NR5A1 is required for functional maturation of Sertoli cells during postnatal development

The orphan nuclear receptor steroidogenic factor 1 (NR5A1 (SF-1)) is expressed in both Sertoli and Leydig cells in the testes. This study investigates the postnatal development of the testes of a gonad-specific Nr5a1 knockout (KO) mouse, in which Nr5a1 was specifically inactivated. The KO testes appeared histologically normal from postnatal day 0 (P0) until P7. However, disorganized germ cells, vacuoles, and giant cells appeared by P14 in the seminiferous tubules of KO but not control mice. Expression of NR5A1 and various factors was examined by immunohistochemistry (IHC). The number of NR5A1-positive Sertoli cells in the KO testes was lower compared with controls at all the developmental stages and decreased to nearly undetectable levels by P21. IHC for anti-Müllerian hormone and p27, immature and mature Sertoli cell markers, respectively, indicated a delay in Sertoli cell maturation in the KO testes. The number of Sertoli cell-expressing factors involved in Sertoli cell differentiation including WT1, SOX9, GATA4, and androgen receptor were lower in the KO testes compared with controls. Furthermore, fewer proliferating cell nuclear antigen-positive proliferative germ cells were observed, and the number of TUNEL-labeled cells was significantly higher in the KO testes compared with controls at P14 and P21, indicating impaired spermatogenesis. IHC for CYP11A1 (SCC) indicated the presence of steroidogenic Leydig cells in the interstitium of the KO testes at all stages examined. These results suggest that NR5A1 is essential for Sertoli cell maturation and therefore spermatogenesis, during postnatal testis development.

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Male Reproductive Function

Textbook of Endocrine Physiology ◽

10.1093/oso/9780199744121.003.0012 ◽

2011 ◽

Author(s):

William J. Kovacs

Keyword(s):

Tight Junctions ◽

Sertoli Cell ◽

Germ Cells ◽

Sertoli Cells ◽

Leydig Cells ◽

Reproductive Function ◽

Rete Testis ◽

Seminiferous Tubules ◽

Male Reproductive Function ◽

Peritubular Tissue

The testes are the source of both germ cells and hormones essential for male reproductive function. The production of both sperm and steroid hormones is under complex feedback control by the hypothalamic-pituitary system. The testis consists of a network of tubules for the production and transport of sperm to the excretory ducts and a system of interstitial cells (called Leydig cells) that express the enzymes required for the synthesis of androgens. The spermatogenic or seminiferous tubules are lined by a columnar epithelium composed of the germ cells themselves as well as supporting Sertoli cells surrounded by peritubular tissue made up of collagen, elastic fibers, and myofibrillar cells. Tight junctions between Sertoli cells at a site between the spermatogonia and the primary spermatocyte form a diffusion barrier that divides the testis into two functional compartments, basal and adluminal. The basal compartment consists of the Leydig cells surrounding the tubule, the peritubular tissue, and the outer layer of the tubule containing the spermatogonia. The adluminal compartment consists of the inner two-thirds of the tubules containing primary spermatocytes and germ cells in more advanced stages of development. The base of the Sertoli cell is adjacent to the basement membrane of the spermatogenic tubule, with the inner portion of the cell engulfing the developing germ cells so that spermatogenesis actually takes place within a network of Sertoli cell cytoplasm. The mechanism by which spermatogonia pass through the tight junctions between Sertoli cells to begin spermatogenesis is unknown. The close proximity of the Leydig cell to the Sertoli cell with its embedded germ cells is thought to be critical for normal male reproductive function. The seminiferous tubules empty into a network of ducts termed the rete testis. Sperm are then transported into a single duct, the epididymis. Anatomically, the epididymis can be divided into the caput, the corpus, and the cauda regions. The caput epididymidis consists of 8 to 12 ductuli efferentes, which have a larger lumen tapering to a narrower diameter at the junction of the ductus epididymidis.

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Minireview: Transcriptional Regulation of Gonadal Development and Differentiation

Endocrinology ◽

10.1210/en.2004-1454 ◽

2005 ◽

Vol 146 (3) ◽

pp. 1035-1042 ◽

Cited By ~ 60

Author(s):

Susan Y. Park ◽

J. Larry Jameson

Keyword(s):

Germ Cells ◽

Sertoli Cells ◽

Leydig Cells ◽

Cell Types ◽

Gonadal Development ◽

Targeted Mutagenesis ◽

Seminiferous Tubules ◽

Molecular Pathways ◽

Theca Cells ◽

Main Cell

The embryonic gonad is undifferentiated in males and females until a critical stage when the sex chromosomes dictate its development as a testis or ovary. This binary developmental process provides a unique opportunity to delineate the molecular pathways that lead to distinctly different tissues. The testis comprises three main cell types: Sertoli cells, Leydig cells, and germ cells. The Sertoli cells and germ cells reside in seminiferous tubules where spermatogenesis occurs. The Leydig cells populate the interstitial compartment and produce testosterone. The ovary also comprises three main cell types: granulosa cells, theca cells, and oocytes. The oocytes are surrounded by granulosa and theca cells in follicles that grow and differentiate during characteristic reproductive cycles. In this review, we summarize the molecular pathways that regulate the distinct differentiation of these cell types in the developing testis and ovary. In particular, we focus on the transcription factors that initiate these cascades. Although most of the early insights into the sex determination pathway were based on human mutations, targeted mutagenesis in mouse models has revealed key roles for genes not anticipated to regulate gonadal development. Defining these molecular pathways provides the foundation for understanding this critical developmental event and provides new insight into the causes of gonadal dysgenesis.

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Developmental stage- and spermatogenic cycle-specific expression of transcription factor GATA-1 in mouse Sertoli cells

Development ◽

10.1242/dev.120.7.1759 ◽

1994 ◽

Vol 120 (7) ◽

pp. 1759-1766 ◽

Cited By ~ 6

Author(s):

K. Yomogida ◽

H. Ohtani ◽

H. Harigae ◽

E. Ito ◽

Y. Nishimune ◽

...

Keyword(s):

Transcription Factor ◽

Developmental Stage ◽

Sertoli Cell ◽

Germ Cells ◽

Sertoli Cells ◽

Transcriptional Activation ◽

Germ Line ◽

Mouse Testis ◽

Seminiferous Tubules ◽

Specific Expression

GATA-1 is an essential factor for the transcriptional activation of erythroid-specific genes, and is also abundantly expressed in a discrete subset of cells bordering the seminiferous epithelium in tubules of the murine testis. In examining normal and germ-line defective mutant mice, we show here that GATA-1 is expressed only in the Sertoli cell lineage in mouse testis. GATA-1 expression in Sertoli cells is induced concomitantly with the first wave of spermatogenesis, and GATA-1-positive cells are uniformly distributed among all tubules during prepubertal testis development. However, the number of GATA-1-positive cells declines thereafter and were found only in the peripheral zone of seminiferous tubules in stages VII, VIII and IX of spermatogenesis in the adult mouse testis. In contrast, virtually every Sertoli cell in mutant W/Wv, jsd/jsd or cryptorchid mice (all of which lack significant numbers of germ cells) expresses GATA-1, thus showing that the expression of this transcription factor is negatively controlled by the maturing germ cells. These observations suggest that transcription factor GATA-1 is a developmental stage- and spermatogenic cycle-specific regulator of gene expression in Sertoli cells.

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Establishment and applications of male germ cell and Sertoli cell lines

Reproduction ◽

10.1530/rep-15-0546 ◽

2016 ◽

Vol 152 (2) ◽

pp. R31-R40 ◽

Cited By ~ 16

Author(s):

Hong Wang ◽

Liping Wen ◽

Qingqing Yuan ◽

Min Sun ◽

Minghui Niu ◽

...

Keyword(s):

Cell Line ◽

Sertoli Cell ◽

Cell Lines ◽

Germ Cells ◽

Sertoli Cells ◽

Molecular Mechanisms ◽

Simian Virus 40 ◽

T Antigen ◽

Seminiferous Tubules ◽

Male Germ Cells

Within the seminiferous tubules there are two major cell types, namely male germ cells and Sertoli cells. Recent studies have demonstrated that male germ cells and Sertoli cells can have significant applications in treating male infertility and other diseases. However, primary male germ cells are hard to proliferatein vitroand the number of spermatogonial stem cells is scarce. Therefore, methods that promote the expansion of these cell populations are essential for their use from the bench to the bed side. Notably, a number of cell lines for rodent spermatogonia, spermatocytes and Sertoli cells have been developed, and significantly we have successfully established a human spermatogonial stem cell line with an unlimited proliferation potential and no tumor formation. This newly developed cell line could provide an abundant source of cells for uncovering molecular mechanisms underlying human spermatogenesis and for their utilization in the field of reproductive and regenerative medicine. In this review, we discuss the methods for establishing spermatogonial, spermatocyte and Sertoli cell lines using various kinds of approaches, including spontaneity, transgenic animals with oncogenes, simian virus 40 (SV40) large T antigen, the gene coding for a temperature-sensitive mutant ofp53, telomerase reverse gene (Tert), and the specific promoter-based selection strategy. We further highlight the essential applications of these cell lines in basic research and translation medicine.

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Differentiation of Fetal Sertoli Cells in the Adult Testis

Reproduction ◽

10.1530/rep-21-0106 ◽

2021 ◽

Author(s):

Tetsuhiro Yokonishi ◽

Blanche Capel

Keyword(s):

Sertoli Cell ◽

Sertoli Cells ◽

Developmental Process ◽

Ips Cells ◽

Cell Maturation ◽

Seminiferous Tubules ◽

Fetal Life ◽

Adult Testis ◽

Testicular Cells

Sertoli cells proliferate and construct seminiferous tubules during fetal life, then undergo differentiation and maturation in the prepubertal testes. In the adult testes, mature Sertoli cells maintain spermatogonia and support spermatogenesis during the entire lifetime. Although Sertoli-like cells have been derived from iPS cells, they tend to remain immature. To investigate whether Sertoli cells can spontaneously acquire the ability to support spermatogenesis when transferred into the adult testis, we transplanted mouse fetal testicular cells into a Sertoli-depleted adult testis. We found that donor E12.5, E14.5 and E16.5 Sertoli cells colonized adult seminiferous tubules and supported host spermatogenesis two months after transplantation, demonstrating that immature fetal Sertoli cells can undergo sufficient maturation in the adult testis to become functional. This technique will be useful to analyze the developmental process of Sertoli cell maturation, and to investigate the potential of iPS-derived Sertoli cells to colonize, undergo maturation, and support spermatogenesis within the testis environment.

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Sertoli cell replacement in explanted mouse testis tissue supporting host spermatogenesis

Biology of Reproduction ◽

10.1093/biolre/ioab104 ◽

2021 ◽

Author(s):

Kazusa Higuch ◽

Takafumi Matsumura ◽

Haruhiko Akiyama ◽

Yoshiakira Kanai ◽

Takehiko Ogawa ◽

...

Keyword(s):

Sertoli Cell ◽

Germ Cells ◽

Sertoli Cells ◽

Diphtheria Toxin ◽

Mouse Testis ◽

Seminiferous Tubules ◽

Functional Assay ◽

Cell Replacement ◽

Replacement Method ◽

Toxin Receptor

Abstract Spermatogenesis takes place in the seminiferous tubules, starting from the spermatogonial stem cell and maturing into sperm through multiple stages of cell differentiation. Sertoli cells, the main somatic cell constituting the seminiferous tubule, are in close contact with every germ cell and play pivotal roles in the progression of spermatogenesis. In this study, we developed an in vitro Sertoli cell replacement method by combining an organ culture technique and a toxin receptor-mediated cell knockout (Treck) system. We used Amh- diphtheria toxin receptor (DTR) transgenic mice, whose Sertoli cells specifically express human DTR, which renders them sensitive to diphtheria toxin (DT). An immature Amh-DTR testis was transplanted with donor testis cells followed by culturing in a medium containing DT. This procedure successfully replaced the original Sertoli cells with the transplanted Sertoli cells, and spermatogenesis originating from resident germ cells was confirmed. In addition, Sertoli cells in the mouse testis tissues were replaced by transplanted rat Sertoli cells within culture conditions, without requiring immunosuppressive treatments. This method works as a functional assay system, making it possible to evaluate any cells that might function as Sertoli cells. It would also be possible to investigate interactions between Sertoli and germ cells more closely, providing a new platform for the study of spermatogenesis and its impairments.

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Functions of TAM RTKs in regulating spermatogenesis and male fertility in mice

Reproduction ◽

10.1530/rep-09-0101 ◽

2009 ◽

Vol 138 (4) ◽

pp. 655-666 ◽

Cited By ~ 36

Author(s):

Yongmei Chen ◽

Huizhen Wang ◽

Nan Qi ◽

Hui Wu ◽

Weipeng Xiong ◽

...

Keyword(s):

Germ Cells ◽

Sertoli Cells ◽

Tyrosine Kinases ◽

Male Fertility ◽

Leydig Cells ◽

Seminiferous Tubules ◽

Wild Type ◽

Male Sterile ◽

Triple Mutant ◽

Insight Into

Mice lacking TYRO3, AXL and MER (TAM) receptor tyrosine kinases (RTKs) are male sterile. The mechanism of TAM RTKs in regulating male fertility remains unknown. In this study, we analyzed in more detail the testicular phenotype of TAM triple mutant (TAM−/−) mice with an effort to understand the mechanism. We demonstrate that the three TAM RTKs cooperatively regulate male fertility, and MER appears to be more important than AXL and TYRO3. TAM−/− testes showed a progressive loss of germ cells from elongated spermatids to spermatogonia. Young adult TAM−/− mice exhibited oligo-astheno-teratozoospermia and various morphological malformations of sperm cells. As the mice aged, the germ cells were eventually depleted from the seminiferous tubules. Furthermore, we found that TAM−/− Sertoli cells have an impaired phagocytic activity and a large number of differentially expressed genes compared to wild-type controls. By contrast, the function of Leydig cells was not apparently affected by the mutation of TAM RTKs. Therefore, we conclude that the suboptimal function of Sertoli cells leads to the impaired spermatogenesis in TAM−/− mice. The results provide novel insight into the mechanism of TAM RTKs in regulating male fertility.

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Spontaneous Spermatogenic Failure in the Marsupial Mouse Antechinus stuartii Macleay (Dasyuridae: Marsupialia)

Australian Journal of Zoology ◽

10.1071/zo9830445 ◽

1983 ◽

Vol 31 (4) ◽

pp. 445 ◽

Cited By ~ 30

Author(s):

JB Kerr ◽

MP Hedger

Keyword(s):

Germ Cells ◽

Sertoli Cells ◽

Cell Maturation ◽

Seminiferous Tubules ◽

Androgen Binding Protein ◽

Antechinus Stuartii ◽

Early Primary ◽

Androgen Binding ◽

Spermatogenic Failure ◽

Sclerophyll Forests

Male Antechinus stuartii were collected from sclerophyll forests in Victoria at regular intervals from February to August. Spermatogenic function was assessed by means of light microscopy of testicular tissues fixed and embedded in epoxy plastic, from which a quantitative analysis of spermatogenesis was determined. Testis cytosols were prepared for assay of androgen-binding protein (ABP) and plasma was collected for androgen assay. Germ cell maturation proceeded normally until May, when failure of spermatogenesis was reflected by depletion of spermatogonia and early primary spermatocytes. However, germ cells of more advanced maturational stages were able to complete the spermatogenic process, yielding mature sperm first observed in the testis late in June. Failure of the seminiferous epithelium to replenish the numbers of early germ cells resulted in progressive depletion of germ cells in later months, leading in August to collapse of the seminiferous tubules which then contained only Sertoli cells and spermatogonia. Plasma androgens exhibited progressive elevation, reaching a peak in July and August, but testicular ABP was always undetectable. The findings suggest that the inability of the testis to maintain spermatogenesis results from intrinsic changes to testicular function exerted at the level of the spermatogonial population.

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Effect of Cryptorchidism on the Histomorphometry, Proliferation, Apoptosis, and Autophagy in Boar Testes

Animals ◽

10.3390/ani11051379 ◽

2021 ◽

Vol 11 (5) ◽

pp. 1379

Author(s):

Xiaorui Fan ◽

Yihui Liu ◽

Meishan Yue ◽

Weidong Yue ◽

Gaoya Ren ◽

...

Keyword(s):

Germ Cells ◽

Sertoli Cells ◽

Proliferating Cell Nuclear Antigen ◽

Molecular Mechanisms ◽

Inguinal Canal ◽

The Body ◽

Nuclear Antigen ◽

Seminiferous Tubules ◽

Mrna Levels ◽

Proliferating Cell

Spontaneous unilateral cryptorchid boars have one testis in the abdomen or inguinal canal, causing its temperature to be at or near the body temperature, which impairs spermatogenesis, although the histomorphometry and molecular mechanisms underlying this process remain unclear. The aim of the present study was to determine the histomorphometry, proliferation, apoptosis, and autophagy alterations in spermatogonia and Sertoli cells in unilateral cryptorchid, scrotal (contrascrotal), and preweaning piglet (preweaning) testes. Histomorphometrical analysis of cryptorchid testes showed that the seminiferous tubules contained only Sertoli cells and a few spermatogonia, but did not contain post-meiotic germ cells. The number of spermatogonia markedly decreased, and the number of Sertoli cells did not change remarkably in cryptorchid testes. TUNEL assay results showed that apoptosis signals were predominantly observed in spermatogonia. In cryptorchid and contrascrotal testes, proliferating cell nuclear antigen (PCNA) and LC3 were located in spermatogonia. The number of PCNA-positive, TUNEL-positive, and LC3-positive germ cells was low, and the protein and mRNA levels of PCNA and LC3 were significantly decreased in cryptorchid testes. Taken together, the number of Sertoli cells did not change remarkably, whereas the number of germ cells decreased in the cryptorchid testes, compared with that in the contrascrotal testes. Insufficient proliferation, excessive apoptosis, and autophagy were involved in the regulation of the decrease in spermatogonia in cryptorchid boar testes.

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318 EFFECT OF 6-N-PROPYL-2-THIOURACIL-INDUCED HYPOTHYROIDISM IN THE HOST MOUSE ON THE DEVELOPMENT OF BOVINE TESTIS XENOGRAFTS

Reproduction Fertility and Development ◽

10.1071/rdv22n1ab318 ◽

2010 ◽

Vol 22 (1) ◽

pp. 315

Author(s):

J. R. Rodriguez-Sosa ◽

G. M. J. Costa ◽

R. Rathi ◽

L. R. França ◽

I. Dobrinski

Keyword(s):

Cell Differentiation ◽

Germ Cell ◽

Sertoli Cell ◽

Germ Cells ◽

Sertoli Cells ◽

Cell Maturation ◽

Germ Cell Differentiation ◽

Host Mouse ◽

Thyroid Hormone Levels ◽

Collection Time

In rodents, thyroid hormones inhibit Sertoli cell proliferation, promote Sertoli cell differentiation, and accelerate lumen formation in the seminiferous tubules. Conversely, transient hypothyroidism prolongs Sertoli cell proliferation, leading to increased Sertoli cell number and testicular size. In order to evaluate whether 6-N-propyl-2-thiouracil (PTU)-induced hypothyroidism in the host mouse would affect seminiferous tubule development and germ cell differentiation, and subsequently increase spermatogenesis in bovine testis xenografts, fragments (∼1 mm3) of testes from 1-wk-old Holstein calves (n = 6) were transplanted ectopically to castrated immunodeficient male mice (n = 6/donor). Mice (n = 3/donor) were treated with 0.1% (w/v) PTU in drinking water for 4 weeks or left as control. At 5 and 7 months after grafting, grafts were analyzed by morphometry and immunohistochemistry for expression of protein gene product 9.5 (PGP 9.5) as a germ cell marker, and Mullerian-inhibiting substance (MIS) and androgen receptor (AR) to assess Sertoli cell maturation. For each variable, averages of each group were compared at each collection point by t-test PTU treatment to the drinking water for 1 month suppressed thyroid hormone levels (T4) in host mice without negative systemic effects (0.3 ± 0.2 v. 4 ± 0.3 μg dL-1 at 4 weeks in treated v. control mice, respectively, P < 0.05). Spermatogenesis in recovered grafts was arrested at meiosis regardless of treatment and collection time. Graft weight was lower in treated mice than in controls (21 ± 4 v. 42 ± 5 and 24 ± 9 v. 51 ± 5 mg, at 5 and 7 months, respectively, P < 0.05). Volume density of the tubular and intertubular compartments, and seminiferous epithelium, was not affected by treatment (P > 0.05); however, treatment reduced lumen density compared to controls (9 ± 2 v. 19 ± 3 and 12 ± 1 v. 24 ± 4%) and tubular diameter (121 ± 3 v. 140 ± 7 and 144 ± 2v. 170 ± 2 (im, at 5 and 7 months, respectively (P < 0.05). Tubule length per milligram was not different at 5 months between control and treated groups (P > 0.05) but was increased at 7 months in the treated grafts (50 ± 1 v. 30 ± 1 cm, P < 0.05). Number of Sertoli cells per milligram was not affected by treatment (P > 0.05). However, Sertoli cell volume was increased in controls (440 ± 19 v. 341 ± 14 and 504 ± 6 v. 388 ± 18 μm3, at 5 and 7 months, respectively, P < 0.05). The number of germ cells per 100 Sertoli cells was not different between groups at any collection time (P > 0.05). Sertoli cells showed variable MIS expression and lack of or weak AR expression regardless of treatment and collection time, indicating an immature phenotype. In conclusion, suppression of thyroid hormone levels in host mice affects seminiferous tubule development in bovine testis xenografts, demonstrating that endocrine manipulation of the mouse host will affect xenografts in a predictable manner. However, treatment did not affect number and differentiation of germ cells. Rather, incomplete Sertoli cell maturation appears to lead to incomplete germ cell differentiation in bovine testis xenografts. Supported by USDA (2007-35203-18213).

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