GnRHa Pretreatment Improves Ovarian Graft Preservation and Function in Mice

Pretreatment with a gonadotropin-releasing hormone agonist (GnRHa) was associated with preserved primordial follicle density, reduced primordial follicle activation after grafting, and attenuated apoptosis in growing follicles compared with combined oral contraceptive (COC) pretreatment and vitrification-only controls in a mouse model of ovarian tissue vitrification and autotransplantation. Ovarian tissue cryopreservation and transplantation represents a critical fertility preservation strategy, but cryoinjury and subsequent ischemia-reperfusion injury typically cause extensive follicle loss and restrict graft longevity. To address these limitations, researchers evaluated the protective effects of GnRHa versus COC pretreatment during two-step vitrification and autotransplantation, tracking molecular markers related to follicular development, apoptosis, and graft revascularization.

The investigation utilized seventy-two six-week-old female C57BL/6 mice assigned across four pretreatment arms (n = 18 per arm: untreated, GnRHa, COC, or saline vehicle). Animals were further divided into pre-transplantation cohorts (fresh control, GnRHa-pretreated vitrification [GV], COC-pretreated vitrification [CV], and vitrification-only [V]; n = 10 per group) and post-transplantation cohorts (fresh-transplanted [FT], GnRHa-pretreated vitrified-transplanted [GVT], COC-pretreated vitrified-transplanted [CVT], and vitrified-transplanted [VT]; n = 8 per group). Ovaries underwent 14 days of GnRHa or COC pretreatment before two-step vitrification and subrenal capsule autotransplantation. Histomorphology, ultrastructure, and molecular markers were evaluated after warming, while follicular density, Ki-67 immunostaining of primordial follicles, cleaved caspase-3 apoptosis, CD31 graft vascular density, and serum hormone concentrations were measured 4 weeks post-grafting.

Both GnRHa and COC pretreatment significantly reduced serum follicle-stimulating hormone (FSH) and estradiol (E₂) levels after the 14-day administration period. Following tissue warming, significant primordial follicle loss was observed in the CV and V groups, with follicle loss being severest in the vitrification-only group, whereas significant primordial follicle loss did not occur in the GV group. The difference in follicle loss between the GV and CV groups was not statistically significant. Markers associated with folliculogenesis and apoptosis showed no significant differences among groups after warming, though ultrastructural damage was mildest in the GV cohort.

Four weeks after subrenal capsule autotransplantation, the GVT group demonstrated a primordial follicle density not significantly different from the fresh-transplanted control group and significantly higher than the vitrified-transplanted group, while the CVT group exhibited intermediate density. Primordial follicle activation, measured by the proportion of Ki-67-positive primordial follicles, did not differ significantly between the GVT and FT cohorts and was significantly lower in GVT than in the CVT and VT groups. Growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15) protein levels in the GVT group were not significantly different from FT but were significantly reduced in VT. Anti-Müllerian hormone (AMH) protein levels were reduced relative to FT across all vitrified-transplanted groups, but remained highest in the GVT group. Cleaved caspase-3 apoptotic scores in growing follicles remained statistically comparable to fresh controls in the GVT group, while showing significant elevation in the CVT and VT groups. Graft vascular density, assessed by CD31 immunostaining, did not differ significantly among any of the transplanted groups.

Endocrine evaluations at four weeks post-transplantation indicated that serum E₂ and FSH levels did not differ significantly between the GVT and FT groups, whereas endocrine function was significantly impaired in both the CVT and VT groups, with E₂ also significantly lower in GVT than in fresh controls. Serum AMH concentrations declined progressively across the FT, GVT, CVT, and VT groups, remaining significantly higher in GVT than in the CVT and VT cohorts. The authors concluded that GnRHa pretreatment preserved primordial follicular density, decreased post-transplantation primordial follicle activation, and attenuated apoptosis in growing follicles, yielding superior structural and functional graft outcomes relative to COC pretreatment and vitrification alone in mice.

Subject of Research: Ovarian tissue vitrification and autotransplantation in mice

Article Title: GnRHa pretreatment is associated with improved follicular preservation and graft function compared with combined oral contraceptive pretreatment after ovarian tissue vitrification and autotransplantation in mice

Article References: Wang, L., Wu, W., Gao, M., Zhou, Z., Yu, H., & Xu, J. (2026). GnRHa pretreatment is associated with improved follicular preservation and graft function compared with combined oral contraceptive pretreatment after ovarian tissue vitrification and autotransplantation in mice. Journal of Ovarian Research. https://doi.org/10.1186/s13048-026-02329-x

Image Credits: AI Generated

DOI: 10.1186/s13048-026-02329-x

Keywords: ovarian tissue cryopreservation, vitrification, autotransplantation, GnRHa, follicular preservation, primordial follicles, mouse model, ovarian graft function, pretreatment, associated, improved, follicular