ABSTRACT
The presence of heparin and a mixture of penicillamine, hypotaurine, and epinephrine (PHE) solution in the in vitro fertilization (IVF) media seem to be a prerequisite when bovine spermatozoa are capacitated in vitro, in order to stimulate sperm motility and acrosome reaction. The present study was designed to determine the effect of the addition of heparin and PHE during IVF on the quality and penetrability of spermatozoa into bovine oocytes and on subsequent embryo development. Sperm quality, evaluated by the integrity of plasma and acrosomal membranes and mitochondrial function, was diminished (P<0.05) in the presence of heparin and PHE. Oocyte penetration and normal pronuclear formation rates, as well as the percentage of zygotes presenting more than two pronuclei, was higher (P<0.05) in the presence of heparin and PHE. No differences were observed in cleavage rates between treatment and control (P>0.05). However, the developmental rate to the blastocyst stage was increased in the presence of heparin and PHE (P>0.05). The quality of embryos that reached the blastocyst stage was evaluated by counting the inner cell mass (ICM) and trophectoderm (TE) cell numbers and total number of cells; the percentage of ICM and TE cells was unaffected (P>0.05) in the presence of heparin and PHE (P<0.05). In conclusion, this study demonstrated that while the supplementation of IVF media with heparin and PHE solution impairs spermatozoa quality, it plays an important role in sperm capacitation, improving pronuclear formation, and early embryonic development.
Subject(s)
Epinephrine/pharmacology , Fertilization in Vitro/veterinary , Heparin/pharmacology , Penicillamine/pharmacology , Taurine/analogs & derivatives , Acrosome Reaction/drug effects , Animals , Cattle , Embryonic Development/drug effects , Fertilization/drug effects , Fertilization in Vitro/methods , Sperm Capacitation/drug effects , Sperm Motility/drug effects , Taurine/pharmacologyABSTRACT
Complexos cumulus-oócito (COC), oócitos desnudos (DO) e DO cocultivados com células do cumulus em suspensão (DO+CC) foram maturados in vitro (MIV) na presença ou ausência de cisteamina (50mM). Observou-se efeito benéfico da cisteamina durante o cultivo de MIV, pois a maturação nuclear no grupo COC cisteamina foi maior do que a do COC controle (P<0,05). No grupo sem a adição de cisteamina, foi observado que a ausência de CC durante o cultivo de MIV prejudicou a maturação nuclear em DO, em relação ao COC (P<0,05), todavia a cisteamina restaurou a capacidade de progressão da meiose em DO, tornando-os semelhantes aos COC (P>0,05). O acoplamento entre oócitos e CC durante MIV demonstrou ser essencial para aquisição da competência do oócito para suportar o desenvolvimento embrionário inicial, pois COC apresentaram maior porcentagem de blastocistos e eclosão quando comparados a DO e DO+CC (P<0,05). A inclusão de cisteamina no cultivo de MIV não restaurou a aquisição da competência em DO e DO+CC, que permaneceram semelhantes aos do grupo-controle (P>0,05). Conclui-se que a cisteamina no meio de MIV melhora as taxas de maturação nuclear em COC e restaura a capacidade de progressão da meiose em DO. Todavia, na concentração utilizada neste estudo, não promove efeito benéfico no desenvolvimento embrionário.(AU)
Cumulus-oocyte complexes (COC), denuded oocytes (DO) and DO co-cultured with cumulus cells in suspension (DO+CC) were in vitro matured (IVM) in the presence or absence of cysteamine (50mM). A beneficial effect of cysteamine was observed during IVM, because the nuclear maturation in the COC cysteamine group was higher than in COC control (P<0.05). In the control group, the absence of CC during IVM impaired nuclear maturation in DO when compared to COC (P<0.05), but cysteamine restored the ability of meiosis progression in DO, making them similar to COC (P>0.05). The coupling between oocytes and CC during IVM proved to be essential for the acquisition of oocyte competence to support early embryonic development, as COC had higher percentages of blastocyst and hatching when compared to DO and DO+DC (P<0.05). However, the inclusion of cysteamine in the IVM culture did not restore the acquisition of competence in DO and DO+DC, which remained similar to the control group (P>0.05). It is concluded that cysteamine in the IVM culture improves the nuclear maturation in COC and restores the progression ability of meiosis in DO. However, in the concentration used in this study, cysteamine does not promote a beneficial effect on embryo development.(AU)
Subject(s)
Animals , Female , Cattle , Cattle/embryology , Cumulus Cells , Meiosis , Maturation-Promoting Factor , Embryonic Development , Cleavage Stage, OvumABSTRACT
Complexos cumulus-oócito (COC), oócitos desnudos (DO) e DO cocultivados com células do cumulus em suspensão (DO+CC) foram maturados in vitro (MIV) na presença ou ausência de cisteamina (50mM). Observou-se efeito benéfico da cisteamina durante o cultivo de MIV, pois a maturação nuclear no grupo COC cisteamina foi maior do que a do COC controle (P<0,05). No grupo sem a adição de cisteamina, foi observado que a ausência de CC durante o cultivo de MIV prejudicou a maturação nuclear em DO, em relação ao COC (P<0,05), todavia a cisteamina restaurou a capacidade de progressão da meiose em DO, tornando-os semelhantes aos COC (P>0,05). O acoplamento entre oócitos e CC durante MIV demonstrou ser essencial para aquisição da competência do oócito para suportar o desenvolvimento embrionário inicial, pois COC apresentaram maior porcentagem de blastocistos e eclosão quando comparados a DO e DO+CC (P<0,05). A inclusão de cisteamina no cultivo de MIV não restaurou a aquisição da competência em DO e DO+CC, que permaneceram semelhantes aos do grupo-controle (P>0,05). Conclui-se que a cisteamina no meio de MIV melhora as taxas de maturação nuclear em COC e restaura a capacidade de progressão da meiose em DO. Todavia, na concentração utilizada neste estudo, não promove efeito benéfico no desenvolvimento embrionário.
Cumulus-oocyte complexes (COC), denuded oocytes (DO) and DO co-cultured with cumulus cells in suspension (DO+CC) were in vitro matured (IVM) in the presence or absence of cysteamine (50mM). A beneficial effect of cysteamine was observed during IVM, because the nuclear maturation in the COC cysteamine group was higher than in COC control (P<0.05). In the control group, the absence of CC during IVM impaired nuclear maturation in DO when compared to COC (P<0.05), but cysteamine restored the ability of meiosis progression in DO, making them similar to COC (P>0.05). The coupling between oocytes and CC during IVM proved to be essential for the acquisition of oocyte competence to support early embryonic development, as COC had higher percentages of blastocyst and hatching when compared to DO and DO+DC (P<0.05). However, the inclusion of cysteamine in the IVM culture did not restore the acquisition of competence in DO and DO+DC, which remained similar to the control group (P>0.05). It is concluded that cysteamine in the IVM culture improves the nuclear maturation in COC and restores the progression ability of meiosis in DO. However, in the concentration used in this study, cysteamine does not promote a beneficial effect on embryo development.