Dynamic changes in microtubular cytoskeleton of human postmature oocytes revert after ooplasm transfer

Abstract

Objective

Study of the influence of ooplasm transfer on the microtubule dynamics in human postmature oocytes.

Design

Prospective experimental study.

Setting

Academic hospital–based fertility center.

Materials(s)

Human in vitro matured (IVM) oocytes (n = 65). Experimental groups: In set 1, sibling oocytes were processed either within 2–3 hours ("young"; n=16) or at 12–14 hours after maturation ("presumably postmature," or PPM; n=14). In set 2, young and PPM oocytes (n = 6 and 10, respectively) were assigned to be ooplasm donors and recipients, respectively. In set 3, PPM oocytes were used as ooplasm donors (n = 2) and recipients (n = 4). Control groups: Metaphase II oocytes from superovulated golden hamsters in set 1; sibling oocytes of ooplasm donor young (n = 4) and PPM oocytes (n = 7) in set 2; and sibling PPM oocytes in set 3 (n = 2).

Intervention(s)

Immunocytochemistry for αtubulin with or without treatment with taxol (Paclitaxel, a microtubule-enhancing agent) in set 1; aspiration and microinjection of ∼20 picolitres ooplasm from donor young and PPM oocytes into recipient PPM oocytes in sets 2 and 3, respectively. Taxol treatment and tubulin immunocytochemistry on ooplasm recipients and control young and PPM sibling oocytes.

Main outcome measure(s)

Morphology and pattern of the microtubules in the spindle and ooplasm as evaluated by confocal microscopy and three-dimensional image reconstructions.

Result(s)

In set 1, taxol-untreated young oocytes had normal spindle morphology and orientation to the oolemma with no microtubules in the ooplasm. Taxol-treated young oocytes revealed markedly broadened spindle poles and minimal or absent ooplasmic microtubules. Taxol-untreated PPM oocytes had variable spindle morphology and a notable increase in cortical ooplasmic microtubules. Taxol treatment of PPM oocytes resulted in a marked increase in ooplasmic microtubules in addition to a broadening of spindle poles and formation of polar asters. In set 2, control young and PPM oocytes had the same findings as the corresponding oocytes in set 1. However, all ooplasm recipient PPM oocytes showed a striking diminution in ooplasmic microtubules, despite the taxol treatment, compared with their sibling PPM control oocytes in set 2 and PPM ooplasm-injected PPM oocytes in set 3.

Conclusion(s)

Postmature oocytes exhibit a dynamic increase in ooplasmic microtubules. However, these changes revert after transfer of ooplasm from young oocytes.