P-445 Recovery of metabolic activity and VEGF secretion after thawing of cryopreserved, slow frozen ovarian tissue

Abstract

Abstract Study question Is slow-frozen ovarian cortical tissue limited in its metabolical activity after thawing, especially with respect to VEGF secretion? Summary answer Slow-frozen ovarian cortical tissue showed decreased metabolic activity and VEGF directly after thawing, but recovered in the course of 48h in in vitro culture. What is known already Cryopreservation of ovarian cortical tissue has become an important method in fertility preservation. Freezing and thawing processes had been optimized to preserve tissue viability. However, the major obstacle of retransplantation is the rapid angiogenesis to accomplish sufficient blood supply. As freezing and thawing entails constraints in cellular metabolic activity, we aimed to assess the metabolic activity and the secretion of the angiogenic factor VEGF of frozen-thawed ovarian cortical tissue over time. Study design, size, duration Ex vivo gained ovarian tissue was cultured over 48h. Thereby, non-frozen fresh tissue was compared to slow-frozen-thawed ovarian tissue. Participants/materials, setting, methods Provided a signed consent, biopsies of ovarian tissue from patients undergoing fertility preservation were cultured without freezing (n = 13) or after slow-freezing and thawing (n = 12). VEGF secretion during 48h was measured by ELISA. Applying CellTiterBlue assay, metabolic activity was assessed in biopsies without freezing or after thawing from the same patients (n = 4-7) after 0, 24 and 48h. Similarly, VEGF secretion was measured after 0, 24 and 48h as well (n = 7) to capture temporarily changes. Main results and the role of chance Although VEGF secretion varies among patients and biopsies, there was a significant lower VEGF content in supernatant from frozen-thawed tissue compared to fresh ovarian cortical tissue after 48h of culture (p < 0.01). Directly after thawing and 24h thereafter, metabolic activity was significantly reduced (p < 0.05) compared to fresh ovarian cortex. In the course of 48h, the metabolic activity recovered (p < 0.01). Similarly, VEGF secretion increased dramatically during 24h after thawing (p < 0.05). Limitations, reasons for caution This in vitro study might not resemble the metabolic and secretory recovery after retransplantation. Wider implications of the findings We showed that ovarian tissue recovers metabolically over 48h. This recovery included an increase in VEGF secretion. As latter induces angiogenesis a culture step after thawing might support VEGF secretion and angiogenesis. However, the recovery period could also be useful as a temporal bridge until sufficient supplies are available. Trial registration number not applicable