Gene expression profile differences in embryos derived from prepubertal and adult Japanese Black cattle during in vitro development

The present study was carried out to compare the gene expression profiles of in vitro-generated embryos derived from adult and prepubertal Japanese Black cattle oocytes using GeneChip Bovine Genome Array (containing 24 072 probe sets representing over 23 000 transcripts). Microarray experiments were performed on populations of 8- to 16-cell stage embryos and blastocysts derived from adult (24–35 months old) versus prepubertal (9–10 months old) Japanese Black cattle oocytes matured and fertilised in vitro. In total, 591 (2.4%) and 490 (2.0%) genes were differentially expressed in prepubertal and adult bovine in 8- to 16-cell and blastocyst stage embryos, respectively. Out of these, 218 and 248 genes were upregulated, while 373 and 242 were downregulated in prepubertal and adult 8- to 16-cell and blastocysts stage embryos, respectively. Gene ontology classification regarding biological process, molecular functions and cellular component revealed diversity in transcript abundances between prepubertal and adult groups in both the distinct developmental stages. Quantitative reverse transcription–PCR validated the expression differences of some selected transcripts as identified by microarray analysis. To our knowledge, this is the first report indicating the significant number of genes differentially expression (>2-fold, P < 0.01) in preimplantition embryos between adult and prepubertal Japanese Black cattle during in vitro development.

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Gene expression profiles and in vitro development following vitrification of pronuclear and 8-cell stage mouse embryos

Molecular Reproduction and Development ◽

10.1002/mrd.20450 ◽

2006 ◽

Vol 73 (6) ◽

pp. 700-708 ◽

Cited By ~ 45

Author(s):

Duangjai Boonkusol ◽

Arpad Baji Gal ◽

Szilard Bodo ◽

Botond Gorhony ◽

Yindee Kitiyanant ◽

...

Keyword(s):

Gene Expression ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Mouse Embryos ◽

Cell Stage ◽

In Vitro Development ◽

Vitro Development

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80 GENE EXPRESSION PROFILES AND IN VITRO DEVELOPMENT FOLLOWING VITRIFICATION OF PRONUCLEAR AND 8 CELL-STAGE MOUSE EMBRYOS

Reproduction Fertility and Development ◽

10.1071/rdv17n2ab80 ◽

2005 ◽

Vol 17 (2) ◽

pp. 190

Author(s):

D. Boonkusol ◽

A. Baji Gal ◽

S.Z. Bodo ◽

B. Gorhony ◽

K. Pavasuthipaisit ◽

...

Keyword(s):

Gene Expression ◽

Real Time ◽

Expression Profiles ◽

Survival Rates ◽

Mouse Embryos ◽

Rt Pcr ◽

Cell Stage ◽

In Vitro Development ◽

Vitro Development

The analysis of differences in gene expression of embryos in response to cryopreservation may explain some of the observed differences in further development. Experiments were conducted to study effects of two vitrification methods, solid surface (SSV) and in-straw vitrification (STR), for pronuclear and 8 cell-stage mouse embryos with regard to gene expression and in vitro development. Both stages of embryos were vitrified by SSV (Dinnyes et al. 2000 Biol. Reprod. 63, 513–518) using 35% ethylene glycol (EG) for vitrification solution (VS) and STR using 40% EG for VS. Warming and rehydration of embryos in the SSV method was performed by placing vitrified droplets into a 37°C 0.3 M trehalose solution for 1 min, and then for 2 min in each of 0.15 M trehalose, 0.075 M trehalose, and base medium at room temperature before culture in CZB medium. Warming of embryos for the in-straw method was performed by holding straws in air for 10 s and then plunging them into 37°C water for 15–20 s. The contents of the straws were expelled into 37°C 0.3 M trehalose solution and embryos were treated as above. No significant differences were found between immediate survival rates of embryos vitrified by SSV and STR in both stages. Blastocyst rates were significantly higher with SSV than with STR and not significantly different from those of the control (Table 1). These results show that SSV was more efficient than STR. Quantification of selected gene transcripts from single embryo (6 embryos/treatment group) was carried out by quantitative real-time RT-PCR. Genes related to oxidative stress (MnSOD and CuSOD), cold stress (CIRP, RBM3, and p53), and β-actin as reference gene were amplified. We found up-regulation of all stress genes at 3 h post-warming in all treatment groups. At 10 h post-warming, a low level of gene expression was found in SSV-treated embryos, but gene expression remained at high level in STR-treated embryos. However, no differences in gene expression were found between blastocysts developed from fresh and vitrified embryos. In conclusion, the real-time RT-PCR method from single embryos opened new opportunities for understanding molecular events following cryopreservation. The continuous up-regulation of stress-related genes at 10 h post-warming might have been an early indicator of reduced viability following STR, as was also indicated by the developmental rate to the blastocyst stage. Table 1. Survival and blastocyst rates of vitrified/warmed pronuclear and 8 cell-stage mouse embryos This work was supported by the Thailand Research Fund (Royal Golden Jubilee Ph.D. scholarship) and a Bio-00017/2002 grant of the Hungarian National Office of Research and Technology.

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171 THE EFFECT OF TRICHOSTATIN A ON THE DEVELOPMENT AND THE GLOBAL GENE EXPRESSION PATTERNS IN MOUSE EMBRYOS DEVELOPING IN VITRO

Reproduction Fertility and Development ◽

10.1071/rdv20n1ab171 ◽

2008 ◽

Vol 20 (1) ◽

pp. 165

Author(s):

X. S. Cui ◽

X. Y. Li ◽

T. Kim ◽

N.-H. Kim

Keyword(s):

Gene Expression ◽

Trichostatin A ◽

Expression Profiles ◽

Expression Patterns ◽

Gene Expression Profiles ◽

Mouse Embryos ◽

Differentially Expressed ◽

Gene Expression Patterns ◽

Cell Stage

Trichostatin A (TSA) is an inhibitor of histone deacetylase and is able to alter gene expression patterns by interfering with the removal of acetyl groups from histones. The aim of this study was to determine the effect of TSA treatment on the development and gene expression patterns of mouse zygotes developing in vitro. The addition of 100 nm TSA to the culture medium did not affect the cleavage of mouse embryos (TSA treatment, 148/150 (99%) v. control, 107/107 (100%)); however, embryos that were treated with TSA arrested at the 2-cell stage (145/148, 98%). We estimated the number of nuclei in control and TSA-treated embryos by propidium iodide staining, taking into account the presence of any cells with two or more nuclei. At 62–63 h post-hCG stimulation, control zygotes had developed to the 4-cell stage and exhibited one nucleus in each blastomere, indicative of normal development. In contrast, we observed tetraploid nuclei in at least one blastomere in 20.8% (11/53) of the embryos that had been treated with TSA. At 28–29 h post-hCG stimulation (metaphase of the 1-cell stage), there was no difference in the mitotic index (as determined by analyzing the microtubule configuration) in the TSA group compared to the control group. At the 2-cell stage, however, we did not observe mitotic spindles and metaphase chromatin in embryos in the TSA treatment group compared to the controls. Interestingly, when embryos were cultured in TSA-free medium from 35 h post-hCG stimulation (S- or early G2-phase of the 2-cell stage) onward, almost all of them (47/50) developed to the blastocyst stage. In contrast, when embryos were cultured in TSA-free medium from 42 h post-hCG stimulation (middle G2-phase of the 2-cell stage) onward, they did not develop to the 4-cell stage. We used Illumina microarray technology to analyze the gene expression profiles in control and TSA-treated late 2-cell-stage embryos. Applied Biosystems Expression System software was used to extract assay signals and assay signal-to-noise ratio values from the microarray images. Our data showed that 897 genes were significantly (P < 0.05; 2-sample t-test) up- or down-regulated by TSA treatment compared to controls. Analysis using the PANTHER classification system (https://panther.appliedbiosystems.com) revealed that the 575 genes that were differentially expressed in the TSA group compared to the control were classified as being associated with putative biological processes or molecular function. Overall, in terms of putative biological processes, more nucleoside, nucleotide, and nucleic acid metabolism, protein metabolism and modification, signal transduction, developmental process, and cell cycle genes were differentially expressed between the TSA and control groups. In terms of putative molecular function, more nucleic acid-binding transcription factor and transferase genes were differentially expressed between the groups. The results collectively suggest that inhibition of histone acetylation in mouse embryos affects gene expression profiles at the time of zygotic genome activation, and this subsequently affects further development.

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198 THE EFFECT OF SOURCE AND IN VITRO MATURATION ON THE ABUNDANCE OF MATERNAL mRNA OF SELECTED GENES IN FOLLICULAR BOVINE OOCYTES AND THEIR INFLUENCE ON IN VITRO DEVELOPMENT

Reproduction Fertility and Development ◽

10.1071/rdv23n1ab198 ◽

2011 ◽

Vol 23 (1) ◽

pp. 199

Author(s):

T. Somfai ◽

K. Imai ◽

M. Kaneda ◽

S. Akagi ◽

S. Haraguchi ◽

...

Keyword(s):

Gene Expression ◽

Embryo Development ◽

In Vitro Maturation ◽

Blastocyst Stage ◽

Cell Stage ◽

In Vitro Development ◽

Before And After ◽

Vitro Development ◽

Bovine Oocytes

The aim of the present study was to investigate the effect of oocyte source and in vitro maturation (IVM) on the expression of selected genes in bovine oocytes and their contribution to in vitro embryo development. Follicular oocytes were collected either by ovum pick-up from live cows or by the aspiration of ovaries of slaughtered cows following storage in Dulbecco’s PBS at 15°C for overnight. In vitro maturation was performed according to the method of (Imai et al. 2006 J. Reprod. Dev. 52, 19–29 suppl.). Gene expression was assessed before and after IVM by real-time PCR. The following genes were investigated: GAPDH, G6PDH, ACTB, H2A, CCNB1, MnSOD, OCT4, SOX2, CX43, HSP70, GLUT8, PAP, GDF9, COX1, ATP1A1, CDH1, CTNNB1, AQP3, DYNLL1, DYNC 1/1, and PMSB1. In brief, mRNA was extracted from 20 oocytes per sample using a Qiagen RNeasy Micro Kit (Qiagen, Valencia, CA). Gene expression was analysed by a Roche Light Cycler 480 device and software (Roche, Indianapolis, IN). Relative expression of each gene was normalized to CCNB1, which in preliminary experiments appeared the most stably expressed irrespective of oocyte source and meiotic stage. Three replications were performed. Data were analysed by paired t-test. In immature ovum pick-up oocytes, genes related to metabolism (GAPDH, G6PDH, GLUT8) and stress (MnSOD, HSP70), and also OCT4, ATP1A1, and DYNC1/1 showed significantly (P < 0.05) higher expression compared with immature oocytes collected from slaughtered-stored ovaries. The expression of GDF9, GLUT8, CTNNB1, and PMSB1 was significantly (P < 0.05) reduced during IVM irrespective of the oocyte source. In a second experiment, IVF IVM oocytes showing an early (at 22 to 25 h after IVF) or late (at 27 to 30 h after IVF) first cleavage were either cultured in vitro or analysed for gene expression at the 2-cell stage. A higher (P < 0.05) rate of early-cleaving oocytes developed to the blastocyst stage compared with the rate of late-cleaving ones (46.2% v. 15.6%, respectively). Nevertheless, only ATP1A1 showed significantly reduced (P < 0.05) expression in late-cleaving embryos compared with early-cleaving ones. Our results suggest that although removal and storage of ovaries and IVM caused a reduction in the relative abundance of several genes in oocytes, in most cases, this did not affect embryo development. Among the genes studied, only ATP1A1 was correlated with in vitro development.

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Genome-Wide Identification of Soybean ABC Transporters Relate to Aluminum Toxicity

International Journal of Molecular Sciences ◽

10.3390/ijms22126556 ◽

2021 ◽

Vol 22 (12) ◽

pp. 6556

Author(s):

Junjun Huang ◽

Xiaoyu Li ◽

Xin Chen ◽

Yaru Guo ◽

Weihong Liang ◽

...

Keyword(s):

Gene Expression ◽

Abc Transporters ◽

Developmental Stages ◽

Aluminum Toxicity ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Purifying Selection ◽

Biotic Stresses ◽

Genome Wide ◽

New Germplasm

ATP-binding cassette (ABC) transporter proteins are a gene super-family in plants and play vital roles in growth, development, and response to abiotic and biotic stresses. The ABC transporters have been identified in crop plants such as rice and buckwheat, but little is known about them in soybean. Soybean is an important oil crop and is one of the five major crops in the world. In this study, 255 ABC genes that putatively encode ABC transporters were identified from soybean through bioinformatics and then categorized into eight subfamilies, including 7 ABCAs, 52 ABCBs, 48 ABCCs, 5 ABCDs, 1 ABCEs, 10 ABCFs, 111 ABCGs, and 21 ABCIs. Their phylogenetic relationships, gene structure, and gene expression profiles were characterized. Segmental duplication was the main reason for the expansion of the GmABC genes. Ka/Ks analysis suggested that intense purifying selection was accompanied by the evolution of GmABC genes. The genome-wide collinearity of soybean with other species showed that GmABCs were relatively conserved and that collinear ABCs between species may have originated from the same ancestor. Gene expression analysis of GmABCs revealed the distinct expression pattern in different tissues and diverse developmental stages. The candidate genes GmABCB23, GmABCB25, GmABCB48, GmABCB52, GmABCI1, GmABCI5, and GmABCI13 were responsive to Al toxicity. This work on the GmABC gene family provides useful information for future studies on ABC transporters in soybean and potential targets for the cultivation of new germplasm resources of aluminum-tolerant soybean.

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Transcriptome analysis of gravitational effects on mouse skeletal muscles under microgravity and artificial 1 g onboard environment

Scientific Reports ◽

10.1038/s41598-021-88392-4 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Risa Okada ◽

Shin-ichiro Fujita ◽

Riku Suzuki ◽

Takuto Hayashi ◽

Hirona Tsubouchi ◽

...

Keyword(s):

Gene Expression ◽

Muscle Mass ◽

Muscle Atrophy ◽

Transcriptome Analysis ◽

Fiber Type ◽

Expression Profiles ◽

Space Station ◽

Gene Expression Profiles

AbstractSpaceflight causes a decrease in skeletal muscle mass and strength. We set two murine experimental groups in orbit for 35 days aboard the International Space Station, under artificial earth-gravity (artificial 1 g; AG) and microgravity (μg; MG), to investigate whether artificial 1 g exposure prevents muscle atrophy at the molecular level. Our main findings indicated that AG onboard environment prevented changes under microgravity in soleus muscle not only in muscle mass and fiber type composition but also in the alteration of gene expression profiles. In particular, transcriptome analysis suggested that AG condition could prevent the alterations of some atrophy-related genes. We further screened novel candidate genes to reveal the muscle atrophy mechanism from these gene expression profiles. We suggest the potential role of Cacng1 in the atrophy of myotubes using in vitro and in vivo gene transductions. This critical project may accelerate the elucidation of muscle atrophy mechanisms.

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Simulated microgravity using the Random Positioning Machine inhibits differentiation and alters gene expression profiles of 2T3 preosteoblasts

AJP Cell Physiology ◽

10.1152/ajpcell.00222.2004 ◽

2005 ◽

Vol 288 (6) ◽

pp. C1211-C1221 ◽

Cited By ~ 94

Author(s):

Steven J. Pardo ◽

Mamta J. Patel ◽

Michelle C. Sykes ◽

Manu O. Platt ◽

Nolan L. Boyd ◽

...

Keyword(s):

Gene Expression ◽

Alkaline Phosphatase ◽

Bone Loss ◽

Simulated Microgravity ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Underlying Mechanism ◽

Bone Biology ◽

Random Positioning Machine

Exposure to microgravity causes bone loss in humans, and the underlying mechanism is thought to be at least partially due to a decrease in bone formation by osteoblasts. In the present study, we examined the hypothesis that microgravity changes osteoblast gene expression profiles, resulting in bone loss. For this study, we developed an in vitro system that simulates microgravity using the Random Positioning Machine (RPM) to study the effects of microgravity on 2T3 preosteoblast cells grown in gas-permeable culture disks. Exposure of 2T3 cells to simulated microgravity using the RPM for up to 9 days significantly inhibited alkaline phosphatase activity, recapitulating a bone loss response that occurs in real microgravity conditions without altering cell proliferation and shape. Next, we performed DNA microarray analysis to determine the gene expression profile of 2T3 cells exposed to 3 days of simulated microgravity. Among 10,000 genes examined using the microarray, 88 were downregulated and 52 were upregulated significantly more than twofold using simulated microgravity compared with the static 1-g condition. We then verified the microarray data for some of the genes relevant in bone biology using real-time PCR assays and immunoblotting. We confirmed that microgravity downregulated levels of alkaline phosphatase, runt-related transcription factor 2, osteomodulin, and parathyroid hormone receptor 1 mRNA; upregulated cathepsin K mRNA; and did not significantly affect bone morphogenic protein 4 and cystatin C protein levels. The identification of gravisensitive genes provides useful insight that may lead to further hypotheses regarding their roles in not only microgravity-induced bone loss but also the general patient population with similar pathological conditions, such as osteoporosis.

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