PARENT SESSION
PLATFORM SESSION 11. GAMETE BIOLOGY AND GAMETOGENESIS II

Monday, August 2, 2004
2:00 PM–4:00 PM
Buchanan A104
Chair: John McCarrey Co-Chair: Barbara Sotolongo

(302) EFFECTS OF MOUSE OVARIAN TISSUE CRYOPRESERVATION ON GRANULOSA CELL-OOCYTE INTERACTION.

Plancha, Carlos¹, Navarro-Costa, Paulo¹, Correia, Sónia², Gouveia-Oliveira, António³, Negreiro, Filipa³, Jorge, Sónia², Cidadão, António¹, Carvalho, Maria², ¹ Biology of Reproduction Unit, Lisbon, Portugal² Reproductive Medicine Unit, Lisbon, Portugal³ Datamedica Lda, Lisbon, Portugal

ABSTRACT- Although regulation of follicular development is dependent on the granulosa-oocyte interface, current ovarian tissue cryopreservation protocols have yet to be assessed in terms of somatic-germ cell interaction. Taking into account the importance of granulosa-oocyte communication in the follicle, this study aimed to investigate for the first time the possible cryo-induced disruption of cell communication via transzonal processes containing filamentous actin (TZPs-Act), in two distinct populations of antral follicles. A total of 93 ovaries from eight to ten-week old F1 (CBAxC57BL/6) eCG-primed mice were divided for fresh, cryopreserved with dimethyl sulfoxide (DMSO) and cryopreserved in glycerol groups. Contents of large and small antral follicles were separately released, both in fresh or post-thaw tissue. The obtained GV-stage oocytes were further scored for the pattern of granulosa-oocyte cell adhesion. The oocytes were stained with phalloidin-TRITC for filamentous actin and TZPs-Act density was assessed by computerized analysis of the intensity of the fluorescence signal emanating from the zona pellucida. Results show that although ovarian tissue cryopreservation reduces the average number of oocytes retrieved per ovary, it increases the relative distribution of granulosa-free oocytes while decreasing that of granulosa-enclosed ones. In addition, a post-thaw decrease in the relative density of TZPs-Act was also recorded. For the assessed parameters, cryopreservation with DMSO instead of glycerol held results closer to those of fresh controls. This study showed that in situ cryopreservation of granulosa-oocyte complexes with current protocols leads to partial disruption of the granulosa-oocyte interface. Given that oocyte maturation seems to depend on the integrity of oocyte-granulosa interactions, this work may be of significance for future studies on ovarian tissue cryopreservation. Furthermore, oocytes exhibited differential patterns of granulosa cell adhesion and TZP-Act relative density as they derived from differently sized antral follicles, suggesting that the granulosa-oocyte interface may be developmentally regulated. *Partially funded by FCT (project POCTI/ESP/43628/2000)

KEY WORDS: granulosa cell-oocyte interaction, ovarian tissue cryopreservation, transzonal processes, actin