Avaliação de embriões ovinos provenientes de oócitos submetidos a estresse calórico durante a maturação in vitro

Santos Junior, Edivaldo Rosas dos; Chaves, Ricardo Macêdo; Silva, José Carlos Ferreira da; Moura, Marcelo Tigre; Bartolomeu, Cláudio Coutinho; Gonçalves, Paulo Bayard Dias; Lima, Paulo Fernandes de; Oliveira, Marcos Antônio Lemos de

doi:10.5216/cab.v14i3.17170

Resumos

Neste trabalho foi avaliado o efeito do estresse calórico durante a maturação de oócitos sobre a produção in vitro de embriões ovinos. Os ovários foram obtidos em abatedouro e os oócitos colhidos de folículos de 2 a 6 mm de diâmetro. Após seleção, os oócitos, em 10 replicações, foram colocados para maturação in vitro (MIV) durante 24 horas. Os oócitos submetidos ao estresse térmico de 41º C durante 3, 6, 12, 18 e 24 horas foram posteriormente transferidos para completar a MIV a 39 ºC, mesma temperatura utilizada para maturação dos oócitos do grupo controle. O desenvolvimento dos embriões foi determinado nos dias 3, 4, 5 e 8 pós-fecundação. A avaliação da qualidade dos embriões foi efetuada através da contagem total de células coradas pelo DAPI e da determinação do número de blastômeros positivos para apoptose através do teste de TUNEL. Observou-se que o estresse térmico diminuiu (P < 0,05) a capacidade de maturação dos oócitos de acordo com o tempo de exposição à temperatura de 41º C. No grupo de oócitos incubados a 39° C, 70,70% maturou, enquanto que nos grupos expostos ao estresse térmico, apenas 45,28%, 35,17%, 12,30%, 9,74% e 4,60% maturaram, respectivamente, após 3, 6, 12, 18 e 24 horas de incubação. A duração de exposição dos oócitos ao estresse calórico foi inversamente proporcional (P < 0,05) à capacidade de desenvolvimento embrionário e diretamente proporcional (P < 0,05) ao número de blastocistos positivos para apoptose. Todavia, o efeito deletério do estresse térmico sobre a clivagem e os embriões nos estádios de 8 a 16 células e de mórula foi crescente (P > 0,05) somente até 18 horas de incubação. Os resultados permitem concluir que o estresse calórico durante a maturação in vitro de oócitos reduz a quantidade e a qualidade dos embriões ovinos produzidos in vitro determinadas pela alta incidência de apoptose.

blastocisto; FIV; MIV; PIV

The aim of this work was to evaluate the effect of heat stress during oocyte maturation on ovine in vitro embryo production. Ovaries were collected at abattoirs and oocytes retrieved from follicles ranging from 2 and 6 mm in diameter. After selection, all oocytes, in 10 replicates, were placed in in vitro maturation (IVM) during 24 hours. The oocytes were submitted to heat stress at 41º C during 3, 6, 12, 18 and 24 hours and were further transferred to 39º C in order to complete IVM, which was the temperature of maturation of control oocytes. Embryonic development was determined on days 3, 4, 5, and 8 post-fertilization. Embryo evaluation was performed as total cell count by DAPI staining and determination of positive blastomere for apoptosis by the TUNEL assay. We observed that heat stress diminishes (P < 0.05) oocyte maturation capacity in accordance with exposure time at 41º C. In the group of oocytes incubated at 39 ºC, 70.70% matured, while in the groups exposed to heat stress at 41º C, only 45.28%, 35.17%, 12.30%, 9.74% and 4.60% matured after 3, 6, 12, 18 and 24 hours of incubation, respectively. The duration of exposure to heat stress was inversely proportional (P < 0.05) to embryonic developmental capacity and directly proportional (P < 0.05) to the number of blastocysts positive to apoptosis. However, the cleavage rate and embryonic development from 8 to 16 cells and morulae stages were affected by heat stress (P > 0.05) only up to 18 hours of incubation. The results allow the conclusion that heat stress during oocyte in vitro maturation reduces the quantity and quality of ovine embryos produced in vitro determined by the high incidence of apoptosis.

blastocyst; IVF; IVM; IVP

ANDREU-VAZQUEZ, C.; LOPEZ-GATIUS, F.; GARCIA-ISPIERTO, I.; MAYA-SORIANO, M.J.; HUNTER, R.H.F.; LOPEZ-BEJAR, M. Does heat stress provoke the loss of a continuous layer of cortical granules beneath the plasma membrane during oocyte maturation? Zygote, v.18, n. 4, p.293-299, 2010.
BADINGA, L.; COLLIER, R.J.; THATCHER, W.W.; WILCOX, C.J. Effects of climatic and management factors on conception rate of dairy cattle in subtropical environment. Journal of Dairy Science, v.68, p.78-85, 1985.
CHIAMENTI, A.; FILHO, C.R.; MOURA, M.T .; PAULA-LOPES, F.F.; NEVES, J.P.; NETO, C.C.; GON"CALVES, P.B.; LIMA, P.F.; OLIVEIRA, M.A. Use of retinyl acetate, retinoic acid and insulin-like growth factor-I (IGF-I) to enhance goat embryo production. Acta Veterinaria Hungarica v.61, p.116-124, 2013.
EALY, A.D.; HOWELL, J.L.; MONTERROSO, V.H.; ARÉCHIGA, C.F.; HANSEN, P.J. Developmental changes in sensitivity of bovine embryos to heat shock and use of antioxidants as thermoprotectants. Journal of Animal Science, v.73, p.1401-1407, 1995.
EDWARDS, J.L.; HANSEN, P.J. Differential responses of bovine oocytes and preimplantation embryos to heat shock. Molecular Reproduction and Development, v.46, p.138-145, 1997.
EDWARDS, J.L.; BOGART, A.N.; RISPOLI, L.A.; SAXTON, A.M.; SCHRICK, F.N. Developmental competence of bovine embryos from heat-stressed ova. Journal of Dairy Science, v.92, p.563-570, 2009.
HANSEN, P.J. Effects of heat stress on mammalian reproduction. Philosophical Transactions of The Royal Society Biological Sciences, v. 364, p. 3341-3350, 2009.
INMET. Instituto Nacional de Meteorologia. Disponível em: <http:www.inmet.gov.br/html/prev_climatica_tempo/prognostico>. Acesso em: 23 abr. 2009.
» link
JU, J.C.; PARKS, J.E.; YANG, X. Thermotolerance of IVM-derived bovine oocytes and embryos after shortterm heat shock. Molecular Reproduction and Development, v.53, p.336-340, 1999.
JU, J.C.; TSENG, J.K. Nuclear and cytoskeletal alterations of in vitro matured porcine oocytes under hyperthermia. Molecular Reproduction and Development, v.68, p.125-133, 2004.
KESKINTEPE, L.; SIMPLICIO, A.A.; BRACKETT, B.G. Caprine blastocyst development after in vitro fertilization with spermatozoa frozen in different extenders. Theriogenology, v.49, n.7, p.1265-1274, 1998.
LAWRENCE, J.L.; PAYTON, R.R.; GODKIN, J.D.; SAXTON, A.M.; SCHRICK, F.N.; EDWARDS, J.L. Retinol improves development of bovine oocytes compromised by heat stress during maturation. Journal of Dairy Science, v.87, p.2449-2454, 2004.
MAYA-SORIANO, M.J.; TABERNER, E.; LOPEZ-BEJAR, M. Retinol improves in vitro oocyte nuclear maturation under heat stress in heifers. Zygote, p.1-8, 2012. Disponível em: http://dx.doi.org/10.1017/S0967199412000135 Acesso em 23 de setembro de 2013.
PAULA-LOPES, F.F.; HANSEN, P.J. Apoptosis is an adaptative response in bovine preimplantation embryos that facilitates survival after heat shock. Biochemical Biophysical Research Communications, v.295, p.37-42, 2002a.
PAULA-LOPES, F.F.; HANSEN, P.J. Heat Shock-Induced Apoptosis in Preimplantation Bovine Embryos. Is a Developmentally Regulated Phenomenon. Biology of Reproduction, v.66, p.1169–1177, 2002b.
PAYTON, R.R.; RISPOLI, L.A.; SAXTON, A.M.; EDWARDS, J.L. Impact of heat stress exposure during meiotic maturation on oocyte, surrounding cumulus cell, and embryo RNA populations. Journal of Reproduction and Development, v.57, n.4, 481-491, 2011.
RIVERA, R. M.; HANSEN, P. J. Development of cultured bovine embryos after exposure to high temperatures in the physiological range. Reproduction, v.121, p.107-115, 2001.
ROTH, Z.; HANSEN, P. J. Involvement of apoptosis in disruption of developmental competence of bovine oocytes by heat shock during maturation. Biology of Reproduction, v.71, p.1898-1906, 2004.
ROTH, Z. Heat stress, the follicle, and its enclosed oocyte: mechanisms and potential strategies to improve fertility in dairy cows. Reproduction in Domestic Animals, v.43, Suplemento 2, p. 238-244, 2008.
SHEHAB-EL-DEEN, M.A.M.M.; LEROY, J.L.M.R.; FADEL, M.S.; SALEH, S.Y.A.; MAES, D.; VAN SOOM, A. Biochemical changes in the follicular fluid of the dominant follicle of high producing dairy cows exposed to heat stress early post-partum. Animal Reproduction Science, v.117, p.189-200, 2010.
SIMPLÍCIO, A. A.; FREITAS, V. J. F.; FONSECA, J. F. Biotécnicas da reprodução como técnicas de manejo reprodutivo em ovinos. Revista Brasileira de Reprodução Animal, v.31, n.2, p.234-246, 2007.
TSENG, J.K.; CHEN, C.H.; CHOU, P.C.; YEH, S.P.; JU, J.C. Influences of follicular size on parthenogenetic activation and in vitro heat shock on the cytoskeleton in cattle oocytes. Reproduction in Domestic Animals, v.39, p.146-153, 2004.
WANG, J.Z.; SUI, H.S.; MIAO, D.Q.; LIU, N.; ZHOU, P.; GE, L.; TAN, J.H. Effects of heat stress during in vitro maturation on cytoplasmic versus nuclear components of mouse oocytes. Reproduction, v.137, p.181-189, 2009.

Datas de Publicação

Publicação nesta coleção
29 Out 2013
Data do Fascículo
Set 2013

Histórico

Recebido
08 Fev 2012
Aceito
13 Nov 2012

This work is licensed under a Creative Commons Attribution 4.0 International License.

[1] ANDREU-VAZQUEZ, C.; LOPEZ-GATIUS, F.; GARCIA-ISPIERTO, I.; MAYA-SORIANO, M.J.; HUNTER, R.H.F.; LOPEZ-BEJAR, M. Does heat stress provoke the loss of a continuous layer of cortical granules beneath the plasma membrane during oocyte maturation? Zygote, v.18, n. 4, p.293-299, 2010.

[2] BADINGA, L.; COLLIER, R.J.; THATCHER, W.W.; WILCOX, C.J. Effects of climatic and management factors on conception rate of dairy cattle in subtropical environment. Journal of Dairy Science, v.68, p.78-85, 1985.

[3] CHIAMENTI, A.; FILHO, C.R.; MOURA, M.T .; PAULA-LOPES, F.F.; NEVES, J.P.; NETO, C.C.; GON"CALVES, P.B.; LIMA, P.F.; OLIVEIRA, M.A. Use of retinyl acetate, retinoic acid and insulin-like growth factor-I (IGF-I) to enhance goat embryo production. Acta Veterinaria Hungarica v.61, p.116-124, 2013.

[4] EALY, A.D.; HOWELL, J.L.; MONTERROSO, V.H.; ARÉCHIGA, C.F.; HANSEN, P.J. Developmental changes in sensitivity of bovine embryos to heat shock and use of antioxidants as thermoprotectants. Journal of Animal Science, v.73, p.1401-1407, 1995.

[5] EDWARDS, J.L.; HANSEN, P.J. Differential responses of bovine oocytes and preimplantation embryos to heat shock. Molecular Reproduction and Development, v.46, p.138-145, 1997.

[6] EDWARDS, J.L.; BOGART, A.N.; RISPOLI, L.A.; SAXTON, A.M.; SCHRICK, F.N. Developmental competence of bovine embryos from heat-stressed ova. Journal of Dairy Science, v.92, p.563-570, 2009.

[7] HANSEN, P.J. Effects of heat stress on mammalian reproduction. Philosophical Transactions of The Royal Society Biological Sciences, v. 364, p. 3341-3350, 2009.

[8] INMET. Instituto Nacional de Meteorologia. Disponível em: <http:www.inmet.gov.br/html/prev_climatica_tempo/prognostico>. Acesso em: 23 abr. 2009.
» link

[9] JU, J.C.; PARKS, J.E.; YANG, X. Thermotolerance of IVM-derived bovine oocytes and embryos after shortterm heat shock. Molecular Reproduction and Development, v.53, p.336-340, 1999.

[10] JU, J.C.; TSENG, J.K. Nuclear and cytoskeletal alterations of in vitro matured porcine oocytes under hyperthermia. Molecular Reproduction and Development, v.68, p.125-133, 2004.

[11] KESKINTEPE, L.; SIMPLICIO, A.A.; BRACKETT, B.G. Caprine blastocyst development after in vitro fertilization with spermatozoa frozen in different extenders. Theriogenology, v.49, n.7, p.1265-1274, 1998.

[12] LAWRENCE, J.L.; PAYTON, R.R.; GODKIN, J.D.; SAXTON, A.M.; SCHRICK, F.N.; EDWARDS, J.L. Retinol improves development of bovine oocytes compromised by heat stress during maturation. Journal of Dairy Science, v.87, p.2449-2454, 2004.

[13] MAYA-SORIANO, M.J.; TABERNER, E.; LOPEZ-BEJAR, M. Retinol improves in vitro oocyte nuclear maturation under heat stress in heifers. Zygote, p.1-8, 2012. Disponível em: http://dx.doi.org/10.1017/S0967199412000135 Acesso em 23 de setembro de 2013.

[14] PAULA-LOPES, F.F.; HANSEN, P.J. Apoptosis is an adaptative response in bovine preimplantation embryos that facilitates survival after heat shock. Biochemical Biophysical Research Communications, v.295, p.37-42, 2002a.

[15] PAULA-LOPES, F.F.; HANSEN, P.J. Heat Shock-Induced Apoptosis in Preimplantation Bovine Embryos. Is a Developmentally Regulated Phenomenon. Biology of Reproduction, v.66, p.1169–1177, 2002b.

[16] PAYTON, R.R.; RISPOLI, L.A.; SAXTON, A.M.; EDWARDS, J.L. Impact of heat stress exposure during meiotic maturation on oocyte, surrounding cumulus cell, and embryo RNA populations. Journal of Reproduction and Development, v.57, n.4, 481-491, 2011.

[17] RIVERA, R. M.; HANSEN, P. J. Development of cultured bovine embryos after exposure to high temperatures in the physiological range. Reproduction, v.121, p.107-115, 2001.

[18] ROTH, Z.; HANSEN, P. J. Involvement of apoptosis in disruption of developmental competence of bovine oocytes by heat shock during maturation. Biology of Reproduction, v.71, p.1898-1906, 2004.

[19] ROTH, Z. Heat stress, the follicle, and its enclosed oocyte: mechanisms and potential strategies to improve fertility in dairy cows. Reproduction in Domestic Animals, v.43, Suplemento 2, p. 238-244, 2008.

[20] SHEHAB-EL-DEEN, M.A.M.M.; LEROY, J.L.M.R.; FADEL, M.S.; SALEH, S.Y.A.; MAES, D.; VAN SOOM, A. Biochemical changes in the follicular fluid of the dominant follicle of high producing dairy cows exposed to heat stress early post-partum. Animal Reproduction Science, v.117, p.189-200, 2010.

[21] SIMPLÍCIO, A. A.; FREITAS, V. J. F.; FONSECA, J. F. Biotécnicas da reprodução como técnicas de manejo reprodutivo em ovinos. Revista Brasileira de Reprodução Animal, v.31, n.2, p.234-246, 2007.

[22] TSENG, J.K.; CHEN, C.H.; CHOU, P.C.; YEH, S.P.; JU, J.C. Influences of follicular size on parthenogenetic activation and in vitro heat shock on the cytoskeleton in cattle oocytes. Reproduction in Domestic Animals, v.39, p.146-153, 2004.

[23] WANG, J.Z.; SUI, H.S.; MIAO, D.Q.; LIU, N.; ZHOU, P.; GE, L.; TAN, J.H. Effects of heat stress during in vitro maturation on cytoplasmic versus nuclear components of mouse oocytes. Reproduction, v.137, p.181-189, 2009.

Brasil

Brasil

Avaliação de embriões ovinos provenientes de oócitos submetidos a estresse calórico durante a maturação in vitro

Evaluation of sheep oocytes submitted to heat stress during in vitro maturation

Resumos

Datas de Publicação

Histórico