Effects of chilling to 0 degrees C on the morphology of meiotic spindles in human metaphase II oocytes.

Objective: To determine the effects of chilling to 0 degrees C on the meiotic spindle of human metaphase II oocytes, as observed by optical sectioning microscopy.

Design: Laboratory study.

Setting: Academic research laboratory in a medical school.

Patient(s): Seventy-two women undergoing infertility treatment donated a total of 108 oocytes.

Intervention(s): Metaphase II oocytes were stripped of their cumulus cells, cooled directly to 0 degrees C, and held for periods of 1 to 10 minutes. They were then fixed at 37 degrees C, stained for immunofluorescence, and examined microscopically.

Main Outcome Measure(s): Morphology of the meiotic spindle in chilled and control oocytes.

Result(s): Microscopic evaluations of 46 chilled oocytes revealed various time-dependent changes in microtubules compared to 9 control oocytes. After 1 minute at 0 degrees C, spindle damage was negligible, but in oocytes cooled for 2 or 3 minutes, there was obvious shortening of the spindle and loss of polarity. Cooling to 0 degrees C for 4 to 9 minutes resulted in increasingly more drastic changes; by 10 minutes the spindles had totally disappeared. Despite depolymerization of microtubular tubulin at 0 degrees C, the chromosomes did not become dispersed, but remained anchored even in the absence of spindles.

Conclusion(s): Even brief exposure of human oocytes to temperatures near 0 degrees C causes profound alterations of the meiotic spindle.