40 Gene Expression Profiling of In Vitro-Produced Blastocysts Derived from In Vitro-Matured Bovine Oocytes Vitrified/Warmed in Media Supplemented with a Biopolymer Produced by an Antarctic Bacterium

2018 ◽  
Vol 30 (1) ◽  
pp. 159 ◽  
Author(s):  
N. Arcarons ◽  
M. Vendrell ◽  
M. Yeste ◽  
M. E. Mercadé ◽  
M. López-Béjar ◽  
...  
Keyword(s):  
Gene Expression ◽  
Expression Patterns ◽  
Rna Extraction ◽  
Mrna Abundance ◽  
The Other ◽  
Developmental Competence ◽  
Crystal Formation ◽  
Control Analysis ◽  
The Impact

Previous reports have demonstrated the beneficial effect of antifreeze glycoprotein supplementation during oocyte vitrification on preventing ice crystal formation and thus enhancing developmental competence after vitrification-warming. Pseudomonas sp. ID1, a bacterium isolated from marine sediment from Antarctica, produces an exopolysaccharide, M1 EPS, as a cold adaptation mechanism. Despite numerous studies on structural and morphological damages induced by cryopreservation in oocytes, few studies have focused on the impact of vitrification on the expression pattern of genes during early embryo development. In the present study, the expression patterns of 6 genes (BAX, BCL2-like 1, DNMT3A, UBE2A, SCLC2A3, and HDAC1) were investigated in Day 8 blastocysts resulting from in vitro-matured oocytes vitrified/warmed in media supplemented with various concentrations of M1 EPS. After 21 h of IVM, 1,062 oocytes were vitrified/warmed in media supplemented with 0, 0.001, 0.01, 0.1, and 1 mg mL−1 M1 EPS. At 24 h of IVM, oocytes were in vitro fertilized and in vitro cultured and the resulting blastocysts were harvested at Day 8 for RNA extraction and qPCR analysis. Fresh, non-vitrified oocytes were used as a control. Analysis of gene expression was performed through Kruskall-Wallis test and followed by Mann-Whitney test, and the level of significance was set at P ≤ 0.05. No significant differences were detected in relative mRNA abundance for SLC2A3, UBE2A, or HDAC-1 between blastocysts derived from vitrified oocytes, regardless of M1 EPS treatment. Expression of DNMT3A was significantly higher in embryos obtained from oocytes vitrified and warmed with 0.1 mg mL−1 M1 EPS compared with other treatment groups. However, no differences in DNMT3A expression were observed when the other vitrified groups were compared. The relative abundance of BAX transcript in embryos from oocytes vitrified in media supplemented with 0.1 mg mL−1 M1 EPS was higher than that in 0 or 0.001 mg mL−1 groups. Embryos from 0.01 and 0.1 mg mL−1 groups showed higher BCL2-like 1 mRNA abundance than those from the 0, 0.001, and 1 mg mL−1 groups. Whereas blastocysts from oocytes vitrified with 0.01 mg mL−1 M1 EPS exhibited the lowest BAX:BCL2-like 1 ratio, no significant differences in BAX:BCL2-like 1 ratio were observed between the other treatments. The significantly lower BAX:BCL2 ratio observed in blastocysts obtained from oocytes vitrified with 0.01 mg mL−1 M1 EPS could be indicative for a better embryo quality. Although optimizing the use of M1 EPS may benefit oocyte cryopreservation protocols, further research is required to clarify the exact mechanism through which it exerts its protective role. This study was supported by the Spanish Ministry of Science and Innovation (Project AGL2016-79802-P and grant CTQ2014-59632-R).

282 ADIPOGENIC DIFFERENTIATION IN VITRO OF PORCINE ADULT MESENCHYMAL STEM CELLS

2009 ◽  
Vol 21 (1) ◽  
pp. 238 ◽  
Author(s):  
E. Monaco ◽  
A. Lima ◽  
S. Wilson ◽  
S. Lane ◽  
M. Bionaz ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Bone Marrow ◽  
Adipose Tissue ◽  
Subcutaneous Adipose Tissue ◽  
Adipogenic Differentiation ◽  
Expression Patterns ◽  
Mrna Abundance ◽  
Subcutaneous Adipose

The quantity and accessibility of subcutaneous adipose tissue in humans make it an attractive alternative to bone marrow as a source of adult stem cells for therapeutic purposes. However, before such a cell source substitution can be proposed, the properties of stem cells derived from adipose tissue (ADSC) and bone marrow (BMSC), and their differentiated progeny must be compared in an animal model, such as swine, that adequately simulates the structure and physiology of humans. The objective of this work was to induce adult porcine stem cells isolated from subcutaneous adipose tissue and bone marrow to differentiate in vitro along the adipogenic lineage and to compare their transcript profile properties. ADSC and BMSC were isolated from subcutaneous adipose tissue and femurs of adult pigs, respectively, and differentiated along the adipogenic lineage using specific inducing medium. Cells were incubated up to 4 weeks with medium replaced every 3 days. Histological staining with Oil Red O was performed at 0, 2, 4, 7, 14, 21, 28 days of differentiation (dd) to confirm the adipogenic differentiation. RNA was also extracted at these time points. qPCR was performed on PPARG, DBI, ACSL1, CD36, CEBPA, DGAT2, ADFP, ADIPOQ, SCD. The geometrical mean of GTF2H3, NUBP, and PPP2CB was used as an internal control. Gene expression was analyzed using a mixed model of SAS with repeated time. The adipogenic differentiation of both ADSC and BMSC was confirmed by the Oil Red O positive staining. The relative mRNA abundance of all the genes at dd0 was similar between the ADSC and BMSC. The relative mRNA abundance of most of the genes was also similar between ADSC and BMSC throughout the adipogenic differentiation. ACSL1 and ADIPOQ had analogous expression patterns among the cell types. ACSL1 had relatively large mRNA abundance before differentiation, but ADIPOQ was barely detectable. As a consequence of differentiation, ACSL1 increased in relative mRNA abundance about 10-fold, whereas ADIPOQ mRNA increased about 1000-fold. Temporal expression patterns of SCD, DGAT2, and ADFP were similar. The increase in gene expression was >800% for SCD, >500% for ADFP, and >50 000% for DGAT2 after 7dd. ADSC had significantly higher expression of those genes compared to BMSC at 14 and 28dd. Both ADIPOQ and DGAT2 were almost undetectable prior to differentiation. mRNA expression of CD36 and DBI was similar with a significantly larger increase in expression of ADSC compared with BMSC. Relative mRNA abundance of CEBPA and PPARG was also larger in ADSC compared with BMSC; however, BMSC had a remarkable increase in temporal expression of those genes throughout adipogenic differentiation. These results suggest both cell types can differentiate towards the adipogenic lineage but with quantitatively different gene expression patterns. More investigation is needed before the ADSC can be considered a practical alternative source for stem cells in future human clinical applications. This research was supported by the Illinois Regenerative Medicine Institute.

99 EXTRACELLULAR VESICLES OF BOVINE OVIDUCTAL FLUID MODIFY THE GENE EXPRESSION ON BOVINE IN VITRO-DERIVED EMBRYOS

2016 ◽  
Vol 28 (2) ◽  
pp. 179 ◽  
Author(s):  
R. Lopera-Vasquez ◽  
M. Hamdi ◽  
V. Maillo ◽  
C. Nunez ◽  
M. Yanez-Mo ◽  
...  
Keyword(s):  
Gene Expression ◽  
Embryo Development ◽  
Extracellular Vesicles ◽  
Membrane Trafficking ◽  
Dna Methyltransferase ◽  
De Novo ◽  
Expression Patterns ◽  
Water Channel ◽  
Developmental Competence

Extracellular vesicles (EVs) act as intercellular communicators through their protein, lipid, and mRNA content. The interaction of EVs from oviducal environment and the first stages of embryo development is currently an enigma. The aim of the present study was to evaluate the developmental competence and the expression profile of bovine blastocysts cultured with previously purified EVs recovered from ampullary and isthmic oviducal fluid (OF) under different centrifugal forces. OF-EVs recovered from oviducts of slaughtered heifers in early luteal phase were quantified with a nanoparticle tracking analysis system, and their integrity and size were assessed by electron microscopy. In vitro-produced zygotes were cultured in SOF+3 mg mL–1 BSA (C–), C– with 3 × 105 OF-EVs/mL from the ampulla (A) and isthmus (I) isolated at 1 × 103 (A10k and I10k, respectively) and 1 × 105 (A100k and I100k, respectively) × g. A control culture group of SOF+5% FCS (C+) was included. Blastocyst development was recorded on Day 7, 8, and 9 (D0: day of fertilization). Blastocysts on Days 7/8 cultured in C–, C+, I10k, and I100k were used to measure the relative mRNA expression of genes related with membrane trafficking (AQP3, AQP11, and ATP1A1), metabolism (LDLR and LDHA), and epigenetics (DNMT3A, IGF2R, GRB10, and SNRPN) by RT-qPCR. One-way ANOVA was used for statistical analysis. The size of ampullary and isthmic OF-EVs was similar with a mean of 220 nm. The concentration of I10k was significantly lower compared with A100k (3.6 × 108 v. 10.5 × 108 EVs/mL, respectively; P < 0.05); however, no differences were found in the rest of the groups with a mean concentration of 7.6 × 108 EVs/mL. EVs and C– groups showed a delayed embryo development at Day 7 compared with C+ (range: 12.0–13.8 v. 20.6%, respectively, P < 0.05); however, it was compensated at Days 8 and 9 (Day 9 range: 28.5–30.8%). The water channel related protein AQP3, associated with blastocoel formation, water, and cryoprotectant movement during cryopreservation, was up-regulated in I10k and I100k blastocysts compared with C+. The lipid receptor LDLR, proposed as a regulator of lipid uptake in blastocysts, was significantly down-regulated in C+ compared with the other groups, a possible consequence of a higher concentration of lipids in the C+ group. The de novo DNA methyltransferase DNMT3A and the imprinting gene SNRPN were down-regulated in the C+ compared with I100k, suggesting alterations in imprinting. In conclusion, bovine isthmic OF-EVs supplementation in in vitro embryo culture has a positive effect on gene expression patterns of developmental related genes compared with serum supplementation, suggesting an association between the oviducal environment and the developing embryo. Funded by the Spanish Ministry of Science and Innovation (AGL2012–37510 and AGL2012–39652-C02–01).

scholarly journals Papillary and Reticular Fibroblasts Generate Distinct Microenvironments that Differentially Impact Angiogenesis

2020 ◽  
Author(s):  
Adèle Mauroux ◽  
Pauline Joncour ◽  
Benjamin Gillet ◽  
Sandrine Hughes ◽  
Corinne Ardidie-Robouant ◽  
...  
Keyword(s):  
Gene Expression ◽  
Skin Fibroblast ◽  
Expression Patterns ◽  
Bioinformatic Analysis ◽  
Vessel Diameter ◽  
Branch Points ◽  
Reticular Dermis ◽  
Angiogenic Potential ◽  
The Impact

ABSTRACTPapillary and reticular dermis show distinct extracellular matrix (ECM) and vascularization, and fibroblasts isolated from these compartments have different gene expression patterns and behaviour in vitro. However, due to lack of relevant models, the contribution of skin fibroblast sub-populations to vascularization remains unknown. We thus cultured human papillary and reticular fibroblasts as cell sheets. Differential transcriptomic analysis was performed by RNA sequencing to characterize their microenvironment. Bioinformatic analysis revealed that each fibroblast population expressed specific angiogenesis and matrisome gene expression signatures resulting in specific ECM that differed both in composition and structure. The impact of secreted and ECM-bound factors was then assessed using 3D angiogenesis assays. When co-cultivated with endothelial cells, the papillary and reticular microenvironments induced the formation of distinct capillary networks mimicking the characteristics of vasculature of native dermis subcompartments (vessel diameter and density, number of branch points). Whereas conditioned media of papillary fibroblasts displayed intrinsic high angiogenic potential, reticular ones only contributed to capillary formation induced by exogenous VEGF. These results show that skin fibroblast populations regulate angiogenesis via both secreted and ECM-bound factors. Our work emphasizes the importance of papillary and reticular fibroblasts, not only for modelling dermis microenvironment but also for its vascularization.

187 IMPRINTED GENE EXPRESSION IN IN VIVO-AND IN VITRO-PRODUCED BOVINE FETUSES AND PLACENTAS

2008 ◽  
Vol 20 (1) ◽  
pp. 173
Author(s):  
F. Perecin ◽  
S. C. Méo ◽  
W. Yamazaki ◽  
C. R. Ferreira ◽  
F. H. Biase ◽  
...  
Keyword(s):  
Gene Expression ◽  
Expression Patterns ◽  
Rna Extraction ◽  
Imprinted Genes ◽  
Total Rna ◽  
Imprinted Gene ◽  
Bovine Fetuses ◽  
Low Efficiency

Some gestational alterations associated with bovine somatic cell nuclear transfer (SCNT) are presumably consequences of abnormal imprinted gene expression. This work aimed to evaluate the expression patterns of imprinted genes IGF2 and IGF2R in bovine fetuses and chorioallantoic membranes derived from in vivo- and in vitro-produced embryos. Fetuses were produced by AI (in vivo group, n = 3), IVF (n = 3), parthenogenesis (n = 3), or SCNT (n = 2). Cows with positive pregnancy diagnosis after ultrasonographic examination were slaughtered between Days 33 and 36 of gestation. The reproductive tract was transported on ice to the laboratory, where fetuses and chorioallantoic fragments were collected and stored in liquid nitrogen. Total RNA extraction was performed using TRIzol, according to manufacturer's instructions, and the reverse transcription reaction was carried out with 1 µg of total RNA, 6.75 µm oligo pd(T)12–18, and 50 U of reverse transcriptase (Improm-II, Promega, Madison, WI, USA). The relative quantification of IGF2 and IGF2R transcripts was done using real-time PCR with SYBR Green dye. The average efficiency of PCR amplifications was estimated for each gene using a linear regression on the logarithm of fluorescence per cycle (Ramakers et al. 2003 Neurosci. Lett. 339, 62–66), and the expression ratios were calculated according to the method described previously by Livak and Schmittgen (2001 Methods 25, 402–408). To verify statistical differences, a pair-wise fixed reallocation randomization test (Pfaffl et al. 2002 Nucl. Acids Res. 30, e36) was used. All expression ratios were normalized by glyceraldehyde 3-phosphate dehydrogenase expression and calibrated by the in vivo group (expression assumed as 1.00 for all genes and tissues). The analysis of relative differences on transcript levels of imprinted genes in fetuses revealed IGF2 down-regulation (P < 0.05) in the SCNT (0.19) and parthenogenetic (0.02) groups when compared to the in vivo group and IVF fetuses (2.02). In chorioallantois, IGF2 was down-regulated (P < 0.001) in parthenotes (0.001) when compared to the in vivo, IVF (3.13), and SCNT (0.98) groups. IGF2R was down-regulated (P < 0.001) in SCNT chorioallantois (0.25) when compared to the in vivo group. Low expression of IGF2 in parthenogenetic fetuses and chorioallantois confirms its imprinted status in bovine. Alterations in the relative frequency of IGF2 and IGF2R transcripts were observed in bovine SCNT-derived fetuses and chorioallantoic membranes, respectively, supporting the hypothesis that abnormalities in the expression of imprinted genes are causes for the low efficiency of SCNT procedures in this species. Such alterations suggest modifications in DNA methylation patterns at IGF2 and IGF2R imprinting centers.

scholarly journals Rescue Potential of Supportive Embryo Culture Conditions on Bovine Embryos Derived from Metabolically Compromised Oocytes

2020 ◽  
Vol 21 (21) ◽  
pp. 8206
Author(s):  
Anouk Smits ◽  
Jo L. M. R. Leroy ◽  
Peter E. J. Bols ◽  
Jessie De Bie ◽  
Waleed F. A. Marei
Keyword(s):  
Embryo Culture ◽  
Embryo Quality ◽  
Negative Impact ◽  
Expression Patterns ◽  
Cellular Metabolism ◽  
Culture Conditions ◽  
Developmental Competence ◽  
Bovine Embryos ◽  
The Impact

Elevated non-esterified fatty acid (NEFA), predominantly palmitic acid (PA), concentrations in blood and follicular fluid are a common feature in maternal metabolic disorders such as obesity. This has a direct negative impact on oocyte developmental competence and the resulting blastocyst quality. We use NEFA-exposure during bovine oocyte in vitro maturation (IVM) as a model to mimic oocyte maturation under maternal metabolic stress conditions. However, the impact of supportive embryo culture conditions on these metabolically compromised zygotes are not known yet. We investigated if the addition of anti-apoptotic, antioxidative and mitogenic factors (namely, Insulin-Transferrin-Selenium (ITS) or serum) to embryo culture media would rescue development and important embryo quality parameters (cell proliferation, apoptosis, cellular metabolism and gene expression patterns) of bovine embryos derived from high PA- or high NEFA-exposed oocytes when compared to controls (exposed to basal NEFA concentrations). ITS supplementation during in vitro culture of PA-exposed oocytes supported the development of lower quality embryos during earlier development. However, surviving blastocysts were of inferior quality. In contrast, addition of serum to the culture medium did not improve developmental competence of PA-exposed oocytes. Furthermore, surviving embryos displayed higher apoptotic cell indices and an aberrant cellular metabolism. We conclude that some supportive embryo culture supplements like ITS and serum may increase IVF success rates of metabolically compromised oocytes but this may increase the risk of reduced embryo quality and may thus have other long-term consequences.

scholarly journals Effects of different oocyte retrieval and in vitro maturation systems on bovine embryo development and quality

Zygote ◽  
2014 ◽  
Vol 23 (3) ◽  
pp. 367-377 ◽  
Author(s):  
Sandra Milena Bernal ◽  
Julia Heinzmann ◽  
Doris Herrmann ◽  
Bernd Timmermann ◽  
Ulrich Baulain ◽  
...  
Keyword(s):  
Gene Expression ◽  
Embryo Development ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Developmental Competence ◽  
Bovine Embryo ◽  
Global Methylation ◽  
Oocyte Developmental Competence

SummaryCyclic adenosine monophosphate (cAMP) modulators have been used to avoid spontaneous oocyte maturation and concomitantly improve oocyte developmental competence. The current work evaluated the effects of the addition of cAMP modulators forskolin, 3-isobutyl-1-methylxanthine (IBMX) and cilostamide during in vitro maturation on the quality and yields of blastocysts. The following experimental groups were evaluated: (i) slicing or (ii) aspiration and maturation in tissue culture medium (TCM)199 for 24 h (TCM24slicing and TCM24aspiration, respectively), (iii) aspiration and maturation in the presence of cAMP modulators for 30 h (cAMP30aspiration) and in vivo-produced blastocysts. In vitro-matured oocytes were fertilized and presumptive zygotes were cultured in vitro to assess embryo development. Cleavage, blastocyst formation, blastocyst cell number, mRNA abundance of selected genes and global methylation profiles were evaluated. Blastocyst rate/zygotes for the TCM24aspiration protocol was improved (32.2 ± 2.1%) compared with TCM24slicing and cAMP30aspiration (23.4 ± 1.2% and 23.3 ± 2.0%, respectively, P<0.05). No statistical differences were found for blastocyst cell numbers. The mRNA expression for the EGR1 gene was down-regulated eight-fold in blastocysts that had been produced in vitro compared with their in vivo counterparts. Gene expression profiles for IGF2R, SLC2A8, COX2, DNMT3B and PCK2 did not differ among experimental groups. Bovine testis satellite I and Bos taurus alpha satellite methylation profiles from cAMP30aspiration protocol-derived blastocysts were similar to patterns that were observed in their in vivo equivalents (P > 0.05), while those from the other groups were significantly elevated. It is concluded that retrieval, collection systems and addition of cAMP modulators can affect oocyte developmental competence, which is reflected not only in blastocyst rates but also in global DNA methylation and gene expression patterns.

80 GLOBAL GENE EXPRESSION PATTERNS OF FRESH AND VITRIFIED IN VITRO-PRODUCED BOVINE BLASTOCYSTS

2013 ◽  
Vol 25 (1) ◽  
pp. 187
Author(s):  
M. J. Sudano ◽  
E. S. Caixeta ◽  
D. M. Paschoal ◽  
T. S. Rascado ◽  
L. F. Crocomo ◽  
...  
Keyword(s):  
Gene Expression ◽  
Messenger Rna ◽  
Expression Patterns ◽  
Rna Extraction ◽  
Global Gene Expression ◽  
Differentially Expressed ◽  
Microarray Data Analysis ◽  
Gene Expression Patterns ◽  
Mrna Levels

Over the past decades, there have been great advances in in vitro production (IVP) systems with improved culture methods and new knowledge regarding embryo genetics, physiology, ultrastructure, and morphology. Nevertheless, a major obstacle for dissemination of this technology is the great sensitivity of IVP embryos to cryopreservation. The objective was to study the global gene-expression patterns of fresh and vitrified IVP bovine embryos. Oocytes (N = 1290) were matured and fertilized in vitro (Day 0). Presumptive zygotes were cultured in SOFaa with 0.5% BSA and 2.5% of FCS. Cleavage and blastocyst production was evaluated after 3 and 7 days under standard culture conditions (at 38.5°C in atmosphere of 5% O2, 5% CO2, and 90% N2). On Day 7, half of the blastocysts were vitrified (n = 94), warmed (Sudano et al. 2011 Theriogenology 75, 1211–1220), and returned for 24 h of additional culture (re-expansion and hatching; hatched was evaluated 12 and 24 h after warming, respectively) when their RNA was extracted (vitrified group). The remaining embryos returned to culture until Day 8 when their RNA was extracted (fresh group). Total RNA extraction of a single blastocyst was performed using the PicoPure Kit (Applied Biosystems®, Foster City, CA, USA). The RNA samples were DNAse treated (Qiagen®, Valencia, CA, USA), and mRNA was amplified (RiboAmp Kit®). The aRNA output was evaluated with a NanoDrop (Thermo®, Wilmington, DE, USA) and Bioanalyzer (Agilent®, Santa Clara, CA, USA). Biotin-labelled and fragmented cRNA were obtained with the 3′IVT Kit (Affymetrix®, Santa Clara, CA, USA) to perform hybridization (N = 6–7, respectively, for vitrified and fresh groups) using the GeneChip Bovine Array (Affymetrix®). Microarray data analysis was performed with the FlexArray 1.6.1.1. Genes with a fold change of at least 2 and a probability of P ≤ 0.05 were considered differentially expressed. Real-time PCR was used to validate microarray results (N = 11–15, respectively, for vitrified and fresh groups). As a control, a pool of 200 blastocysts was submitted or not to mRNA amplification followed by the reverse transcription and qPCR of 17 genes. For statistical analyses, PROC GLIMMIX, PROC LOGISTIC, and PROC CORR were used. Cleavage and blastocyst production rates were 86.8 ± 1.0 and 32.5 ± 1.9%, respectively. Re-expansion and hatching/hatched rates were 69.3 and 19.3%, respectively. Messenger RNA abundance of amplified and nonamplified RNA had a high correlation (r = 0.89, P < 0.01). The microarray analysis indicated 383 differentially expressed genes (P ≤ 0.05) between fresh and vitrified blastocysts. Genes involved in apoptosis (PRDX2), heat shock (HSPA5), maternal recognition of pregnancy (IFNT2 and PAG2), and cell differentiation and placenta formation (KRT18) were downregulated in vitrified embryos. According to qPCR analysis, mRNA abundance of IFNT2, PRDX2, and KRT18 was downregulated, whereas HSPA5 mRNA levels were upregulated in vitrified blastocysts. Messenger RNA abundance of PAG2 was not different (P = 0.46) between fresh and vitrified embryos. In conclusion, vitrification alters the expression profile of the genes IFNT2, PRDX2, KRT18, and HSPA5 that can be related with embryo postcryopreservation survival capacity. FAPESP and LNBio-CNPEM are acknowledged.

Sodium butyrate improves the cloned yak embryo viability and corrects gene expression patterns

Zygote ◽  
2013 ◽  
Vol 23 (1) ◽  
pp. 19-26 ◽  
Author(s):  
Xian-rong Xiong ◽  
Dao-liang Lan ◽  
Jian Li ◽  
Yong Wang ◽  
Jin-cheng Zhong
Keyword(s):  
Gene Expression ◽  
Expression Patterns ◽  
Formation Rate ◽  
Cell Number ◽  
Developmental Competence ◽  
Gene Expression Patterns ◽  
Embryo Viability ◽  
Cell Stage ◽  
Expression Levels

SummaryInterspecies somatic cell nuclear transfer (iSCNT), a powerful tool in basic scientific research, has been used widely to increase and preserve the population of endangered species. Yak (Bos grunniens) is one of these species. Development to term of interspecies cloned yak embryos has not been achieved, possibly due to abnormal epigenetic reprogramming. Previous studies have demonstrated that treatment of intraspecies cloned embryos with (NaBu) significantly improves nuclear–cytoplasmic reprogramming and viability in vitro. Therefore, in this study, we evaluated the effect of optimal NaBu concentration and exposure time on preimplantation development of yak iSCNT embryos and on the expression patterns of developmentally important genes. The results showed that 8-cell rate, blastocyst formation rate and total cell number increased significantly compared with their untreated counterparts when yak iSCNT embryos were treated with 5 nM NaBu for 12 h after activation, but that the 2-cell stage embryo rate was not significantly different. The treatment of NaBu also increased significantly the expression levels of Oct-4 and decreased the expression levels of HDAC-2, Dnmt-1 and IGF-1; the expression patterns of these genes were more similar to that of their bovine–yak in vitro fertilization (BY-IVF) counterparts. The results described above indicated that NaBu treatment improved developmental competence in vitro and ‘corrected’ the gene expression patterns of yak iSCNT embryos.

Lipid profile of bovine blastocysts exposed to insulin during in vitro oocyte maturation

2018 ◽  
Vol 30 (9) ◽  
pp. 1253 ◽  
Author(s):  
Denise Laskowski ◽  
Göran Andersson ◽  
Patrice Humblot ◽  
Marc-André Sirard ◽  
Ylva Sjunnesson ◽  
...  
Keyword(s):  
Gene Expression ◽  
Lipid Profile ◽  
Oocyte Maturation ◽  
Profile Analysis ◽  
Expression Patterns ◽  
Sensitive Period ◽  
Desi Ms ◽  
In Vitro Oocyte Maturation ◽  
The Impact

Insulin is a key hormone with important functions in energy metabolism and is involved in the regulation of reproduction. Hyperinsulinaemia is known to impair fertility (for example, in obese mothers); therefore, we aimed to investigate the impact of elevated insulin concentrations during the sensitive period of oocyte maturation on gene expression and lipid profiles of the bovine Day-8 embryo. Two different insulin concentrations were used during in vitro oocyte maturation (INS10 = 10 µg mL−1 and INS0.1 = 0.1 µg mL−1) in order to observe possible dose-dependent effects or thresholds for hyperinsulinaemia in vitro. By investigating gene expression patterns by an mRNA microarray in combination with lipid profile analysis by desorption electrospray ionisation-mass spectrometry (DESI-MS) of embryos derived from insulin-treated oocytes, we gained further insights regarding molecular responses of embryos to insulin provocation during the first days of development. Lipid metabolism appeared to be influenced on multiple levels according to gene expression results but the profiles collected in positive-ion mode by DESI-MS (showing mostly ubiquinone, cholesteryl esters and triacylglycerols) did not differ significantly from controls. There are parallels in follicular development of ruminants and humans that make this bovine model relevant for comparative research on early human embryonic development during hyperinsulinaemia.

scholarly journals Human Cerebrospinal Fluid Modulates Pathways Promoting Glioblastoma Malignancy

2021 ◽  
Vol 11 ◽  
Author(s):  
Anna Carrano ◽  
Natanael Zarco ◽  
Jordan Phillipps ◽  
Montserrat Lara-Velazquez ◽  
Paola Suarez-Meade ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cerebrospinal Fluid ◽  
Expression Patterns ◽  
Therapeutic Implications ◽  
Transcriptional Changes ◽  
The Central Nervous System ◽  
And Migration ◽  
The Impact

Glioblastoma (GBM) is the most common and devastating primary cancer of the central nervous system in adults. High grade gliomas are able to modify and respond to the brain microenvironment. When GBM tumors infiltrate the Subventricular zone (SVZ) they have a more aggressive clinical presentation than SVZ-distal tumors. We suggest that cerebrospinal fluid (CSF) contact contributes to enhance GBM malignant characteristics in these tumors. We evaluated the impact of human CSF on GBM, performing a transcriptome analysis on human primary GBM cells exposed to CSF to measure changes in gene expression profile and their clinical relevance on disease outcome. In addition we evaluated the proliferation and migration changes of CSF-exposed GBM cells in vitro and in vivo. CSF induced transcriptomic changes in pathways promoting cell malignancy, such as apoptosis, survival, cell motility, angiogenesis, inflammation, and glucose metabolism. A genetic signature extracted from the identified transcriptional changes in response to CSF proved to be predictive of GBM patient survival using the TCGA database. Furthermore, CSF induced an increase in viability, proliferation rate, and self-renewing capacity, as well as the migratory capabilities of GBM cells in vitro. In vivo, GBM cells co-injected with human CSF generated larger and more proliferative tumors compared to controls. Taken together, these results provide direct evidence that CSF is a key player in determining tumor growth and invasion through the activation of complex gene expression patterns characteristic of a malignant phenotype. These findings have diagnostic and therapeutic implications for GBM patients. The changes induced by CSF contact might play a role in the increased malignancy of SVZ-proximal GBM.

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