Spontaneous antral follicle formation and metaphase II oocyte from a non-stimulated prepubertal ovarian tissue xenotransplant.

BACKGROUND: Current strategies in cancer treatment have markedly increased the rates of remission and survival for cancer patients, but are often associated with subsequent sterility. While there are various options available to an adult female depending on the patient's particular situation, the only realistic option for preserving fertility in prepubertal females is to cryopreserve ovarian tissue. This is the first report of a morphologically mature oocyte collected from non-stimulated prepubertal ovarian tissue xenotransplants. METHODS: Ovarian tissue from a 6 year old patient suffering from nephroblastoma was removed and cryopreserved for fertility preservation. The frozen-thawed ovarian tissue fragments were xenotransplanted to bilaterally oophorectomized severe combined immunodeficiency (SCID) mice to assess follicle development. RESULTS: Antral follicle formation occurred post-xenotransplantation in a single ovarian fragment without exogenous hormone stimulation. A morphologically maturing oocyte was harvested from these follicles. CONCLUSIONS: Prepubertal human ovarian follicles and oocytes can be matured after xenotransplantation even without exogenous hormone stimulation. These results indicate that tissue collected from prepubertal patients can support fertility in cancer survivors.

Preliminary observations on whole-ovary xenotransplantation as an experimental model for fertility preservation.

Ovarian tissue preservation and retransplantation is a promising strategy to restore fertility in cancer survivors. Ischaemia accompanying ovarian tissue grafting, however, can lead to significant follicle loss. Transplantation of the whole ovary by vascular anastomosis has been considered as an alternative to prevent widespread ischaemic damage. In this study, the feasibility and function of transplanting whole ovary with intact vasculature were evaluated, with the goal of developing a xenograft model for studies using donated human ovaries. Whole-swine ovaries with vascular pedicles were perfused and transplanted as intact ovaries by anastomosis into irradiated ovariectomized nude rats (n = 10). The observation period was between 1 and 4 weeks. Fresh swine ovaries served as controls (n = 10). Ovarian stroma and follicle populations were assessed through histological examination in both transplanted and control ovaries. Most of the transplanted whole ovaries (n = 6) maintained stromal quality and all preantral follicle classes were represented, although follicle numbers decreased compared with fresh control. Four transplanted ovaries were fibrotic after 1-4 weeks within the nude rat. Our results demonstrate transplantation of whole-pig ovary into nude rats is possible and support development of this xenograft model system for human studies.

Miscarriage determination in first trimester based on alpha-fetoprotein extracted from sanitary pads.

OBJECTIVE: To determine if high alpha-fetoprotein (AFP) level in vaginal blood collected on a sanitary pad can assist with detecting an active miscarriage. DESIGN: A prospective cohort study. SETTING: Academic medical center. PATIENT(S): Five groups were evaluated: women with active miscarriage, pregnancy of unknown location, completed miscarriage or extrauterine pregnancy (EUP), ongoing pregnancy, and undergoing elective dilation and curettage (D&C). INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): For each patient, AFP level in the vaginal blood collected on a sanitary pad was quantified. RESULT(S): The vaginal blood AFP median levels (and their ranges) were 3.7 IU/mL (0.5-739.2) and 4,542 IU/mL (15.6-100,000) in the active miscarriage (n = 16) and the elective D&C (n = 24) groups, respectively. Alpha-fetoprotein was detected in all elective D&C and active miscarriage cases except in 1 case. In the ongoing pregnancy group (n = 35), only 2 of 35 specimens showed detectable AFP levels. In the pregnancy of unknown location (n = 12) and the completed miscarriage or EUP (n = 10) groups, no AFP was detected. Receiver operating characteristic analysis demonstrated 93.7% sensitivity and 97.8% specificity for the detection of an active miscarriage (cutoff 0.61 IU/mL; area under the curve 0.96). CONCLUSION(S): Alpha-fetoprotein can be extracted from vaginal blood collected on sanitary pads. A high level of vaginal AFP can assist with the same-day detection of an active miscarriage. This novel test is useful in differentiating active miscarriages from ongoing pregnancies, completed miscarriages, and EUPs and, therefore, it reduces uncertainty, anxiety level, and number of repeat office visits.

A step towards the automation of intracytoplasmic sperm injection: real time confirmation of mouse and human oocyte penetration and viability by electrical resistance measurement.

OBJECTIVE: To evaluate if oocyte penetration and viability can be confirmed by an electrical resistance increase. Automated (robotic) intracytoplasmic sperm injection (ICSI) requires confirmation of oolemma penetration before sperm injection. Visual assessment using image processing algorithms have been developed but remain unreliable. We hypothesized that an increase in electrical resistance upon oolemma piercing during ICSI can serve as an objective tool to confirm oocyte penetration and viability. DESIGN: Experimental study. SETTING: Research laboratory in an academic center. PATIENTS/ANIMALS: Oocytes from female mice and women undergoing oocyte retrieval procedure. INTERVENTION: Oolemma piercing attempts with the ICSI pipette were performed by advancing the pipette towards mature (metaphase II) oocytes collected from 6 to 12-week-old mice and immature (germinal vesicle stage and metaphase I) oocytes donated by women who underwent oocyte retrieval. Electrical resistance was measured using a conventional electrophysiological setup that includes an electrical resistance meter and two electrical wires located in the lumina of the holding and ICSI pipettes. MAIN OUTCOME MEASURE(S): The measure of interest was the change in electrical resistance (ΔR) before and after advancing the ICSI pipette in an attempt to penetrate an oocyte. The experiments of resistance measurements were done in 3 steps: Step 1 (proof of concept), penetrated vs. non-penetrated mouse oocytes. Step 2, mouse oocytes with visually intact oolemma vs. fragmented mouse oocytes. Step 3, human oocytes with visually intact oolemma vs. fragmented human oocytes. For each group, median and range (in parenthesis) of ΔR were determined in MΩ. Mann-Whitney test was performed to compare the two groups in each step. RESULTS: In Step 1, the penetrated mouse oocytes showed a statistically significant resistance increase compared to the non-penetrated ones (n = 20, median ΔR = 7.79 [2.57 - 106.00] vs. n = 15, median ΔR = 0.10 [-0.06 - 0.69], respectively. In Step 2, the mouse oocytes with visually intact oolemma showed a statistically significant resistance increase compared to the fragmented ones (n = 45, median ΔR = 6.5 [0.1 - 191.7] vs. n = 13, median ΔR = 0.1 [-0.3 - 2.2], respectively. In Step 3, the human oocytes with visually intact oolemma showed a statistically significant resistance increase compared to the fragmented ones (n = 96, median ΔR = 1.92 [-0.05 - 6.70] vs. n = 17, median ΔR = 0.11 [0.00 - 0.30], respectively. CONCLUSIONS: An electrical resistance increase can serve as a reliable tool to confirm oocyte penetration and viability, independent of optical visualization. Following further validation and safety assessment, this technology can potentially be integrated into manual and robotic ICSI systems.