P–709 Dual stimulation in-vitro-maturation (Duostim IVM) for overcoming oocyte maturation arrest, resulting in embryo transfer and livebirth

2021 ◽  
Vol 36 (Supplement_1) ◽  
Author(s):  
S Hatirnaz ◽  
E Hatirnaz ◽  
M Dahan ◽  
B Ata ◽  
A Basbug ◽  
...  
Keyword(s):  
Embryo Transfer ◽  
Oocyte Maturation ◽  
Luteal Phase ◽  
In Vitro Maturation ◽  
Follicular Phase ◽  
Embryo Cryopreservation ◽  
Ovulatory Cycle ◽  
Maturation Arrest ◽  
Immature Oocytes

Abstract Study question Does luteal phase followed by follicular phase letrozole priming and dual oocyte retrieval for in-vitro maturation (IVM) overcome oocyte maturation arrest (OMA)? Summary answer Oocyte maturation, fertilization,embryo cryopreservation and livebirth can be achieved with letrozole priming IVM in rare cases of OMA. What is known already OMA is an intractable problem resulting in only immature oocytes being collected and to date no succesful treatment exists. Attempts to mature oocytes collected in stimulated IVF cycles with OMA have so far failed. Cases with OMA can be due to intrinsic oocyte defects, intrafollicular factors or resistance to stimulation. Study design, size, duration Six women with OMA in ≥ 2 prior stimulated IVF cycles were treated between March 2019 and December 2020. Participants/materials, setting, methods Participants had total of 18 (range 2 - 6) prior IVF cycles yielding only 166 immature oocytes. Letrozole 5mg was given days 15–18 of ovulatory cycle; SC decapeptyl 0.1mg trigger given at follicles 12 mm, 38 hours<OPU. After menstruation, letrozole 5mg days 3–7; SChCG 250ug when follicles=12 mm 38 hours<OPU. After in-vitro-maturation oocytes reaching MII were fertilized. Embryos from luteal collection were frozen and fresh embryo transfer was attempted after follicular phase collection. Main results and the role of chance Six women underwent DuoStim IVM, median (quartiles) 3.5 (0 - 9) GV and 0.5 (0 - 2) MI oocytes were collected from luteal phase OC and 0 (0 - 0) GV and 2(0 – 4.5) MI oocytes were collected from follicular phase OC. They had a total of 166 immature oocytes collected in prior IVF cycles. There were no MII oocytes at the time of collection in any cycles.0 (0 – 3.5) oocytes matured from luteal phase OC and 1 (0 – 4) from follicular phase OC. 0 (0 – 1.5) embryos were available from luteal phase and 0 (0 - 2) from follicular phase OC.Two subjects (29 and 33 years old) underwent fresh DET and the 29 year old with 2 previous failed IVF cycles achieved a livebirth (50% per ET and 16.7% per started cycle). None of the women who did not have an embryo for fresh transfer from the follicular phase collection had an embryo from the luteal phase collection. The same 29 year old has 2 luteal phase and 2 more follicular phase embryos vitrified. Limitations, reasons for caution OMA is a rare condition with a variety of etiologies. Different etiologies can require different managements. Wider implications of the findings: It may be possible to overcome OMA with letrozole IVM in rare cases. This case is the first recorded live birth. The value of dual stimulation overcoming OMA remains uncertain. Trial registration number This study is approved by the local ethical commitee of Medicana Samsun International Hospital by a Grant number of 02/05.02.2020: registration is not required due to retrospective status

scholarly journals 303 EFFECT OF SERUM SUPPLEMENTATION AND ESTRUS CYCLE STAGE ON IN VITRO NUCLEAR MATURATION OF CANINE OOCYTES

2005 ◽  
Vol 17 (2) ◽  
pp. 302
Author(s):  
F.Y. Heru ◽  
H.J. Oh ◽  
M.K. Kim ◽  
J. Goo ◽  
M.S. Hossein ◽  
...  
Keyword(s):  
Luteal Phase ◽  
In Vitro Maturation ◽  
Germinal Vesicle ◽  
Follicular Phase ◽  
Estrus Cycle ◽  
Immature Oocytes ◽  
Nuclear Maturation ◽  
Serum Supplementation ◽  
Maturation Rate

The present study investigated the effects of the estrus cycle stage and serum supplementation on nuclear maturation of canine oocytes. Ovaries were collected from a private clinic after ovariohysterectomy and classified into follicular, luteal, or anestrus stages through a combination of ovarian morphology and vaginal cytology. A total of 2214 oocytes from 196 ovaries (903 oocytes from 96 anestrus ovaries, 609 oocytes from 36 follicular ovaries, and 702 oocytes from 64 luteal ovaries) were used for experiments. The oocyte retrieval per ovary was 10, 19, and 12 for anestrus, follicular and luteal-phase ovaries, respectively. In Exp. 1, immature oocytes were cultured for 72 h in TCM-199 alone or TCM-199 supplemented with 10% canine anestrus (CAS), estrus (CES), or diestrus (CDS) serum or fetal bovine serum (FBS). In Exp. 2, immature oocytes were cultured for 72 h in TCM-199 supplemented with 0, 5, 10, or 20% CES. After staining with Hoechst 33342, chromatin state and position as well as spindle formation were evaluated to determine the stage of meiosis: germinal vesicle (GV) stage, germinal vesicle breakdown (GVBD), metaphase I (MI) stage, metaphase II (MII) stage. The experiments with anestrus and luteal-phase oocytes were repeated eight times and follicular-phase oocytes were repeated six times. Data were subjected to analysis of variance (ANOVA) and protected least significant difference (LSD) test to determine differences among experimental groups by using the Statistical Analysis System (SAS, SAS Institute, Inc., Cary, NC, USA) program. Statistical significance was determined where P value was less than 0.05. In Exp. 1, the in vitro maturation of oocytes up to MII stage was higher when oocytes were collected from ovaries in follicular phase. The maturation rate up to MII stage was 0.0 to 1.7%, 1.3 to 10.2%, and 1.0 to 3.2% for the oocytes collected from the anestrus, follicular, and luteal-phase ovaries, respectively, depending on the culture media used. In basic TCM media only, 0.0, 1.3, and 2.3% oocytes reached the MII stage for anestrus, follicular, and luteal-phase oocytes, respectively. A significantly higher rate of maturation was obtained when oocytes collected from follicular phase were cultured in TCM-199 supplemented with 10% CES (10.2%), compared to 10% CAS (4.0%), CDS (2.7%), FBS (1.3%), or the control (1.3%). In Exp. 2, supplementing with 10% CES induced the highest (P < 0.05) maturation rate to the MII stage in oocytes collected from follicular-stage ovaries (11.5%) compared to supplementing with 0% (1.0%), 5% (1.3%), or 20% CES (5.1%). Supplementing with CES (5, 10, or 20%) did not have a significant effect on nuclear maturation of canine oocytes collected from anestrus or luteal-stage ovaries. In conclusion, supplementing in vitro maturation medium with 10% CES increased nuclear maturation of canine oocytes, and canine oocytes collected from follicular-stage ovaries are the most suitable to complete nuclear maturation in vitro. This study was supported by grants from the Biogreen 21-1000520030100000.

36 EFFECT OF cAMP MODULATORS DURING OOCYTE IN VITRO MATURATION ON GAP JUNCTIONAL ACTIVITY OF VITRIFIED BOVINE OOCYTES

2016 ◽  
Vol 28 (2) ◽  
pp. 148
Author(s):  
C. A. S. Monteiro ◽  
G. R. Leal ◽  
H. F. R. A. Saraiva ◽  
A. J. R. Camargo ◽  
P. M. S. Rosa ◽  
...  
Keyword(s):  
Oocyte Maturation ◽  
Fluorescence Intensity ◽  
In Vitro Maturation ◽  
Staining Pattern ◽  
Control Group ◽  
Immature Oocytes ◽  
Junctional Activity ◽  
Gap Junctional ◽  
Bovine Oocytes

Oocyte cryopreservation is a strategic tool for in vitro embryo production, but low rates of cryosurvival are reported for bovine oocytes. Simulated physiological oocyte maturation system (Albuz et al. 2010 Hum. Reprod. 25, 12) uses cAMP modulators to increase oocyte competence by the extension of meiosis block and gap junctional communications activity. The aim of this study was to investigate the effect of simulated physiological oocyte maturation system on gap junctional activity of vitrified bovine oocytes. Oocytes from slaughterhouse ovaries were divided into 4 groups: C (control: fresh immature oocytes); V (vitrified immature oocytes); PM-V (vitrified oocytes after a 2-h pre-in vitro maturation phase – in the presence of AMPc modulators, 100 μM Forskolin, and 500 μM IBMX); and PM (fresh immature oocytes subjected to pre-in vitro maturation). Viable oocytes (n = 404 obtained from 4 replicates) were stained with Calcein-AM using the protocol of Thomas et al. (2004 Biol. Reprod. 71(4), 1142–1149) in order to measure gap junctions activity. Images were captured in fluorescence microscope, and fluorescence intensity was analysed with ImageJ software. Mean fluorescence intensity of each group was normalized to control group to obtain relative intensity value. Means were compared by Kruskal-Wallis test and Dunn post-test. A second analysis was performed considering the percentage of each staining pattern (low, middle, and high intensity) for each group. Results were analysed using Fisher exact test. All statistical analysis were performed in GraphPad Instat program with 5% significance level. Results demonstrated that all treatments induced an increase (P < 0.05) in fluorescence intensity (V: 1.76 ± 1.13; PM-V: 1.58 ± 0.98; PM: 1.38 ± 0.94) compared with control (C: 1.00 ± 0.48). Regarding the staining patterns analyses, immature vitrified oocytes (V group) differed from control group in middle and low patterns (G1, calibrator – high: 11.2%ab, middle: 43.8%a, low: 44.9%a; G2 – high: 8.2%ab, middle: 63.9%b, low: 27.9%b; G3 – high: 16.3%a, middle: 42.3%a, low: 41.3%a; G4 – high: 6.7%b, middle: 53.9%ab, low: 39.3a). In conclusion, unexpectedly, vitrification also increased gap junctional activity, as was found for pre-in vitro maturation group. However, staining pattern analysis results showed only vitrified group was different from control, suggesting vitrified and pre-in vitro maturation groups could have gap activity affected by different ways. This research was supported by FAPERJ (E26/111.61/2013) and CAPES.

Collection and in vitro maturation of Mazama gouazoubira (brown brocket deer) oocytes obtained after ovarian stimulation

Zygote ◽  
2021 ◽  
pp. 1-7
Author(s):  
Luciana Diniz Rola ◽  
Eveline dos Santos Zanetti ◽  
Maite del Collado ◽  
Ellen de Fátima Carvalho Peroni ◽  
José Maurício Barbanti Duarte
Keyword(s):  
Oocyte Maturation ◽  
Ovarian Stimulation ◽  
In Vitro Maturation ◽  
Wildlife Conservation ◽  
Estradiol Benzoate ◽  
Ex Situ ◽  
Follicular Aspiration ◽  
Mazama Gouazoubira

Summary In vitro production of embryos has gained prominence as a tool for use in wildlife conservation programmes in situ and ex situ. However, the development of this technique depends on steps that include ovarian stimulation, collection and oocyte maturation. The purpose of this study was to assess the feasibility of an ovarian stimulation protocol for follicular aspiration, the efficiency of videolaparoscopy for follicular aspiration and test a medium for in vitro oocyte maturation for the species Mazama gouazoubira. Five females were submitted to repeated ovarian stimulation (hormone protocol using controlled internal drug release), and estradiol benzoate on D0 and eight injections of follicle-stimulating hormone, once every 12 h, from D4 onwards at 30-day intervals. Fourteen surgical procedures were performed in superstimulated females, resulting in the collection of 94 oocytes and an average of 17.1 ± 9.1 follicles observed, 13.5 ± 6.6 follicles aspirated and 7.2 ± 3.7 oocytes collected per surgery. After collection, the oocytes were submitted to in vitro maturation for 24 h and stained with Hoechst 33342 dye to evaluate their nuclear status; 64.5% of the oocytes reached MII and 16.1% were spontaneously activated by parthenogenesis. The nuclear status of oocytes that did not undergo in vitro maturation was evaluated; 80.9% were found to be immature.

scholarly journals Feasibility of Secondary Follicle Isolation, Culture and Achievement of In-Vitro Oocyte Maturation from Superovulated Ovaries: An Experimental Proof-of-Concept Study Using Mice

2021 ◽  
Vol 10 (13) ◽  
pp. 2757
Author(s):  
Xia Hao ◽  
Amandine Anastácio ◽  
Kenny A. Rodriguez-Wallberg
Keyword(s):  
Oocyte Maturation ◽  
In Vitro Maturation ◽  
Follicular Development ◽  
Clinical Situation ◽  
Experimental Proof ◽  
Secondary Follicle ◽  
17Β Estradiol ◽  
Follicle Size ◽  
And Control

Fertility preservation through ovarian stimulation, aiming at cryopreserving mature oocytes or embryos, is sometimes unsuccessful. This clinical situation deserves novel approaches to overcome infertility following cancer treatment in patients facing highly gonadotoxic treatment. In this controlled experimental study, we investigated the feasibility of in-vitro culturing secondary follicles isolated from superovulated ovaries of mice recently treated with gonadotropins. The follicle yields of superovulated ovaries were 45.9% less than in unstimulated controls. Follicles from superovulated ovaries showed faster growth pace during the initial 7 days of culture and secreted more 17β-estradiol by the end of culture vs controls. Parameters reflecting the outcome of follicular development and oocyte maturation competence in vitro were similar between superovulated and control groups, with a similar follicle size at the end of culture and around 70% survival. Nearly half of cultured follicles met the criteria for in-vitro maturation in both groups and approximately 60% of those achieved a mature MII oocyte, similarly in both groups. Over 60% of obtained MII oocytes displayed normal-looking spindle and chromosome configurations, without significant differences between the groups. Using a validated follicle culture system, we demonstrated the feasibility of secondary follicle isolation, in-vitro culture and oocyte maturation with normal spindle and chromosome configurations obtained from superovulated mice ovaries.

Salivary oestradiol and progesterone after in vitro fertilization and embryo transfer using different luteal support regimens

1990 ◽  
Vol 2 (4) ◽  
pp. 351 ◽  
Author(s):  
YF Wong ◽  
EP Loong ◽  
KR Mao ◽  
PP Tam ◽  
NS Panesar ◽  
...  
Keyword(s):  
In Vitro Fertilization ◽  
Embryo Transfer ◽  
Luteal Phase ◽  
Biologically Active ◽  
Luteal Support ◽  
Vitro Fertilization ◽  
Group A ◽  
Group B ◽  
Ivf Et

Salivary oestradiol (E2) and progesterone (P) levels have been shown to reflect the biologically active fractions in the serum. The luteal-phase status of stimulated cycles was investigated after in vitro fertilization and embryo transfer (IVF-ET). Thirty patients were randomly allocated to one of three luteal therapy groups: group A had no support, group B had intramuscular P and group C had intramuscular P and human chorionic gonadotrophin (hCG). One pregnancy was achieved in group A, two in group B and three in group C. Significant correlations between salivary and serum levels of E2 and of P in matched samples during luteal phase were found. Salivary E2 levels from luteal day 8 through day 14 and P levels from day 3 through day 14 were significantly higher in the pregnant than in the nonpregnant cycles. Among the nonpregnant cycles, salivary E2 and P levels were significantly higher in group C than in group A or B. These findings suggest that, in stimulated cycles for IVF-ET, determination of salivary E2 and P levels may be used as reliable alternatives to serum concentrations for assessing the luteal phase. Also, the additional hCG has an enhanced luteotrophic effect, as reflected by the higher salivary E2 and P levels, which may lead to a better pregnancy rate.

Varying the patterns and concentrations of gonadotrophin-releasing hormone stimulation does not alter the ratio of LH and FSH released from perifused sheep pituitary cells

1986 ◽  
Vol 109 (2) ◽  
pp. 155-161 ◽  
Author(s):  
J. E. A. McIntosh ◽  
R. P. McIntosh
Keyword(s):  
Dose Response ◽  
Dose Interval ◽  
Follicular Phase ◽  
Pituitary Cells ◽  
Ovulatory Cycle ◽  
Short Term ◽  
Releasing Hormone ◽  
Dissociated Cells ◽  
Gonadotrophin Releasing Hormone

ABSTRACT Our aim was to determine whether release of LH and FSH can be controlled differentially by the characteristics of applied signals of stimulatory gonadotrophin-releasing hormone (GnRH) alone, free of the effects of steroid feedback or other influences from the whole animal. The outputs of both gonadotrophins were significantly correlated (r≈0·90; P<0·0005) when samples of freshly dispersed sheep pituitary cells were perifused in columns for 7 h with medium containing a range of concentrations of GnRH in various patterns of pulses. Hormone released in response to the second, third and fourth pulses from every column was analysed in detail. Dose–response relationships for both LH and FSH were very similar when cells were stimulated with 5–8500 pmol GnRH/1 in 5-min pulses every hour. When GnRH was delivered in pulses at a maximally stimulating level, the outputs of both hormones increased similarly with increasing inter-pulse intervals. Efficiency of stimulation (release of gonadotrophin/unit stimulatory GnRH) decreased (was desensitized) with increasing pulse duration in the same way for both hormones. Thus, varying the dose, interval and duration of GnRH pulses did not alter the proportions of LH and FSH released in the short-term from freshly dissociated cells. However, the same cell preparations released more LH relative to FSH when treated with maximally stimulating levels of GnRH for 3 h in the presence of 10% serum from a sheep in the follicular phase of its ovulatory cycle compared with charcoal-treated serum. Because there was no gonadotrophin synthesis under the conditions used in vitro these results suggest that changes in the LH/FSH ratio seen in whole animals are more likely to result from differential clearance from the circulation, ovarian feedback at the pituitary, differential synthesis in intact tissue or another hormone influencing FSH secretion, rather than from differences in the mechanism of acute release controlled by GnRH. J. Endocr. (1986) 109, 155–161

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