Ovarian tissue health after laparoscopic unilateral oophorectomy: A porcine model for establishing optimized fertility preservation techniques in children.

BACKGROUND: The only pre-treatment fertility preservation option for prepubertal girls, who are at risk for infertility due to their diagnosis or treatment, is surgical removal of ovarian tissue for cryopreservation (OTC). We investigated ovarian tissue health following isolation with an ultrasonic advanced energy device (UAED), that has a previously reported thermal spread of ≤2 cm. METHODS: The ovaries of eight Yucatan minipigs were isolated by laparoscopy (1) close dissection with the UAED located up to 2 mm away from the ovarian capsule, (2) far dissection with the UAED located >2 cm away, or by (3) laparotomy for control ovaries using cold scissors. Ovarian cortex tissues were cultured for 4 days to assess tissue health. RESULTS: Ovarian cortex tissue isolated using a UAED produced an altered metabolic ratio in both the far and close dissection compared to control (p < 0.001). There was an increase in folliculogenesis in the control samples over samples isolated with far and close dissection (p < 0.0001), and a reduction in estradiol production in experimental groups (p < 0.0001). CONCLUSIONS: This model defines differences in ovarian tissue health among different isolation techniques. Ongoing work will further define the standard of care surgical technique for OTC.

Proteomic analyses of decellularized porcine ovaries identified new matrisome proteins and spatial differences across and within ovarian compartments.

Premature ovarian insufficiency (POI) affects approximately 1% of women. We aim to understand the ovarian microenvironment, including the extracellular matrix (ECM) and associated proteins (matrisome), and its role in controlling folliculogenesis. We mapped the composition of the matrisome of porcine ovaries through the cortical compartment, where quiescent follicles reside and the medullary compartment, where the larger follicles grow and mature. To do this we sliced the ovaries, uniformly in two anatomical planes, enriched for matrisome proteins and performed bottom-up shotgun proteomic analyses. We identified 42 matrisome proteins that were significantly differentially expressed across depths, and 11 matrisome proteins that have not been identified in previous ovarian protein analyses. We validated these data for nine proteins and confirmed compartmental differences with a second processing method. Here we describe a processing and proteomic analysis pipeline that revealed spatial differences and matrisome protein candidates that may influence folliculogenesis.