, which influence developmental potential both during preimplantation and fetal development. The objective of this study was to define the effects of culture conditions on: bovine embryonic development to blastocyst stage, blastocyst cell number, apoptosis and expression patterns of a panel of developmentally important genes. Bovine embryos were culturedin vitroin three culture media containing amino acids, namely potassium simplex optimization medium (KSOMaa), Charles Rosenkrans 1 (CR1aa) and synthetic oviductal fluid (SOFaa). Apoptosis in blastocysts was determined by TUNEL assay and expression profiles of developmentally important genes were assayed by real-time PCR.In vivo-produced bovine blastocysts were used as controls for experiments determining gene expression patterns. While the cleavage rates did not differ, embryos cultured in SOFaa had higher rates of development to blastocyst stage (P< 0.05). Mean cell numbers and percentages of apoptotic cells per blastocyst did not differ among the groups. Expression of the heat shock protein 70 (Hsp70) gene was significantly up-regulated in both CR1aa and KSOMaa when compared with SOFaa (P< 0.001). DNA methyltransferase 3a (Dnmt3a) expression was higher in embryos cultured in CR1aa than in those cultured in SOFaa (P< 0.001). Expression of interferon tau (IF-τ) and insulin-like growth factor II receptor (Igf-2r) genes was significantly up-regulated in KSOMaa when compared with CR1aa (P< 0.001). Gene expression did not differ betweenin vivo-derived blastocysts and theirin vitro-derived counterparts. In conclusion, SOFaa supports higher development to blastocyst stage than KSOMaa and CR1aa, and the culture conditions influence gene expression.

55 STUDY ON THE INTERSPECIFIC NUCLEAR TRANSFER OF PRZEWALSKI'S GAZELLES AND BOVINES

2013 ◽  
Vol 25 (1) ◽  
pp. 175
Author(s):  
Y. Gao ◽  
L. Cheng ◽  
G. Su ◽  
Z. Wei ◽  
G. Li
Keyword(s):  
Embryo Development ◽  
Somatic Cells ◽  
Oocyte Nucleus ◽  
Control Group ◽  
Blastocyst Development ◽  
Affymetrix Microarray ◽  
Developmental Potential ◽  
Reconstructed Embryos ◽  
Deacetylase Inhibitor ◽  
Bovine Oocytes

Przewalski’s gazelle (Procapra przewalskii), also known as Platts antelope, is an endangered species only found in China. It belongs to the Artiodactyla order, Bovidae family, antelope subfamily, and Gazella genus. In this study, 5 experiments were designed to examine the developmental potential of Przewalski’s gazelle somatic cells transplanted into bovine enucleated oocytes. Enucleation was conducted by Hoechst 33342 staining of the oocytes and guided by a fluorescent microscope to ensure the removal of the nuclei. The gazelle cells were then transferred to the enucleated oocytes and electrically fused to reconstructed embryos. The study resulted in 5 major findings. (1) When gazelle-bovine reconstructed embryos were treated with the deacetylase inhibitor valproic acid (VPA), at different concentrations and for different times, treatment of the cloned embryos with VPA at 0.5 mM for 24 h significantly increased the 8- to 16-cell-stage embryo development [61.9% (96/155) v. 33.8% (46/136) control]. However, the morula [1.3% (2/155) v. 1.5% (2/155); P > 0.05] and blastocyst (0.7% v. 1.5%; P > 0.05) development were similar to that of the control. In the intraspecific (bovine-bovine) control group, the cleavage, morula and blastocyst development of 3 cloned embryos were 72.6% (127/175), 28.0% (49/175), and 23.4% (41/175). (2) Octamer-binding transcription factor 4 (Oct-4), as a developmental potential and expression marker, was transfected to gazelle cells. When Oct-4-eGFP-confected cells were transferred, the cloned embryo development did not improve either with or without VPA treatment. (3) When the gazelle-bovine embryos were treated with the deacetylase inhibitor trichostatin A (TSA) for 24 h at 10 ng mL–1, blastocyst development was significantly higher than in the control group [3.6% (6/168) v. 0.8% (1/125); P < 0.05]. (4) When a reverse NT protocol, in which the oocyte nucleus was removed after the cell nucleus was fused to the oocyte, was used for NT, the cloned embryo development did not improve. (5) The gazelle-bovine and bovine-bovine cloned embryos at 8- to 16-cell stages, gazelle cells, bovine cells, and bovine oocytes transcriptomes were analyzed by Affymetrix microarray (Affymetrix Microarray Inc., Santa Clara, CA, USA) and repeated twice. A total of 643 genes were activated in gazelle-cattle embryos compared with oocytes, whereas 1527 genes were activated in bovine-bovine clones. A total of 1010 genes that were exclusively expressed in gazelle somatic cells were still expressed in the interspecies cloned embryos. In conclusion, TSA treatment of Przewalski’s gazelle somatic cells transferred into enucleated bovine oocytes improved development of cloned embryos to the blastocyst stage, although still with low efficiency. Data from microarray analyses of the gazelle-cattle embryos showed that over 1000 gazelle-specific genes were still expressed in the interspecific cloned embryos. This work was supported by the National Basic Research Program of China (no. 2012CB22306).

152 POST-HATCHING EMBRYO DEVELOPMENT IN VITRO UNTIL DAY 15 IN AGAR GEL DILUTED IN PBS OR IN MilliQ WATER

2010 ◽  
Vol 22 (1) ◽  
pp. 234
Author(s):  
G. M. Machado ◽  
E. S. Caixeta ◽  
M. M. Franco ◽  
R. Rumpf ◽  
M. A. N. Dode
Keyword(s):  
Gene Expression ◽  
Embryo Development ◽  
Solute Carrier Family ◽  
Final Size ◽  
Agar Gel ◽  
Interferon Tau ◽  
Developmental Potential ◽  
Solute Carrier ◽  
Chi Square Analysis

Post-hatching (PH) in vitro embryo culture is a procedure that allows the establishment of more accurate tools for evaluating embryo developmental potential. An important step in the PH system is the tunnels preparation, since they will hold the developing embryo in advanced stages during culture. However, if the diluent used to dissolve agarose for tunnels preparation can influence the diffusion of nutrients from culture medium to the gel compromising embryo development, is not known. The aim of this study was to compare developmental kinetics and gene expression of Day 15 embryos cultured in the PHD system using PBS and MilliQ water to dilute the agar gel used for tunnels construction. Bovine oocytes obtained from slaughterhouse ovaries were matured, fertilized, and cultured in vitro for 8 days in SOFaaci with 5% fetal bovine serum (FBS) at 39°C in 5% CO2 in air. On Day 9, PHD (Brandão et al. 2004 Biol. Reprod. 71, 2048-2055) medium was added in culture droplets and on Day 11 embryos were measured. Only morphologically normal with ≥0.5 mm of diameter Day 11 blastocysts were placed individually in the tunnels. The tunnels were produced using agar gel 2.4% diluted either in PBS or MilliQ water, supplemented with 10% FBS. Morphology and length of embryos were evaluated on Day 11, Day 12.5, Day 14, and Day 15. Elongated embryos in Day 15 were used for extraction of total RNA using Trizol Reagent to verify gene expression by qPCR. A total of 4 pools containing 3 Day 15 embryos in each pool were used to evaluate the expression of genes involved in glucose metabolism [glucose-6-phosphate dehydrogenase (G6PDH), solute carrier family 2 member 1 (SLC2A1), solute carrier family 2 member 3 (SLC2A3), phosphoglycerate kinase 1 (PGK1)], placenta development [placenta-specific 8 (PLAC8), keratin proteins 8 (KRT8)], heat stress [heat shock transcription factors 1(HSF1)], and maternal recognition of pregnancy [interferon tau (IFNT)]. Embryo growth and gene expression data were examined by the t-test for parametric or Mann-Whitney for non-parametric (P < 0.05). From a total of 2,933 oocytes used, 1,216 (41%) reached the blastocyst stage on Day 8 and of those 72% hatched on Day 9 (n = 873). On Day 11, 39.5% (345/873) of the hatched blastocysts had diameter ≥0.5 mm and 286 were loaded into the MilliQ water (154) and PBS tunnels (132). No difference was observed in the proportion of elongated embryos between PBS (54%, n = 71) and MilliQ water treatment (42%, n = 65) by chi-square analysis. Final size at Day 15 as well as increase in length of the embryos between Days 11 and 15 were similar in PBS (2.37 ± 0.13 mm; 1.68 ± 0.13 mm) and MilliQ water tunnels (3.03 ± 0.23 mm; 2.33 ± 0.22 mm). Regarding gene expression analysis, only SLC2A1 had a tendency (P = 0.08) to be higher expressed in embryos cultured in PBS tunnels. The results suggested that water can be use to construct agar gel tunnels for the PHD system without interfering in embryo developmental kinetics until Day 15 of culture. Embrapa, CNPq, CAPES.

137 SPLITTING OF IVP BOVINE BLASTOCYST AFFECTS MORPHOLOGY AND GLOBAL GENE EXPRESSION OF RESULTING DEMI-EMBRYOS DURING ELONGATION

2015 ◽  
Vol 27 (1) ◽  
pp. 161
Author(s):  
A. E. Velásquez ◽  
D. Veraguas ◽  
J. F. Cox ◽  
F. O. Castro ◽  
L. l. Rodriguez
Keyword(s):  
Gene Expression ◽  
Embryo Development ◽  
Average Length ◽  
Global Gene Expression ◽  
Differentially Expressed ◽  
Control Group ◽  
Molecular Characteristics ◽  
Bovine Embryo ◽  
Developmental Potential

Embryo splitting has been used since the early 1980s to produce identical twins and increase the pregnancy rate per available embryo. However, very little is known about the effect of splitting on embryo development and competence. Indeed splitting could provoke a negative effect on embryo survival and it can be presumed that each demi-embryo might respond differently to the injury. In this sense, even when embryos are genetically and morphologically identical at the moment of splitting, their developmental potential and molecular characteristics might change as a consequence of the intense manipulation or epigenetic differences due to the interaction with the environment. We have proposed an approach to evaluate the effect of blastocyst splitting on the morphological and gene expression in in vivo development up to the filamentous stage. For that, the effect of splitting on bovine embryo development was evaluated during the elongation period by transferring split and nonsplit IVF-derived blastocysts to cattle recipients and collecting them at Day 17 of development. The number of collected embryos, embryo size, and global gene expression was compared between both groups. Collected elongated embryos derived from split blastocyst were compared with time matched collected control embryos. From 14 transferred hemi-embryos, 5 (35.7%) were collected while 9 elongated from 17 controls were recovered (52.9%). Neither the recovery rate nor the average length of the elongated embryos was significantly different between the two treatments. However, when embryos were rated depending on their size, more than 50% of embryos from the control group had a length surpassing 100 mm, while only 33% of the split embryos reached that size. Global gene expression was performed using 2-colour microarray-based gene expression analysis. This was a whole-genome microarray study comparing 10 individual elongated embryos derived from split and nonsplit IVF blastocysts. Genes were considered differentially expressed if the fold change is greater than 2 (up or down-regulation) with P ≤ 0.05. A total of 29 585 transcripts were detected in all embryos. From those, 449 (1.5%) were differentially expressed between elongated embryos derived from split and nonsplit IVF blastocysts, among them, 248 (0.83%) genes were down-regulated and 201 (0.67%) genes were up-regulated in split embryos. Gene ontology analysis identified deregulated genes related with intrinsic component of membrane (ELOVL7, GJA1, LAPTM4B, LDLR, SLC18A2, SLC1A3, SLC38A5, TSPAN13), lipid transporter activity (RBP4, APOA1, MTTP), and organophosphate ester transport (GJA1, GJB1, ATP9B). In conclusion, we showed that splitting affect the in vivo developmental capability and gene expression profile during the elongation period of bovine embryos. However, further studies are needed to determine the long-term effect of this technique to produce viable offspring. This work was partially supported by Fondecyt No. 11100082 and Fondequip No. EQM12113 from the Ministry of Education of Chile.

94 EFFICIENT BOVINE EMBRYO SPLITTING FOR GENE EXPRESSION AND IN VIVO DEVELOPMENT STUDIES

2014 ◽  
Vol 26 (1) ◽  
pp. 161
Author(s):  
A. Velasquez ◽  
D. Veraguas ◽  
F. O. Castro ◽  
J. F. Cox ◽  
L. l. Rodriguez-Alvarez
Keyword(s):  
Gene Expression ◽  
Expression Pattern ◽  
Gene Expression Pattern ◽  
Blastocyst Stage ◽  
Embryo Morphology ◽  
Bovine Embryo ◽  
Gene Markers ◽  
Developmental Potential

It is known that embryos produced in vitro are less competent than their in vivo-derived counterparts. When embryos are produced or manipulated in vitro, their developmental potential decreases significantly, which impinges upon the production of viable offspring. In bovines, embryos that will be transferred to a surrogate mother are selected at the blastocysts stage using noninvasive methods, such as their morphological features. However, many of those embryos are not able to implant or to maintain a normal pregnancy because embryo morphology does not reflect its developmental potential and a correct gene expression pattern that support a normal development. It seems that the ideal method for embryo selection would be based on the screening of gene markers that correlate with successful pregnancy after embryo transfer. In that sense, we have proposed an approach to characterise gene expression pattern of early (Day 7) bovine blastocysts and to correlate this gene expression with further developmental potential in vivo, i.e. upon elongation until Day 17. For that, it was established an efficient method to produce identical and viable hemi-embryos by splitting IVF bovine blastocysts in order to set the expression profile of certain genes in one hemi-embryo at blastocyst stage, while the counterpart embryo elongates in vivo for 10 days. A total of 129 blastocysts were split. Six groups of blastocysts were used for splitting and the results compared: 1) Day-7 early blastocysts (n = 20); 2) Day-7 expanded blastocysts (n = 25); 3) Day-7 hatched blastocysts (n = 17); 4) Day-8 early blastocysts (n = 10); 5) Day-8 expanded blastocysts (n = 12); and 6) Day-8 hatched blastocysts (n = 45). Hemi-embryos derived from day-8 grade I and well expanded blastocysts had the greatest survival rate, in vitro re-expansion (67.7%; P < 0.05) and both hemi-embryos conserved a normal morphology with a total cell number over 80 after 6 h in culture. Also both hemi-embryos at blastocyst stage showed homogeneous expression pattern of the genes OCT4, SOX2, NANOG, CDX2, ACTB, and GAPDH (P < 0.05). Finally, the in vivo survival of hemi-embryos was assessed and compared with nonsplit embryos (control) by transferring to recipient cow and collecting at Day 17 of development. For this, hemi-embryos derived from Day-8 hatched blastocyst were used. From 14 transferred hemi-embryos, 5 (35.7%) were collected, and 9 elongated from 17 controls were recovered (52.9%). Also the elongation rate was significantly lower in hemi-embryos than in control; the length of hemi-embryos had a range between 1 and 5 cm, whereas 60% of the control embryos were longer than 10 cm. Our results provide an initial approach to study the correlation among the gene expression characteristics of early bovine embryos with their further development. However, it seems that embryo splitting hampers their elongation in vivo. It might be possible that the development of split embryos is retarded because of manipulation. This work was partially supported by Fondecyt grant no. 11100082 from the Ministry of Education of Chile.

Ubiquitin proteasome pathway gene expression varies in rhesus monkey oocytes and embryos of different developmental potential

2007 ◽  
Vol 31 (1) ◽  
pp. 1-14 ◽  
Author(s):  
Namdori R. Mtango ◽  
Keith E. Latham
Keyword(s):  
Gene Expression ◽  
Rhesus Monkey ◽  
Early Embryo ◽  
Developmental Competence ◽  
Cell Physiology ◽  
Developmental Potential ◽  
Ubiquitin Proteasome Pathway ◽  
Pathway Gene ◽  
Ubiquitin Proteasome ◽  
Proteasome Pathway

Protein degradation via the ubiquitin-proteasome pathway (UPP) plays a key role in diverse aspects of cell physiology and development. In the early embryo, the UPP may play an important role in the transition from maternal to embryonic control of development. Disruptions in the UPP could thus compromise embryo developmental potential. Additionally, species-specific requirements may dictate diverse patterns of regulation of the UPP components. To investigate the expression of UPP components in a nonhuman primate embryo model, to compare expression between a primate and nonprimate species, and to determine whether disruption of this pathway may contribute to reduced developmental potential, we examined the expression of >50 mRNAs encoding UPP components in rhesus monkey oocytes and embryos. We compared this expression between the rhesus monkey and mouse embryo and between rhesus monkey oocytes and embryos of high, intermediate, and low developmental potential. We report here the temporal patterns of UPP gene expression in oocytes and during preimplantation development, including striking differences between the rhesus monkey and mouse. We also report significant differences in UPP gene expression correlating with oocyte and embryo developmental competence and associated with altered regulation of maternally inherited mRNAs encoding these proteins.

scholarly journals Insulin exposure during in vitro bovine oocyte maturation changes blastocyst gene expression and developmental potential

2017 ◽  
Vol 29 (5) ◽  
pp. 876 ◽  
Author(s):  
Denise Laskowski ◽  
Ylva Sjunnesson ◽  
Patrice Humblot ◽  
Marc-André Sirard ◽  
Göran Andersson ◽  
...  
Keyword(s):  
Gene Expression ◽  
Oocyte Maturation ◽  
Quantitative Polymerase Chain Reaction ◽  
Blastocyst Stage ◽  
Specific Gene ◽  
Bovine Oocyte ◽  
Developmental Potential ◽  
Metabolic Imbalance ◽  
Negative Effect

Metabolic imbalance impairs fertility, because changes in concentrations of metabolites and hormones in the blood and follicular fluid create an unfavourable environment for early embryonic development. Insulin is a key metabolic hormone known for its effects on fertility: insulin concentrations are increased during energy balance disturbances in diabetes or metabolic syndrome. Still, insulin is frequently used at supraphysiological concentrations for embryo in vitro culture with unknown consequences for the developmental potential of the offspring. In the present study we investigated the effects of insulin exposure during in vitro bovine oocyte maturation on developmental rates, embryo quality and gene expression. Supplementation of the maturation media with insulin at 10 or 0.1 µg mL–1 decreased blastocyst rates compared with an insulin-free control (19.8 ± 1.3% and 20.4 ± 1.3% vs 23.8 ± 1.3%, respectively; P < 0.05) and led to increased cell numbers (nearly 10% more cells on Day 8 compared with control; P < 0.05). Transcriptome analysis revealed significant upregulation of genes involved in lipid metabolism, nuclear factor (erythroid-derived 2)-like 2 (NRF2) stress response and cell differentiation, validated by quantitative polymerase chain reaction. To conclude, the results of the present study demonstrate that insulin exposure during in vitro oocyte maturation has a lasting effect on the embryo until the blastocyst stage, with a potential negative effect in the form of specific gene expression perturbations.

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