Germ cells within the male and female gonads develop from an apparently homogeneous bi-potential population of PGC. In both male and female human fetal gonads germ cells initially proliferate but thereafter their fates diverge with female germ cells entering meiotic prophase as early as 11 weeks gestation (9 weeks post conception; ) whereas meiotic entry is does not occur until puberty in males. In rodents male germ cells removed from the testicular environment and placed in vitro can spontaneously enter meiosis emphasising the common developmental potential of the germ cells in the two sexes and the critical role played by the testicular environment in modifying germ cell fate [25, 26].
In the current study we have used OCT4 immunohistochemistry to identify the subpopulation of putative 'germ stem' cells in human fetal testes and ovaries. Studies from our own laboratory  have already established that expression of OCT4 is restricted to the gonocyte population of human testicular germ cells whilst others  have reported that this protein is localised to oogonia in the ovary. Expression of DAZL and VASA was compared to OCT4 as studies in knockout mice have reported that both genes are essential for germ cell maturation in that species [14, 19]. Analysis of ovarian extracts revealed a striking increase in the total amount of both DAZL and VASA mRNA and protein in the ovary as development progressed into the 2nd trimester. A similar trend in expression was noted in the testes but was less striking. Expression of OCT4 was maintained in both ovary and testis at all gestational ages examined consistent with persistence of a putative germ 'stem cell' population for many months in the human gonads although the extent to which these cells can acquire/maintain pluripotency in vitro remains unresolved (reviewed in ). In the 2nd trimester testis all seminiferous cords contained a mixture of OCT4 positive and OCT4 negative germ cells a situation which is different to that in rodents where the expression of OCT4 is down regulated synchronously in all germ cells within the cords before birth [29, 30]. In the human fetal ovary expression of OCT4 protein was restricted to oogonia in the peripheral zone of the organ, in agreement with the observations of Stoop et al.. In mice expression of Oct4 mRNA is extinguished in a rostro-caudal wave coincident with meiotic entry of germ cells .
Co-incident and overlapping patterns of expression of these germ cell specific proteins were revealed by fluorescent co-immunolocalisation on fixed specimens from a range of gestational ages. Previous studies have immunolocalised DAZL in a small number of samples recovered in the 2nd trimester [32–34] but we believe this is the first study to document expression in 1st trimester samples and to compare expression with that of OCT4 and VASA. In the 1st trimester, DAZL was detected in the nuclear compartment in OCT4 positive germ cells in both sexes. In a previous study we immunolocalised DAZL to the nuclei of male germ cells in a single sample at 17 weeks gestation and in the cytoplasm of female germ cells at 15 weeks . Other studies have claimed that DAZL is present in both the cytoplasm and nucleus of male germ cells at 20–21 weeks . We have extended these observations and demonstrated that DAZL is localised to the nuclear compartment in OCT4 positive cells (gonocytes and oogonia). In the ovaries localisation of DAZL to the cytoplasmic compartment appeared to be transient occurring as OCT4 expression but thereafter the amount of protein declined and was low in the largest ovarian germ cells, i.e. those that had formed into primordial follicles and those approaching that stage. This was evident when a morphometric analysis was performed showing that the germ cells immunopositive for DAZL alone were smaller than those containing both DAZL and VASA, which were in turn smaller than those only immunopositive for VASA.
Two studies on human populations have provided preliminary evidence that expression of DAZL is important for normal functioning of the human germ line. The first study reported a strong association between several common single nucleotide polymorphisms (SNPs) and age at menopause in a sample population of 324 women . In a second study a patient with premature ovarian failure at age 34 was found to contain a homozygous mutation of DAZL (Arg to Gly at 115) in a region of the protein critical for RNA binding . The situation in men is complicated by the presence of multiple copies of the closely related DAZ gene on the Y chromosome, deletions of which have been frequently documented as a cause of male infertility . However a patient with a homozygous mutation in DAZL (Asn 10 Cys) was reported to be azoospermic .
The marked increase in expression of VASA between 9 and 14 weeks gestation suggested to us that this was coincident with the entry of female germ cells into meiosis. However increases in both protein and mRNA were also demonstrated in male germ cells suggesting that the onset of expression of VASA was associated the maturation of the gonocytes into prespermatogonia rather than meiotic entry per se. In the ovary VASA was detected in the cytoplasm of in slightly larger, more mature germ cells than DAZL, and was also present in oocytes within primordial follicles. Previous studies that have detected expression of VASA in cytoplasm of germ cells within the fetal ovary and testes at 17 weeks gestation [9, 21, 37]. Co-staining of sections with OCT4 and VASA has extended these findings by demonstrating that VASA protein is not expressed in gonocytes or oogonia in the 1st trimester and provides a useful method for identifying the different populations of germ cells present within the 2nd trimester gonads.
Examination of the 3' UTR of the Mvh mRNA has revealed a number of putative Dazl-binding sites that are conserved between human, rat and mouse . Our data provide some additional support for a role for DAZL in initiating expression of VASA although further studies are required preferably using isolated human germ cells. DAZL has also been implicated in the regulation of other conserved germ-cell RNA-binding proteins including PUM2 , the human homolog of Pumilio that is required for maintenance of germ line stem cells in Drosophila and Caenorhabditis elegans.