Conclusion

Mitogenic growth factors are thought to play key regulatory roles in preimplantation embryo development by promoting nuclear and cytoplasmic maturation, increasing fecundity and rates of fertilization, optimizing cellular cleavage, and synchronizing embryonic and maternal maturation. Our results, in the context of our earlier work, strongly support the contention that mammalian embryos are particularly sensitive to the presence of mitogenic growth factors and that adequate levels and appropriately timed expression of these factors are crucial for successful preimplantation embryo development. This necessary role in development indicates that EGF and TGFa expression and in turn EGFR activation are essential for ensuring appropriate cell proliferation and differentiation and thus promote survival and normal preimplantation development.

Continue reading

Advertisements

Discussion – part 2

Our findings are consistent with results that indicate a lower rate of development and a greater incidence of programmed cell death (that is, apoptosis) in IVF and cloned embryos than in vivo-derived embryos. To a variable extent, these earlier observations were dependent on culture conditions, which likely affect gene expression and nuclear reprogramming. Our experimental design controlled for this variable by applying identical culture conditions to all 3 embryo groups. In this way, we could compare and contrast results between fertilized and cloned embryos in order to specifically address the role of mito-genic growth factors in preimplantation embryo development. Continue reading

Discussion

In the present study we determined that the development of mouse embryos derived by either natural mating or IVF was negatively affected by immunoneutralization of mitogenic growth factors and their receptor. Treatment with immunoneutralizing antibodies to EGF, TGFa, and EGFR decreased the rate of development of 1-cell-stage embryos to blastocysts, reduced the total number of differentiated cells in the ICM and TE, decreased cell size, reduced the ICM:TE ratio, and increased the fragmentation of nuclear DNA (that is, apoptosis). These differences were accentuated, in most cases, when embryos were treated simultaneously with neutralizing antibodies to both EGF and TGFa. In vitro-fer-tilized embryos appeared to be more sensitive than in vivo-de-rived embryos to immunoneutralizing antibodies. Therefore, the negative effect of neutralizing antibodies to mitgoegnic growth factors on the growth and differentiation characteristics of fertilized embryos indicates a direct relationship between the actions of EGF and TGFa and normal development of mouse embryos. Continue reading

Number of cells and differential labeling

Number of cells and differential labeling. As the total number of embryonic cells decreases, the developmental potential of preimplantation mammalian embryos decreases. Compared with that after control antibody treatments, the total number of cells (mean + SE) in in vivo-derived embryos was significantly (P < 0.05) less after treatment with neutralizing antibodies to EGF (68.8 + 0.9 cells), TGFa (67.8 + 1.6 cells), and EGFR (68.0 + 2.2 cells) and combined treatment with antibodies to both EGF and TGFa (59.4 + 0.7 cells; Table 3 and Figure 1 A, D, and G). Similar relative results were observed after treatment of in vitro-fertilized embryos with neutralizing antibodies to EGF (63.8 + 0.7 cells), TGFa (63.0 + 1.2 cells), and EGFR (63.6 + 1.3 cells) and combined treatment with antibodies to both EGF and TGFa (57.4 + 0.5). Cell size was decreased after immunoneutralization treatment (Figure 1 D). Continue reading

Apoptosis

Apoptosis. Apoptosis, or programmed cell death, is an important parameter by which to assess the developmental competence of preimplantation mammalian embryos. Apoptotic cells were detected in all groups of embryos after treatment with neutralizing antibodies (Table 2 and Figure 1 B and E). Compared with that after control antibody treatment, the numbers of cells (mean + SE) with apoptotic nuclei among in vivo-derived embryos were significantly (P < 0.05) greater after treatment with neutralizing antibodies to EGF (7.3 + 0.4 cells), TGFa (7.7 + 0.5 cells), and EGFR (7.5 + 0.3 cells) and combined treatment with antibodies to both EGF and TGFa (10.3 + 0.3 cells). Similar relative and statistically significant (P < 0.01) results were observed after treatment of in vitro-fertilized embryos with neutralizing antibodies to EGF (9.4 + 0.3 cells), TGFa (9.6 + 0.4 cells), and EGFR (9.4 + 0.2) and combined treatment with antibodies to both EGF and TGFa (10.8 + 0.5 cells). In both groups of embryos, after treatment with neutralizing antibodies, 92% to 94% of apoptotic cells were in the ICM (Figure 1 B and E). Continue reading