Developmental regulation and modulation of apoptotic genes expression in sheep oocytes and embryos cultured in vitro with L-carnitine

Reprod Domest Anim. 2016 Dec;51(6):1020-1029. doi: 10.1111/rda.12789. Epub 2016 Sep 30.

Abstract

The objective of this study was to find out the impact of L-carnitine (10 mM) on developmental regulation of preimplantation sheep embryos cultured in vitro when supplemented in maturation medium and post-fertilization medium separately. Subsequent objective was to observe the L-carnitine-mediated alteration in expression of apoptotic genes (Bcl2, Bax, Casp3 and PCNA) in sheep oocytes and developing embryos produced in vitro. Oocytes matured with L-carnitine showed significantly (p < .05) higher cleavage (67.23% vs 43.12%), morula (47.65% vs 28.58%) and blastocysts (32.12% vs 13.24%) percentage as compared to presumptive zygotes cultured with L-carnitine during post-fertilization period. So it is suggested to use L-carnitine during maturation than post-fertilization period. Antiapoptotic and proliferative effects of L-carnitine were confirmed by inducing culture medium with actinomycin D (apoptotic agent) and TNFα (antiproliferative agent), respectively, with and without L-carnitine. Oocytes and embryos cultured with actinomycin D and TNFα showed developmental arrest with significant (p < .05) decrease in morula and blastocysts percentage but supplementation of L-carnitine to actinomycin D and TNFα induced culture medium showed similar result as that of control. L-carnitine supplementation during IVM significantly (p < .05) upregulated the expression of Bcl2 and PCNA genes in majority of the developmental stages. Although L-carnitine upregulated the expression of Bax in initial developmental stages but downregulated at latter part, whereas the expression of Casp3 was upregulated upto 16-cell stage but after that there was no difference in expression. Expression of GAPDH gene was not affected by L-carnitine supplementation. In conclusion, L-carnitine acted as an antiapoptotic and proliferative compound during embryo development and supplementation of L-carnitine during IVM altered the expression of apoptotic genes in the developmental stages of embryos.

MeSH terms

  • Animals
  • Carnitine / chemistry
  • Carnitine / pharmacology*
  • Dactinomycin / pharmacology
  • Embryo Culture Techniques / veterinary*
  • Embryo, Mammalian / drug effects
  • Embryonic Development / drug effects
  • Gene Expression Regulation, Developmental
  • Oocytes / drug effects*
  • Sheep / embryology*
  • Transcriptome
  • Tumor Necrosis Factor-alpha / pharmacology

Substances

  • Tumor Necrosis Factor-alpha
  • Dactinomycin
  • Carnitine