BACKGROUND: The aim of this study was to assess effects of cysteamine as an anti-oxidant on rate of in vitro maturation of oocyte and determination of its effects on spindle size and shape. METHODS: Pre-mature mice were primed with pregnant mare stimulating gonadotrophin (PMSG) and germinal vesicle (GV) stage oocytes were obtained 48 h after. The oocytes were cultured in tissue culture medium (TCM 199) with 50, 100, 200 and 500 microM cysteamine. Experiments also included a group of ovulated oocytes (in vivo matured) after priming with PMSG and human chorionic gonadotrophin. For spindle immuno-cytochemistry of metaphase II (MII), oocytes alpha and beta tubulin antibody were performed. Chromosome staining was performed with Hoechst. RESULTS: Our results showed that the rate of GV breakdown (GVBD) and MII increased in all cysteamine groups except in group cultured with 500 microM cysteamine. Immuno-cytochemistry analysis showed that spindle area in all in vitro Maturation oocytes increased when compared to in vivo oocytes. Spindle shape and size (spindle area) in 100 microM cysteamine in comparison to in vivo group was insignificant (P > 0.05). CONCLUSION: Our results revealed that cysteamine improved IVM rate in a dose dependant manner. The size and shape of spindle in the presence of given concentration of cysteamine was similar to in vivo. Therefore, cysteamine improved microtubule organization, rate of GVBD and MII development.
BACKGROUND: The aim of this study was to assess effects of cysteamine as an anti-oxidant on rate of in vitro maturation of oocyte and determination of its effects on spindle size and shape. METHODS: Pre-mature mice were primed with pregnant mare stimulating gonadotrophin (PMSG) and germinal vesicle (GV) stage oocytes were obtained 48 h after. The oocytes were cultured in tissue culture medium (TCM 199) with 50, 100, 200 and 500 microM cysteamine. Experiments also included a group of ovulated oocytes (in vivo matured) after priming with PMSG and human chorionic gonadotrophin. For spindle immuno-cytochemistry of metaphase II (MII), oocytes alpha and beta tubulin antibody were performed. Chromosome staining was performed with Hoechst. RESULTS: Our results showed that the rate of GV breakdown (GVBD) and MII increased in all cysteamine groups except in group cultured with 500 microM cysteamine. Immuno-cytochemistry analysis showed that spindle area in all in vitro Maturation oocytes increased when compared to in vivo oocytes. Spindle shape and size (spindle area) in 100 microM cysteamine in comparison to in vivo group was insignificant (P > 0.05). CONCLUSION: Our results revealed that cysteamine improved IVM rate in a dose dependant manner. The size and shape of spindle in the presence of given concentration of cysteamine was similar to in vivo. Therefore, cysteamine improved microtubule organization, rate of GVBD and MII development.