P-396 Improving post-ovarian transplant primordial follicle survival by neovascularizing approaches
S Celik, M Erden, N Y Rho, W Molla, K H OktayAbstract
Study question
Can post-ovarian-grafting ischemic primordial follicle loss be reduced with neovascularizing agents FTY-720 (sphingosine-1-phosphate analogue), Salera (human chorio-amionic allograft) and Flex-HD Pliable (extracellular matrix scaffold)?
Summary answer
FTY-720 reduces post-transplant primordial follicle loss, particularly when used in conjunction with FlexHD-Pliable. Paradoxically, Salera results in follicular growth activation in ovarian grafts.
What is known already
Sphingosine-1-phosphate is a sphingolipid which was previously shown to reduce post-xenografting ischemia and primordial follicle loss in a human ovarian xenograft modeI. However, sphingosine-1-phosphate has a short half-life and is not approved for clinical use. Its synthetic analog FTY-720, on the other hand, is FDA-approved for alternate medical indications. FlexHD-Pliable is an acellular dermal matrix composed entirely of reticular dermis which is used in plastic surgery to bridge tissue defects and enable revascularization. Salera is a dehydrated human amnion and chorion allograft, rich with growth factors, and is used to enhance wound healing with vascularizing potential.
Study design, size, duration
Slow frozen ovarian cortical pieces (5x5 mm) from four cadaveric organ donors (27-32 years old) were thawed and xenotransplanted subcutaneously to 8–10-week-old severe-combined-immunodeficiency mice. The grafts were harvested 13 days later, after following interventions: (i) vehicle injections (control group) on days 9–12; (ii) FlexHD-Pliable alone; (iii) FTY-720 alone (1 mg/kg/day on days 9–12); (iv) FlexHD-Pliable pre-incubated with FTY-720 prior to xenografting (FlexHD-Pliable+FTY-720); (v) FlexHD-Pliable+Salera; and (vi) FlexHD-Pliable preincubated with FTY-720 +Salera.
Participants/materials, setting, methods
Paraffin-embedded tissues were serially sectioned at 5 µm and stained with H&E. Every fifth section from each block was evaluated by observers, blinded to each other’s assessments. Primordial follicle density was determined as total number of primordial follicles per donor divided by the total tissue surface examined, and was expressed per mm². Primordial follicle activation was assessed by the total primary/primordial follicle ratio.
Main results and the role of chance
Primordial follicle density was similar between the vehicle control and FlexHD-Pliable-alone groups but was significantly higher in FTY-720-alone and FlexHD-Pliable+pre-incubation FTY-720 groups compared to the controls (0.66/mm2 vs. 0.30/mm2, p = 0.0065 and 0.73/mm2 vs. 0.30/mm2, p = 0.0015, respectively). Primordial follicle density was also significantly higher in the FlexHD-Pliable preincubated with FTY-720 than in the FlexHD-Pliable-alone group (0.73/mm2 vs. 0.43/mm2, p = 0.019). Compared to vehicle, FTY-720 treatment alone reduced primordial follicle loss by 120%. Preincubation of FlexHD-Pliable with FTY-720 further enhanced primordial preservation, reducing follicle loss by 143.3%. However, FlexHD-Pliable+Salera combination reduced primordial follicle density compared to FTY-720 treatment alone (0.15/mm2 vs. 0.66/mm2, p = 0.0164) and FlexHD-Pliable+FTY-720 preincubation groups (0.15/mm2 vs. 0.73/mm2, p = 0.0052). Primordial follicle density was not significantly different among FlexHD-Pliable+Salera+FTY-720 preincubation vs. FTY-720 alone vs. FlexHD-Pliable+FTY-720 preincubation groups. In the FlexHD-Pliable+Salera group, the primary/primordial follicle ratio was significantly higher than all other groups, including the FlexHD-Pliable+Salera+FTY-720 group (5.79 vs. 0.66-1.80). These results suggest that administration of Salera paradoxically triggers the primordial follicle growth initiation in vivo, but this activation can be blocked by FTY-720. Overall, our experiment show that FTY-720 preserves primordial follicle density in ovarian grafts while blocking follicle activation, and FlexHD-Pliable scaffold may enhance this effect.
Limitations, reasons for caution
Though highly promising, the findings from this xenograft model will need to be confirmed in clinical studies before FTY-720 + FlexHD-Pliable combination can be recommended to enhance ovarian tissue transplant outcomes.
Wider implications of the findings
Although ovarian tissue transplantation is no longer considered experimental by several organizations, its success is limited by post-transplant ischemic loss of primordial follicles. FTY-720 and FlexHD-Pliable hold promise to improve ovarian transplant longevity and success. The mechanisms by Salera induces follicle activation need to be investigated.
Trial registration number
No