COMPARISON OF GENE EXPRESSION IN INDIVIDUAL PREIMPLANTATION BOVINE EMBRYOS PRODUCED BY IN VITRO FERTILIZATION OR SOMATIC CELL NUCLEAR TRANSFER

Authors: DE A. CAMARGO, L - EMBRAPA; Powell, Anne; DO VALE FILHO, V - U OF MINAS GERAIS, BRAZIL; Wall, Robert

Submitted to: Journal Of Reproduction, Fertility And Development
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/14/2005
Publication Date: 5/1/2005
Citation: De A. Camargo, L.S., Powell, A.M., Do Vale Filho, V.R., Wall, R.J. 2005. Comparison of gene expression in individual preimplantation bovine embryos produced by in vitro fertilization or somatic cell nuclear transfer. Journal of Reproduction, Fertility and Development. 17:469-487.

Interpretive Summary: Bovine somatic cell nuclear transfer (cloning) is associated with a higher incidence of developmental abnormalities and neonatal viability problems than other artificial reproductive technologies. It is likely that these defects are the result of inappropriate gene expression, possibly associated with imperfect reprogramming of the cloned embryos nucleus. To determine if unusual patterns of gene expression could be detected in the first week of development, the expression of 11 important genes were compared in cloned bovine embryos, those produced by in vitro fertilized embryos and in vivo fertilized embryos harvested from cows at a week of age. It was found that the cloned embryos and the in vitro fertilized embryos, both produced of the laboratory, had nearly identical patterns of gene expression. Their pattern of expression was also not dramatically different when compared to 'normal,' in vivo produced, embryos expression. However, the in vivo produced embryos expressed the 11 genes a much higher levels then did those produced in vitro. There results suggest that culturing embryos in the laboratory for a week may have more influence on their gene expression than the procedures used to produce them (cloning vs. fertilization). At least based on these 11 genes it is not possible to explain why the cloned pregnancies fail at a higher rate then those established with in vitro fertilized embryos.

Technical Abstract: In vitro fertilization (IVF) and somatic cell nuclear transfer (SCNT) have been implicated in a variety of developmental abnormalities. Aberrant gene expression is likely to account for much of the diminished viability and developmental abnormalities observed. In this study, expression of multiple genes in IVF and SCNT bovine blastocyst stage embryos were evaluated and compared to in vivo produced embryos. Eleven genes expressed at and following maternal-zygotic transcription transition were evaluated in individual blastocysts by real-time PCR following RNA amplification. A subset of those genes were also evaluated in individual IVF and SCNT 8-cells embryos. A fibroblast specific gene, expressed by nuclear donor cells, was also evaluated in IVF and SCNT embryos. The observed gene expression pattern at the 8-cell stage was not different between IVF and SCNT embryos (P>0.05). IVF and SCNT blastocysts expression was lower (P<0.01) for all genes than for their in vivo produced counterparts, except for lactate dehydrogenase isoenzyme A (P<0.001). The patterns of gene expression of the IVF and SCNT blastocysts were indistinguishable. Neither SCNT 8-cell nor blastocysts stage embryos expressed the gene used as a fibroblast marker, collagen VI'1. For the genes evaluated, the level of expression was influenced more by the environment than by the method used to produce them. These results support the notion that if developmental differences observed in IVF and SCNT produced fetuses and neonates are the result of aberrant gene expression during the preimplantation stage, those differences in expression are subtle.