Human sperm aster formation and chromatin configuration in rabbit oocytes following intracytoplasmic sperm injection using a Piezo-Micromanipulator

ABSTRACT

In human fertilization, the sperm centrosome nucleates a radial array of microtubules called the sperm aster. The sperm aster is responsible for apposition of male and female pronuclei, and later gives rise to the first meiotic spindle. The objective of this study was to determine microtubule assembly and chromatin configuration in rabbit oocytes following intracytoplasmic injection with human sperm by piezo-driven pipette. Oocytes were collected from superovulated dose 14-15 h after hCG injection and were fertilized by injection of a single human sperm into the ooplasm of each oocyte without additional activation treatment. Four hours post heterologous intracytoplamic sperm injection [ICSI], rabbit eggs were fixed and microtubule organization and chromatin configuration were examined by immunofluorescence microscopy. In unfertilized oocytes, microtubules were present only in the metaphase-arrested second meiotic spindle. Following human sperm injection, an aster of microtubules formed adjacent to the sperm head, around mid-piece, and sperm aster was enlarged and assembled around male and female pronuclei. During pronuclear centration, male and female pronuclei were surrounded by a microtubule array without nucleation sites. With fertile human sperm, the sperm aster formation rate was 54.6%. From our data we concluded that human spermatozoa can be injected successfully into rabbit oocytes, resulting in a reasonable survival rate, and that rabbit oocytes provide a reliable tools for assessing human sperm centrosomal function using the Piezo-ICSI system