ReviewThe regulation of spermatogenesis by androgens
Introduction
Male fertility is dependent upon the successful perpetuation of spermatogenesis, the multi-step process of male germ cell expansion and development that occurs within the seminiferous tubules of the testes. Although other hormones facilitate the process of spermatogenesis, only the steroid hormone testosterone is essential to maintain spermatogenesis. Testosterone actions in the testis in relation to the regulation of spermatogenesis have been discussed in recent reviews [1], [2], [3], [4], [5], [6]. Here, we summarize the spermatogenesis processes regulated by testosterone, cell specific actions of testosterone as well as the intracellular pathways and genes that are controlled by testosterone signaling in the testis.
Spermatogenesis occurs in the seminiferous tubules of the testis. The seminiferous tubules are composed of three major cell types: peritubular myoid (PTM) cells, Sertoli cells and germ cells. PTM cells surround the external wall of the tubule and contract to force sperm down the tubule. Sertoli cells relay external signals and provide factors required for the proliferation and differentiation of germ cells. PTM cells cooperate with Sertoli cells to produce the basement membrane of the seminiferous tubule and provide the niche for spermatogonial stem cells (SSCs) that produce the germ cells that will develop into sperm [7], [8]. The cytoplasm of Sertoli cells extends from the basement membrane to the lumen of the tubule surrounding the developing germ cells. Leydig cells are present in the interstitial space between the tubules and produce testosterone, which diffuses into the seminiferous tubules, as well as blood vessels in the interstitial space (Fig. 1).
The division of SSCs along the basement membrane of the seminiferous tubule initiates the spermatogenesis process. The proliferation of SSCs results in either the production of two new stem cells to retain the stem cell pool or undifferentiated spermatogonia that are destined to develop into sperm. The undifferentiated spermatogonia undergo a series of mitotic divisions with incomplete cytokinesis to form chains of spermatogonia. Once the chains reach a length of 16 or 32 cells, they undergo differentiation en mass to become differentiated spermatogonia that are committed to becoming sperm. The differentiated spermatogonia undergo a series of divisions with a final mitosis resulting in the production of preleptotene spermatocytes that initiate the process of meiosis. At the conclusion of meiosis, haploid round spermatids are produced that undergo differentiation into elongated spermatids and then finally spermatozoa (Fig. 1).
Spermatogenesis is supported by somatic Sertoli cells that surround and nurture the developing germ cells. Sertoli cells contribute to the niche required to maintain the renewal of spermatogonial stem cells so that developing germ cells can be produced continuously. Sertoli cells also provide growth factors and nutrients for the developing germ cells. Specialized adhesion junctions are formed between adjacent Sertoli cells that in total form the blood testis barrier (BTB) near the basement membrane of the seminiferous tubules. The BTB divides the seminiferous tubule into basal and adluminal compartments. During the initial preleptotene stage of meiosis, spermatocytes “pass through” the BTB moving from the basement membrane to adluminal compartment. Once through the BTB, the germ cells continue to develop into spermatozoa in a defined, protected microenvironment. However, because the BTB denies germ cells in the adluminal compartment access to factors supplied by the circulatory system, the Sertoli cell must provide for the needs of the more mature germ cells [9], [10].
Testosterone is the major androgen in the testis that regulates spermatogenesis. Testosterone is produced by the Leydig cell in response to stimulation with luteinizing hormone (LH) and acts as a paracrine factor that diffuses into the seminiferous tubules. Androgen effects are mediated by the androgen receptor (AR, also denoted NR3C4), which is a 110 kD protein localized to the nucleus and cytoplasm. There are no functional receptors for androgen in germ cells [11], [12], [13], [14]. Instead the testosterone that diffuses into Sertoli cells binds to the AR present in the cytoplasm and nucleus to initiate the functional responses required to support spermatogenesis. In the testis, testosterone also interacts with AR expressed in Leydig, PTM cells, arteriole smooth muscle and vascular endothelial cells.
Because of the localized production of testosterone from Leydig cells, testosterone levels in the testes of men and rodents are 25–125-fold higher than that present in serum [15], [16], [17], [18], [19]. The physiological importance of high testosterone levels in the testis is not fully understood. However, it has been established that sperm production decreases exponentially once testosterone levels in the testis fall below 70 mM [20]. The high levels of testosterone in the testis cannot be explained by a sequestration mechanism to “deactivate” the hormone because at least two thirds of testicular testosterone is free or weakly bound to albumen and is bioavailable. Only one third of testosterone is tightly bound by sex hormone binding globulin (SHBG) or androgen binding protein (ABP) [21], [22]. Thus, the bioavailable testosterone in the testis greatly exceeds the kD for AR binding of approximately 1 mM [23].
Section snippets
Developmental patterns of AR expression
In humans and rodents, AR is expressed in PTM cells at high levels from the fetal period throughout adulthood [24], [25], [26], [27]. Adult Leydig cells also express AR at a constant level [28]. Sertoli cells do not express AR in fetal life [29]. In humans, AR is first detectable in the nuclei of a few Sertoli cells at the age of 5 months. Labeling is weak until 4 years of age and increases thereafter [24], [25], [26], [27]. In monkeys, androgen binding activity of Sertoli cells cultured from
Testosterone regulation of essential spermatogenesis processes
Testosterone is required for at least four critical processes during spermatogenesis: maintenance of the BTB, meiosis, Sertoli–spermatid adhesion and sperm release (indicated by the numbers 1, 2, 3, and 4 in Fig. 1).
Classical testosterone signaling
AR is the only specific receptor for androgens that has been identified. However, AR is capable of transmitting testosterone signals by at least 2 mechanisms, the classical and non-classical pathways. In the classical signaling pathway, testosterone that diffuses through the cell membrane interacts with AR that is often sequestered in the cytoplasm by heat shock proteins (Fig. 2, pathway 1). After binding androgen, a conformational change in AR allows the receptor to disengage from the heat
Mouse cell specific AR knock out and over expression models
To examine the functions of testosterone signaling in various cells in the testis, several groups have developed mouse models in which AR expression is ablated in specific cell types in the testis. Studies of the cell specific AR knock out mice plus gain of function mouse models have been the subject of recent reviews [1], [4], [73] and are summarized below as well as in Table 1.
Animal models of testosterone depletion and enhancement
Until the employment of RiboTag mouse + RNA seq strategies (see Section 6.2) studies of AR regulated gene expression have provided some important but somewhat limited information regarding testosterone-mediated gene expression that is required for spermatogenesis. Mice in which testosterone levels were depleted or enhanced in all cells were the first models used to assay gene expression in the testis. These models included normal 8 day-old neonatal mice injected with testosterone propionate (TP)
Summary and conclusions
Testosterone is required for processes that are critical for spermatogenesis including maintaining the BTB, supporting the completion of meiosis, the adhesion of elongated spermatids to Sertoli cells and the release of sperm. Previously, studies of testosterone and AR actions have been centered on the Sertoli cell but increasingly testosterone actions in other cell types including PTM and vascular smooth muscle cells are being found to affect processes that occur in the seminiferous tubules.
Acknowledgements
This work was supported by the UK Medical Research Council (L.B.S.) and the Magee Womens Research Institute (W.H.W.).
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