Lifestyle and fertility-specific quality of life affect reproductive outcomes in couples undergoing in vitro fertilization.

Objective: A Mediterranean dietary pattern, sleeping habits, physical activity, and lifestyle appear to affect reproductive health. There are few reports about whether fertility-specific quality of life (QOL) is linked to infertility treatment outcomes. The aim of this study is to investigate when lifestyle factors and fertility-specific QOL are comprehensively considered, which factors influence assisted reproductive technology (ART) outcomes. Methods: This prospective cohort includes 291 women undergoing a first ART treatment at multiple centers in Japan and was designed to evaluate the influence of diet, physical activity, sleeping pattern, computer use duration, and fertility-specific quality of life tool (FertiQoL) score on ART treatment outcomes using a questionnaire. The primary endpoint was the good-quality blastocyst rate per oocyte retrieval and the secondary endpoints were a positive pregnancy test and gestational sac (GS) detection. Results: The good-quality blastocyst rate per oocyte retrieval tended to be negatively associated with frequent fish consumption. After all embryo transfer (ET) cycles, a positive pregnancy test tended to be positively associated with longer sleep and longer computer use (OR = 1.6, 95% CI = 0.9-2.7 and OR = 1.7, CI = 1.0-2.8, respectively) and negatively associated with a smoking partner (OR = 0.6, CI = 0.3-1.0). GS detection was positively and significantly associated with frequent olive oil intake and longer computer use (OR = 1.7, CI = 1.0-3.0 and OR = 1.7, CI = 1.0-3.0, respectively). After ET cycles with a single blastocyst, a positive pregnancy test was positively and significantly associated with longer computer use (OR = 2.0, CI = 1.1-3.7), while GS detection was significantly more likely in women with longer computer use (OR = 2.1, CI = 1.1-3.8) and tended to be more likely in women with a higher FertiQoL Total scaled treatment score (OR = 1.8, CI = 1.0-3.3). p < 0.05 was considered statistically significant and 0.05 ≤ p <0.01 as tendency. Conclusions: Olive oil may be an important factor in dietary habits. Fertility-specific QOL and smoking cessation guidance for partners are important for infertile couples.

Exploring the Impact of Endometrioma Aspiration and Dienogest Combination Therapy on Cyst Size, Inflammatory Cytokines in Follicular Fluid and Fertility Outcomes.

Endometriomas (chocolate cysts) are cystic lesions that can develop on ovaries, and are characterized by the presence of ectopic endometrial tissue or similar tissue. Such lesions can cause a decline in the number and quality of oocytes, and lead to implantation failure. In this study, we retrospectively assessed the efficacy of repeated endometrioma aspiration and dienogest combination therapy in patients suffering endometriosis-associated infertility with endometriomas. A comparison was made between a treated group that underwent combination therapy followed by controlled ovarian hyperstimulation (COH) (n = 30) and a control group that did not undergo treatment (n = 40), at the IVF Osaka Clinic from September 2019 to September 2021. There were no differences in patient background between the two groups. A reduction in endometrioma size continued for 12 months after treatment. The numbers of follicles that developed to 15 mm or greater in size following COH and mature oocytes were significantly lower in the treated group compared to those in the control group. The levels of inflammatory cytokines in the follicular fluid significantly decreased in the treated group (p < 0.05). In patients in the treated group who underwent a second ova retrieval, the results were compared between those in the first ova retrieval (immediately after the end of treatment) and those in the second ova retrieval (four months after the first retrieval). The numbers of follicles following COH, retrieved, mature and fertilized ova were significantly increased in the second ova retrieval.

Exogenous GSH Supplementation to Raw Semen Alters Sperm Kinematic Parameters in Infertile Patients.

Glutathione is an important antioxidant found in all mammalian cells. Sperm motility is positively correlated with seminal reduced glutathione (GSH) levels, and infertile men are known to have lower GSH levels. Studies on GSH supplementation in improving sperm functions in infertility patients are limited. Here, we re-investigate the effect of exogenous GSH supplementation on human sperm motility and kinematic parameters. Residual semen samples from 71 infertility patients who came for routine semen analysis for infertility assessment were studied. Liquefied raw semen was supplemented with GSH (0-10 mM) for 1 h. The untreated sample was the blank control. Only a 5 mM concentration was tested in all 71 samples. After two washes, the sperm was incubated and then analyzed for sperm motility and kinematic parameters by computer-assisted semen analysis (CASA), followed by adenosine triphosphate (ATP), reactive oxygen species (ROS) levels, free thiols, and DNA damage analyses. At 2 hrs post-treatment, GSH supplementation significantly altered many of the kinematics, compared to the control. Straight line velocity (VSL) (p = 0.0459), curvilinear velocity (VCL) (p < 0.0001), average path velocity (VAP) (p < 0.0001), and lateral head amplitude (ALH) (p < 0.0001) were decreased, whereas straightness (STR) (p = 0.0003), linearity (LIN) (p = 0.0008), and beat cross frequency (BCF) (p = 0.0291) were increased in 5 mM group. Wobble (WOB) (p = 0.4917), motility (MOT) (p = 0.9574), and progressive motility (PROG) (p = 0.5657) were unchanged. ATP level was significantly increased in the 5 mM group (p < 0.05). It is concluded that exogenous GSH supplementation does alter sperm kinematics in humans. These altered kinematic parameters together with increased energy (ATP) may have a positive role in influencing the success rates of ART procedures.

Mitochondria Transfer from Adipose Stem Cells Improves the Developmental Potential of Cryopreserved Oocytes.

Although it is not a well-established technology, oocyte cryopreservation is becoming prevalent in assisted reproductive technologies in response to the growing demands of patients' sociological and pathological conditions. Oocyte cryopreservation can adversely affect the developmental potential of oocytes by causing an increase in intracellular oxidative stresses and damage to the mitochondrial structure. In this study, we studied whether autologous adipose stem cell (ASC) mitochondria supplementation with vitrified and warmed oocytes could restore post-fertilization development that decreased due to mitochondrial damage following cryopreservation. ASC mitochondria showed similar morphology to oocytes' mitochondria and had a higher ATP production capacity. The vitrified-warmed oocytes from juvenile mice were supplemented with ASC mitochondria at the same time as intracellular sperm injection (ICSI), after which we compared their developmental capacity and the mitochondria quality of 2-cell embryos. We found that, compared to their counterpart, mitochondria supplementation significantly improved development from 2-cell embryos to blastocysts (56.8% vs. 38.2%) and ATP production in 2-cell embryos (905.6 & 561.1 pmol), while reactive oxygen species levels were comparable. With these results, we propose that ASC mitochondria supplementation could restore the quality of cryopreserved oocytes and enhance the embryo developmental capacity, signifying another possible approach for mitochondrial transplantation therapy.

Stoichiometric CT number calibration using three-parameter fit model for ion therapy.

PURPOSE: Treatment planning for ion therapy involves the conversion of computed tomography number (CTN) into a stopping-power ratio (SPR) relative to water. The purpose of this study was to create a CTN-to-SPR calibration table using a stoichiometric CTN calibration model with a three-parameter fit model for ion therapy, and to demonstrate its effectiveness by comparing it with a conventional stoichiometric CTN calibration model. METHODS: We inserted eight tissue-equivalent materials into a CTN calibration phantom and used six CT scanners at five radiotherapy institutes to scan the phantom. We compared the theoretical CTN-to-SPR calibration tables created using the three-parameter fit and conventional models to the measured CTN-to-SPR calibration table in three tissue types: lung, adipose/muscle, and cartilage/spongy bone. We validated the estimated SPR differences in all cases and in a worst-case scenario, which revealed the largest estimated SPR difference in lung tissue. RESULTS: For all cases, the means ± standard deviations of the estimated SPR difference for the three-parameter fit method model were -0.1 ± 1.0%, 0.3 ± 0.7%, and 2.4 ± 0.6% for the lung, adipose/muscle, and cartilage/spongy bone, respectively. For the worst-case scenario, the estimated SPR differences of the conventional and the three-parameter fit models were 2.9% and -1.4% for the lung tissue, respectively. CONCLUSIONS: The CTN-to-SPR calibration table of the three-parameter fit model was consistent with that of the measurement and decreased the calibration error for low-density tissues, even for the worst-case scenario.

Use of a vaginal probiotic suppository and antibiotics to influence the composition of the endometrial microbiota.

The aim of the present study was to determine the status of the endometrial microbiota in patients with repeated implantation failure (RIF) and to assess treatment strategies for nondominant Lactobacillus (NLD) cases. A total of 392 patients with RIF were enrolled in this prospective cohort study (UMIN-CTR 000038582) and underwent endometrial microbiota analysis. Patients diagnosed with NLD were treated with a combination of oral and vaginal probiotics or oral prebiotics and antibiotics. The outcome was evaluated through re-analysis of the endometrial microbiota following treatment, and the results are presented as cure rates. NLD represented 44.9 % of the total endometrial microbiota in patients with RIF. The most commonly detected bacterium was Gardnerella vaginalis. The cure rates in the oral probiotics + oral prebiotics, antibiotics, oral probiotics + oral prebiotics + antibiotics, vaginal probiotic suppository, and vaginal probiotic suppository + antibiotics groups were 29.5, 33.33, 33.33, 43.6, and 78.6 %, respectively. Significant improvements were noted in the vaginal probiotic suppository + antibiotics group. Moreover, we revealed that approximately half of patients with RIF had NLD. Thus, the combination of a vaginal probiotic suppository and antibiotics may represent an effective treatment for NLD cases.

Development of a CT number calibration audit phantom in photon radiation therapy: A pilot study.

PURPOSE: In photon radiation therapy, computed tomography (CT) numbers are converted into values for mass density (MD) or relative electron density to water (RED). CT-MD or CT-RED calibration tables are relevant for human body dose calculation in an inhomogeneous medium. CT-MD or CT-RED calibration tables are influenced by patient imaging (CT scanner manufacturer, scanning parameters, and patient size), the calibration process (tissue-equivalent phantom manufacturer, and selection of tissue-equivalent material), differences between tissue-equivalent materials and standard tissues, and the dose calculation algorithm applied; however, a CT number calibration audit has not been established. The purposes of this study were to develop a postal audit phantom, and to establish a CT number calibration audit process. METHODS: A conventional stoichiometric calibration conducts a least square fit of the relationships between the MD, material weight, and measured CT number, using two parameters. In this study, a new stoichiometric CT number calibration scheme has been empirically established, using three parameters to harmonize the calculated CT number with the measured CT number for air and lung tissue. In addition, the suitable material set and the minimal number of materials required for stoichiometric CT number calibration were determined. The MDs and elemental weights from the International Commission on Radiological Protection Publication 110 were used as standard tissue data, to generate the CT-MD and CT-RED calibration tables. A small-sized, CT number calibration phantom was developed for a postal audit, and stoichiometric CT number calibration with the phantom was compared to the CT number calibration tables registered in the radiotherapy treatment planning systems (RTPSs) associated with five radiotherapy institutions. RESULTS: When a least square fit was performed for the stoichiometric CT number calibration with the three parameters, the calculated CT number showed better agreement with the measured CT number. We established stoichiometric CT number calibration using only two materials because the accuracy of the process was determined not by the number of used materials but by the number of elements contained. The stoichiometric CT number calibration was comparable to the tissue-substitute calibration, with a dose difference less than 1%. An outline of the CT number calibration audit was demonstrated through a multi-institutional study. CONCLUSIONS: We established a new stoichiometric CT number calibration method for validating the CT number calibration tables registered in RTPSs. We also developed a CT number calibration phantom for a postal audit, which was verified by the performances of multiple CT scanners located at several institutions. The new stoichiometric CT number calibration has the advantages of being performed using only two materials, and decreasing the difference between the calculated and measured CT numbers for air and lung tissue. In the future, a postal CT number calibration audit might be achievable using a smaller phantom.

Tolerance levels of mass density for CT number calibration in photon radiation therapy.

Computed tomography (CT) data are required to calculate the dose distribution in a patient's body. Generally, there are two CT number calibration methods for commercial radiotherapy treatment planning system (RTPS), namely CT number-relative electron density calibration (CT-RED calibration) and CT number-mass density calibration (CT-MD calibration). In a previous study, the tolerance levels of CT-RED calibration were established for each tissue type. The tolerance levels were established when the relative dose error to local dose reached 2%. However, the tolerance levels of CT-MD calibration are not established yet. We established the tolerance levels of CT-MD calibration based on the tolerance levels of CT-RED calibration. In order to convert mass density (MD) to relative electron density (RED), the conversion factors were determined with adult reference computational phantom data available in the International Commission on Radiological Protection publication 110 (ICRP-110). In order to validate the practicability of the conversion factor, the relative dose error and the dose linearity were validated with multiple RTPSes and dose calculation algorithms for two groups, namely, CT-RED calibration and CT-MD calibration. The tolerance levels of CT-MD calibration were determined from the tolerance levels of CT-RED calibration with conversion factors. The converted RED from MD was compared with actual RED calculated from ICRP-110. The conversion error was within ±0.01 for most standard organs. It was assumed that the conversion error was sufficiently small. The relative dose error difference for two groups was less than 0.3% for each tissue type. Therefore, the tolerance levels for CT-MD calibration were determined from the tolerance levels of CT-RED calibration with the conversion factors. The MD tolerance levels for lung, adipose/muscle, and cartilage/spongy-bone corresponded to ±0.044, ±0.022, and ±0.045 g/cm3 , respectively. The tolerance levels were useful in terms of approving the CT-MD calibration table for clinical use.

Mitochondrial function in immature bovine oocytes is improved by an increase of cellular cyclic AMP.

Although in vitro maturation (IVM) of oocytes is important for assisted reproduction, the rate of development of embryos from IVM oocytes is lower than from their in vivo counterparts. It has been shown that an artificial increase of intracellular cAMP before culture significantly improves oocyte developmental competence in cattle and mice. Here, we revealed that forskolin and 3-isobutyl-1-methylxanthine treatment of prophase-stage oocytes induced the expression of genes required for glycolysis, fatty acid degradation, and the mitochondrial electron transport system and improved mitochondrial functions and ATP levels in oocytes without involving nuclear maturation. We propose the existence of a comprehensive energy-supply system in oocytes under follicle-stimulating hormone stimulation as a potential explanation of how oocytes acquire developmental competence.

Influence of Negative Emotional Expressions on the Outcomes of Shared Decision Making During Oncofertility Consultations in Japan.

This report examines how negative emotional expressions (NEE) influence the consequences of shared decision making (SDM) in oncofertility treatment among 32 young female cancer patients and 19 family members. Using a cross-sectional observational study, results showed that NEE influence the outcome consequences of SDM related to patients' decisions about desired treatment(s) and that the absence of negative emotional reactions to information from doctors was related to willingness to receive the desired treatment. This suggests that healthcare providers need to be sensitive to NEE of patients and their families, and highlights the need for psychological counseling before oncofertility consultation.

Tolerance levels of CT number to electron density table for photon beam in radiotherapy treatment planning system.

The accuracy of computed tomography number to electron density (CT-ED) calibration is a key component for dose calculations in an inhomogeneous medium. In a previous work, it was shown that the tolerance levels of CT-ED calibration became stricter with an increase in tissue thickness and decrease in the effective energy of a photon beam. For the last decade, a low effective energy photon beam (e.g., flattening-filter-free (FFF)) has been used in clinical sites. However, its tolerance level has not been established yet. We established a relative electron density (ED) tolerance level for each tissue type with an FFF beam. The tolerance levels were calculated using the tissue maximum ratio (TMR) and each corresponding maximum tissue thickness. To determine the relative ED tolerance level, TMR data from a Varian accelerator and the adult reference computational phantom data in the International Commission on Radiological Protection publication 110 (ICRP-110 phantom) were used in this study. The 52 tissue components of the ICRP-110 phantom were classified by mass density as five tissues groups including lung, adipose/muscle, cartilage/spongy-bone, cortical bone, and tooth tissue. In addition, the relative ED tolerance level of each tissue group was calculated when the relative dose error to local dose reached 2%. The relative ED tolerances of a 6 MVFFF beam for lung, adipose/muscle, and cartilage/spongy-bone were ±0.044, ±0.022, and ±0.044, respectively. The thicknesses of the cortical bone and tooth groups were too small to define the tolerance levels. Because the tolerance levels of CT-ED calibration are stricter with a decrease in the effective energy of the photon beam, the tolerance levels are determined by the lowest effective energy in useable beams for radiotherapy treatment planning systems.

Successful fertility preservation following ovarian tissue vitrification in patients with primary ovarian insufficiency.

STUDY QUESTION: Is ovarian tissue cryopreservation using vitrification followed by in vitro activation (IVA) of dormant follicles a potential approach for infertility treatment of patients with primary ovarian insufficiency (POI)? SUMMARY ANSWER: Our vitrification approach followed by IVA treatment is a potential infertility therapy for POI patients whose ovaries contain residual follicles. WHAT IS KNOWN ALREADY: Akt (protein kinase B) stimulators [PTEN (phosphatase with TENsin homology deleted in chromosome 10) inhibitor and phosphatidyinositol-3-kinase (PI3 kinase) stimulator] activate dormant primordial follicles in vitro and ovarian fragmentation disrupts the Hippo signaling pathway, leading to the promotion of follicle growth. We treated POI patients with a combination of ovarian vitrification, fragmentation and drug treatment, followed by auto-transplantation, and reported successful follicle growth and pregnancies. STUDY DESIGN, SIZE, DURATION: Prospective clinical study of 37 infertile women with POI between 12 August 2011 and 1 November 2013. We enrolled 10 new patients since the previous publication. PARTICIPANTS/MATERIALS, SETTING, METHODS: POI patients were originally selected based on a history of amenorrhea for more than 1 year and elevated serum FSH levels of >40 mIU/ml (n = 31) but this was later changed to >4 months, age <40 years and serum FSH levels of >35 mIU/ml (n = 6) (mean 71.8 ± 30.8, range 35.5-197.6) so as to include patients with a shorter duration of amenorrhea. Under laparoscopic surgery, ovariectomy was performed and ovarian cortices were dissected into strips for vitrification. Some pieces were examined histologically. After warming, two to three strips were fragmented into smaller cubes before culturing with Akt stimulators for 2 days. After washing, ovarian cubes were transplanted beneath the serosa of Fallopian tubes under laparoscopic surgery. Follicle growth was monitored by ultrasound and serum estrogen levels. After oocyte retrieval from mature follicles, IVF was performed. MAIN RESULTS AND THE ROLE OF CHANCE: Among 37 patients, 54% had residual follicles based on histology. Among patients with follicles, 9 out of 20 showed follicle growth in auto-grafts with 24 oocytes retrieved from six patients. Following IVF and embryo transfer into four patients, three pregnancies were detected based on serum hCG, followed by one miscarriage and two successful deliveries. For predicting IVA success, we found that routine histological analyses of ovarian cortices and shorter duration from initial POI diagnosis to ovariectomy are valid parameters. LIMITATIONS, REASONS FOR CAUTION: Although our findings suggest that the present vitrification protocol is effective for ovarian tissue cryopreservation, we have not compared the potential of vitrification and slow freezing in follicle growth after grafting. We chose the serosa of Fallopian tubes as the auto-grating site due to its high vascularity and the ease to monitor follicle growth. Future studies are needed to evaluate the best auto-grafting sites for ovarian tissues. Also, future studies are needed to identify biological markers to indicate the presence of residual follicles in POI to predict IVA treatment outcome. WIDER IMPLICATIONS OF THE FINDINGS: In POI patients, ovarian reserve, namely the pool of residual follicles, continues to diminish with age. If one ovary is cryopreserved at an earlier stage of POI, patients could undergo additional non-invasive infertility treatments before the final decision for the IVA treatment. Furthermore, in the cases of unmarried POI patients, cryopreservation of ovarian tissues allows their fertility preservation until they desire to bear children. STUDY FUNDING/COMPETING INTERESTS: This work was supported by Grant-In-Aid for Scientific Research (Research B: 24390376, Challenging Exploratory Research: 24659722, and Innovative Areas, Mechanisms regulating gamete formation in animals: 26114510) and by research funds from the Smoking Research Foundation, and the Takeda Science Foundation. None of the authors has a conflict of interest. TRIAL REGISTRATION NUMBER: UMIN000010828.

Hippo signaling disruption and Akt stimulation of ovarian follicles for infertility treatment.

Primary ovarian insufficiency (POI) and polycystic ovarian syndrome are ovarian diseases causing infertility. Although there is no effective treatment for POI, therapies for polycystic ovarian syndrome include ovarian wedge resection or laser drilling to induce follicle growth. Underlying mechanisms for these disruptive procedures are unclear. Here, we explored the role of the conserved Hippo signaling pathway that serves to maintain optimal size across organs and species. We found that fragmentation of murine ovaries promoted actin polymerization and disrupted ovarian Hippo signaling, leading to increased expression of downstream growth factors, promotion of follicle growth, and the generation of mature oocytes. In addition to elucidating mechanisms underlying follicle growth elicited by ovarian damage, we further demonstrated additive follicle growth when ovarian fragmentation was combined with Akt stimulator treatments. We then extended results to treatment of infertility in POI patients via disruption of Hippo signaling by fragmenting ovaries followed by Akt stimulator treatment and autografting. We successfully promoted follicle growth, retrieved mature oocytes, and performed in vitro fertilization. Following embryo transfer, a healthy baby was delivered. The ovarian fragmentation-in vitro activation approach is not only valuable for treating infertility of POI patients but could also be useful for middle-aged infertile women, cancer patients undergoing sterilizing treatments, and other conditions of diminished ovarian reserve.

Cysteamine supplementation during in vitro maturation (IVM) of rabbit oocyte improves the developmental capacity after intracytoplasmic sperm injection.

PURPOSE: Current approaches to in vitro maturation (IVM) may result in low efficiency and inadequate quality of the oocytes due to insufficient cytoplasmic maturation. Although positive effects of the cysteamine supplementation in IVM medium for oocyte nuclear maturation or male pronuclear formation have been confirmed, it is still controversial whether the cysteamine addition affects embryo development after IVM. We aimed here to confirm the effect of cysteamine addition into IVM medium for subsequent embryo development in vitro. METHODS: We administered the cysteamine to the IVM culture of rabbit immature oocytes at various concentrations and observed the developmental rate, speed to reach blastocyst stage and cell numbers at the blastocyst stage. RESULTS: Cysteamine supplementation improved developmental rate to blastocyst stage of the IVM oocytes. On the other hand, addition of glutathione (GSH) inhibitor buthionine sulfoximine inhibited GSH accumulation in the oocytes and subsequent embryo development to the blastocyst stage. CONCLUSIONS: Controlling the GSH quantity of IVM oocytes may be an important factor for success of embryo development, and it is quite probable that a cysteamine supplementation can contribute to an increase of GSH content in oocyte.

[The possibility of the fertility preservation in young cancer patients based on practice of oncofertility treatment].

Young female cancer patients face various problems, including a decrease in their quality of life(QOL)due to early menopause or loss of fertility after remission. Chemotherapy and radiotherapy can cause loss of reproductive function in young women due to adverse effects such as ovarian failure. The frequency of ovarian failure depends on the age of the patient, the anticancer agents used, and the dose of each agent. In these patients, improvement of the post-treatment QOL and fertility preservation can be achieved by measures such as protection of ovarian function against the effects of anticancer agents. Ovum freezing or fertilized egg freezing are also becoming fertility preservation methods for these patients. However, ovarian hyperstimulation to obtain ova is time consuming and sometimes considered taboo depending on the type of cancer. A self solution to problems occurring frequently at the same time is demanded from the patient, and the patient is forced to deal with too many choices in too little time. It is often less than one month until the cancer treatment begins after an underlying disease is diagnosed, since chemotherapy cannot be delayed. In these cases, cryopreservation of ovarian tissue is currently proposed for fertility preservation. In this manuscript, I will discuss a topic about fertility preservation in young cancer survivors including recent knowledge regarding cryopreservation of ovarian tissue.

Cryopreservation of female germ cells and ovarian tissues for fertility preservation.

To preserve the fertility of patients who undergo chemotherapy and/or radiotherapy, procedures for cryopreservation of female germ cells have been investigated. Cyropreservation methods differ according to follicle stage because the mammalian ovary contains a large number of oocytes at different growth stages. Follicles at very early stages, for example the primordial and primary stages, are usually cryopreserved within ovarian cortical tissue because they need surrounding somatic cells for subsequent development. In contrast, fully-grown oocytes in Graafian follicles are cryopreserved without any other cells at the metaphase II stage. Recently, ultra-rapid cooling was incorporated into cryopreservation procedures for human ovaries. In this review, we describe oocyte freezing, the development of ultra-rapid cooling systems for ovarian tissues, freezing of human ovaries, and ovarian transplantation.

Effects of vitrification solutions and equilibration times on the morphology of cynomolgus ovarian tissues.

This study assessed the effects of vitrification solutions and equilibration times on morphology of cynomolgus ovarian tissues. Ovarian cortical sections (0.1-0.2 cm thickness) of seven cynomolgus monkeys were randomly allocated to either a control group or one of six vitrification groups. Ovarian tissue sections were vitrified ultra-rapidly by placing them directly into liquid nitrogen using two different vitrification solutions (VSEGP: 5.64 mol/l ethylene glycol+5% (w/v) polyvinylpyrrolidone+0.5 mol/l sucrose; and VSED: 3.22 mol/l ethylene glycol+2.56 mol/l dimethylsulphoxide+0.5 mol/l sucrose) after three different exposure times (5-20 min). After warming, follicle morphology was analysed using light and transmission electron microscopy. The proportion of morphologically normal follicles vitrified using VSED after a 5-min exposure was lower (P<0.05) than those vitrified by other conditions. The proportion of normally structured mitochondria in oocytes of preantral follicles vitrified after a 5-min exposure to VSED (56%) was lower (P<0.01) than those vitrified by other conditions (78-88%). Following tissue vitrification with VSED, the surface ratio of lysosome was increased compared with non-vitrified oocytes (1.64% versus 1.11%; P<0.05). These results indicate that VSEGP can support the morphology of vitrified preantral follicles and oocytes.

Heterotopic autotransplantation of ovarian cortex in cynomolgus monkeys.

Abstract In recent years, removal of ova or ovaries before chemotherapy or radiation therapy has been investigated in young female cancer patients to avoid the adverse effects of treatment. Orthotopic autotransplantation of ovarian cortex has advantages such as easy collection of ova and the possibility of spontaneous pregnancy. Although children have been born after successful orthotopic autotransplantation into the residual ovaries, some patients cannot undergo this procedure such as those who need bilateral ovariectomy or pelvic radiation therapy, therefore it is still necessary to investigate suitable heterotopic autotransplantation sites. The present study was performed in primates (cynomolgus monkeys) with the objective of determining the optimum site for heterotopic autotransplantation of ovarian cortex to enhance the clinical application of this method. The retroperitoneal iliac fossa and omentum were selected as sites for heterotopic autotransplantation. Two cynomolgus monkeys were subjected to laparotomy under anesthesia. After resection of the bilateral adnexae, the ovaries were cut into 0.5 cm cubes that were transplanted. Blood levels of follicle-stimulating hormone, luteinizing hormone, estradiol, and progesterone were monitored, and monkeys with a regular estrus cycle underwent superovulation and egg collection. In both monkeys studied, recovery of a regular estrus cycle was confirmed after heterotopic autotransplantation of ovarian tissue. MII phase ova were successfully collected from tissues transplanted into the retroperitoneal iliac fossa or omentum. Development to the early blastocyst stage was confirmed after microfertilization. We established an appropriate method of heterotopic autotransplantation using ovarian cortex into the retroperitoneal iliac fossa or omentum in primates.

Effect of aspiration vacuum on the developmental competence of immature human oocytes retrieved using a 20-gauge needle.

In-vitro maturation (IVM) of immature oocytes has been proposed as a potential alternative to conventional IVF treatment following ovarian stimulation. However, the effects of the oocyte retrieval conditions on subsequent development have not been well understood. This study assessed the effects of different aspiration vacuums during oocyte retrieval on the developmental competence of immature oocytes following IVM, IVF and embryo transfer, retrospectively. Immature oocytes were aspirated with 20-gauge needles with a vacuum of 180 or 300 mmHg. Immature oocytes were cultured in IVM medium for 26 h. All mature oocytes were inseminated by intracytoplasmic sperm injection (ICSI). Embryo transfer was carried out 2 or 3 days after ICSI. The percentage of cumulus-cell enclosed oocytes and of transferable embryos per retrieved oocytes in 180 mmHg (69.7% and 23.8%, respectively) were significantly higher (P < 0.01) than those in 300 mmHg (46.2% and 12.8%, respectively). The ongoing pregnancy rate per retrieval cycle in 180 mmHg (30%) was higher (P < 0.01) than that in 300 mmHg (4.3%). The data indicate that lower pressure of vacuum aspiration with a 20-gauge needle improves the developmental competence of immature oocytes following IVM, IVF and embryo transfer.

Effect of different sites for cryopreserved ovarian tissue implantation in rabbit.

BACKGROUND: Autotransplantation of frozen-thawed ovarian tissue has proven to be an effective method to restore endocrine function and fertility. But it remains to be studied which site and which method is most effective and practical. We therefore implanted small pieces of cryopreserved ovarian tissues into different sites in rabbits to find the optimal position. METHODS: Fifteen New Zealand white female rabbits were randomly divided into three groups. In group 1, fresh ovarian tissues were implanted into the mesometrium and ovarian bursa. In group 2, cryopreserved ovarian tissues were implanted into the mesometrium and ovarian bursa. In group 3, cryopreserved ovarian tissues were implanted into the preserved ovary. RESULTS: There were no significant differences among the three groups as to the proportions of normal and morphologically changed follicles in implanted ovarian tissues. The implanted ovarian tissues in the three groups did not show any evident changes in histology and ultrastructure, and all resumed follicle development and revealed maturescent follicles. CONCLUSIONS: Cryopreservation and implantation of small pieces of ovarian tissues are feasible. Generally, the mesometrium, ovarian bursa and ovary are all available sites for implantation and have similar rates of acceptance, despite some differences in the details of implantation.