Derivation and characterization of pluripotent embryonic germ cells in chicken

Embryonic germ (EG) cell lines established from primordial germ cells (PGCs) are undifferentiated and pluripotent stem cells. To date, EG cells with proven germ-line transmission have been completely established only in the mouse with embryonic stem (ES) cells. We isolated PGCs from 5.5-day-old (stage 28) chicken embryonic gonads and established a putative chicken EG cell line with EG culture medium supplemented with stem cell factor (SCF), leukemia inhibitory factor (LIF), basic fibroblast growth factor (bFGF), interleukin-11 (IL-11), and insulin-like growth factor-I (IGF-I). These cells grew continuously for ten passages (4 months) on a feeder layer of mitotically active chicken embryonic fibroblasts. After several passages, these cells were characterized by screening with the periodic acid-Schiff reaction, anti-SSEA-1 antibody, and a proliferation assay. The chicken EG cells maintained characteristics of gonadal PGCs and undifferentiated stem cells. When cultured in suspension, the chicken EG cells successfully formed an embryoid body and differentiated into a variety of cell types. The chicken EG cells were injected into stage X blastodermal layer and produced chimeric chickens with various differentiated tissues derived from the EG cells. Chicken EG cells will be useful for the production of transgenic chickens and for studies of germ cell differentiation and genomic imprinting.