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How to spot a primordial germ cell

Marking primordial germ cells enables monitoring their generation from embryonic stem cells

By | September 16, 2003

Germline cells transmit genetic information from generation to generation, making them the focus of reproductive studies. In mice, primordial germ cells (PGCs) appear in gastrulating embryos at embryonic day 7.25 and have been generated in vitro from the outer germinal membrane of an ovum (epiblast), provided the epiblast is cocultured with cells expressing bone morphogenic protein (BMP) 4 and 8b. How the founder population of PGCs is separated from the rest of the pluripotent epiblast has been unclear because of technical difficulties in distinguishing PGCs from embryonic stem (ES) cells, since standard PGC markers (OCT4 and alkaline phosphatase) are also positive in ES cells. In the September 15 PNAS, Yayoi Toyooka and colleagues at the Mitsubishi Kagaku Institute of Life Sciences use the mouse vasa homolog (Mvh) as a molecular marker for PGCs and show that ES cells can form these germ cells in vitro (PNAS, DOI:10.1073/pnas.1932826100, September 15, 2003).

Toyooka et al. targeted the Mvh gene in an ES cell line with hybrid Mvh-GFP and Mvh-lacZ genes—MVH is a helicase that is specific for differentiating PGCs—and generated knock-in ES cell lines in which PGCs markers could be easily monitored by GFP and LacZ expression. When these cells were cultured in vitro as aggregates to form embryoid bodies, approximately 10% of the cells expressed GFP or LacZ. Purification of GFP- or LacZ-positive cells by flow cytometry also showed them to express GCNA1 and SYCP-3—markers specific to PGCs after they colonize the fetal gonad—suggesting these MVH-positive PGCs were equivalent to those found in the fetal gonad. The authors then tested the inducing effects of BMPs on PGC emergence by coculturing the knock-in ES cells with cells expressing either BMP4 or BMP8b. They observed that BMP4 increased the number of MVH-positive cells, while BMP8b did not, indicative of BMP4 being the critical protein for PGC emergence in a cell mass.

This suggests that the ES cell line used could generate PGCs in culture, but it gave no clue as to the developmental potential of these cells. Therefore, Toyooka et al. performed transplantation experiments with purified LacZ-positive ES-derived cells. After coculture with gonadal cells and transplantation into the male gonad, ES-derived PGCs differentiated into spermatogenic cells and, ultimately, sperm.

"We have demonstrated that ES cells are able to differentiate into germ-line cells in vitro" and "have the potential to undergo meiosis and produce sperm," conclude the authors. The easy detection of differentiating PGCs could facilitate advances in reproductive engineering.

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Comments

Avatar of: reza ranjbar

reza ranjbar

Posts: 1

November 5, 2006

i like to know every thing about pgcs,,,,,,

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