Relative transcript abundance in porcine cumulus cells collected from different size follicles

2014 ◽  
Author(s):  
Carla Moros Nicolas ◽  
Maria Jose Izquierdo Rico ◽  
Y Li ◽  
Rakel Romar ◽  
H Funahashi
Keyword(s):  
Transcript Abundance ◽  
Cumulus Cells ◽  
Relative Transcript Abundance

299 EFFECT OF ESTRADIOL-17β AND FOLLICLE STIMULATING HORMONE ON THE IN VITRO MATURATION OF PORCINE OOCYTES DERIVED FROM SMALL FOLLICLES

2015 ◽  
Vol 27 (1) ◽  
pp. 238
Author(s):  
Y. Li ◽  
C. Moros ◽  
M. J. Izquierdo-Rico ◽  
R. Romar ◽  
H. Funahashi
Keyword(s):  
In Vitro Maturation ◽  
Egf Receptor ◽  
Transcript Abundance ◽  
Cumulus Cells ◽  
Fsh Receptor ◽  
Egf Receptors ◽  
Relative Transcript Abundance ◽  
Maturation Rate ◽  
Positive Effect

In porcine cumulus-oocyte complexes (COC) from middle follicles (MF: 3–6 mm in diameter), FSH is known to induce the resumption of meiosis and accompanied by transactivate of the EGF receptor and activation of MAPK3/1 in the cumulus cells. The aim of the current study was to examine the effect of oestradiol-17β (E2: 0.1 μg mL–1) or FSH on in vitro maturation (IVM) of porcine oocytes derived from small follicles (SF: 1–2 mm in diameter). The COC were aspirated from MF of porcine ovaries obtained at slaughterhouse and cultured for IVM in mPOM (with 1 mM dibutyryl cAMP, 10 IU mL–1 of eCG, and 10 IU mL–1 of hCG for 20 h and then without those for 24 h in an atmosphere of 5% CO2 in air at 39°C) after washing 3 times. The COC from SF, which were aspirated at the same time with COC from MF, were precultured in the absence or presence of E2 or E2 plus FSH for 6 h before IVM culture. After the culture, oocytes were denuded from cumulus cells with 0.1% (vol/vol) hyaluronidase and the meiotic stage was observed. Relative transcript abundance of FSH and EGF receptors of CC was also examined by real-time RT–PCR just after preincubation for 6 h. Statistical analysis of data from 3 to 5 replicates was analysed by ANOVA and Tukey's multiple comparison tests. Maturation rate of oocytes from SF (40.6 ± 3.1%) was significantly lower than that of oocytes from MF controls (78.8 ± 2.8%, P < 0.01). Preincubation in the presence of E2 alone and E2 plus 0.005 IU of FSH significantly increases the maturation rate of oocytes from SF (56.8 ± 1.5 and 55.7 ± 3.1%, respectively, P < 0.01), although the rate was still lower than MF controls. However, in the presence of E2 plus a higher concentration of FSH (0.05 and 0.5 IU), oocyte maturation rate was similar (36.3 ± 2.4 and 33.7 ± 1.9%, respectively) to SF controls and lower than those of E2 alone and E2 plus 0.005 IU of FSH groups. Relative transcript abundance of FSH receptor of CC increased (P < 0.01) during preincubation in the presence of E2, but decreased in the presence of 0.05 IU of FSH. There were no significant differences in the transcript abundance of EGF receptors among treatments during preincubation (P = 0.09). In conclusion, preincubation of COC from SF in the presence of E2 alone and E2 plus 0.005 IU of FSH improves the maturation rate of the oocytes, whereas the presence of FSH more than 0.05 IU mL–1 concealed the positive effect. These effects may be yielded by change in the relative transcript abundance of FSH receptor of COC through the treatments.

Relative transcript abundance in porcine cumulus cells collected from different sized follicles

2020 ◽  
Author(s):  
Carla Moros‐Nicolás ◽  
Mª José Izquierdo‐Rico ◽  
Yang Li ◽  
Leopoldo González‐Brusi ◽  
Raquel Romar ◽  
...  
Keyword(s):  
Transcript Abundance ◽  
Cumulus Cells ◽  
Relative Transcript Abundance

scholarly journals Analysis of Cadmium Root Retention for Two Contrasting Rice Accessions Suggests an Important Role for OsHMA2

Plants ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 806
Author(s):  
Moez Maghrebi ◽  
Elena Baldoni ◽  
Giorgio Lucchini ◽  
Gianpiero Vigani ◽  
Giampiero Valè ◽  
...  
Keyword(s):  
Transcript Abundance ◽  
Root Cells ◽  
Expression Levels ◽  
Cd Tolerance ◽  
Relative Contribution ◽  
Relative Transcript Abundance ◽  
Potential Capacity ◽  
Cd Distribution ◽  
Cd Exposure ◽  
Cd Availability

Two rice accessions, Capataz and Beirao, contrasting for cadmium (Cd) tolerance and root retention, were exposed to a broad range of Cd concentrations (0.01, 0.1, and 1 μM) and analyzed for their potential capacity to chelate, compartmentalize, and translocate Cd to gain information about the relative contribution of these processes in determining the different pathways of Cd distribution along the plants. In Capataz, Cd root retention increased with the external Cd concentration, while in Beirao it resulted independent of Cd availability and significantly higher than in Capataz at the lowest Cd concentrations analyzed. Analysis of thiol accumulation in the roots revealed that the different amounts of these compounds in Capataz and Beirao, as well as the expression levels of genes involved in phytochelatin biosynthesis and direct Cd sequestration into the vacuoles of the root cells, were not related to the capacity of the accessions to trap the metal into the roots. Interestingly, the relative transcript abundance of OsHMA2, a gene controlling root-to-shoot Cd/Zn translocation, was not influenced by Cd exposure in Capataz and progressively increased in Beirao with the external Cd concentration, suggesting that activity of the OsHMA2 transporter may differentially limit root-to-shoot Cd/Zn translocation in Capataz and Beirao.

Detection of abscisic acid and relative transcript abundance in Syntrichia caninervis Mitt.

2021 ◽  
pp. 1-8
Author(s):  
Yigong Zhang ◽  
Ayibaiheremu Mutailifu ◽  
Yi Zhang ◽  
Honglan Yang ◽  
Daoyuan Zhang
Keyword(s):  
Abscisic Acid ◽  
Transcript Abundance ◽  
Syntrichia Caninervis ◽  
Relative Transcript Abundance

scholarly journals Predictive value of bovine follicular components as markers of oocyte developmental potential

2014 ◽  
Vol 26 (2) ◽  
pp. 337 ◽  
Author(s):  
Satoko Matoba ◽  
Katrin Bender ◽  
Alan G. Fahey ◽  
Solomon Mamo ◽  
Lorraine Brennan ◽  
...  
Keyword(s):  
Follicular Fluid ◽  
Cumulus Cell ◽  
Principal Component ◽  
Transcript Abundance ◽  
Cumulus Cells ◽  
Blastocyst Stage ◽  
Developmental Potential ◽  
Thecal Cells ◽  
Total Fatty Acids

The follicle is a unique micro-environment within which the oocyte can develop and mature to a fertilisable gamete. The aim of this study was to investigate the ability of a panel of follicular parameters, including intrafollicular steroid and metabolomic profiles and theca, granulosa and cumulus cell candidate gene mRNA abundance, to predict the potential of bovine oocytes to develop to the blastocyst stage in vitro. Individual follicles were dissected from abattoir ovaries, carefully ruptured under a stereomicroscope and the oocyte was recovered and individually processed through in vitro maturation, fertilisation and culture. The mean (± s.e.m.) follicular concentrations of testosterone (62.8 ± 4.8 ng mL–1), progesterone (616.8 ± 31.9 ng mL–1) and oestradiol (14.4 ± 2.4 ng mL–1) were not different (P > 0.05) between oocytes that formed (competent) or failed to form (incompetent) blastocysts. Principal-component analysis of the quantified aqueous metabolites in follicular fluid showed differences between oocytes that formed blastocysts and oocytes that degenerated; l-alanine, glycine and l-glutamate were positively correlated and urea was negatively correlated with blastocyst formation. Follicular fluid associated with competent oocytes was significantly lower in palmitic acid (P = 0.023) and total fatty acids (P = 0.031) and significantly higher in linolenic acid (P = 0.036) than follicular fluid from incompetent oocytes. Significantly higher (P < 0.05) transcript abundance of LHCGR in granulosa cells, ESR1 and VCAN in thecal cells and TNFAIP6 in cumulus cells was associated with competent compared with incompetent oocytes.

214 EFFECT OF OVARIAN SUPERSTIMULATION ON EXPRESSION OF GENES ASSOCIATED WITH THE OOCYTE DEVELOPMENTAL COMPETENCE OF NELORE COWS

2013 ◽  
Vol 25 (1) ◽  
pp. 255
Author(s):  
R. A. Satrapa ◽  
E. M. Razza ◽  
A. G. Pupulim ◽  
A. C. S. Castilho ◽  
B. Loureiro ◽  
...  
Keyword(s):  
Animal Health ◽  
Rna Extraction ◽  
Transcript Abundance ◽  
Cumulus Cells ◽  
Blastocyst Stage ◽  
Developmental Competence ◽  
Control Group ◽  
Oocyte Competence ◽  
Embryo Yield ◽  
Expression Of Genes

The P36 protocol has contributed to the genetic improvement of Brazilian herd through its successful use in embryo transfer programs. We aimed to investigate the effect of P36 protocol on embryo yield and mRNA expression of genes correlated with the competence of cumulus–oocyte complex (COC): receptors of FSH (FSHR), EGF (EGFR), and pentraxin 3 (PTX3) in cumulus cells; receptors of LH (LHR) and angiotensin 2 (AT2) in granulosa cells; and GDF9, BMP15, and histone H2A (H2A) in oocytes. Multiparous Nelore cows were allocated in control and P36 groups. Control group (non-superovulated, n = 15) received a progesterone intravaginal device (P4, 1.0 g, Primer®, Tecnopec, Sao Paulo, Brazil) and 2.5 mg of oestradiol benzoate (EB, IM, BER-BE®, Syntex, Buenos Aires, Argentina) at a random day of the oestrous cycle (Day 0). A PGF2α analogue (150 mg d-cloprostenol, IM, Prolise®, RARS SRL) was administered (Day 8) and Primer® was removed. The P36 group (n = 10) received a Primer® and 2.0 mg of EB (Day 0). The FSH treatment (160 mg Folltropin®, Bioniche Animal Health, Ontario, Canada) was initiated at decreasing doses: 40, 30, 20, and 10% of the total dose twice daily for 4 days (Day 5). The PGF2α analogue was administered (Day 8) and after 36 h primer was removed. Animal slaughter to ovary collection was performed 12 h after Primer® removal (Day 9). Some of the oocytes were matured (TCM199), fertilized with Nelore semen (n = 6), and cultured (SOF-synthetic oviduct fluid) to the blastocyst stage. Embryos were removed from culture (Day 6), allocated in 5 pools with 5 embryos in each group, and subjected to RNA extraction. Remaining oocytes were denuded from cumulus and zona pellucida (vortex and Protease®, Sigma-Aldrich, St. Louis, MO, USA). Pools of 20 oocytes and of their respective cumulus cells (n = 6 pools; control group and n = 4 pools, P36 group) were subjected to RNA extraction (RNeasy kit, Qiagen, Valencia, CA, USA). Gene expression was performed by real-time RT-PCR using oligo-dT in reverse transcription and bovine-specific primers. Expression of cyclophilin A was used as endogenous control. Change to developmental rates to the blastocyst stage and transcript abundance were compared by t-test and significance was considered when P < 0.05. Blastocyst rates were also similar (P > 0.05) in groups P36 (40/99; 40%) and control (16/43; 37%). Expression of H2A, EGFR, FSHR, and PTX3 in cumulus cells did not differ (P > 0.05) among treatment groups. The expression of GDF9 and BMP15 in cumulus cells was higher (P < 0.05) in the P36 group, but in oocytes these transcripts were more expressed in the control group (P < 0.05). Although important genes (GDF9 and BMP15) were less expressed in oocytes from superstimulated cows, the maintenance of H2A in oocytes, as well as PTX3, EGFR, and FSHR, and the increases in GDF9 and BMP15 expression in cumulus cells do not seem to affect oocyte competence due to the similar embryo yield of both groups. Supported by FAPESP.

217 TRANSCRIPT ABUNDANCE OF CATHEPSIN GENES IN CUMULUS CELLS AS A MARKER OF CATTLE OOCYTE QUALITY

2013 ◽  
Vol 25 (1) ◽  
pp. 257
Author(s):  
E. Warzych ◽  
A. Wolc ◽  
A. Cieslak ◽  
D. Lechniak-Cieslak
Keyword(s):  
Real Time Pcr ◽  
Copy Number ◽  
Transcript Abundance ◽  
Cumulus Cells ◽  
Oocyte Quality ◽  
Mtdna Copy Number ◽  
Cdna Synthesis ◽  
Fa Composition ◽  
Grade 3

Dynamics of follicular growth and atresia is closely connected with apoptosis. Cathepsins (CTS) are involved in diverse biological functions, whereas one member of this family, cathepsin B, plays a major regulatory role in the process of apoptosis. Oocyte quality is a complex trait shaped by the follicular components (e.g. cumulus cells, CC; follicular fluid, FF). A negative relationship between relative transcript abundance (RA) of CTSB, CTSS, and CTSZ genes in CC with the quality of corresponding oocytes was reported in cattle (Bettegowda et al. 2008). Fatty acid (FA) composition of the FF and mtDNA copy number in the oocyte are other markers of oocyte quality. Therefore, in this study, we analysed relations between selected traits of the 3 follicular components (FF, CC, oocyte) within the individual follicle with the focus on oocyte quality. The experiment was based on cumulus–oocyte-complexes (COC) and FF obtained from individual follicles of slaughterhouse ovaries. Each follicle was measured and assigned into 1 of 3 classes (small <6 mm; medium 6 to 8 mm; large >8 mm). The COC morphology (grades 1 to 4) was evaluated according to Stojkovic et al. (2001). The following analyses were performed: CC, mRNA abundance of CTSB, CTSS, CTSZ, and CTSK genes (real-time PCR, 100 replicates, ACTB as a reference gene); the oocyte, mtDNA copy number (real-time PCR, 93 replicates, COX1 gene); and FF and FA composition (gas chromatography). The following procedures were employed: total RNA isolation, mirVana Paris Kit (Ambion); total DNA isolation, High Pure PCR Template Preparation Kit (Roche, Indianapolis, IN, USA); cDNA synthesis, Transcriptor High Fidelity cDNA Synthesis Kit (Roche); and the standard curve method, to analyse the qPCR data. For statistical analysis, the Kruskal-Wallis test as well as Spearman rank correlation were applied. The highest RA of CTSB gene was noted in CC from the grade 3 COC (P < 0.05), whereas that of CTSK and CTSZ genes in CC from the grade 4 COC (P < 0.01). Because grade 3 and 4 COC are not suitable for IVM, we assumed that high RA of CTS gene in CC may indicate reduced quality of the corresponding oocyte. Surprisingly, the highest RA for CTSB gene was observed in CC from the medium follicles (P < 0.05). Significant (P < 0.05) correlations were estimated between the following: RA of CTSB gene in CC and mtDNA copy number in the oocyte (r = 0.27), RA of CTSB gene in CC and C18.3 n-3 concentration in FF (r = 0.32), RA of CTSZ gene in CC and C18.3 n-3 concentration in FF (r = 0.37), as well as RA of CTSZ gene in CC and n-3 concentration in FF (r = 0.34). Although an increase in RA of the CTS genes in CC was accompanied by the inferior oocyte morphology, it was also correlated with higher mtDNA copy number in the oocyte and FA content in FF. The last 2 features have been previously attributed to oocytes of better quality, what contrasts with the high RA of the CTS genes. Thus, higher RA of CTS genes within CC may not mark the bovine oocyte of reduced quality. Funding–National Science Center, grant no. N N302 604438.

126 Interference of mastitis with ovulation and oocyte and granulosa cell quality in dairy cows

2019 ◽  
Vol 31 (1) ◽  
pp. 188
Author(s):  
G. Santos ◽  
M. P. Bottino ◽  
A. P. C. Santos ◽  
R. E. Orlandi ◽  
L. M. S. Simões ◽  
...  
Keyword(s):  
Dairy Cows ◽  
Follicular Fluid ◽  
Rate Control ◽  
Cumulus Cell ◽  
Transcript Abundance ◽  
Cumulus Cells ◽  
Subclinical Mastitis ◽  
Oocyte Quality ◽  
Ovulation Rate ◽  
Intravaginal Device

The objective of this study was to evaluate the effect of mastitis diagnosed by somatic cell count (SCC) on follicular growth, ovulation, oocytes and cumulus cells quality and the concentration and size of exosomes in follicular fluid of dairy cows. In the study, crossbred cows (Bos taurus-Holstein×Bos indicus-Gir) were classified for analysis as control (SCC &lt;200.000 cells mL−1) and mastitis (SCC &gt;400.000 cells mL−1) groups. In Experiment 1 (follicular dynamics), cows (n=57: control=31; mastitis=26) received a progesterone intravaginal device (Sincrogest®, Ourofino Saude Animal, Cravinhos, Brazil) and 2mg of oestradiol benzoate (Sincrodiol®, Ourofino Saude Animal) injected IM. Eight days later (D8), the progesterone device was removed and cows received IM 500mg of cloprostenol (Sincrocio®, Ourofino Saude Animal), 1mg of oestradiol cypionate (SincroCP®, Ourofino Saude Animal) and 300IU of eCG (SicroeCG®, Ourofino Saude Animal). Ultrasound exams (Mindray 4900, probe linear de 5MHz, Shenzhen, China) were performed every 24h from removal of the progesterone-releasing intravaginal device (D8) until 48h later. Thereafter, evaluations were performed every 12h, until ovulation or up to 96h after removal of the progesterone-releasing intravaginal device. In Experiment 2 (oocyte, cumulus complexes, and follicular fluid evaluation), cows (n=26: control=13; mastitis=13) were submitted to follicular aspiration (ovum pickup) for oocyte quality and cumulus cells transcript evaluation. Transcript abundance of apoptosis markers (BCL2, BAX, PI3K, PTEN, FOXO3) was determined by real-time RT-PCR. Moreover, 7 days after the ovum pickup session, the dominant follicle was aspirated and follicular fluid samples were obtained. Exosomes were isolated from the follicular fluid by serial centrifugations, which were also performed for evaluation of particle size and concentration. Statistical analyses were performed using the SAS (SAS Institute Inc., Cary, NC, USA), and the GLIMMIX procedure was used to determine significant differences between groups. Gene expression and exosome data were submitted to the Student’s t-test. Ovulation rate [control 77.4% (24/31) and mastitis 57.7% (15/26); P=0.09] and viable oocytes rate [control 59.1% (130/220) and mastitis 41.9% (125/298); P=0.01] were higher in control animals. Additionally, there was a greater number of degenerate oocytes (control 6.7±1.2 and mastitis 13.3±5.5; P=0.001) in subclinical mastitis cows. There was greater abundance (P=0.003) of BAX cumulus cell transcripts and exosome mean (P=0.03) was smaller in subclinical mastitis cows. However, BCL2, PI3K, PTEN, nd FOXO3 cumulus cell transcripts was similar between treatments. In conclusion, ovulation rate, oocyte quality, and exosome diameter were smaller in cows with SCC &gt;400.000 cells mL−1, demonstrating that subclinical mastitis can influence the fertility of dairy cows.

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