Meiotic competence of equine oocytes and pronucleus formation after intracytoplasmic sperm injection (ICSI) as related to granulosa cell apoptosis.
Follicle atresia and granulosa cell apoptosis may be related to oocyte meiotic and developmental competence. We analyzed the relationships among granulosa cell apoptosis, initial cumulus morphology, oocyte nuclear maturation in vitro, and pronucleus formation after intracytoplasmic sperm injection (ICSI) in the horse. For each follicle, the size was measured and granulosa cells were used for DNA laddering analysis. Oocytes were evaluated for cumulus morphology, cultured for in vitro maturation, and submitted to ICSI. Apoptosis was categorized as absent, intermediate, or advanced according to the relative concentrations of two DNA fragments at 900 and 360 base pairs (bp). In 98 oocyte-follicle pairs, 52 oocytes were classified as expanded (Exp), 39 as compact (Cp), and 7 as having a partial (P) cumulus. Advanced apoptosis was detected in 55% (54/98) of follicles; 37% (36/98) of follicles showed an intermediate level of apoptosis; and 8 follicles (8%) were nonapoptotic. Follicle size was not significantly correlated with granulosa cell apoptosis (P > 0.05). Significantly more Exp than Cp oocytes originated from follicles with advanced apoptosis (P < 0.001). The proportion of oocytes maturing in vitro was significantly higher in oocytes issuing from apoptotic follicles than in oocytes issuing from healthy follicles (P < 0.05). The proportion of normally (two pronuclei) or abnormally fertilized oocytes (one or greater than two pronuclei, or partially decondensed sperm) did not differ in relation to granulosa cell apoptosis. We conclude that, in the mare, granulosa cell apoptosis is related to cumulus expansion and an increase in oocyte meiotic competence but has no effect on the proportion of meiotically competent oocytes that activate after ICSI. These results provide selection criteria for horse oocytes used in assisted reproductive techniques so that embryo production may be maximized.
Dell'Aquila
Maria Elena
ME
Department of Animal Production--Section of Reproductive Biology and Veterinary Obstetrics, University of Bari, 70010 Valenzano, Bari, Italy. e.dellaquila@veterinaria.uniba.it
Albrizio
Maria
M
Maritato
Filippo
F
Minoia
Paolo
P
Hinrichs
Katrin
K
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D002467:Cell Nucleus / Q000502:physiology*; D002469:Cell Separation; D002843:Chromatin / Q000235:genetics; D004247:DNA / Q000235:genetics; D053938:DNA Fragmentation / Q000502:physiology; D005260:Female; D005307:Fertilization in Vitro; D006107:Granulosa Cells / Q000502:physiology*; D006736:Horses; D008297:Male; D008540:Meiosis / Q000502:physiology*; D009707:Nucleosomes / Q000502:physiology; D009865:Oocytes / Q000502:physiology*; D006080:Ovarian Follicle / Q000033:anatomy & histology / Q000502:physiology; D020554:Sperm Injections, Intracytoplasmic; D013079:Sperm Maturation / Q000502:physiology; D015053:Zygote / Q000502:physiology
D002843:Chromatin; D009707:Nucleosomes; D004247:DNA