Moment of Addition of LH to the Culture Medium Improves In Vitro Survival and Development of Secondary Goat Pre-antral Follicles

SummaryThe main aim of the present study was to examine the hypothesis that an increase in the number of granulosa cells surrounding developing bovine oocytes results in both high ATP levels and an increase in the acetylation level of H4K12 in oocytes grown in vitro. Oocyte–granulosa cell complexes (OGCs) were collected from early antral follicles (EAFs, 0.4–0.7 mm in diameter), and individually cultured on 96-well plates with or without additional granulosa cell mass that had been prepared from other OGCs. After 16 days of culture, we examined: (i) the rate of antrum formation of the OGCs; (ii) the diameter, maturation, and fertilization rate of the oocytes; and (iii) the ATP content and acetylation level of H4K12 in the oocytes grown in vitro. Granulosa cell mass added to the culture medium contributed to the development of OGCs with a higher rate of antrum formation and oocyte growth. Furthermore, the addition of granulosa cells increased the ATP content and acetylation level of H4K12 in oocytes grown in vitro compared with those developed without addition of granulosa cells. In addition, there was a positive correlation between the ATP content in oocytes grown in vitro and the number of granulosa cells in the corresponding OGCs. The results suggest that granulosa cells play a role not only in the development of OGCs and the growth of oocytes, but also in the determination of ATP content and the acetylation of H4K12 in the oocytes developed in vitro.

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117 Kit ligand enhances growth and nuclear maturation of buffalo oocytes in vitro

Reproduction Fertility and Development ◽

10.1071/rdv31n1ab117 ◽

2019 ◽

Vol 31 (1) ◽

pp. 184

Author(s):

M. N. Islam ◽

M. H. Alam ◽

A. Khatun ◽

M. A. Hashem ◽

M. Moniruzzaman

Keyword(s):

Serum Sodium ◽

Bovine Serum ◽

Culture Medium ◽

Fetal Bovine Serum ◽

Survival Rates ◽

Sodium Pyruvate ◽

Antral Follicles ◽

Kit Ligand ◽

Fetal Bovine

This study aimed to investigate the effect of Kit ligand (KL), a growth factor that regulates folliculogenesis in mammalian ovaries, on growth of buffalo oocytes in early antral follicles in vitro. Cumulus-oocyte complexes were dissected from early antral follicles (1mm) of slaughtered buffaloes and cultured in Dulbecco’s minimum essential medium supplemented with fetal bovine serum, sodium pyruvate, gentamycin, hypoxanthine, dexamethasone, cysteine, polyvinylpyrolidione, l-ascorbic acid, oestradiol-17β, and androstenedione in a 96-well culture plate at 38.5°C under an atmosphere of 5% CO2 in air for 6 days. The culture medium was supplemented with 0, 50, and 100 ng/mL KL (recombinant human SCF, Cat. No. H8416, R&D Systems, Minneapolis, MN, USA). Sixty oocytes were cultured in each group with 6 replications. In vitro-grown oocytes were cultured for maturation in tissue culture medium-199 supplemented with 5% fetal bovine serum, sodium pyruvate, gentamycin, and 100 ng/mL FSH at 38.5°C for 24h under an atmosphere of 5% CO2 in air. The oocytes were then stained with aceto-orcein and examined under a differential interference contrast microscope. Data were analysed using SAS/STAT version 9.1.3 for Windows (SAS Institute Inc., Cary, NC, USA) by one-way ANOVA and means compared with Tukey’s HSD test. The mean diameter of oocytes measured at the time of seeding on the culture substrate was 100.6±0.4μm (n=180). After 6 days of culture, the diameters of oocytes increased to 110.8±0.5, 114.0±0.5, and 115.0±0.6µm in 0, 50, and 100 ng/mL KL-treated groups, respectively. The survival rates were 60.0±6, 81.2±1.2, and 92.0±4.9% in 0, 50, and 100 ng/mL KL-supplemented oocytes at Day 6. Moreover, KL pretreatment enhanced maturation of buffalo oocytes dose dependently. A small proportion of oocytes (8.4%) treated with 50 ng/mL KL reached the MII stage. This number increased to 25% when oocytes were treated with 100 ng/mL KL. These results show that KL enhances growth, viability, and meiotic progression of buffalo oocytes in vitro.

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Effect of bone morphogenetic protein-7 (BMP-7) on in vitro survival of caprine preantral follicles

Pesquisa Veterinária Brasileira ◽

10.1590/s0100-736x2010000400004 ◽

2010 ◽

Vol 30 (4) ◽

pp. 305-310 ◽

Cited By ~ 10

Author(s):

Valdevane R. Araújo ◽

Cleidson M. Gomes da Silva ◽

Deborah M. Magalhães ◽

Gerlane Modesto da Silva ◽

Sônia N. Báo ◽

...

Keyword(s):

Oocyte Diameter ◽

Cortical Tissue ◽

Minimum Essential Medium ◽

Preantral Follicles ◽

Morphogenetic Protein ◽

Transmission Electron ◽

Survival And Development ◽

Survival And Growth ◽

In Vitro Survival

This study was conducted in order to verify the effect of different concentrations of BMP-7 in the in vitro survival and development of caprine preantral follicles. Fragments of caprine ovarian cortical tissue were cultured for 1 or 7 days in Minimum Essential Medium (MEM+) supplemented with different concentrations of BMP-7 (1, 10, 50 or 100ng/ml). Non-cultured fragments or those cultured for 1 or 7 days were processed for classical histology and transmission electron microscopy (TEM). Parameters such as follicular survival, activation and growth were evaluated. The results showed that, after 1 or 7 days of culture, the percentage of morphologically normal follicles was significantly reduced in all treatments when compared with fresh control, except at 1ng/ml of BMP-7 for 1 day. In addition, the concentration of 10ng/ml of BMP-7 significantly increases follicular diameter from day 1 to 7 of culture. There was no influence of the other concentrations of BMP-7 regarding to the follicular and oocyte diameter. Ultrastructure studies confirmed follicular integrity after 7 days of culture in 1ng/ml BMP-7. In conclusion, small concentrations of BMP-7 can improve the survival and growth of caprine preantral follicles during in vitro culture.

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Amburana cearensis leaf extract maintains survival and promotes in vitro development of ovine secondary follicles

Zygote ◽

10.1017/s0967199415000179 ◽

2015 ◽

Vol 24 (2) ◽

pp. 277-285 ◽

Cited By ~ 10

Author(s):

R.S. Barberino ◽

V.R.P. Barros ◽

V.G. Menezes ◽

L.P. Santos ◽

V.R. Araújo ◽

...

Keyword(s):

Culture Medium ◽

Antioxidant Properties ◽

Control Medium ◽

Cell Culture Medium ◽

Preantral Follicle ◽

Acid Control ◽

Survival And Development ◽

Alternative Culture ◽

Vitro Development

SummaryThe antioxidant properties of Amburana cearensis extract may be a useful substitute for standard cell culture medium. Thus, the aim of this study was to evaluate the effect of this extract, with or without supplementation, on in vitro survival and development of sheep isolated secondary follicles. After collection of the ovaries, secondary follicles were isolated and cultured for 18 days in α-MEM+ supplemented with bovine serum albumin, insulin, transferrin, selenium, glutamine, hypoxanthine and ascorbic acid (control medium) or into medium composed of different concentrations of A. cearensis extract without supplements (Amb 0.1; 0.2 or 0.4 mg/ml) or A. cearensis extract supplemented with the same substances described above for α-MEM+ supplementation. The A. cearensis supplemented medium was named Amb 0.1+; 0.2+ or 0.4+ mg/ml. There were more morphologically normal follicles in Amb 0.1 or Amb 0.4 mg/ml than in the control medium (α-MEM+) after 18 days of culture. Moreover, the percentage of antrum formation was significantly higher in Amb 0.1 or Amb 0.2 mg/ml than in α-MEM+ and Amb 0.1+ mg/ml, and similar to the other treatments. All A. cearensis extract media induced a progressive and significant increase in follicular diameter throughout the culture period. In conclusion, this study showed that 0.1 mg/ml of this extract, without supplementation, maintains follicular survival and promotes the development of ovine isolated secondary follicles in vitro. This extract can be an alternative culture medium for preantral follicle development.

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STUDIES ON THE EXTRACELLULAR CULTIVATION OF AN INTRACELLULAR PARASITE (AVIAN MALARIA)

Journal of Experimental Medicine ◽

10.1084/jem.96.5.465 ◽

1952 ◽

Vol 96 (5) ◽

pp. 465-476 ◽

Cited By ~ 31

Author(s):

William Trager

Keyword(s):

Malic Acid ◽

Avian Malaria ◽

Culture Medium ◽

Coenzyme A ◽

Intracellular Parasite ◽

Sodium Pyruvate ◽

Plasmodium Lophurae ◽

Survival And Development ◽

Development In Vitro

The extracellular survival and development in vitro of the erythrocytic stages of Plasmodium lophurae were favored by the addition to the culture medium of l-malic acid and concentrates rich in coenzyme A. In a concentrated extract of duck erythrocytes supplemented with these two substances in addition to adenosinetriphosphate, sodium pyruvate, and certain other materials of like nature, only 5 to 10 per cent of the extracellular parasites had become abnormal after 3 days of cultivation.

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Serum free embryo culture medium improves in vitro survival of bovine blastocysts to vitrification

Theriogenology ◽

10.1016/j.theriogenology.2007.12.015 ◽

2008 ◽

Vol 69 (8) ◽

pp. 1013-1021 ◽

Cited By ~ 47

Author(s):

E. Gómez ◽

A. Rodríguez ◽

M. Muñoz ◽

J.N. Caamaño ◽

C.O. Hidalgo ◽

...

Keyword(s):

Embryo Culture ◽

Culture Medium ◽

Serum Free ◽

Embryo Culture Medium ◽

In Vitro Survival

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Effect of estradiol during culture of bovine oocyte–granulosa cell complexes on the mitochondrial DNA copies of oocytes and telomere length of granulosa cells

Zygote ◽

10.1017/s0967199412000603 ◽

2012 ◽

Vol 22 (4) ◽

pp. 431-439 ◽

Cited By ~ 9

Author(s):

M. Endo ◽

K. Kimura ◽

T. Kuwayama ◽

Y. Monji ◽

H. Iwata

Keyword(s):

Mitochondrial Dna ◽

Telomere Length ◽

Granulosa Cells ◽

Culture Medium ◽

Mt Dna ◽

Cell Complexes ◽

Antral Follicles ◽

Mitochondrial Dna Copy Number ◽

Significant Difference

SummaryDuring the development of oocytes from early antral follicles (EAFs) to antral follicles (AFs), the mitochondrial DNA copy number (Mt DNA number) increases, and granulosa cells markedly proliferate. This study examined the effect of supplementation of culture medium with estradiol-17β (E2) on the in vitro growth of oocytes, and increases in the Mt DNA number, and telomere length during the in vitro culture of oocytes derived from EAFs (0.4–0.7 mm in diameter). The E2 supplementation improved antrum formation and the ratio of oocytes reaching the metaphase II (MII) stage, and there was a significant difference in these values between addition E2 concentrations of 10 μg/ml and 0.1 μg/ml. When the oocytes were cultured in the medium containing 10 μg/ml E2, the Mt DNA number determined by real-time polymerase chain reaction (PCR) significantly increased, and the ratio of the Mt DNA number at the end of culture to the Mt DNA number at the beginning of the culture was greatly different among cows, and could be predicted by the degree of the difference between the Mt DNA number of oocytes derived from EAFs and that of oocytes derived from AFs (3–6 mm in diameter). When oocytes were cultured for 16 days in a medium containing 10 μg/ml E2 or 0.1 μg/ml E2, the Mt DNA number of oocytes grown in vitro did not differ, but the telomere length of the granulosa cells was significantly greater in the 10 μg/ml E2 group than in the 0.1 μg/ml group. In conclusion, E2 supplementation in culture medium improved the growth of oocytes derived from EAFs, and a high E2 concentration increased the telomere length of the granulosa cells.

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