Abstract
The last 100 years have seen a concerning decline in male reproductive health associated with decreased sperm production, sperm function and male fertility. Concomitantly, the incidence of defects in reproductive development, such as undescended testes, hypospadias and testicular cancer has increased. Indeed testicular cancer is now recognised as the most common malignancy in young men. Such cancers develop from the pre-invasive lesion Carcinoma in Situ (CIS), a dysfunctional precursor germ cell or gonocyte which has failed to successfully differentiate into a spermatogonium. It is therefore essential to understand the cellular transition from gonocytes to spermatogonia, in order to gain a better understanding of the aetiology of testicular germ cell tumours. MicroRNA (miRNA) are important regulators of gene expression in differentiation and development and thus highly likely to play a role in the differentiation of gonocytes. In this study we have examined the miRNA profiles of highly enriched populations of gonocytes and spermatogonia, using microarray technology. We identified seven differentially expressed miRNAs between gonocytes and spermatogonia (down-regulated: miR-293, 291a-5p, 290-5p and 294*, up-regulated: miR-136, 743a and 463*). Target prediction software identified many potential targets of several differentially expressed miRNA implicated in germ cell development, including members of the PTEN, and Wnt signalling pathways. These targets converge on the key downstream cell cycle regulator Cyclin D1, indicating that a unique combination of male germ cell miRNAs coordinate the differentiation and maintenance of pluripotency in germ cells.
Highlights
Over the last 100 years there has been a substantial increase in diseases of the male reproductive system including developmental abnormalities, poor semen quality and testicular cancer, especially in developed countries [1,2,3]
In order to interrogate the changes in miRNA expression between gonocytes and spermatogonia, these cell populations were isolated and confirmed as germ cells
Analysis with qRT-PCR revealed a number of significant differences in the expression germ/pluripotency/stem cell markers, including Oct3/ 4 and Ngn3 which were up-regulated in the spermatogonia cell population and, Nanog and Plzf which were down-regulated in the spermatogonia cell population (Fig. 1C)
Summary
Over the last 100 years there has been a substantial increase in diseases of the male reproductive system including developmental abnormalities, poor semen quality and testicular cancer, especially in developed countries [1,2,3]. There is concern that exposure to environmental toxicants in utero, especially endocrine disruptors, augments a genetic predisposition for testicular germ cell tumours such as defects in kit signalling (KITLG itself and SPRY an inhibitor of kit stimulated MAPK signalling), apoptosis (BAK), sex determination (DMRT1) and telomere regulation (TERT) [5,6]. This is pertinent given that the gonocyte-to-spermatogonia transition occurs in late gestation in humans [7] and that Carcinoma in Situ (CIS) cells have previously been identified as arising from arrested/dysfunctional gonocytes [8]. A study of 130 non-redundant proteins in Sertoli cell-specific Dicer knockout mice, demonstrated that 50 proteins were up-regulated and three proteins were downregulated, indicating that miRNA are primarily involved in the inhibition of protein translation [13]
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