In vitro buffalo embryo production under heat stress: Role of granulosa cells co-culture as a monolayer

Effect ofheat shock and co-culture of monolayer granulosa cells (GCs) on in vitroembryo developmental competence in Egyptian buffalo was the aim of the present study.Cumulus oocytes complexes (COCs) were collected from ovaries (n= 448) of cyclicslaughtered buffalo cows. Good quality immature oocytes (n= 1512) were subjected to in vitro maturation and fertilization. Post 18-22 h of fertilization, presumptive embryos were randomly assigned into four groups: (G1) No heat shock (38.5^OC), (G2) Heat shock (40.5°C) and (G3) Co-culture with GCs monolayer and heat shock or (G4) Co-culture with GCs monolayer and no heat shock. Heat-shockedembryos were exposed to temperature of40.5°Cfor the first two hours of culture before exposing to 38.5°C afterward up to the end of culture perio            d (8 days). The data were analyzed by General LinearUnivariate model using SPSS.
Oocyte expansionrate was 90.8±1.1%. This percentage is higher than nuclear maturation rate (oocytes at Telophase and Metaphase II) as detected byaceto-orcein staining by about 17%. Cleavage rate as recorded at day 3 post fertilization (day 0) was higher (p≤0.05) for G1 and G4 (71.1±10.5% and 70.5±7.9 with GCs, respectively).On the other hand, cleavage rate was dramatically decreased (P ≤0.05) for embryos of G2 (43.7±7.0%) compared with that of G3 (80.2±7.0%). In addition, embryos of G3showed approximately the same rate ofdeveloped embryos(Morula and blastocyst stages at day 8 of culture) as of G1 (50.9±5.3 and51.7±7.9%, respectively). In conclusion, using co-culture of GCs as a monolayer enhances cleavage rate and subsequent embryo development of buffalo embryos under heat stress condition.