Biology session

Chicken primordial germ cells (PGCs) are the precursors of the germ cells, which later become mature sperm or egg cells during differentiation. These cells are located in the epiblast of a stage X embryo. Later the PGCs translocate anteriorly to the germinal crescent and begin to divide. Within 48-56 hours they migrate through the vascular system and reach the developing genital ridges where they become gonadal germ cells.

Chicken PGCs are obtained from embryonic blood of 2-day old embryos. After isolation it is possible to maintain the cells in long-term in vitro culture or freeze them.

In chickens, as in all the known bird species, cells in females contain ZW, whereas males ZZ sex chromosomes. Previously, it was only possible to culture the male PGCs, but nowadays researchers created a specific medium, which supported also the female PGC culture. Our aim was to investigate the difference between the male and female cell lines established in our laboratory recently.

During my work, I examined the expression profile of pluripotency related markers and miRNAs in female and male PGC cultures. Using this method we were able to monitor the changes in the expression level and characterize the different cell lines.

We determined the gender of the donor embryos by sex PCR. Pooled PGC cultures were produced by mixing 5 male (pooled male) and 5 female (pooled female) individual cell lines. After 2 weeks the pools were frozen. After thawing the cells we could continue the culture. Samples were collected from these two pooled female and male cell lines at different time points. The expression of Cvh, cPouv, cNanog and miR-302a expression was investigated by quantitative PCR and using immunostaining.

Based on the results, there is no significant difference between the expression of germ and stem cell specific markers in male and female cultures. To prove the functionality and migrating capability of the cell lines we injected back labelled cells to 3-day old recipient embryos. The incorporation was successful in case of both sexes. In the future, I would like to examine the mixed (containing male and female PGCs) cell cultures too.

Our work is important due to gene preservation purposes. We can create germline chimeras if we inject PGCs into recipient animals. Thanks to this methodology conservation of threatened bird species, bird species on verge of extinction and preservation of valuable indigenous breeds are possible.