Vitrificação de embriões bovinos produzidos in vivo e in vitro após lipólise química
Carregando...
Data
2009-12-17
Autores
Paschoal, Daniela Martins [UNESP]
Título da Revista
ISSN da Revista
Título de Volume
Editor
Universidade Estadual Paulista (Unesp)
Resumo
O objetivo desta dissertação foi avaliar a viabilidade e a resistência à criopreservação após a realização de lipólise química utilizando-se o forskolin. Embriões Bos taurus indicus (OZ) e Bos taurus indicus X Bos taurus taurus (OMZT) produzidos in vitro, foram comparados em meio de cultivo contendo ou não soro fetal bovino (SFB). Para o experimento1, 1.052 oócitos de graus I e II foram colhidos de ovários de vacas Bos taurus indicus de abatedouro, dividindo em 7 réplicas. Os oócitos foram maturados em TCM 199 acrescido de SFB, Piruvato de Sódio, FSH, LH e Penicilina/Estreptomicina. A fertilização foi realizada com sêmen de touro Nelore (Bos taurus indicus) ou com sêmen de touro Simental (Bos taurus taurus), em meio HTF acrescido de BSA, Piruvato de Sódio, Cafeína, Heparina, Penicilinamina, Hipotaurina, Epinefrina e Penicilina/Estreptomicina. Os prováveis zigotos foram cultivados em meio SOFaa, acrescido de BSA, Piruvato de Sódio, Penicilina/Estreptomicina, suplementado ou não com SFB (Grupo 2,5%SFB e Grupo 0%SFB). No D6 os blastocistos foram novamente divididos em 2 sub-grupos com ou sem adição de 10μM de forskolin (total de 4 grupos - 2,5%SFB, 0%SFB, 2,5%+F e 0%+F). Para o experimento 2 os embriões foram expostos a solução de vitrificação 1 (SV1 = 5M de Etileno Glicol) durante 3 minutos e transferidos para uma gota com 15μL da solução de vitrificação 2 (SV2 = composta por 7M de Etileno Glicol, 0.5M de Galactose e 18% (w/v) de Ficoll 70) sendo, em seguida envasados em palhetas francesas estéreis de 0,25 mL. Para o aquecimento, as palhetas foram mantidas em ar por 8 segundos e depois em água a 37°C por 15 segundos. Como controle dos experimentos, embriões bovinos produzidos in vivo coletados de vacas Nelores foram utilizados. A taxa de clivagem e de produção de blastocisto no experimento 1, não diferiu entre os tratamentos dentro dos grupo...
The main objective of this dissertation was to evaluate the viability and cryotolerance of bovine embryos treated with forskolin to induce chemical lipolysis. In vitro produced Bos taurus indicos (OZ) and Bos taurus indicus X Bos taurus taurus (OMZT) embryos were compared in culture conditions containing or not fetal calf serum (FCS). For Experiment 1, 1.052 grade I and II Bos taurus indicos oocytes were obtained from slaughterhouse ovaries and divided in 7 replicates. The oocytes were matured in TCM 199 supplemented with FCS, Sodium Piruvate, FSH, LH and Penicillin/Streptomycin. Fertilization was performed using semen from one Nelore bull (Bos taurus indicos) and one Simental bull (Bos taurus taurus) in HTF medium supplemented with BSA, Sodium Pyruvate, Caffeine, Heparine, Penicilamine, Hipotaurine, Epinephrine and Penicillin/Streptomycin. The presumptive zygotes were cultured in SOFaa with BSA, Sodium Pyruvate and Penicillin/Streptomycin supplemented or not with FCS (Group 2.5%FCS and Group 0%FCS). On day 6 embryos from both groups were divided in two sub-groups containing or not 10μM of forskolin (total of 4 groups - 2.5%FCS, 0%FCS, 2.5%+F and 0%+F). For Experiment 2 the embryos were exposed to a vitrification solution 1 (SV1 = 5M of Ethylene Glycol) during 3 minutes and transferred to a vitrification solution 2 (SV2 = 7M Ethylene Glycol + 0.5M Glucose + 18% Ficoll). While in the SV2 embryos were packaged in 0.25 mL straws, incubated for 1 minute in Liquid N2 vapor and plunged into Liquid N2. For warming the straws were held in air for 8 seconds and plunged in 37oC water bath for 15 seconds. As a control, in vivo produced embryos collected from Nelore cows were used. The cleavage and blastocyst production rates, in Experiment 1, was not different among groups of OZ or OMZT embryos (P>0.05). However, a significant difference was observed in blastocyst production rates when... (Complete abstract click electronic access below)
The main objective of this dissertation was to evaluate the viability and cryotolerance of bovine embryos treated with forskolin to induce chemical lipolysis. In vitro produced Bos taurus indicos (OZ) and Bos taurus indicus X Bos taurus taurus (OMZT) embryos were compared in culture conditions containing or not fetal calf serum (FCS). For Experiment 1, 1.052 grade I and II Bos taurus indicos oocytes were obtained from slaughterhouse ovaries and divided in 7 replicates. The oocytes were matured in TCM 199 supplemented with FCS, Sodium Piruvate, FSH, LH and Penicillin/Streptomycin. Fertilization was performed using semen from one Nelore bull (Bos taurus indicos) and one Simental bull (Bos taurus taurus) in HTF medium supplemented with BSA, Sodium Pyruvate, Caffeine, Heparine, Penicilamine, Hipotaurine, Epinephrine and Penicillin/Streptomycin. The presumptive zygotes were cultured in SOFaa with BSA, Sodium Pyruvate and Penicillin/Streptomycin supplemented or not with FCS (Group 2.5%FCS and Group 0%FCS). On day 6 embryos from both groups were divided in two sub-groups containing or not 10μM of forskolin (total of 4 groups - 2.5%FCS, 0%FCS, 2.5%+F and 0%+F). For Experiment 2 the embryos were exposed to a vitrification solution 1 (SV1 = 5M of Ethylene Glycol) during 3 minutes and transferred to a vitrification solution 2 (SV2 = 7M Ethylene Glycol + 0.5M Glucose + 18% Ficoll). While in the SV2 embryos were packaged in 0.25 mL straws, incubated for 1 minute in Liquid N2 vapor and plunged into Liquid N2. For warming the straws were held in air for 8 seconds and plunged in 37oC water bath for 15 seconds. As a control, in vivo produced embryos collected from Nelore cows were used. The cleavage and blastocyst production rates, in Experiment 1, was not different among groups of OZ or OMZT embryos (P>0.05). However, a significant difference was observed in blastocyst production rates when... (Complete abstract click electronic access below)
Descrição
Palavras-chave
Bovino - Reprodução, Criopreservação, Vitrification, Forskolin
Como citar
PASCHOAL, Daniela Martins. Vitrificação de embriões bovinos produzidos in vivo e in vitro após lipólise química. 2009. 132 f. Dissertação (mestrado) - Universidade Estadual Paulista, Faculdade de Medicina Veterinária e Zootecnia de Botucatu, 2009.