264 TESTOSTERONE IN THE OOCYTE CULTURE DOES NOT ALTER SEX-RATIO OF IN VITRO PRODUCED BOVINE EMBRYOS

2009 ◽  
Vol 21 (1) ◽  
pp. 229
Author(s):  
C. Díez ◽  
P. Bermejo-Alvarez ◽  
A. Gutiérrez-Adan ◽  
J. N. Caamaño ◽  
M. Muñoz ◽  
...  
Keyword(s):  
Sex Ratio ◽  
Cumulus Cells ◽  
Basic Medium ◽  
Bovine Embryos ◽  
Experimental Conditions ◽  
Vitro Fertilization ◽  
Grant Support ◽  
High Concentrations

The production of sex-known offspring is a main objective in reproductive biotechnology. It has been reported that bovine ova developed in follicles with high concentrations of testosterone in vivo yielded significantly more male embryos in vitro (Grant V et al. 2008 Biol. Reprod. 78, 812–815). In this work we aimed to test the effects of testosterone on sex ratio of bovine embryos produced in fully in vitro conditions. Immature bovine cumulus–oocyte complexes (COCs; n = 750) from slaughterhouse ovaries were cultured in 199 HNaCO3 with polyvinyl alcohol (PVA) 0.1 mg mL–1 as a basic medium. Culture was made in two steps, a 24 h meiotic arrest (roscovitine 25 μm), and a subsequent in vitro maturation period with FSH-LH for 24 h. Testosterone (T-86500, Sigma-Aldrich, St. Louis, MO, USA) was added throughout the entire oocyte culture at 0, 30, 300, and 1500 nm. After in vitro fertilization (Day 0), zygotes were freed of cumulus cells by pipetting, and subsequently cultured in SOF + 6 g L–1 BSA up to Day 3. At this time, embryo development was recorded, and all embryos having 3 or more cells were treated with pronase to remove the zona pellucida. Zona-free embryos were washed in PBS containing PVA 0.1 mg mL–1 and individually frozen at –80°C until sex analysis by PCR (Bermejo-Alvarez P et al. 2008 Biol. Reprod. doi:10.1095/biolreprod.108.070169). A total of 252 embryos from 5 replicates were sexed. Data for development and sex-ratio are presented as % LSM ± SD. There were no interactions between testosterone treatment, embryonic sex, and embryonic stage analyzed. Testosterone did not affect development rates (P > 0.05) at any stage: cleavage (47.8 ± 6.8, 56.5 ± 6.8; 50.9 ± 6.8; 62.2 ± 6.8), 3 to 4 cells (40.6 ± 5.2, 45.8 ± 5.2; 37.8 ± 5.2; 47.7 ± 5.2) and >5 cells rates (24.5 ± 4; 27.3 ± 4; 21.3 ± 4; 25.3 ± 4) for 0, 30, 300, and 1500 nm testosterone, respectively. Cumulative percentages of male embryos were as follows: 53 ± 8 (n = 56), 42.6 ± 8 (n = 52), 53.6 ± 6 (n = 81) and 57.6 ± 8 (n = 63) for 0, 30, 300, and 1500 nm groups respectively (P > 0.05). These results show that the testosterone effects on oocyte ability to select Y-chromosome bearing spermatozoa are not reproducible in vitro under the present experimental conditions. Grant support: MEC, project AGL2008-01530; RTA2008-0082; M. Muoz is supported by FICYT.

scholarly journals Features of the preparation of biological material for genome editing in cattle

2019 ◽  
Vol 191 (12) ◽  
pp. 40-44
Author(s):  
A. Barkova ◽  
M. Modorov ◽  
G. Isaeva ◽  
A. Krivonogova
Keyword(s):  
Genome Editing ◽  
Biological Material ◽  
Genetic Material ◽  
Cumulus Cells ◽  
Vitro Fertilization ◽  
Expected Time ◽  
Donor Material ◽  
Material Transportation

Abstract. To carry out genome editing in cattle, an effective and well-functioning system for obtaining gametes, fertilizing eggs and their cryopreservation is necessary. Aim of the work: review and research of present-day existing methods of obtaining, insemination and cryopreservation of donor material, in order to provide genome editing in cows. Methods and materials. The work is completed according to the theme No. 0532-2019-0001 “Development of complex technology of marker-based genome selection of agricultural animals” within State Order of Ministry of Education and Science of the Russian Federation. The analysis of open scientific literature on the issues of in vitro fertilization in animals, cryopreservation of oocytes and embryons, sperm preparation and methods of insemination of cows’ oocytes, and cryopreservation of oocytes and embryons of animals is done. Features of the preparation of biological material of cattle for genome editing by microinjection into ooplasm are described. Results of research and duscussion. At present time there are two ways to obtain donor material from cattle: from live animals and taking ovaries after slaughtering cows. Material transportation is carried out at a temperature of 30–37 °C depending on the distance to the laboratory and expected time period of transportation. Oocyte-cumulus complexes can be removed by ovarian dissection and aspiration of visible follicles. In both cases, immature eggs are predominantly obtained. Subsequent ripening is carried out in vitro using special media in a CO2 incubator. The culture medium for oocyte maturation should contain hormones that mimic the peak of LH (luteinizing hormone), which occurs in vivo during the maturation of oocytes before ovulation. To accumulate a certain number of eggs at the stage of MII, it is recommended to carry out their cryopreservation by the method of vitrification, having previously released the oocyte from the cumulus cells. After thawing, oocytes need to be incubated for 2–3 hours 38.5 °C in 5–6.5% CO2 to restore the spindle. In order to make editing more effective, the introduction of genetic material is recommended to be carried out in parallel with the fertilization method “icsi”. In humans, mice and rabbits, an injection of sperm into the cytoplasm is sufficient to activate the oocyte, however, in cattle, just micro-injection of the sperm is not enough and often the male pronucleus does not form. To solve the problem, various methods are used, including freezing-thawing of sperm, resulting in damage of a membrane, or addition of heparin-glutathione into the medium that increases decondensation of the sperm DNA.

74 IN VIVO SURVIVAL OF DOMESTIC CAT OOCYTES AFTER VITRIFICATION, INTRACYTOPLASMIC SPERM INJECTION, AND TRANSFER TO RECIPIENTS

2008 ◽  
Vol 20 (1) ◽  
pp. 118 ◽  
Author(s):  
M. C. Gómez ◽  
N. Kagawa ◽  
C. E. Pope ◽  
M. Kuwayama ◽  
S. P. Leibo ◽  
...  
Keyword(s):  
Intracytoplasmic Sperm Injection ◽  
Polar Body ◽  
Oocyte Retrieval ◽  
Cumulus Cells ◽  
Domestic Cat ◽  
Minimal Amount ◽  
Vitro Fertilization ◽  
First Polar Body

The ability to cryopreserve female gametes efficiently holds immense economic and genetic implications. The purpose of the present project was to determine if domestic cat oocytes could be cryopreserved successfully by use of the Cryotop method. We evaluated (a) cleavage frequency after in vitro fertilization (IVF) v. intracytoplasmic sperm injection (ICSI) of in vivo- and in vitro-matured oocytes after vitrification, and (b) fetal development after transfer of resultant embryos into recipients. In vivo-matured cumulus–oocyte complexes (COCs) were recovered from gonadotropin-treated donors at 24 h after LH treatment, denuded of cumulus cells, and examined for the presence of the first polar body (PB). In vitro-matured COCs were obtained from ovaries donated by local clinics and placed into maturation medium for 24 h before cumulus cells were removed and PB status was determined. Oocytes were cryopreserved by the Cryotop method (Kuwayama et al. 2005 Reprod. Biomed. Online 11, 608–614) in a vitrification solution consisting of 15% DMSO, 15% ethylene glycol, and 18% sucrose. For IVF, oocytes were co-incubated with 1 � 106 motile spermatozoa mL–1 in droplets of modified Tyrode's medium in 5% CO2/air at 38�C (Pope et al. 2006 Theriogenology 66, 59–71). For ICSI, an immobilized spermatozoon was loaded into the injection pipette, which was then pushed through the zona pellucida into the ooplasm. After a minimal amount of ooplasm was aspirated into the pipette, the spermatozoon was carefully expelled, along with the aspirated ooplasm. After ICSI, or at 5 or 18 h post-insemination, in vivo- and in vitro-matured oocytes, respectively, were rinsed and placed in IVC-1 medium (Pope et al. 2006). As assessed by normal morphological appearance after liquefaction, the survival rate of both in vivo- and in vitro-matured oocytes was >90% (93–97%). For in vitro-matured oocytes, cleavage frequencies after IVF of control and vitrified oocytes were 73% (16/22) and 53% (30/57), respectively, as compared to 68% (19/28) after ICSI of vitrified oocytes (P > 0.05). For in vivo-matured oocytes, cleavage frequencies after IVF of control and vitrified oocytes were 55% (18/33) and 35% (6/17), respectively, compared to 50% (10/20) after ICSI of vitrified oocytes (P > 0.05). At 18–20 h after ICSI, 18 presumptive zygotes and four 2-cell embryos derived from vitrified in vitro-matured oocytes and 19 presumptive zygotes produced from seven in vivo-matured and 12 in vitro-matured vitrified oocytes were transferred by laparoscopy into the oviducts of two recipients at 24–26 h after oocyte retrieval. The two recipients were 9-month-old IVF/ET-derived females produced with X-sperm sorted by flow cytometry. At ultrasonography on Day 22, both recipients were pregnant, with three live fetuses observed in one recipient and one live fetus seen in the second recipient. On Day 63 and Day 66 of gestation, four live kittens were born, without assistance, to the two recipients. The one male and three female kittens weighed an average of 131 g. In summary, in vivo viability of zygotes/embryos produced by ICSI of cat oocytes vitrified by the Cryotop method was demonstrated by the birth of live kittens following transfer to recipients.

scholarly journals Pregnancies after co-culture of cumulus cells with bovine embryos derived from in-vitro fertilization of in-vitro matured follicular oocytes

Reproduction ◽  
1988 ◽  
Vol 83 (2) ◽  
pp. 753-758 ◽  
Author(s):  
K. Goto ◽  
Y. Kajihara ◽  
S. Kosaka ◽  
M. Koba ◽  
Y. Nakanishi ◽  
...  
Keyword(s):  
In Vitro Fertilization ◽  
Cumulus Cells ◽  
Bovine Embryos ◽  
Vitro Fertilization

Factors affecting the sex ratio of in-vitro and in-vivo produced bovine embryos

1999 ◽  
Vol 51 (1) ◽  
pp. 396 ◽  
Author(s):  
G Carbonneau ◽  
H Twagiramungu ◽  
N Morin ◽  
C Brisson ◽  
J Durocher ◽  
...  
Keyword(s):  
Sex Ratio ◽  
Bovine Embryos ◽  
Factors Affecting

211 IN VIVO MATURATION AND IN VITRO FERTILIZATION OF ALPACA OOCYTES

2011 ◽  
Vol 23 (1) ◽  
pp. 204 ◽  
Author(s):  
W. Huanca ◽  
R. L. Condori ◽  
M. A. Chileno ◽  
J. Cainzos ◽  
J. J. Becerra ◽  
...  
Keyword(s):  
Cumulus Cells ◽  
Hypodermic Needle ◽  
High Humidity ◽  
Cleavage Rate ◽  
Sodium Pyruvate ◽  
Vitro Fertilization ◽  
Collection Rate ◽  
Follicular Response

The objectives of the study were to evaluate the ovarian follicular response, cumulus–oocyte complex (COC) collection rate, fertilization, and culture of COC collected from alpacas after treatment with 2 different gonadotropins. Female alpacas were assigned to Group 1 (n = 8), 200 mg of FSH (Folltropin, Bioniche, Belleville, Ontario, Canada) divided b.i.d. for 3 days, plus a single IM dose of 1000 IU of hCG (Chorulon, Intervet, Salamanca, Spain) 24 h after the last FSH treatment; or Group 2 (n = 10), 750 IU of eCG (Folligon, Intervet) as a single dose, plus a single IM dose of 1000 IU of hCG on Day 3 after eCG treatment (Day 0 = start of the superstimulatory treatment). At 20 to 22 h post-hCG treatment, the ovaries were surgically exposed and COC were aspirated from follicles ≥6 mm and evaluated. The COC with a homogeneous cytoplasm and 2 or more layers of cumulus cells were transferred to plates with a 40-μL drop of TCM-199 maturation medium supplemented with 10% FCS (vol/vol) plus 0.5 μg mL–1 of FSH, 10 μg mL–1 of hCG, 0.2 mM sodium pyruvate, 50 μg mL–1 of gentamicin, and 1 μg mL–1 of oestradiol under mineral oil with 10 to 12 oocytes/drop and maturated 24 h at 39°C in an atmosphere of 5% CO2 and high humidity. After maturation, COC were removed and fertilized in vitro using epididymal sperm. Testes were collected from mature males from a slaughterhouse and transported to the laboratory. The caudal epididymide was isolated. A prick was made on the convoluted tubules with a sterile hypodermic needle and the fluid, rich in spermatozoa, was aspirated in syringes containing 2 mL of Tris-fructose egg yolk extender. Motile spermatozoa were obtained by centrifugation at 600 × g on a Percoll discontinuous gradient (45.0:22.5%) for 10 min. The supernatant was then removed by aspiration and the pellet was resuspended in TL-HEPES and centrifuged again at 300 × g for 5 min. The pellet was resuspended in TL-stock. Gametes were co-incubated for 18 h at 39°C with 5% CO2 and high humidity. Presumptive zygotes were cultured in KSOM medium supplemented with 1 mM glutamine, 0.3 mM sodium pyruvate, 50 μg mL–1 of gentamicin, EDTA, essential and nonessential amino acids, and BSA for 3 days and cultured in SOF medium for 7 days. Embryo development was evaluated at 72 h and 7 days. Data were subjected to ANOVA. The number of follicles ≥6 mm did not differ at the time of COC collection (19.3 ± 5.7 and 21.5 ± 7.3), and the number of COC collected was 16.7 ± 5.3 and 17.3 ± 6.6 for the FSH group and the eCG group, respectively. The cleavage rate was 45.2 and 42.1% for the FSH group and the eCG group, respectively, at 72 h of culture, and the blastocyst stage at Day 7 (22.2 v. 19.3) did not differ between treatments. In conclusion, the FSH and eCG treatments did not differ in the ovarian follicular response, COC collection rate, fertilization, and culture of COC. Both gonadotropins can be used in the IVF protocol for alpacas. Grant 064 FINCyT-PIBAP 2008 and Grant 032-2009 PROCYT–CONCYTEC.

191 IN VITRO CULTURE DURING OOCYTE MATURATION AND FERTILIZATION INFLUENCES THE TRANSCRIPTOME PROFILES OF BOVINE BLASTOCYSTS

2015 ◽  
Vol 27 (1) ◽  
pp. 186
Author(s):  
A. Gad ◽  
U. Besenfelder ◽  
V. Havlicek ◽  
M. Hölker ◽  
F. Rings ◽  
...  
Keyword(s):  
Lipid Metabolism ◽  
Oocyte Maturation ◽  
Microarray Data ◽  
In Vitro Maturation ◽  
Culture Conditions ◽  
Control Group ◽  
Bovine Embryos ◽  
Vitro Fertilization

Early embryonic development, the period from oocyte maturation until blastocyst formation, is the most critical period of mammalian development. It is well known that in vitro maturation, fertilization, and culture of bovine embryos is highly affected by culture conditions. However, the stage-specific effect of culture environment is poorly understood. Therefore, we aimed to examine the effect of in vitro culture conditions during oocyte maturation and fertilization on the transcriptome profile of the resulting blastocysts. Bovine oocytes were matured in vitro and then either directly transferred to synchronized recipients, fertilized, and cultured in vivo (Vitro_M), or transferred after in vitro fertilization (Vitro_F), or at zygote stage (Vitro_Z) and blastocysts were collected at Day 7 by uterine flushing. For in vivo or in vitro fertilization, the same frozen-thawed commercial bull semen has been used. Complete in vitro (IVP) and in vivo produced blastocysts were used as controls. Gene expression patterns between each blastocyst group and in vivo produced blastocyst group were compared using EmbryoGENE's bovine microarray (EmbryoGENE, Québec, QC, Canada) over six replicates of each group (10 blastocyst/replicate). Microarray data were statistically analysed using the Linear Models for Microarray Data Analysis (LIMMA) package under the R program (The R Project for Statistical Computing, Vienna, Austria). Results showed that, the longer the embryos spent under in vitro conditions, the higher was the number of differentially expressed genes (DEG, fold-change = 2 with adjusted P-value = 0.05) compared with in vivo control group. The Vitro_M group showed the lowest number of DEG (149); in contrast IVP group represented 841, DEG, respectively compared to in vivo control group. Ontological classification of DEG showed that lipid metabolism was the most significant function influenced by in vitro maturation conditions. More than 55% of DEG in the Vitro_M group were involved in the lipid metabolism process and most of them showed down-regulation compared to in vivo control group. On the other hand, Vitro_F and Vitro_Z groups showed nearly similar numbers of DEG (584 and 532, respectively) and the majority of these genes in both groups were involved in cell-death- and cell-cycle-related functions. Pathway analysis revealed that retinoic acid receptor activation pathways were the common ones in the Vitro_M and Vitro_F groups. However, different signalling pathways were commonly dominant in the Vitro_F and Vitro_Z groups. This study provides the transcriptome elasticity of bovine embryos exposed to different environments during maturation, fertilization, and culture periods of development.

86 EFFECT OF CRYOPROTECTANT EXPOSURE, VITRIFICATION, AND WARMING TIME OF BOVINE CUMULUS OOCYTE COMPLEXES ON IN VITRO FERTILIZATION AND EMBRYONIC DEVELOPMENT

2011 ◽  
Vol 23 (1) ◽  
pp. 148
Author(s):  
J. R. Prentice ◽  
J. Singh ◽  
R. J. Mapletoft ◽  
M. Anzar
Keyword(s):  
Calf Serum ◽  
Cumulus Cells ◽  
Control Group ◽  
Time Interval ◽  
Crystal Formation ◽  
Ice Crystal ◽  
Vitro Fertilization ◽  
Vitrification Solution ◽  
High Concentrations

Despite the importance of cryoprotectants for avoiding ice crystal formation, the high concentrations required for vitrification may be toxic to bovine oocytes. During warming (thawing), the removal of permeating cryoprotectants from cells can lead to osmotic injury, and the most appropriate time interval for warming and cryoprotectant removal from vitrified oocytes is currently uncertain. The present study aimed to evaluate the effect of cryoprotectant exposure, vitrification, and warming time of bovine cumulus oocyte complexes (COC) on fertilization and ability to develop as embryos in vitro. Follicles <8 mm in diameter were aspirated from slaughterhouse-derived bovine ovaries. Cumulus oocyte complexes with ≥3 layers of cumulus cells and a uniform cytoplasm were selected, washed 3 times in Dulbecco’s PBS + 5% newborn calf serum (CS), and randomly divided into 4 groups: 1) control group: no treatment; 2) VS1 group: COC were exposed to vitrification solution 1 [VS1: 7.5% ethylene glycol (EG) and 7.5% dimethyl sulfoxide (DMSO) in TCM-199 + 20% CS] for 5 min; 3) VS1+VS2 group: COC were exposed to VS1 for 5 min followed by vitrification solution 2 (VS2: 15% EG, 15% DMSO, and 0.5 M sucrose in TCM-199 + 20% CS) for 30 s; and 4) vitrified group: COC were exposed to VS1 and VS2, and then vitrified in liquid nitrogen using cryotops. The COC in VS1, VS1+VS2, and vitrified groups were exposed to a warming solution (0.5 M sucrose in TCM-199) for 1 or 5 min. The COC from all groups were in vitro matured (IVM) for 22 h in TCM-199 containing 5% CS, 5 μg mL–1 LH, 0.5 μg mL–1 FSH, and 0.05 μg mL–1 gentamicin at 38.5°C, 5% CO2, and high humidity, incubated with frozen–thawed sperm in Brackett-Oliphant capacitating medium for 18 h, and the presumptive zygotes were cultured in Charles Rosenkrans 1 amino acids (CR1aa) + 5% CS for 9 days. Data were analysed using Proc Glimmix in SAS® 9.2 (SAS Institute Inc., Cary, NC, USA). Cleavage and blastocyst rates in the vitrified group (25 and 2%, respectively) were significantly lower (P < 0.0001) than in control (75 and 27%), VS1 (68 and 19%), or VS1+VS2 (63 and 22%) groups. Cleavage and blastocyst rates did not differ among non-vitrified groups (P > 0.05). In VS1, VS1+VS2, and vitrified groups, warming time had no effect on cleavage or blastocyst rates (P > 0.05). In conclusion, although cryoprotectant exposure and warming times had no apparent adverse effect, vitrification of bovine COC drastically reduced cleavage and blastocyst rates. Further studies are required to understand how vitrification of bovine COC affects subsequent fertilization and embryo development. This study was supported by the Canadian Animal Genetic Resources Program, Agriculture and Agri-Food Canada.

Impact of in vitro fertilization of bovine oocytes with sex-sorted frozen–thawed spermatozoa on developmental kinetics, quality and sex ratio of developing embryos

Zygote ◽  
2010 ◽  
Vol 18 (3) ◽  
pp. 185-194 ◽  
Author(s):  
J. Peippo ◽  
M. Räty ◽  
K. Korhonen ◽  
M. Eronen ◽  
K. Kananen ◽  
...  
Keyword(s):  
In Vitro Fertilization ◽  
Sex Ratio ◽  
Embryo Development ◽  
Bovine Embryos ◽  
Embryo Production ◽  
Male Embryo ◽  
Vitro Fertilization ◽  
Bovine Oocytes

SummaryWe studied whether bovine embryos developing after in vitro fertilization (IVF) with sex-sorted spermatozoa differed in developmental kinetics, quality and sex ratio from embryos produced with unsorted spermatozoa. Abattoir-derived oocytes were fertilized with X-sorted, Y-sorted or unsorted spermatozoa from a single bull. To evaluate economical use of the sex-sorted spermatozoa, washed spermatozoa from a single straw (2 million spermatozoa) were used to fertilize each batch of collected oocytes without any further isolation steps. Concentration of the unsorted spermatozoa was adjusted accordingly. Fertilizations were assessed by staining sperm asters at 10 hpi and pronuclei at 20 hpi. Embryo development and morphological quality were monitored on days 2, 7, 8 and 9 of the development (IVF = day 0). All embryos were sexed using PCR. Following fertilization, penetration and subsequent cleavage rates were compromised in the X-sorted group compared with the Y-sorted and unsorted groups (penetration: 58.0% vs. 89.8% and 90.0%, cleavage: 65.3% vs. 81.5% and 75.0%). The use of the sex-sorted spermatozoa did not, however, reduce the proportion of transferable embryos (sex-sorted 29.6% vs. unsorted 27.7%) or their quality (quality 1: sex-sorted 36.0% vs. unsorted 19.9%). The Y-sorted spermatozoa produced more transferable embryos of better quality than the X-sorted spermatozoa (days 7–8: 31.9% vs. 26.4%, quality 1: 38.9% vs. 30.6%). On average, out of 10 transferable embryos, nine were of the predicted sex in the X- and Y-sorted spermatozoa groups. These results indicate that low numbers of X- and Y-sorted spermatozoa can be used successfully for female and male embryo production in vitro.

Steroid secretion by cumulus cells isolated from human preovulatory follicles

1985 ◽  
Vol 108 (3) ◽  
pp. 407-413 ◽  
Author(s):  
Torbjörn Hillensjö ◽  
Anita Sjögren ◽  
Björn Strander ◽  
Nieves Andino
Keyword(s):  
Cumulus Cells ◽  
Human Oocyte ◽  
Functional Difference ◽  
Fertilization Program ◽  
Hormonal Stimulation ◽  
Vitro Fertilization ◽  
Follicular Maturation ◽  
Preovulatory Follicles

Abstract. The secretion of progesterone, testosterone, and oestradiol by intact human oocyte-cumulus complexes in vitro was examined in incubations lasting 6–24 h. The complexes were aspirated from preovulatory follicles in 32 women who, due to tubal disease, were participating in an in vitro fertilization program. In 12 of the women follicular maturation was induced with clomiphene and human chorionic gonadotrophin (hCG), in 13 women with human menopausal gonadotrophin (hMG) and hCG and in 7 women with a combination of clomiphene-hMG plus hCG. The net secretion of steroids into the fertilization medium was studied before (0-6 h) and after (6–24 h) the addition of sperm, by RIA of aliquots removed at specific times. A high and sustained secretion of progesterone was found both before and after insemination. Testosterone secretion remained at a low and constant level while a net release of oestradiol was found mainly during the first hours of incubation. The release of steroids, particularly progesterone, varied according to the mode of hormonal stimulation in vivo and was highest in complexes from clomiphene-hMG-treated women, probably reflecting different maturity of the aspirated follicles. In a second series of experiments the dispersed cumulus cells were recovered after fertilization and cultured as monolayers for 2–4 days. The cells underwent spontaneous luteinization and secreted high amounts of progesterone. These results extend previous work in animals showing that also in the human the periovulatory cumulus cells are steroidogenically active. The results also suggest a functional difference in the cumulus cells related to the mode of ovulation induction.

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