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157 EFFECT OF MELATONIN ON EMBRYO QUALITY OF BOVINE OOCYTES SUBJECTED TO HEAT SHOCK

Alves, N. G ; Ascari, I. J ; Camargo, L. S. A ; Jasmin, J ; Quintão, C. C. R ; Oliveira, J. A. S ; Souza, E. D ; Assunção, C. M. A ; Mendes, V. R. A ; Lima, R. R

Reproduction fertility and development, 2016, Vol.28 (2), p.208 [Periódico revisado por pares]

CSIRO Publishing

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  • Título:
    157 EFFECT OF MELATONIN ON EMBRYO QUALITY OF BOVINE OOCYTES SUBJECTED TO HEAT SHOCK
  • Autor: Alves, N. G ; Ascari, I. J ; Camargo, L. S. A ; Jasmin, J ; Quintão, C. C. R ; Oliveira, J. A. S ; Souza, E. D ; Assunção, C. M. A ; Mendes, V. R. A ; Lima, R. R
  • Assuntos: apoptosis ; blood serum ; carbon dioxide ; computer software ; cows ; estrus ; experimental design ; fluorometry ; follicle-stimulating hormone ; heat ; humidity ; in vitro fertilization ; linear models ; melatonin ; oocytes ; oxygen ; Poisson distribution ; slaughterhouses ; trophoblast ; zygote
  • É parte de: Reproduction fertility and development, 2016, Vol.28 (2), p.208
  • Notas: http://dx.doi.org/10.1071/RDv28n2Ab156
  • Descrição: The aim of this study was to evaluate the effect of different concentrations of melatonin added to in vitro maturation (IVM) medium of oocytes subjected to heat shock on embryo quality. Immature oocytes aspirated from ovaries obtained from a slaughterhouse were selected and randomly allocated in factorial experiment design (3×2). Three concentrations of melatonin (0, 10-6, and 10-4 M; M5250, Sigma, St. Louis, MO, USA) were added to the IVM medium and 2 incubation conditions (conventional: 24h at 38.5°C and 5% CO2; heat shock: 12h at 41°C followed by 12h at 38.5°C and 5% CO2) were tested, resulting in treatments: M1 (0 M; 38.5°C; n=15), M2 (10-6 M; 38.5°C; n=15), M3 (10-4 M; 38.5°C; n=15), M4 (0 M; 41°C; n=15), M5 (10-6 M; 41°C; n=15), and M6 (10-4 M; 41°C; n=15). The IVM was performed in a Nunc plate (144444 – Thermo, Fisher Scientific Inc., Pittsburgh, PA, USA) containing 400µL of TCM-199 (Invitrogen, Carlsbad, CA, USA) supplemented with 20µgmL-1 of FSH (Pluset®, Calier Laboratories, Barcelona, Spain) and 10% oestrus cow serum. Oocytes were IVF in FERT-TALP medium for 20–22h and incubated at 38.5°C and 5% CO2. After IVF, the presumptive zygotes were denuded and cultured in CR2aa medium supplemented with 2.5% FCS (Nutricell, Campinas, Brazil) in an incubator at 38.5°C under 5% CO2, 5% O2, and 90% N2, and saturated humidity for 8 days. Blastocysts with 8 days post-fertilization from different treatments were fixed in 4% paraformaldehyde in PBS for 1h and analysed by TUNEL assay (Deadend™ Fluorometric TUNEL System, Promega, Madison, WI, USA) to evaluate embryonic quality. Data were analysed by generalised linear models considering the Poisson distribution and using the Proc Genmod of SAS software (version 9.1; SAS Institute Inc., Cary, NC, USA) considering effects of melatonin concentration, incubation conditions, and interaction between the factors. Values shown are the mean±s.e.m. The interaction between melatonin concentration and incubation conditions was no significant (P>0.05). The total number of cells was not affected (P>0.05) by melatonin, but it was decreased (P<0.05) by heat shock (M1=117±6.7a; M2=118±4.2a; M3=120±6.3a; M4=102±6.2b; M5=106±5.7b; M6=108±8.9b). Melatonin and heat shock did not affect (P>0.05) the index of embryo apoptotic cells (M1=15.3%±2.0; M2=15.5%±1.3; M3=13.6%±2.0; M4=14.9%±1.5; M5=13.3%±1.3; M6=13.5%±1.2) and the index of trophoblast cells (M1=74.6%±2.3; M2=75.0%±1.7; M3=75.2%±1.9; M4=78.4%±2.3; M5=76.4%±3.0; M6=75.2%±2.6). The melatonin and heat shock affected the index of the inner cell mass (ICM; P<0.05), and the heat shock reduced the index of the ICM of oocytes not treated with melatonin (M1=25.4%±2.3a; M2=25.0%±1.7a; M3=24.8%±1.8a; M4=21.6%±2.3b; M5=23.6%±3.0a; M6=24.8%±2.6a). In conclusion, melatonin supplementation to the medium IVM of oocytes subjected to heat shock had no effect on blastocyst total cell number, general apoptotic index, or index of the trophoblast cells, but increased index of the ICM. Research was supported by Fapemig, CNPq, Embrapa, and CAPES.
  • Editor: CSIRO Publishing
  • Idioma: Inglês
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  • Realização: ABCD-USP USP   Logos de Redes Sociais: Freepik

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