Additional effect of epidermal growth factor during in vitro maturation for individual bovine oocytes using a chemically defined medium

Zygote ◽  
2004 ◽  
Vol 12 (2) ◽  
pp. 143-150 ◽  
Author(s):  
Takahiro Oyamada ◽  
Hiroshi Iwayama ◽  
Yutaka Fukui
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Defined Medium ◽  
Control Medium ◽  
Chemically Defined Medium ◽  
Blastocyst Formation ◽  
Epidermal Growth ◽  
The Individual ◽  
Bovine Oocytes

This study was performed to establish an individual bovine oocyte-IVP system using a chemically defined simple medium (mSOFaa containing 1 mg/ml polyvinyl alcohol: PVA) and to investigate the effects of epidermal growth factor (EGF) during oocyte maturation on in vitro maturation, fertilization and embryonic development. Cumulus–oocyte complexes were collected from bovine ovaries and were matured in mSOFaa containing PVA (control medium) supplemented with 0, 1, 10 or 50 ng/ml of EGF. Two further groups (TCM199 and mSOFaa, supplemented with 10% fetal calf serum were also included. In this study, mSOFaa containing PVA were used as a basic medium for fertilization and embryo development in vitro. Experiments were conducted in both group- and individual-IVP systems. In the group-IVP system, the proportion of matured oocytes (MII) in the control medium (62.7%±5.0%) was significantly (p<0.05) lower than in all other treatments, and in the individual-IVP system, the addition of 1 ng/ml EGF significantly (p<0.05) increased the maturation rate (1 ng/ml EGF vs control: 76.2%±5.4% vs 57.1%±14.4%). The addition of EGF did not affect the proportions of penetrated and normally fertilized oocytes in either individual- or group-culture systems. In the group-IVP system, no significant difference among treatments was found in the rate of blastocyst formation, whereas in the individual-IVP system the control medium supplemented with 10 ng/ml EGF resulted in a significantly (p<0.05) higher the rate of blastocyst formation (20.0±5.2%) than that in the control medium (6.2%±3.5%). These results indicate that bovine oocytes can successfully develop to blastocysts in an individual-IVP system using a single chemically defined medium, and that the group-IVP system also resulted in a similar level of blastocyst formation to that in a standard multiple-media system in our laboratory. The effect of EGF during oocyte maturation medium differed depending on whether embryos were cultured individually or in groups.

143 EFFECT OF FSH OR EPIDERMAL-GROWTH-FACTOR-LIKE PEPTIDE SUPPLEMENTATION TO MATURATION MEDIUM ON DEVELOPMENTAL COMPETENCE OF BOVINE OOCYTES DERIVED FROM FULLY DEVELOPED FOLLICLES INDUCED BY SUPER-STIMULATION

2017 ◽  
Vol 29 (1) ◽  
pp. 180
Author(s):  
T. Yamanouchi ◽  
S. Sugimura ◽  
H. Matsuda ◽  
M. Ohtake ◽  
Y. Goto ◽  
...  
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Formation Rate ◽  
Calf Serum ◽  
Developmental Competence ◽  
Blastocyst Formation ◽  
Maturation Medium ◽  
Epidermal Growth ◽  
Bovine Oocytes

Bovine oocytes obtained by ovum-pick-up (OPU) following follicle growth treatment (FGT) have improved quality and competence (Imai et al. 2008 Reprod. Fertil. Dev. 20, 182). However, the effect of the presence of FSH or epidermal growth factor (EGF) like peptide during in vitro maturation (IVM) on the developmental competence of FGT oocytes has not been well known. This study was undertaken to examine the developmental competence of FGT oocytes following IVM in the presence of FSH (recombinant human FSH) or EGF-like peptide (amphiregulin; Areg) and IVF. Japanese Black cows (n = 17) were used as donors. Five days after arbitrary OPU (opu group), follicles ≥8 mm in diameter were aspirated again, a controlled internal drug release (CIDR) was inserted into the vagina, and then pFSH was injected twice a day from the evening of Day 6 to the morning of Day 10 with decreasing doses (total of 20 AU; 4, 4, 3, 3, 2, 2, 1, 1 AU/day). On the evening of Day 8, PGF2α (0.5 mg of cloprostenol) was administered. On Day 11, oocytes were aspirated from follicles with ≥5 mm in diameter of the treated donors by OPU (fgt group). The cumulus-oocyte complexes (COC) were cultured in the absence (opu-cont and fgt-cont groups) or presence of 0.1 IU mL−1 FSH (opu-fsh and fgt-fsh groups) or 100 ng mL−1 Areg (opu-areg and fgt-areg groups) in IVM medium (mTCM199 containing 5 mg mL−1 BSA) for 20 to 22 h (1 COC/5 µL, total of 162–171 COC per group), and then co-cultured with 3 × 106 sperm/mL for 6 h. The presumptive zygotes were continued to culture in mCR1aa supplemented with 5% newborn calf serum for 216 h (1 zygote/5 µL) using micro-well culture dishes (Dai-Nippon-Print). When repeating this opu-fgt session in the same cow, an interval at least for 50 days was kept, and the session was performed 28 times. Statistical analysis was carried out by Mann-Whitney’s U-test (between opu and fgt groups) or Steel-Dwass test after Kruskal-Wallis test (among all groups). The number of follicles ≥5 mm increased in the fgt than opu group (17.8 v. 2.9; P < 0.01). The number of COC collected was not different between the opu and fgt groups (23.1 v. 19.6; P > 0.05). The blastocyst formation rate was higher in the fgt than opu group (36.9 v. 23.1%; P < 0.01). Within 6 groups, the blastocyst formation rate was higher in the fgt-fsh (43.3%; P < 0.01) and fgt-areg (39.5%; P < 0.05) groups than the opu-cont (16.3%) group. The rate in the fgt-fsh group was also higher than that in the opu-fsh group (43.3 v. 18.7%; P < 0.01). These results suggested that FGT improved the developmental competence of bovine oocytes, probably through improving the ability of the COC to react against FSH/Areg.

The effects of 17β-estradiol and protein supplement on the response to purified and recombinant follicle stimulating hormone in bovine oocytes

Zygote ◽  
2002 ◽  
Vol 10 (1) ◽  
pp. 65-71 ◽  
Author(s):  
Atef Ali ◽  
Marc-André Sirard
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Follicle Stimulating Hormone ◽  
Defined Medium ◽  
Protein Supplement ◽  
Maturation Medium ◽  
17Β Estradiol ◽  
Epidermal Growth ◽  
Bovine Oocytes

During in vitro maturation of bovine oocytes, effects of gonadotropins (FSH, LH) and growth factors such as epidermal growth factor (EGF) vary among studies. Now that we can use defined or semi-defined medium, it becomes possible to evaluate recombinant products to assess their roles. Therefore, this study was designed to evaluate the effect of purified porcine (pFSH) or recombinant human (r-hFSH; 5, 50, or 500 ng/ml) follicle stimulating hormone, luteinising hormone (LH; 50, 500 or 5000 ng/ml) and epidermal growth factor (EGF; 5, 10, 30 or 50 ng/ml) on subsequent embryonic development of in vitro matured bovine oocytes. In addition, the presence of bovine serum albumin (BSA; 8 mg/ml) as a protein supplement during in vitro maturation (IVM) was studied. For all treatments, cumulus-oocyte complexes were matured in defined maturation medium consisting of synthetic oviduct fluid. Addition of LH to the maturation medium at all concentrations studied did not increase the proportion of oocytes developing to the morula and blastocyst stages. However, morula and blastocyst yield were improved (p < 0.05) after addition of EGF (30 ng/ml) as compared with maturation medium alone (29.3% vs 18.0%, respectively). Addition of r-hFSH to the maturation medium in the presence of 17β-estradiol (E2) significantly (p < 0.0001) increased the morula and blastocyst rate compared with maturation medium alone (40.3% vs 19.3%, respectively). The presence of BSA alone during IVM significantly reduced the developmental competence of oocytes as reflected by the morula and blastocyst yield. These results demonstrate the essential role of FSH, EGF and E2 on the kinetics of nuclear and cytoplasmic maturation that are essential for the formation of an egg capable of fertilisation and development. Also, supplementation of r-hFSH and E2 during IVM under our conditions increases morula and blastocyst yield following in vitro fertilisation and in vitro culture in defined medium. Finally, the presence of BSA as the only protein supplement during IVM may be detrimental to oocyte maturation.

PSX-B-13 Effect of epidermal growth factor and prolactin on the quality of bovine oocytes aging in vitro

2021 ◽  
Vol 99 (Supplement_3) ◽  
pp. 327-327
Author(s):  
Ekaterina Shedova ◽  
Galina Singina ◽  
Irina Y Lebedeva ◽  
Aleksandr Lopukhov
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Control Group ◽  
Tunel Staining ◽  
Apoptosis Resistance ◽  
Prolonged Culture ◽  
Epidermal Growth ◽  
Bovine Oocytes

Abstract The evaluation of factors responsible for the protection of the oocytes attained the metaphase-II stage from aging is importance for successful in vitro embryo reproduction. The aim of the present research was to study dose-dependent effects of epidermal growth factor (EGF) and prolactin (PRL) on the quality of bovine oocytes after their aging in vitro. Bovine cumulus-enclosed oocytes (CEOs) were matured in vitro for 20 h in TCM 199 containing 0.2 mM sodium pyruvate, 10% fetal calf serum (FCS), 10 μg/ml FSH and LH. At the end of in vitro maturation, oocytes were transferred to TCM 199 supplemented with 10% FCS (aging medium) and cultured for additional 24 h in the absence (Control) and in presence of EGF (10 and 50 ng/ml) and PRL (20 and 50 ng/ml). After prolonged culture oocytes were used for apoptosis detection (TUNEL staining, n=251) and the state of chromosomes evaluation (Tarkowski’s cytogenetic method, n=359). The data from 3–4 replicates were analyzed by ANOVA. At the end of prolonged culture (24 h) the rate of apoptotic oocytes in the Control group was 47.4±8.5%. EGF at concentration of 10 ng/ml and PRL at both doses decreased this rate to 15.0–22.1% (p &lt; 0.05). Furthermore, PRL (not EGF) reduced the frequency of abnormal chromosome modifications (decondensation, adherence, clumping) at concentrations of 20–50 ng/ml from 58.7±2.1% (Control) to 41.2±1.9 and 45.6±2.7% respectively (p &lt; 0.01). Thus, EGF and PRL is able to maintain the apoptosis resistance of bovine oocytes during their prolonged in vitro culture as well as PRL have the decelerating effect on abnormal modifications of M-II chromosomes. The research was supported by RFBR (17-29-08035) and the Ministry of Science and Higher Education of Russia.

Effect of epidermal growth factor on secondary palatal epithelium in vitro: tissue isolation and recombination studies

Development ◽  
1980 ◽  
Vol 58 (1) ◽  
pp. 93-106
Author(s):  
Mary S. Tyler ◽  
Robert M. Pratt
Keyword(s):  
Cell Death ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Programmed Cell Death ◽  
Dna Synthesis ◽  
Thymidine Incorporation ◽  
Control Medium ◽  
Epidermal Growth ◽  
Stimulation Of

Previous studies have shown that epidermal growth factor (EGF), a peptide of m.w. 6045, can specifically inhibit in organ culture the cessation of DNA synthesis and programmed cell death that normally occur in the presumptive fusion zone (PFZ) of the secondary palatal epithelium. The aim of this study was to determine if EGF acts directly on the epithelium to exert its effect and if there is a requirement for the underlying mesenchyme. Palatal processes from 13- and 14-day Swiss Webster embryonic mice were enzymatically separated into epithelium and mesenchyme which were then cultured alone or in transfilter recombination for up to 72 h. Tissues were examined by transmission- and scanning-electron microscopy and DNA synthesis was monitored autoradiographically using [3H]thymidine incorporation. In isolated epithelium cultured in control medium, cell death occurred in the PFZ and DNA synthesis did not occur in the oral and nasal epithelial regions. EGF (20–50 ng/ml) did not prevent cell death in the PFZ and failed to stimulate DNA synthesis in the isolated epithelium; EGF, however, did have an effect on epithelial cell morphology. In the presence of mesenchyme and EGF, there was extensive proliferation in the entire epithelium and cell death within the PFZ was not evident. The results indicate that the stimulation of DNA synthesis in the palatal epithelium by EGF requires the presence of the underlying mesenchyme and that EFG alone is not sufficient to inhibit programmed cell death within the PFZ of the isolated palatal epithelium.

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