Long-time cooling of human ovarian tissue before cryopreservation as obvious procedure: stimulation of follicular development and neo-vascularisation.

BACKGROUND

The positive effect of cooling on tissue cells is known. The aim of this research was to study the intensiveness of neo-vascularisation and follicular development in ovarian tissue after 24 hours cooling to 5 degrees C before cryopreservation.

METHODS

Fifty six pieces from 7 patients were divided into the following four groups: Group 1: pieces cultured just after operation, Group 2: pieces cooled after operation to 5 degrees C for 24 hours and then cultured, Group 3: pieces frozen-thawed just after operation and then cultured, Group 4: pieces cooled after operation to 5 degrees C for 24 hours, frozen, thawed, and then cultured. Culture of ovarian pieces was performed in a chorioallantoic membrane (CAM)-system for 5 days. The efficacy of the tissue cooling was evaluated by the development of follicles and intensiveness of neo-vascularisation (by Desmin).

RESULTS

For Group 1, 2, 3, and 4, mean density of follicles per 1 mm3 was 10.1, 11.1, 9.8, and 12.0, respectively (P1-2, 3-4 < 0.05). For these groups 91%, 92%, 90%, and 90% preantral follicles were morphologically normal (P1-2, 3-4 > 0.1). The immunohistochemical analysis showed that the intensiveness of neo-vascularisation observed in ovarian tissue of Group 2 (pre-cooled before culture) and Group 4 (pre-cooled before cryopreservation) was drastically increased.

CONCLUSIONS

The 24 hour cooling to 5 degrees C before cryopreservation is beneficial for cryopreservation of human ovarian tissue.