In search of rat stem Leydig cells: Identification, isolation, and lineage-specific development

Abstract

Leydig cells (LCs) are thought to differentiate from spindle-shaped precursor cells that exhibit some aspects of differentiated function, including 3␤-hydroxysteroid dehydrogenase (3␤HSD) activity. The precursor cells ultimately derive from undifferentiated stem LCs (SLCs), which are postulated to be present in testes before the onset of precursor cell differentiation. We searched for cells in the neonatal rat testis with the abilities to: (i) proliferate and expand indefinitely in vitro (self renew); (ii) differentiate (i.e., 3␤HSD and ultimately synthesize testosterone); and (iii) when transplanted into host rat testes, colonize the interstitium and subsequently differentiate in vivo. At 1 week postpartum, spindle-shaped cells were seen in the testicular interstitium that differed from the precursor cells in that they were 3␤HSD-negative, luteinizing hormone (LH) receptor (LHR)-negative, and platelet-derived growth factor receptor ␣ (PDGFR␣)-positive. These cells were purified from the testes of 1-week-old rats. The cells contained proteins known to be involved in LC development, including GATA4, c-kit receptor, and leukemia inhibitory factor receptor. The putative SLCs expanded over the course of 6 months while remaining undifferentiated. When treated in media that contained thyroid hormone, insulin-like growth factor I, and LH, 40% of the putative SLCs came to express 3␤HSD and to synthesize testosterone. When transplanted into host rat testes from which LCs had been eliminated, the putative SLCs colonized the interstitium and subsequently expressed 3␤HSD, demonstrating their ability to differentiate in vivo. We conclude that these cells are likely to be the sought-after SLCs.c-kit ͉ leukemia inhibitory factor ͉ platelet-derived growth factor receptor ␣ ͉ puberty ͉ steroidogenesis L eydig cells (LCs) are the primary source of testosterone in the male, and their differentiation in the testes during puberty is a signature event in the development of the male body plan. It is hypothesized, but far from proven, that LCs first arise from undifferentiated stem cells [stem LCs (SLCs)] (1-3). It has been suggested that, in rats, the putative SLCs are present in the testis at birth, and that by 11 days postpartum, at least some of their progeny express LC-specific genes and thus become committed to the LC lineage (4, 5).The committed cells subsequently undergo phased transitions through progenitor and immature stages and ultimately to terminally differentiated adult LC stage (6). In particular, progenitor LCs (PLCs) form during days 12-28 postpartum (presumably from SLCs). The PLCs proliferate and also exhibit some aspects of differentiated function, including 3␤-hydroxysteroid dehydrogenase (3␤HSD) activity (7). Luteinizing hormone (LH) receptors (LHRs) first appear as the PLCs differentiate, suggesting that SLCs are likely to be independent of LH control (8). The development of the steroidogenic capacity of PLCs requires stimulation by LH (9). The mitotic activity of PLCs gradually is reduced, and the cells enlarge...

Role of androgen and gonadotrophins in the development and function of the Sertoli cells and Leydig cells: Data from mutant and genetically modified mice

“…Results showed that there was a marked but transient increase in Pdgfa and Leukemia inhibitory factor (Lif) transcript levels 24h after treatment with EDS, at a time when Leydig cell ablation was nearly complete and regeneration starting . Both LIF and PDGFA are reported to stimulate Leydig cell stem cell proliferation (Ge et al, 2006), consistent with a role in Leydig cell regeneration after EDS. Surprisingly there was a significant decrease in Dhh levels 24h after EDS treatment which recovered by 48h .…”

“…In the absence of DHH and PDGFA adult Leydig cells fail to develop but this may be related to a failure of stem cell development or proliferation (Clark et al, 2000;Gnessi et al, 2000;Park et al, 2007). It has been shown that the adult testis contains stem cells which will give rise to Leydig cells (Ge et al, 2006) but the origin of these cells and where they reside within the normal testis remains unknown. Surprisingly, the immediate progenitors of the adult Leydig cells also remain in some doubt.…”

Steroidogenesis and steroidogenic gene expression in postnatal fetal rat Leydig cells

“…We further explored the role of cAMP-dependent signaling as well as PDGFα and LIF, the paracrine factors that control Leydig cell development (Gnessi et al, 2000;Ge et al, 2006), in the regulation of PFLC proliferation. We observed that PDGFα significantly (1.8-fold over control, P<0.05) stimulated the proliferation of PFLCs.…”

Isolation, characterization, andin vitroandin vivodifferentiation of putative thecal stem cells

“…Leydig cells can remain undifferentiated in vitro for months and can respond to growth factors to differentiate into steroidogenic precursor cells (5). After transplantation into the testes of Leydig cell-deficient mice, these stem cells recolonize normally in the host testicular interstitium and restore the serum testosterone concentration (4).…”

“…In contrast to theca cells, their male counterparts, Leydig cells, have been studied in detail because their putative stem cells can be isolated from mouse and rat testes by flow cytometric sorting (4,5). Leydig cells can remain undifferentiated in vitro for months and can respond to growth factors to differentiate into steroidogenic precursor cells (5).…”