Supervisors info:
Λουτράδης Δημήτριος, Καθηγητής, Ιατρική, ΕΚΠΑ
Δρακάκης Πέτρος, Αναπληρ. Καθηγητής, Ιατρική, ΕΚΠΑ
Ντόμαλη Αικατερίνη, Επίκ. Καθηγήτρια, Ιατρική, ΕΚΠΑ
Summary:
In mammals, the formation of the zygote becomes apparent after the coalescence of the female and the male pronucleus in the ampulla of the fallopian tube and it is considered successful only when the implantation is achieved in the endometrium of the female. Throughout this track, pre-embryo undergoes many structural changes. Initially, the fertilized ovum is divided into 2, 4 and 8 cells, subsequently it is compacted by establishing the morula and finally it creates a cavity, forming the blastocyst.
During the third cell division of the zygote, approximately three days after the fertilization, the pre-embryo of 8 cells occurs, which is characterized as pluripotent. The significance of studying this particular embryonic stage is the fact that maternal developmental control is replaced by the activation and the expression of zygotic genes. In mouse, zygotic gene activation takes place at the 2-cell embryonic stage.
The cell cycle gene RPLP0 (ribosomal protein with a PO subunit) encodes human 60s ribosomal protein P0. It is a phosphoprotein based on the large arm of chromosome 12. It interacts with the acidic ribosomal proteins P1 and P2 to form the pentameric complex consisting of the dimers of P1 and P2 and the monomer of P0.
The P0 protein is homologous to the Escherichia coli prokaryotic L10 and it is detected in the cytoplasm and nucleus, but not in the kernel. The RPLP0 gene is very important for the proper functioning of protein synthesis.
In this diploma thesis, the expression of the RPLP0 gene in mouse precursors in the 8 cell stage is compared with the expression of the RPLP0 gene in dead mice embryos. The purpose of this procedure is to identify the potential molecular and cellular mechanisms associated with a normal embryonic development and therefore to be used as molecular biomarkers to predict the progression of the implanted embryo. Furthermore, the aim is to correlate the expression of the RPLP0 gene with the quality of the embryos.
Overall, 81 embryos were obtained from female mice, of which 18 embryos reached 8 cells and the 63 embryos were isolated from them. Subsequently, total RNA was isolated from the embryonic cells, followed by cDNA synthesis, and finally the study of RPLP0 gene expression by the Real Time PCR method.
According to the results, the RPLP0 gene is expressed both in dead embryos and embryos at the 8-cell stage. However, the gene expression in embryos at the 8-cells stage occurs in fewer cycles. Therefore, they have greater expression.
The comprehension of this mechanism and the establishment of an embryonic transcriptional profile at the 8-cell developmental stage, based on the expression or not of specific genes, could be a useful molecular tool in IVF to predict the progression of the implanted embryo.
Keywords:
RPLP0 gene, Mouse, Mus musculus, RT-PCR, Cellular cycle, Eight-cell stage, Fertilization
