Metabolic and secretory recovery of slow frozen-thawed human ovarian tissue in vitro

Abstract

Within the options available for fertility preservation, cryopreservation of ovarian cortical tissue has become an important technique. Freezing and thawing procedures have been optimized to preserve tissue integrity and viability. However, the improvement of the tissue retransplantation is currently of great interest. Rapid angiogenesis is needed at the retransplantation site to accomplish sufficient blood supply to provide oxygen and nutrients. Many studies address this issue. However, we need to understand the physiology of the thawed tissue to gain further understanding of the complexities of the procedure. As freezing and thawing generally impairs cellular metabolism, we aimed to characterize the changes in metabolic activity and secretion of the angiogenic factor vascular endothelial growth factor-A (VEGF-A) of frozen-thawed ovarian cortical tissue over time. Biopsy punches of ovarian cortical tissue from patients undergoing fertility preservation were maintained in culture without freezing or after a slow-freezing and thawing procedure. VEGF-A secretion was measured after 48 h by ELISA. To examine temporary changes, metabolic activity was assessed for both fresh and frozen-thawed tissue of the same patient. Metabolic activity and VEGF-A secretion were measured at 0, 24 and 48 h in culture. Thawed ovarian cortical tissue secreted significantly less VEGF-A compared to fresh ovarian cortical tissue within 48 h of culture. After thawing, metabolic activity was significantly reduced compared to fresh ovarian cortex but over the course of 48 h, the metabolic activity recovered. Similarly, VEGF-A secretion of thawed tissue increased significantly over 48 h. Here, we have shown that it takes 48 h for ovarian cortical tissue to recover metabolically after thawing, including VEGF-A secretion.

Keywords: VEGF; angiogenesis; cryobiology; cryopreservation; fertility preservation; metabolic activity; ovarian tissue; slow freeze; vascular endothelial growth factor; vascularization.