Stem Leydig Cells Research Articles

078 Evaluation of Paracrine Factors Critical for Human Leydig Stem Cell Function

Impaired testosterone production as a result of Leydig cell loss or dysfunction can occur in men with testicular failure. Although several testosterone formulations are available, none are capable of replicating the physiological pattern of testosterone secretion. We have shown in our recent study conducted in murine models that, Leydig stem cell transplantation along with peritubular myoid cells and Sertoli cells could be used to physiologically increase serum testosterone thereby potentially minimizing the adverse effects. However, in order to optimize the function of Leydig stem cells, we need to understand the paracrine factors released by myoid and Sertoli cells. In the present study we evaluated the significance of paracrine factors secreted by human peritubular myoid cells and Sertoli cells on Leydig stem cell function. A total of 5 men with testicular failure underwent testis biopsies for sperm retrieval. Using an IRB approved protocol, about 10mg of testicular tissue from each of these men were processed for Leydig stem cell isolation, culture and characterized.

Fibroblast Growth Factor 1 Promotes Rat Stem Leydig Cell Development.

Fibroblast growth factor 1 (FGF1) is reported to be expressed in the testis. How FGF1 affects stem Leydig cell development remains unclear. Here, we report the effects of FGF1 on rat stem Leydig cell development in an ethane dimethane sulfonate (EDS)-treated model. FGF1 (100 ng/testis) significantly increased serum testosterone level, increased PCNA-positive Leydig cell percentage and Leydig cell number, but down-regulated the expression of Lhcgr, Star, Cyp11a1, Hsd3b1, Cyp17a1, and Hsd11b1 in Leydig cells per se, after its daily intratesticular injection from post-EDS day 14 for 14 days. Primary culture of the seminiferous tubules showed that FGF1 stimulated EdU incorporation to stem Leydig cells but blocked the differentiation into the Leydig cell lineage, possibly via FGFR1-mediated mechanism. In conclusion, FGF1 promotes stem Leydig cell proliferation but blocks its differentiation.

Stem Leydig cell regeneration in the adult rat testis is inhibited after a short-term triphenyltin exposure

Triphenyltin (TPT) is an organotin compound and may be an endocrine disruptor, impairing the male reproductive system. However, the effect of short-term TPT exposure on stem Leydig cell regeneration later on remains unknown. Here, we show that TPT affects stem Leydig cell regeneration in the adult rat testis. Adult male Sprague Dawley rats were gavaged with TPT (0, 0.5, 1.0, 2.0 mg/kg body weight/day) for 10 days, followed by a single intraperitoneal injection of ethane dimethane sulfonate (EDS, 75 mg/kg body weight) to eliminate Leydig cells. Testis parameters and hormone levels were investigated on post-EDS days 21, 35, and 56. TPT significantly reduced serum testosterone levels, decreased Leydig cell number and cell size, and down-regulated its specific gene and protein expression at 1.0 and 2.0 mg/kg even 56 days after cession of treatment. TPT lowered PCNA-labeling index of progenitor Leydig cells on post-EDS day 21. TPT also lowered AKT1 and AKT2, and ERK1/2 phosphorylation on post-EDS day 56. This study reveals that a short-term exposure to TPT blocks stem Leydig cell regeneration in the long term thus delaying spermatogenesis.

Fibroblast growth factor 16 stimulates proliferation but blocks differentiation of rat stem Leydig cells during regeneration.

ObjectivesWe aim to investigate the effects of fibroblast growth factor 16 (FGF16) on Leydig cell regeneration in ethane dimethane sulphonate (EDS)‐treated rat testis.MethodsWe intraperitoneally inject 75 mg/kg EDS to adult male Sprague Dawley rats and then intratesticularly inject FGF16 (0, 10 and 100 ng/testis/day) from post‐EDS day 14 for 14 days. We investigate serum hormone levels, Leydig cell number, gene and protein expression in vivo. We also explore the effects of FGF16 treatment on stem Leydig cell proliferation in vitro.ResultsFGF16 lowers serum testosterone levels (21.6% of the control at a dose of 100 ng/testis) without affecting the levels of luteinizing hormone (LH) and follicle‐stimulating hormone (FSH) on post‐EDS day 28 in vivo. FGF16 increases Leydig cell number at doses of 10 and 100 ng/mg without affecting Sertoli cell number, increases the percentage of PCNA‐positive Leydig cells, and down‐regulates the expression of Leydig cell genes (Lhcgr, Scarb1, Star, Cyp11a1, Cyp17a1 and Hsd17b3) and Sertoli cell genes (Fshr, Dhh and Sox9) and their proteins in vivo. FGF16 increases phosphorylation of AKT1 and AKT2 as well as EKR1/2 in vivo, indicating that it possibly acts via AKT1/ATK2 and ERK1/2 pathways. FGF16 also lowers medium testosterone levels and down‐regulates the expression of Leydig cell genes (Lhcgr, Scarb1, Star, Cyp11a1, Cyp17a1 and Hsd17b3) but increases EdU incorporation into stem Leydig cells in vitro.ConclusionsThese data suggest that FGF16 stimulates stem and progenitor Leydig cell proliferation but blocks their differentiation, thus lowering testosterone biosynthesis.

Fetal Leydig cells dedifferentiate and serve as adult Leydig stem cells.

Previous studies have established that fetal Leydig cells (FLCs) and adult Leydig cells (ALCs) show distinct functional characteristics. However, the lineage relationship between FLCs and ALCs has not been clarified yet. Here, we reveal that a subset of FLCs dedifferentiate at fetal stages to give rise to ALCs at the pubertal stage. Moreover, the dedifferentiated cells contribute to the peritubular myoid cell and vascular pericyte populations in the neonatal testis, and these non-steroidogenic cells serve as potential ALC stem cells. We generated FLC lineage-specific Nr5a1 (Ad4BP/SF-1) gene-disrupted mice and mice lacking the fetal Leydig enhancer (FLE) of the Nr5a1 gene. Phenotypes of these mice support the conclusion that most of the ALCs arise from dedifferentiated FLCs, and that the FLE of the Nr5a1 gene is essential for both initial FLC differentiation and pubertal ALC redifferentiation.

Effects of spermatogenic cycle on Stem Leydig cell proliferation and differentiation

We reported previously that stem Leydig cells (SLC) on the surfaces of rat testicular seminiferous tubules are able to differentiate into Leydig cells. The proliferation and differentiation of SLCs seem likely to be regulated by niche cells, including nearby germ and Sertoli cells. Due to the cyclical nature of spermatogenesis, we hypothesized that the changes in the germ cell composition of the seminiferous tubules as spermatogenesis proceeds may affect tubule-associated SLC functions. To test this hypothesis, we compared the ability of SLCs associated with tubules at different stages of the cycle to differentiate into Leydig cells in vitro. SLCs associated with stages IX-XI were more active in proliferation and differentiation than SLCs associated with stages VII-VIII. However, when the SLCs were isolated from each of the two groups of tubules and cultured in vitro, no differences were seen in their ability to proliferate or differentiate. These results suggested that the stage-dependent local factors, not the SLCs themselves, explain the stage-dependent differences in SLC function. TGFB, produced in stage-specific fashion by Sertoli cells, is among the factors shown in previous studies to affect SLC function in vitro. When TGFB inhibitors were included in the cultures of stages IX-XI and VII-VIII tubules, stage-dependent differences in SLC development were reduced, suggesting that TGFB may be among the paracrine factors involved in the stage-dependent differences in SLC function. Taken together, the findings suggest that there is dynamic interaction between SLCs and germ/Sertoli cells within the seminiferous tubules that may affect SLC proliferation and differentiation.

Oncostatin M inhibits differentiation of rat stem Leydig cells invivo and invitro.

Oncostatin M (OSM) is a pleiotropic cytokine within the interleukin six family of cytokines, which regulate cell growth and differentiation in a wide variety of biological systems. However, its action and underlying mechanisms on stem Leydig cell development are unclear. The objective of the present study was to investigate whether OSM affects the proliferation and differentiation of rat stem Leydig cells. We used a Leydig cell regeneration model in rat testis and a unique seminiferous tubule culture system after ethane dimethane sulfonate (EDS) treatment to assess the ability of OSM in the regulation of proliferation and differentiation of rat stem Leydig cells. Intratesticular injection of OSM (10 and 100 ng/testis) from post‐EDS day 14 to 28 blocked the regeneration of Leydig cells by reducing serum testosterone levels without affecting serum luteinizing hormone and follicle‐stimulating hormone levels. It also decreased the levels of Leydig cell‐specific mRNAs (Lhcgr, Star, Cyp11a1, Hsd3b1, Cyp17a1 and Hsd11b1) and their proteins by the RNA‐Seq and Western blotting analysis. OSM had no effect on the proliferative capacity of Leydig cells in vivo. In the seminiferous tubule culture system, OSM (0.1, 1, 10 and 100 ng/mL) inhibited the differentiation of stem Leydig cells by reducing medium testosterone levels and downregulating the expression of Leydig cell‐specific genes (Lhcgr, Star, Cyp11a1, Hsd3b1, Cyp17a1 and Hsd11b1) and their proteins. OSM‐mediated action was reversed by S3I‐201 (a STAT3 antagonist) or filgotinib (a JAK1 inhibitor). These data suggest that OSM is an inhibitory factor of rat stem Leydig cell development.

Subcutaneous Leydig Stem Cell Autograft: A Promising Strategy to Increase Serum Testosterone.

Exogenous testosterone therapy can be used to treat testosterone deficiency; however, it has several adverse effects including infertility due to negative feedback on the hypothalamic–pituitary–gonadal (HPG) axis. Leydig stem cell (LSC) transplantation could provide a new strategy for treating testosterone deficiency, but clinical translatability of injecting stem cells inside the testis is not feasible. Here, we explore the feasibility of subcutaneously autografting LSCs in combination with Sertoli and myoid cells to increase testosterone. We also studied whether the grafted LSCs can be regulated by the HPG axis and the molecular mechanism behind this regulation. LSCs were isolated from the testes of 12‐week‐old C57BL/6 mice, and subcutaneously autografted in combination with Sertoli cells and myoid cells. We found that LSCs alone were incapable of self‐renewal and differentiation. However, in combination with Sertoli cells and myoid cells, LSCs underwent self‐renewal as well as differentiation into mature Leydig cells. As a result, the recipient mice that received the LSC autograft showed testosterone production with preserved luteinizing hormone. We found that testosterone production from the autograft was regulated by hedgehog (HH) signaling. Gain of function and loss of function study confirmed that Desert HH (DHH) agonist increased and DHH antagonist decreased testosterone production from autograft. This study is the first to demonstrate that LSCs, when autografted subcutaneously in combination with Sertoli cells and myoid cells, can increase testosterone production. Therefore, LSC autograft may provide a new treatment for testosterone deficiency while simultaneously preserving the HPG axis. Stem Cells Translational Medicine 2019;8:58–65

Evaluation of paracrine factors critical for human leydig stem cell function

Evaluation of paracrine factors critical for human leydig stem cell function

The Leydig Cells of the Testis Originate from the Microvascular Pericytes

This review offers some clarifying thoughts about the nature and origin of the fetal and adult Leydig cells, supporting the conception that the pericytes (the periendothelial cells) and the smooth muscle cells of the microvasculature, that represent the main omnipresent adult stem cell population of the mammalian organism, are the Leydig cell ancestors. Our attention is specifically dedicated to the numerous contradictions as well as ambiguities concerning the hypotheses that the mesenchymal stromal cells (MSCs), the neural crest stem cells (NCSCs) and the peritubular myoid stem cells (PMCs) represent the stem ancestors of the Leydig cells. In effect, it becomes evident that the only pluripotent stem cell-like cells in the vertebrate body, including the testis, are the pericytes. The pericytes are derivate of the embryonal epiblast and retain its pluripotency within the microvascular niches where they are disseminated during the embryo- and fetogenesis and are stored as a resting adult stem cell population for tissue generation, maintenance, repair and regeneration. The pluripotency of the epiblast and the pericytes themselves are responsible and explain the neural features of the Leydig cells. Thus, both NCSCs and PMCs are not the ancestors of the pericytes, respectively of the Leydig cells. Biomed Rev 2017; 28: 1-21. Keywords: Leydig cells, origin, microvascular pericytes, smooth muscle cells, Leydig stem cells

Bisphenol A stimulates differentiation of rat stem Leydig cells invivo and invitro.

Bisphenol A (BPA) is widely used in consumer products and a potential endocrine disruptor linked with sexual precocity. However, its action and underlying mechanisms on male sexual maturation is unclear. In the present study, we used a unique invivo ethane dimethane sulfonate (EDS)-induced Leydig cell regeneration model that mimics the pubertal development of Leydig cells and an invitro stem Leydig cell differentiation model to examine the roles of BPA in Leydig cell development in rats. Intratesticular exposure to doses (100 and 1000 pmol/testis) of BPA from post-EDS day 14-28 stimulated Leydig cell developmental regeneration process by increasing serum testosterone level and Leydig cell-specific gene (Lhcgr, Star, Cyp11a1, Hsd3b1, Cyp17a1, Hsd17b3, and Hsd11b1) and their protein expression levels. BPA did not alter serum luteinizing hormone and follicle-stimulating hormone levels as well as the proliferative capacity of Leydig cells invivo. Invitro study demonstrated that BPA (100 nmol/L) stimulated the differentiation of stem Leydig cells by increasing medium testosterone levels and up-regulating Leydig cell-specific gene (Lhcgr, Cyp11a1, Hsd3b1, Cyp17a1, and Hsd17b3) and their proteins but did not affect their proliferation measured by EdU incorporation. In conclusion, BPA stimulates the differentiation of stem Leydig cells in rat testes, thus possibly causing sexual precocity in the male.

Aldosterone Blocks Rat Stem Leydig Cell Development In Vitro.

Aldosterone (ALDO) is a primary endogenous mineralocorticoid, appearing as the main hormone controlling sodium and water homeostasis. Its emerging role in the development of many organs has gained interest over the past few years. In the testis, Leydig cells contain mineralocorticoid receptors and ALDO stimulates androgen synthesis via the mineralocorticoid receptors in rat adult Leydig cells. Although ALDO pharmacologically promoted the Leydig cell function, its role in Leydig cell development was unclear. In the present study, we investigated effects of ALDO on rat stem Leydig cell (SLC) proliferation and differentiation. Using an in vitro culture system of the seminiferous tubules from Leydig cell-depleted testis and EdU (a modified thymidine analog) incorporation into the SLC for flurorescent labeling to judge its DNA synthesis and measurement of medium testosterone production, steroidogenesis-related gene and protein expression, we found that: (1) ALDO suppressed EdU incorporation into SLCs at 100 nM via mineralocorticoid receptor-mediated mechanism and (2) ALDO reduced Leydig cell number. In conclusion, ALDO pharmacologically blocked rat SLC development.

Interleukin 6 inhibits the differentiation of rat stem Leydig cells.

Inflammation causes male hypogonadism. Several inflammatory cytokines, including interleukin 6 (IL-6), are released into the blood and may suppress Leydig cell development. The objective of the present study was to investigate whether IL-6 affected the proliferation and differentiation of rat stem Leydig cells. Leydig cell-depleted rat testis (invivo) and seminiferous tubules (invitro) with ethane dimethane sulfonate (EDS) were used to explore the effects of IL-6 on stem Leydig cell development. Intratesticular injection of IL-6 (10 and 100 ng/testis) from post-EDS day 14 to 28 blocked the regeneration of Leydig cells, as shown by the lower serum testosterone levels (21.6% of the control at 100 ng/testis dose), the down-regulated Leydig cell gene (Lhcgr, Star, Cyp11a1, Cyp17a1, and Hsd17b3) expressions, and the reduced Leydig cell number. Stem Leydig cells on the surface of the seminiferous tubules were induced to enter the Leydig cell lineage invitro in the medium containing luteinizing hormone and lithium. IL-6 (1, 10, and 100ng/ml) concentration-dependently decreased testosterone production and Lhcgr, Cyp11a1, Cyp17a1, Hsd17b3 and Insl3 mRNA levels. The IL-6 mediated effects were antagonized by Janus kinase 1 (JAK) inhibitor (filgotinib) and Signal Transducers and Activators of Transcription 3 (STAT3) inhibitor (S3I-201), indicating that a JAK-STAT3 signaling pathway is involved. In conclusion, our results demonstrated that IL-6 was an inhibitory factor of stem Leydig cell development.

Parathyroid Hormone-Related Protein Promotes Rat Stem Leydig Cell Differentiation.

The regulatory factors for stem Leydig cell development are largely unknown. Herein, we reported that parathyroid hormone-related protein (PTHrP) may be a factor to regulate this process. The effects of PTHrP on rat stem Leydig cell proliferation and differentiation were investigated using a stem Leydig cell culture system and an ethane dimethane sulfonate (EDS)-treated in vivo Leydig cell regeneration model. PTHrP (1,000 pg/ml) significantly increased medium testosterone level and up-regulated STAR, CYP17A1, and 17β-HSD3 expressions. Co-treatment with PKA inhibitor H-89 or PKC inhibitor U73122 reversed PTHrP-mediated increase of testosterone production in vitro. Intratesticular injection of PTHrP (100 ng/testis) into the Leydig cell-depleted testis from post-EDS day 7 to 21 significantly increased serum testosterone level, up-regulated LHCGR, SCARB1, CYP11A1, 11β-HSD1, and CYP17A1 expressions. It also enlarged Leydig cell size without affecting PCNA-labeled Leydig cell number. This indicates that PTHrP promotes stem Leydig cell differentiation. PTHrP in vivo increased CREB and p-CREB levels, suggesting that PTHrP acts via a PKA-CREB signaling pathway. In conclusion, PTHrP stimulates stem Leydig cell differentiation without affecting its proliferation, showing its novel action and mechanism on rat stem Leydig cell development.

Transdifferentiation of adult rat stem Leydig cells into prostatic and uterine epithelium, but not epidermis.

Stem Leydig cells (SLCs), precursors of testicular Leydig cells that secrete testosterone required for male sexual differentiation, spermatogenesis, and fertility, were recently identified in rat testes. Various types of stem cells have shown the ability to differentiate into other tissues, but there is no information on the plasticity of adult rat SLCs (rSLCs). This study investigated the ability of rSLCs to transdifferentiate into cell types from all three germ layers-prostatic epithelium (endoderm), uterine epithelium (mesoderm), and epidermis (ectoderm)-under the influence of inductive mesenchyme from fetal and neonatal tissues. To differentiate rSLCs into cells of other lineages, mesenchyme from green fluorescent protein (GFP)-expressing mice was used. Tissue recombinants of urogenital sinus mesenchyme (a potent prostate inducer) and rSLCs grafted into adult male hosts formed ductal structures resembling prostate after 5weeks. Prostate epithelium was of rSLC origin as determined by absence of GFP expression, and expressed characteristic markers of prostatic epithelium. Similarly, uterine mesenchyme + rSLCs tissue recombinants contained a simple columnar epithelium that was histologically similar to normal uterine epithelium and expressed typical uterine epithelial markers, but was of rSLC origin. In contrast, epidermal tissue was absent in fetal dermis + rSLCs recombinants, suggesting rSLCs did not form skin epithelium. Thus, rSLCs can transdifferentiate into uterine and prostatic epithelium, mesodermal, and endodermal derivatives, respectively, but they may have a limited transdifferentiation potential, as shown by their inability to form epidermis, an ectodermal derivative.

Transplanted human p75-positive stem Leydig cells replace disrupted Leydig cells for testosterone production

Previous studies have demonstrated that rodent stem Leydig cell (SLC) transplantation can partially restore testosterone production in Leydig cell (LC)-disrupted or senescent animal models, which provides a promising approach for the treatment of hypogonadism. Here, we isolated human SLCs prospectively and explored the potential therapeutic benefits of human SLC transplantation for hypogonadism treatment. In adult human testes, p75 neurotrophin receptor positive (p75+) cells expressed the known SLC marker nestin, but not the LC lineage marker hydroxysteroid dehydrogenase-3β (HSD3β). The p75+ cells which were sorted by flow cytometry from human adult testes could expand in vitro and exhibited clonogenic self-renewal capacity. The p75+ cells had multi-lineage differentiation potential into multiple mesodermal cell lineages and testosterone-producing LCs in vitro. After transplantation into the testes of ethane dimethane sulfonate (EDS)-treated LC-disrupted rat models, the p75+ cells differentiated into LCs in vivo and secreted testosterone in a physiological pattern. Moreover, p75+ cell transplantation accelerated the recovery of serum testosterone levels, spermatogenesis and reproductive organ weights. Taken together, we reported a method for the identification and isolation of human SLCs on the basis of p75 expression, and demonstrated that transplanted human p75+ SLCs could replace disrupted LCs for testosterone production. These findings provide the groundwork for further clinical application of human SLCs for hypogonadism.

Subcutaneous leydig stem cell autograft in human: a novel approach to increase serum testosterone

Subcutaneous leydig stem cell autograft in human: a novel approach to increase serum testosterone

Subcutaneous leydig stem cell autograft in mice: a novel approach to increase serum testosterone

Subcutaneous leydig stem cell autograft in mice: a novel approach to increase serum testosterone

Insights into the Development of the Adult Leydig Cell Lineage from Stem Leydig Cells.

Adult Leydig cells (ALCs) are the steroidogenic cells in the testes that produce testosterone. ALCs develop postnatally from a pool of stem cells, referred to as stem Leydig cells (SLCs). SLCs are spindle-shaped cells that lack steroidogenic cell markers, including luteinizing hormone (LH) receptor and 3β-hydroxysteroid dehydrogenase. The commitment of SLCs into the progenitor Leydig cells (PLCs), the first stage in the lineage, requires growth factors, including Dessert Hedgehog (DHH) and platelet-derived growth factor-AA. PLCs are still spindle-shaped, but become steroidogenic and produce mainly androsterone. The next transition in the lineage is from PLC to the immature Leydig cell (ILC). This transition requires LH, DHH, and androgen. ILCs are ovoid cells that are competent for producing a different form of androgen, androstanediol. The final stage in the developmental lineage is ALC. The transition to ALC involves the reduced expression of 5α-reductase 1, a step that is necessary to make the cells to produce testosterone as the final product. The transitions along the Leydig cell lineage are associated with the progressive down-regulation of the proliferative activity, and the up-regulation of steroidogenic capacity, with each step requiring unique regulatory signaling.

PD08-11 LEYDIG STEM CELL AUTOGRAFT IN MICE: A NOVEL APPROACH TO INCREASE SERUM TESTOSTERONE WHILE PRESERVING FERTILITY

You have accessJournal of UrologyInfertility: Basic Research & Pathophysiology II1 Apr 2017PD08-11 LEYDIG STEM CELL AUTOGRAFT IN MICE: A NOVEL APPROACH TO INCREASE SERUM TESTOSTERONE WHILE PRESERVING FERTILITY HIMANSHU ARORA, Marilia Sanches Santos Rizzo Zutti, Bruno Nahar, Joshua M. Hare, and Ranjith Ramasamy HIMANSHU ARORAHIMANSHU ARORA More articles by this author , Marilia Sanches Santos Rizzo ZuttiMarilia Sanches Santos Rizzo Zutti More articles by this author , Bruno NaharBruno Nahar More articles by this author , Joshua M. HareJoshua M. Hare More articles by this author , and Ranjith RamasamyRanjith Ramasamy More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2017.02.548AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Leydig cell loss or dysfunction is associated with impaired testosterone production. Exogenous testosterone supplementation can be used to treat low testosterone, however it has several adverse effects including infertility due to negative feedback on the hypothalamic-pituitary-gonadal axis. We studied testosterone production in mouse models following autograft in skin with Leydig stem cells isolated from testes. METHODS A total of 10 wild-type adult C57/BL6 mice were included in the study. Orchiectomy was conducted in seven mice (4 experimental and 3 negative controls) and the remaining three were used as positive controls. Leydig stem cells were harvested from testis by collagenase/trypsin digestion. Cells from each mouse were allowed to grow separately in the media containing DMEM, FBS (10%), P/S, ITS, Dexamethasone, EGF, PDGF-AA. After 10 days following orchiectomy, 1 X 106 cells from four animals were autografted in the subcutaneous tissue. After four weeks, grafts and blood were harvested. We evaluated testosterone production, graft morphology, and expression of Leydig cell markers. RESULTS We successfully isolated and cultured up to 1 million Leydig stem cells / testis from all 7 animals. These cells were differentiated and converted into functional adult Leydig cells in vitro. Stem cell property of cultured cells was confirmed by IF and qPCR in which the expression of PDGFR-a was high in regular media vs differentiation induction media and expression of 3BHSD was low in regular media vs differentiation induction media. The autografts were able to survive in animals for at least one month. H&E staining showed the presence of Leydig stem cells subcutaneously. Testosterone levels were almost doubled in autograft mice as compared to negative controls. CONCLUSIONS Our results indicate that Leydig stem cells can be isolated and cultured from wild-type mice. Leydig stem cell autograft can a novel therapeutic approach to increasing serum testosterone while simultaneously preserving fertility. © 2017FiguresReferencesRelatedDetails Volume 197Issue 4SApril 2017Page: e192-e193 Advertisement Copyright & Permissions© 2017MetricsAuthor Information HIMANSHU ARORA More articles by this author Marilia Sanches Santos Rizzo Zutti More articles by this author Bruno Nahar More articles by this author Joshua M. Hare More articles by this author Ranjith Ramasamy More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...