A confocal microscopy analysis of the spindle and chromosome configurations of human oocytes cryopreserved at the germinal vesicle and metaphase II stage.
BACKGROUND
Routine oocyte cryopreservation remains an elusive technique in the wide range of assisted reproductive technologies available. This study examines the effect of a cryopreservation protocol on the spindle and chromosome configurations of human oocytes cryopreserved at the germinal vesicle (GV) and metaphase II (MII) stage.


METHODS
GV oocytes were randomly assigned to one of three groups: (i) control oocytes matured in vitro to MII stage (n = 156); (ii) oocytes cryopreserved at the GV stage and then matured in vitro (n = 90); (iii) oocytes cryopreserved at the MII stage (n = 147). Following cryopreservation and in-vitro maturation, immunostaining of tubulin and chromatin was performed, before visualization using confocal microscopy.


RESULTS
A statistically significant increase was observed in the survival rate in group 2 (73.3%, 66/90) compared to group 3 (55.7%, 82/147) (P < 0.007). Exposure of oocytes to the cryoprotective solutions without freezing had no effect on the structure of their second meiotic spindle. However, statistically significant differences were observed on both spindle and chromosome configurations of oocytes from group 2 (5.2 and 5.2% respectively) and group 3 (16.2 and 18.8% respectively) compared with group 1 oocytes (71.6 and 82.0% respectively) (P < 0.001 in all cases).


CONCLUSIONS
The protocol followed results in high rates of survival and potential for in-vitro maturation, but has a deleterious effect on the organization of the meiotic spindle of human oocytes cryopreserved at both the GV and MII stages.
Boiso
Irene
I
Servei de Medicina de la Reproducció, Dept Obtetrícia i Ginecologia, Institut Universitari Dexeus, P. Bonanova 89-91, Barcelona 08017, Spain. ireboi@iudexeus.uab.es
Martí
Mercè
M
Santaló
Josep
J
Ponsá
Montse
M
Barri
Pere N
PN
Veiga
Anna
A
D000328:Adult; D002470:Cell Survival / Q000187:drug effects; D016922:Cellular Senescence / Q000187:drug effects; D002875:Chromosomes / Q000648:ultrastructure*; D015925:Cryopreservation; D003451:Cryoprotective Agents / Q000494:pharmacology; D005260:Female; D006801:Humans; D008677:Metaphase; D018613:Microscopy, Confocal; D009865:Oocytes / Q000166:cytology* / Q000502:physiology / Q000648:ultrastructure*; D008941:Spindle Apparatus / Q000187:drug effects / Q000648:ultrastructure*
D003451:Cryoprotective Agents