Km. Saunders et Je. Parks, Effects of cryopreservation procedures on the cytology and fertilization rate of in vitro-matured bovine oocytes, BIOL REPROD, 61(1), 1999, pp. 178-187
The survival and developmental capacity of bovine oocytes after cryopreserv
ation are greatly impaired, possibly due to organelle damage caused by free
zing procedures. Distributions of chromosomes, microtubules, and microfilam
ents in bovine oocytes matured in vitro were examined after cooling, ethyle
ne glycol (EG) exposure, or freezing. Oocytes were incubated after treatmen
t for 20 min or 1 or 3 h, fixed, and evaluated using specific fluorescent p
robes. Abnormal cytological features increased over control levels after co
oling or EC exposure and rewarming. Changes observed in oocytes during pref
reezing manipulations included chromosome dispersal and clumping, microtubu
le depolymerization and alteration of spindle structure, and formation of c
raters and discontinuity in cytoskeletal actin staining. Freezing also led
to an increase in the occurrence of cytological abnormalities. Less than 31
% of frozen-thawed oocytes contained a normal chromosome arrangement 3 h po
stthaw (versus 90% of controls). Only 7-14% of frozen-thawed oocytes had no
rmal spindles (versus 59-71% of controls). Normal distribution of filamento
us actin was observed in less than 30% of oocytes postthaw (versus 62-89% o
f controls). These results indicate that the steps in a conventional freezi
ng procedure cause irreversible alterations in multiple cytological compone
nts of bovine oocytes, demonstrating the need for improved strategies for p
reventing cellular damage during cryopreservation procedures.