Spontaneous in vitro maturation of oocytes prior to ovarian tissue cryopreservation in natural cycles of oncologic patients

Purpose

To evaluate the recovery rate and spontaneous in vitro maturation (IVM) of immature oocytes enclosed within or released from follicles during the processing of ovarian tissue prior to its cryopreservation.

Methods

Thirty-three oncologic patients who had not previously undergone chemo or radiotherapy underwent ovarian tissue cryopreservation (OTC) during natural menstrual cycles. Immature oocytes, enclosed within follicles or released during ovarian cortex processing, were collected and matured spontaneously in vitro for 48 h. Nuclear maturation was assessed every 24 h and the ability of the IVM oocytes to display a normal activation response following parthenogenetic activation was evaluated. The following outcome measures were also evaluated: disease, age, FSH, LH, E2, P4 and AMH serum levels, menstrual cycle day, recovery and spontaneous IVM and parthenogenetic activation rates.

Results

Oocytes recovered per patient were 3.3 ± 0.7 (1.8–4.7 oocytes, 95CI), regardless of the menstrual phase. The mean number of IVM oocytes per patient was 1.3 ± 0.2 oocytes (95CI: 0.8–1.8), regardless of menstrual phase (p = 0.86) and oocyte origin (p = 0.61). Forty-one percent of oocytes extruded the second polar body and formed one pronucleus after parthenogenetic activation.

Conclusion

Twenty-one of the 33 women (63.6 %) requesting OTC produced at least one mature oocyte.     

Retrieval of immature oocytes followed by in vitro maturation and vitrification: a case report on a new strategy of fertility preservation in women with borderline ovarian malignancy

BACKGROUND: We report a novel fertility preservation strategy in a woman with borderline ovarian tumors involving retrieval of immature oocytes, in vitro maturation (IVM) and subsequent cryopreservation. CASE: A 43-year-old woman underwent laparotomy for cystic ovarian masses on day 18 of her menstrual cycle. A diagnosis of bilateral borderline ovarian tumors was made following histological frozen section analysis. Left salpingo-oophorectomy, right ovarian cystectomy, omentectomy and lymph node sampling were performed. All visible follicles on the surface of the removed ovary were aspirated. Four immature oocytes were retrieved and underwent IVM. Three oocytes matured after 48 h and were cryopreserved. CONCLUSION: Immature oocytes can be successfully isolated from the oophorectomy specimen regardless of the day of menstrual cycle, matured in vitro and cryopreserved, providing a possible strategy for fertility preservation in this group of women.     

A novel system for in vitro maturation of human oocytes

Objective: To compare in vitro maturation of cumulus-free oocytes in glucose-free medium (P1) and standard medium (TC199).

Design: Prospective, cohort study.

Setting: Assisted reproductive technology program.

Patient(s): One hundred eight patients undergoing ICSI.

Intervention(s): Germinal vesicle–stage or metaphase I–stage oocytes were allocated to culture with P1 or TC199. Metaphase II oocytes were fixed for immunofluorescence analysis or fluorescence in situ hybridization at 24 or 48 hours (or both). Media were compared by performing conditional logistic regression analysis that controlled for egg-specific factors.

Main Outcome Measure(s): Proportion of mature oocytes and appearance of normal spindle-chromosome cytoarchitecture.

Result(s): At 24 hours, more P1 oocytes than TC199 oocytes reached metaphase II (59.7% vs. 44.9%). At 48 hours, 71.7% of P1 oocytes and 61.0% of TC199 oocytes reached metaphase II, but this difference was not significant. Metaphase II oocytes in P1 were 34.3% more likely than those in TC199 to have a bipolar spindle with aligned chromosomes. Compared with oocytes at the germinal vesicle stage at 0 hour, those at metaphase I at 0 hour were more likely to progress to metaphase II (72.6% vs. 46.1% at 24 hours; 84.1% vs. 60.6% at 48 hours).

Conclusion(s): P1 is superior to TC199 for in vitro maturation of granulosa-free human oocytes.     

卵母细胞体外成熟预防卵巢过度刺激综合征的应用

目的:探讨卵母细胞体外成熟预防卵巢过度刺激综合征(ovarian hyperstim- ulation syndrome,OHSS)的效果。方法:51例IVF长方案超排卵过程中出现OHSS早期征象时,按患者意愿分成两组,实验组于当天注射人绒毛膜促性腺激素(hCG)36h后取卵,经体外培养成熟行体外受精、胚胎移植;对照组则继续按常规用药方案及受精方案行胚胎移植。测定两组出现OHSS早期征象当日的E_2、统计FSH用药天数、用药量、受精率及临床妊娠率。结果:实验组与对照组在出现OHSS早期征象当日的血E_2水平、受精率、优质胚胎率、临床妊娠率无统计学差异;但实验组的FSH总用量、FSH用药天数、获卵数方面显著低于对照组(P<0.05)。实验组1例发生中度OHSS,对照组则发生3例重度OHSS和2例中度OHSS,重度OHSS发生率差异极显著(P<0.01)。结论:超排卵过程中,出现OHSS早期征象时改行卵母细胞体外成熟培养技术可有效地降低OHSS的发生率。     

Clinical implications for immunoscintigraphy in patients with ovarian malignancy: a preliminary study using monoclonal antibody 791T/36

Twelve patients with suspected primary and recurrent carcinoma of the ovary have undergone immunoscintigraphy with an antitumour monoclonal antibody in order to assess the impact of the technique on patient management. Consistent tumour uptake of radiolabelled antibody was visualized in malignant tumours after subtraction of the blood background activity. Eight patients were imaged before surgery and in all of these the sites of uptake visualized on the images agreed with the surgical findings. In one patient with recurrent disease the imaging information was used as an aid for establishing the extent of external beam radiotherapy. A repeat study in this patient 6 months later revealed a reduction in the size of the tumour, which still concentrated labelled antibody and confirmed the viability of repeat investigations. Immunoscintigraphy was capable of providing diagnostic information which may offer an alternative to second-look surgery in these patients.     

低氧培养对未成熟卵体外成熟及胚胎发育潜能的影响

目的:研究辅助生殖技术超促排卵治疗中获得的未成熟卵在低氧培养体系中的发育潜能。方法:卵胞质内单精子显微注射(ICSI)或体外受精-胚胎移植中(IVF-ET)短时受精中挑选出的GV(germinal vesicle)期卵母细胞80枚,MI(metaphase I)期卵母细胞124枚,随机分为实验组和对照组,实验组置于低氧分压(体积分数5%)的三气培养箱中、对照组置于高氧分压(体积分数20%)常规培养箱中进行体外成熟培养(IVM)及随后的囊胚和胚胎培养。IVM后成熟的卵母细胞进行ICSI受精,观察受精率、卵裂率及囊胚形成率。结果:实验组GV期未成熟卵体外培养至MII卵的成熟率、双原核(2PN)率、卵裂率、囊胚形成率及优质囊胚率与对照组无统计学差异(P0.05)。但实验组MI期卵培养至MII卵的成熟率[96.8%(60/62)]、2PN率[80%(48/60)]、囊胚形成率[43.8%(21/48)]及优质囊胚率[20.8%(10/48)]均高于对照组[分别为58.1%(36/62),52.8%(19/36),11.8%(2/17)和0.0%(0/17)],差异有统计学意义(P0.05),而2种培养体系的卵裂率无统计学差异。结论:超促排卵中获得的未成熟卵,特别是MI期裸卵在低氧培养体系中进行体外成熟及后续的胚胎培养,有利于获得优质囊胚。     

Clinical implications for immunoscintigraphy in patients with ovarian malignancy: a preliminary study using monoclonal antibody 791T/36

Summary. Twelve patients with suspected primary and recurrent carcinoma of the ovary have undergone immunoscintigraphy with an anti-tumour monoclonal antibody in order to assess the impact of the technique on patient management. Consistent tumour uptake of radiolabelled antibody was visualized in malignant tumours after subtraction of the blood background activity. Eight patients were imaged before surgery and in all of these the sites of uptake visualized on the images agreed with the surgical findings. In one patient with recurrent disease the imaging information was used as an aid for establishing the extent of external beam radiotherapy. A repeat study in this patient 6 months later revealed a reduction in the size of the tumour, which still concentrated labelled antibody and confirmed the viability of repeat investigations. Immunoscintigraphy was capable of providing diagnostic information which may offer an alternative to second-look surgery in these patients.     

常规体外受精-胚胎移植周期中改行未成熟卵母细胞体外培养的临床研究

目的 探讨常规体外受精-胚胎移植(IVF-ET)周期中改行未成熟卵母细胞体外培养(IVM)的临床疗效果.方法 回顾性分析2008年1月至2009年6月在温州医学院附属第一医院生殖医学中心行常规IVF治疗,在促排卵5-7 d后,双侧卵泡数目≥20个,或用药8-13 d后,卵泡发育缓慢且两侧卵泡数日≥15个,平均最大卵泡直径≤12 mm的155个周期,根据患者意愿,即刻停药取卵、改行IVM治疗60个周期(A组),继续按照IVF常规治疗95个周期(B组).比较两组促性腺激素(Gn)的用量、周期取消率、平均获卵数、受精率、优质胚胎率、移植周期数、卵巢过度刺激综合征(OHSS)的发生率及妊娠结局.结果 周期移植率A组为92%(55/60),B组为63%(60/95),两组比较,差异有统计学意义(P<0.05).平均Gn用昔、平均获卵数、卵裂率、OHSS发生率A组分别为(1030±468)U、(10±6)个、82.2%(231/281)、0,B组分别为(1544±338)U、(14±4)个、94.0%(502/534)、35%(21/60),两组比较,筹异均有统计学意义(P<0.05);两组的受精率和优质胚胎率比较,差异无统计学意义(P>0.05).A组29例临床妊娠,每移植周期的临床妊娠率为53%(29/55),多胎妊娠发生率为14%(4/29);B组28例临床妊娠,每移植周期的临床妊娠率为47%(28/60),多胎妊娠发牛率为32%(9/28);两组比较,差异无统计学意义(P>0.05).结论 常规IVF周期巾改行IVM,不仅可以避免OHSS的发生和减少促排卵药物的使用,还可获得较高的临床妊娠率.     

In vitro maturation of ovine oocytes using different maturation media: effect of human menopausal serum

Objective  

To compare the effect of human menopausal serum with estrous sheep serum, estrous goat serum, ovine follicular fluid and bovine follicular fluid on in vitro maturation, in vitro fertilization and embryo development of sheep oocytes     

Influence of atmospheric versus reduced oxygen concentration on development of human blastocysts in vitro: a prospective study on sibling oocytes

Numerous studies show the beneficial effect of reduced oxygen on the culture of animal embryos in vitro. However, few similar studies have been carried out in humans, and the conclusions from these were contradictory. Using sibling human oocytes, a prospective study was carried out to analyse the effect of 5 and 20% oxygen on prolonged development of embryos. The outcomes measured were fertilization rate and proportion of morphologically optimal embryos, blastocysts and optimal blastocysts developing on day 5. The results were analysed separately for the group of IVF (n = 988 oocytes) and ICSI (n = 928 oocytes) cycles. It was found that low oxygen did not influence fertilization, but in comparison with 20% oxygen, it resulted in a significantly higher proportion of embryos being optimal on day 3 after IVF (59 versus 43.2%; P < 0.001) as well as after ICSI cycles (51.2 versus 28.5%; P < 0.001). In both methods, the lower oxygen concentration improved the blastulation rate (73.2 versus 63.1%; P < 0.05 and 67.4 versus 54.7%; P < 0.001) and increased the proportion of embryos reaching the stage of expanded blastocyst with normal inner cell mass on day 5 (31.1 versus 14.6%; P < 0.001 and 18.9 versus 11.4%; P < 0.01). The ratio of successful embryo development to optimal blastocyst stage on day 5 of culture, calculated for two oxygen concentrations, was 2.1 for IVF and 1.7 for ICSI, in favour of lower oxygen tension.     

Effect of single-oocyte culture system on in vitro maturation and developmental competence in mice

Purpose

To investigate whether single-culture systems influence the quality of in vitro-matured oocytes, we examined the maturation and developmental competence of oocytes obtained by grouped in vitro maturation (IVM) or single IVM.

Methods

In vitro-matured oocytes were obtained using the culture drop (CD) method for the grouped IVM experiments, and the CD and hanging drop (HD) method for the single IVM experiments. To evaluate oocyte developmental competence, we performed in vitro fertilization and culture, and counted the number of blastocysts. To evaluate the oocyte cytoplasmic maturation, we measured the maturation promoting factor (MPF) expression levels.

Results

Oocytes cultured singly had lower maturity and developmental competence than the grouped IVM oocytes. However, enhanced oocyte fertility and blastocyst quality was achieved by the HD single IVM method. Additionally, the MPF activity level increased in all culture methods, compared to the control; however, it lagged behind nuclear maturation.

Conclusions

These results suggest that the HD method is efficient for single IVM.     

In vitro maturation (IVM) of oocytes in patients with resistant ovary syndrome and in patients with repeated deficient oocyte maturation

Purpose

To assess the efficiency of IVM in patients with repeated ART failure due to resistant ovary syndrome or due to deficient oocyte maturation.

Methods

Clinical and laboratory data were obtained retrospectively from 28 patients who underwent 49 cycles of IVM between 2010 and 2017; nine patients had resistant ovary syndrome and 19 patients had repeated deficient oocyte maturation.

Results

Nine patients with resistant ovary syndrome underwent 24 IVM cycles. In those, an average of 11.5?±?10.4 cumulus-oocyte complexes (COC) was retrieved, and IVM resulted in 3.4?±?3.1 mature oocytes. After ICSI and transfer of 23 cleavage-stage embryos, eight pregnancies were obtained, resulting in five healthy live births. The live birth rate was 16.7% per started cycle and 33.3% per patient.Nineteen patients with a history of deficient oocyte maturation underwent 25 IVM cycles. An average of 10.6?±?9.2 COC was retrieved, and after IVM, 1.3?±?2.1 oocytes were mature. No mature oocytes were obtained in 11 cycles. In ten cycles with mature oocytes, none of them fertilized after ICSI. Out of four cycles with fertilized oocytes, only one good-quality embryo was obtained. No live births were obtained after IVM in patients with a history of deficient oocyte maturation.

Conclusions

Based on our experience, IVM is a valuable approach in patients with resistant ovary syndrome, but should not be recommended for patients with deficient oocyte maturation.

     

In vitro maturation of oocytes as a strategy for fertility preservation

In vitro fertilization and embryo cryopreservation are regarded as the only established method for the preservation of fertility in female cancer patients. However, a possible delay in the treatment of cancer and exposure to supraphysiologic estrogen levels caused by ovarian stimulation raise concerns for patients and physicians. In vitro maturation avoids treatment delay or exposure to increased estradiol levels associated with in vitro fertilization. In vitro maturation combined with embryo or oocyte vitrification provides options that have been unavailable earlier, such as immature oocyte collection in the luteal phase, for some patients and improves the services provided by a fertility preservation program.     

In vitro maturation of oocytes: an option for fertility preservation in women

Although female cancer incidence may be on rise, antineoplastic regimens have become more successful. As a result, an increasing number of women with cancer survive to endure the long-term consequences of chemotherapy. One of the most important long-term consequences of cancers treatments in young female is premature ovarian failure and infertility. Because of the increasing survival rates, many of these young women are seeking methods to preserve their fertility. Currently, embryo/oocytes cryoporeservation obtained after ovarian stimulation appears to provide the best fertility preservation option. However, patients may not have sufficient time to undergo ovarian stimulation prior to chemotherapy and/or the hormones used in ovarian stimulation are contra-indicated for estrogen-dependant tumors. In vitro maturation of oocytes (IVM) has been suggested to avoid ovarian stimulation and time requirement in patients with cancer, and can be combined with ovarian tissue cryobanking. In this review, we will discuss the position of IVM in the strategy of fertility preservation in young women.