RESUMO
In this study, the effect of heat shock on frozen-thawed blastocysts was evaluated using in vitro-produced (IVP) bovine embryos. In experiment 1, the effects of 6 h of heat shock at 41.0 C on fresh blastocysts were evaluated. HSPA1A expression as a reflection of stress was increased by heat shock (P < 0.05), but the expressions of the quality markers IFNT and POU5F1 were not affected. In experiment 2, frozen-thawed blastocysts were incubated at 38.5 C for 6 h (cryo-con) or exposed to heat shock at 41.0 C for 6 h (cryo-HS). Then, blastocysts were cultured at 38.5 C until 48 h after thawing (both conditions). Cryo-HS blastocysts exhibited a decreased recovery rate: HSPA1A expression was dramatically increased compared with that in fresh or cryo-con blastocysts at 6 h, and IFNT expression was decreased compared with that in cryo-con blastocysts at 6 h (both P < 0.05). Cryo-con blastocysts at 6 h also exhibited higher HSPA1A expression than fresh blastocysts (P < 0.05). At 48 h after thawing, the number of hatched blastocysts and blastocyst diameter were lower in cryo-HS blastocysts (P < 0.05). Cryo-con blastocysts showed lower POU5F1 levels at 48 h than fresh, cryo-con or cryo-HS blastocysts at 6 h (P < 0.05), but their POU5F1 levels were not different from those of cryo-HS blastocysts at 48 h. These results indicated that application of heat shock to frozen-thawed blastocysts was highly damaging. The increase in damage by the interaction of freezing-thawing and heat shock might be one reason for the low conception rate in frozen-thawed embryo transfer in summer.
Assuntos
Blastocisto/citologia , Criopreservação/veterinária , Ectogênese , Regulação da Expressão Gênica no Desenvolvimento , Matadouros , Animais , Biomarcadores/metabolismo , Blastocisto/metabolismo , Bovinos , Sobrevivência Celular , Técnicas de Cultura Embrionária/veterinária , Feminino , Fertilização in vitro/veterinária , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Temperatura Alta/efeitos adversos , Interferon Tipo I/genética , Interferon Tipo I/metabolismo , Masculino , Fator 3 de Transcrição de Octâmero/genética , Fator 3 de Transcrição de Octâmero/metabolismo , Proteínas da Gravidez/genética , Proteínas da Gravidez/metabolismoRESUMO
The ATP-binding cassette sub-family B member 1 (ABCB1) plays a critical role in maintaining the metabolic capability of cells as an efflux transporter that pumps xenobiotics out of cells. We investigated the effects of highly expressed ABCB1 on the development and viability of cryopreserved bovine embryos. The ABCB1 level in cultured bovine embryos was decreased during development to blastocyst-stage compared to germinal vesicle- and second metaphase-stage oocytes. When bovine embryos were cultured with forskolin and/or rifampicin, the ABCB1 level was significantly increased in blastocysts but embryo development was not significantly improved. After embryo cryopreservation, highly ABCB1-expressed blastocysts exhibited significant increases in viability and hatching rates. The high viability of the cryopreserved blastocysts was accompanied by a significant increase in cell proliferation during culture for 48 h. Thus, ABCB1 is expressed in bovine oocytes and embryos, and the cellular quality of bovine blastocysts is improved by the enhancement of ABCB1 expression.
