Oocyte cryopreservation is offered to women of various age groups for both health and social reasons. Oocytes derived from either controlled ovarian stimulation or in vitro maturation (IVM) are cryopreserved via vitrification. As maternal age is a significant determinant of oocyte quality, there is limited data on the age-related susceptibility of oocytes to the vitrification-warming procedure alone or in conjunction with IVM. In the present study, metaphase II oocytes obtained from 2, 6, 9, and 12 month old Swiss albino mice either by superovulation or IVM were used. To understand the association between maternal age and oocyte cryotolerance, oocytes were subjected to vitrification-warming and compared to non vitrified sibling oocytes. Survived oocytes were evaluated for mitochondrial potential, spindle integrity, relative expression of spindle checkpoint protein transcripts, and DNA double-strand breaks. Maturation potential and vitrification-warming survival were significantly affected ( p < 0.001 and p < 0.05, respectively) in ovulated oocytes from the advanced age group but not in IVM oocytes. Although vitrification-warming significantly increased spindle abnormalities in ovulated oocytes from advanced maternal age ( p < 0.01), no significant changes were observed in IVM oocytes. Furthermore, Bub1 and Mad2 transcript levels were significantly higher in vitrified-warmed IVM oocytes ( p < 0.05). In conclusion, advanced maternal age can have a negative impact on the cryosusceptibility of ovulated oocytes but not IVM oocytes in mice.

Cyclophosphamide (CY) exposure is known to affect the ovary and impair fertility. Clinically, treatment is generally given over multiple doses, but research models have generally used single doses. The relative effects of administering multiple small doses of CY in the prepubertal period are not elucidated. Two-week-old early-prepubertal Swiss albino female mice were administered with either large single (200 mg/Kg x 1; CY200X1) or small multiple (75 mg/Kg x 4; CY75X4) CY doses, thus a 50% higher total dose. Surviving females were assessed for estrous cyclicity, ovarian follicle reserve, oocyte functional competence, and postnatal assessment of first-generation (F1) pups. Exposure to CY75X4 reduced the loss of ovarian follicles (p < 0.05), and body weight (p < 0.001), and resulted in a larger population of cycling females (p < 0.01) with higher oocyte yield (p < 0.05) compared to CY200X1. Although CY200X1 exposed cycling females had comparable oocyte quality, and fertility index, the postnatal mortality was higher in F1 pups (p < 0.05) in comparison to the CY75X4 group. Although both strategies affect oocyte quality and functional competence similarly, CY75X4, despite the higher overall dose, results in reduced follicle loss, produces higher oocyte/blastocyst yield, and exhibits lower postnatal mortality rates, suggesting a potential advantage over CY200X1 for later fertility and offspring health. The differences in effects of the two treatment models show the need for designing animal model studies that more closely mimic the clinical administration of gonadotoxic therapies such as cyclophosphamide.

Background and Aims: Response of preimplantation embryos to varying oxygen tensions, when cultured in vitro, has shown conflicting results. Though physiological or low oxygen culture is gaining popularity in IVF programs, how oxygen tension influences the behavior of embryos derived from pathological conditions is unknown. Since there has been an increasing trend in the incidence of diabetes in children and young adults, this study is aimed to look at the functional ability of embryos derived from experimentally induced hyperglycemic mice at the prepubertal phase of life to varying oxygen tension in vitro. Method: The experimentally induced hyperglycemic state was monitored in the prepubertal phase (3 to 6 weeks), followed by oocyte quality assessment, in vitro maturation potential, mitochondrial status, and spindle integrity analysis. Oocytes were then fertilized and cultured at high/atmospheric ([Formula: see text] 19%) and low/physiological ([Formula: see text]5%) oxygen tension until 96 h in vitro. Embryo quality and developmental progression, blastocyst rate, apoptosis, inner cell mass (ICM) outgrowth proliferation, and expression of Hif1[Formula: see text] were assessed. Results: Prepubertal hyperglycemia did not affect the ovarian reserve significantly; however, the maturation potential (P<0.001), mitochondrial integrity (P<0.01), and meiotic spindle assembly (P<0.05–0.001) in oocytes were altered significantly. Importantly, such oocytes showed better functional ability during the preimplantation phase of development when cultured at low oxygen tension. Hif1[Formula: see text] transcripts in ICM outgrowths were significantly higher (P<0.05) in low oxygen tension compared to atmospheric oxygen tension. Conclusion: Embryo culture in low oxygen tension appears beneficial for embryos derived from hyperglycemic mothers with respect to developmental competence and retaining genetic integrity. ART clinics need to customize their embryo culture strategies based on patient’s physiological and pathological conditions.

Reproductive abnormalities in women with a history of childhood diabetes are believed to be partially attributed to hyperglycemia. Prolonged hyperglycemia can negatively affect ovarian function and fertility during reproductive life. To address this in an experimental setting, the present study used streptozotocin-induced hyperglycemic prepubertal mouse model. The impact of prolonged hyperglycemic exposure during prepubertal life on ovarian function, oocyte quality, and functional competence was assessed in adult mice. The ovarian reserve was not significantly altered; however, the in vitro maturation potential (P < 0.001), mitochondrial integrity (P < 0.01), and meiotic spindle assembly (P < 0.05-0.001) in oocytes were significantly affected in hyperglycemic animals in comparison to control groups. The results from the study suggest that prepubertal hyperglycemia can have adverse effects on the oocyte functional competence and spindle integrity during the reproductive phase of life. Because these changes can have a significant impact on the genetic integrity and developmental potential of the embryos and fetus, the observation warrants further research both in experimental and clinical settings.

Corticosteroids are increasingly being used during the peri-implantation period to treat women with repeated IVF failure and recurrent miscarriage. However, the direct effects of prednisolone (PRDL), one of the commonly used corticosteroids on early embryo development is not understood. To mimic the possible clinical scenario and to understand the embryonic response to direct PRDL exposure, this pilot study was conducted in a mouse model. Cleavage stage embryos exposed to 3 and 30 µM PRDL in vitro were assessed for peri-implantation developmental potential, genetic integrity, inner cell mass (ICM) proliferation and pluripotency markers in the proliferated ICM cells. Exposure to 30 µM PRDL delayed the embryonic progression beyond compaction ( P < 0.05) in comparison to vehicle control and, had reduced total cell number ( P < 0.001) than all other groups. In addition, 30 µM PRDL exposure resulted in poor hatching potential ( P < 0.05) and increased apoptosis in blastocysts ( P < 0.05) compared to 3 µM PRDL. On the other hand, completely formed ICM outgrowths were significantly higher ( P < 0.05) in 3 µM PRDL compared to control. However, no significant differences were observed in the expression of pluripotency genes. In conclusion, the trend observed in embryos exposed to PRDL in vitro provides important information concerning the use of this drug when treating patients at the peri-implantation phase of IVF cycles. However, the clinical value of this observation on human embryo development needs further research.

This study investigated if in vitro maturation (IVM) before or after vitrification would be more successful for prepubertal oocytes. To mimic prepubertal conditions in an experimental setup, oocytes were collected from healthy 14, 21 and 28day old Swiss albino mice. The germinal vesicle (GV) stage oocytes and in vitro matured MII oocytes were subjected to vitrification-warming. Both structural (meiotic spindle morphology, mitochondrial integrity, cortical granules) and functional (sperm zona binding, fertilization) characteristics were assessed in oocytes after warming. This study demonstrated that IVM was more detrimental to prepubertal oocytes than to young adults. Further, vitrification of the IVM oocytes resulted in an increase in the number of abnormal meiotic spindles, a change in the cortical distribution pattern, a reduction in sperm zona binding and the fertilization rate. Importantly, oocyte integrity was better when prepubertal oocytes were vitrified before, rather than after, IVM. The above observations support GV stage vitrification for prepubertal oocytes requiring fertility preservation. Understanding the mechanisms behind the differing outcomes for oocytes from immature females will help in refining current protocol, thereby retaining the oocytes' maximum structural and functional integrity Further investigation is necessary to determine whether human prepubertal oocytes also behave in a similar way. It is to be noted here, with great emphasis, that a major limitation of this study is that the oocytes' abilities were tested only until fertilisation, as a consequence of which the study cannot reveal the developmental potentials of the embryos beyond fertilisation.