FORMATION OF 5α-REDUCED PRODUCTS FROM TESTOSTERONE IN VITRO BY GERM CELLS FROM IMMATURE RATS

ABSTRACT In the incubation study of [3H] testosterone with tissue homogenates in the presence of NADPH, it was shown that the rate of production of the sum of all 5α-reduced products by 30-day-old rat testes (50 to 100 nmoles/g tissue or 100 mg protein/h) was of the order of twenty times that by the ventral prostate (2 to 4 nmoles/g tissue/h). The activities of the C19-steroid Δ4-5α-reductase of the seminiferous tubules, germ cells and tubular non-germ cells isolated from immature rats (7 to 10, 11 to 13 and 5 to 7 nmoles/100 mg protein/h, respectively) were of similar order to those of the prostate, while much less activity was found in the spleen and muscle. It was also noted that the activity of the 5α-androstane-3α-, and -3β-hydroxysteroid dehydrogenase seemed to be higher in each tissue of immature rat testes than in the prostate tissue. The seminiferous tubules were isolated by collagenase treatment and germ cells were separated from non-germ cells by tubular cell culture at 34°C for 2 days. Thus, germ cells of immature rat testes seem to have the characteristics of androgen responsive tissue.

Download Full-text

Immature rat seminiferous tubules reconstructed in vitro express markers of Sertoli cell maturation after xenografting into nude mouse hosts

MHR Basic science of reproductive medicine ◽

10.1093/molehr/gap081 ◽

2009 ◽

Vol 16 (2) ◽

pp. 97-110 ◽

Cited By ~ 19

Author(s):

K. Gassei ◽

J. Ehmcke ◽

M.A. Wood ◽

W.H. Walker ◽

S. Schlatt

Keyword(s):

Sertoli Cell ◽

Nude Mouse ◽

Cell Maturation ◽

Seminiferous Tubules ◽

Immature Rat

Download Full-text

Stem cell factor protects germ cells from apoptosis in vitro

Journal of Cell Science ◽

10.1242/jcs.113.1.161 ◽

2000 ◽

Vol 113 (1) ◽

pp. 161-168 ◽

Cited By ~ 4

Author(s):

W. Yan ◽

J. Suominen ◽

J. Toppari

Keyword(s):

Stem Cell ◽

Germ Cells ◽

Stem Cell Factor ◽

Primordial Germ Cells ◽

Survival Function ◽

Seminiferous Tubules ◽

Testicular Development ◽

Dna Laddering ◽

Survival Effect

Stem cell factor (SCF) plays an important role in migration, adhesion, proliferation, and survival of primordial germ cells and spermatogonia during testicular development. However, the function of SCF in the adult testis is poorly described. We have previously shown that, in the presence of SCF, there were more type A spermatogonia incorporating thymidine at stage XII of rat seminiferous tubules cultured in vitro than in the absence of SCF, implying that the increased DNA synthesis might result from enhanced survival of spermatogonia. To explore the potential pro-survival function of SCF during spermatogenesis, the seminiferous tubules from stage XII were cultured in the presence or absence of SCF (100 ng/ml) for 8, 24, 48, and 72 hours, respectively, and apoptosis was analyzed by DNA laddering and in situ 3′-end labeling (ISEL) staining. Surprisingly, not only spermatogonia, but also spermatocytes and spermatids, were protected from apoptosis in the presence of SCF. Apoptosis took place much later and was less severe in the SCF-treated tubules than in the controls. Based on previous studies showing that FSH prevents germ cells from undergoing apoptosis in vitro, and that SCF level is increased dramatically in response to FSH stimulation, we also tested if the pro-survival effect of FSH is mediated through SCF by using a function-blocking monoclonal antibody, ACK-2, to block SCF/c-kit interaction. After 24 hours of blockade, the protective effect of FSH was partially abolished, as manifested by DNA laddering and ISEL analyses. The present study demonstrates that SCF acts as an important survival factor for germ cells in the adult rat testis and FSH pro-survival effect on germ cells is mediated partially through the SCF/c-kit pathway.

Download Full-text

Derivation of male germ cells from induced pluripotent stem cells in vitro and in reconstituted seminiferous tubules

Cell Proliferation ◽

10.1111/j.1365-2184.2012.00811.x ◽

2012 ◽

Vol 45 (2) ◽

pp. 91-100 ◽

Cited By ~ 36

Author(s):

S. Yang ◽

J. Bo ◽

H. Hu ◽

X. Guo ◽

R. Tian ◽

...

Keyword(s):

Stem Cells ◽

Induced Pluripotent Stem Cells ◽

Germ Cells ◽

Pluripotent Stem Cells ◽

Seminiferous Tubules ◽

Male Germ Cells ◽

Induced Pluripotent

Download Full-text

Inhibition of prostatic growth in rats by 6-methylene-4-pregnene-3,20-dione

Journal of Endocrinology ◽

10.1677/joe.0.0950311 ◽

1982 ◽

Vol 95 (3) ◽

pp. 311-313 ◽

Cited By ~ 19

Author(s):

Vladimir Petrow ◽

G. M. Padilla ◽

Keith Kendle ◽

A. Tantawi

Keyword(s):

Body Weight ◽

Seminal Vesicles ◽

Ventral Prostate ◽

Irreversible Inhibitor ◽

Immature Rats ◽

Daily Dose ◽

Prostatic Growth ◽

Rat Prostate

6-Methylene-4-pregnene-3,20-dione, a potent irreversible inhibitor of rat prostate 5α-reductase in vitro, markedly inhibited the growth of the ventral prostate and seminal vesicles in immature rats when administered s.c. in a daily dose of 100 mg/kg body weight for 35 days. Kidney and testes weights were reduced, together with some reduction in body weight. In the mature rat treated with this steroid in a dose of 40 mg/kg body weight per day for 11 days, marked regression in ventral prostate and seminal vesicles was observed.

Download Full-text

Male reproductive toxicity and b-luteinizing hormone gene expression in sexually mature and immature rats exposed to 2-bromopropane

Human & Experimental Toxicology ◽

10.1191/096032799678839536 ◽

1999 ◽

Vol 18 (11) ◽

pp. 683-690 ◽

Cited By ~ 9

Author(s):

X Wu ◽

A S Faqi ◽

J Yang ◽

X Ding ◽

X Jiang ◽

...

Keyword(s):

Gene Expression ◽

Dose Level ◽

Germ Cells ◽

Serum Testosterone Level ◽

Seminiferous Tubules ◽

Control Group ◽

Dependent Manner ◽

Immature Rats ◽

The Absolute ◽

Sexually Mature

1 The reproductive effects of 2-bromopropane (2-BP) in sexually mature and immature male Sprague-Dawley rats were investigated. The animals were randomly divided into three treatment groups and one control group each of which comprised six mature and six immature rats. The treated groups were injected s.c. 200, 600 and 1800 mg/kg of 2-BP on 5 days a week for 5-7 weeks and the control group received the vehicle. 2 The absolute and relative testis weights were significantly reduced in 600 and 1800 mg/kg b.w. dose groups in both mature and immature rats. The absolute epididymis, prostate, seminal vesicle, and pituitary weights and the relative epididymis weights, however, were significant only at the highest dose level used in both mature and immature rats. 3 The sperm concentration and sperm viability in epididymal duct decreased and the percentage of abnormal sperm increased in a dose-dependent manner in both mature and immature rats. Additionally, serum testosterone level was significantly decreased in all dose groups in mature rats, and was significantly reduced only in the group treated with the middle and highest dose in immature rats. 4 In both mature and immature rats treated with 200 and 600 mg/kg, the seminiferous tubules were atrophied and all types of germ cells were decreased in number. At the highest dose level, the effect was more marked showing severely atrophied seminiferous tubules and a complete loss of all types of germ cells. 5 The mating, pregnancy and fertility indices were significantly reduced in the 600 and 1800 mg/kg groups. Additionally, at the highest dose group the number of implantations and viable fetuses per litter were reduced and the resorption rate was increased significantly. 6 In the mature rats, the b-LH gene expression increased significantly in the 1800 mg/kg group when compared to the control group. 7 It can be concluded that 2-BP induces alterations in both neuro-endocrine axis and the reproductive tract under the present experimental conditions. The no observed adverse effect level (NOAEL) in this study could be estimated to be lower than 200 mg/kg/b.w. based on the alteration in testicular morphology as well as on sperm parameters observed at the dose level of 200 mg/kg.

Download Full-text

VEGF/VEGFR2 Signaling Regulates Germ Cell Proliferation in vitro and Promotes Mouse Testicular Regeneration in vivo

Cells Tissues Organs ◽

10.1159/000440949 ◽

2016 ◽

Vol 201 (1) ◽

pp. 1-13 ◽

Cited By ~ 12

Author(s):

Ruhui Tian ◽

Shi Yang ◽

Yong Zhu ◽

Shasha Zou ◽

Peng Li ◽

...

Keyword(s):

Cell Proliferation ◽

Germ Cell ◽

Germ Cells ◽

Seminiferous Tubules ◽

Testicular Development ◽

Male Germ Cells ◽

Vegfr2 Signaling ◽

Proliferation In Vitro ◽

Germ Cell Proliferation

Vascular endothelial growth factor (VEGF) plays fundamental roles in testicular development; however, its function on testicular regeneration remains unknown. The objective of this study was to explore the roles VEGF/VEGFR2 signaling plays in mouse germ cells and in mouse testicular regeneration. VEGF and the VEGFR2 antagonist SU5416 were added to culture medium to evaluate their effects on spermatogonial stem cell line (C18-4 cells) proliferation. Testicular cells obtained from newborn male ICR mice were grafted into the dorsal region of male BALB/c nude mice. VEGF and SU5416 were injected into the graft sites to assess the effects of the VEGF and VEGFR2 signaling pathways on testicular reconstitution. The grafts were analyzed after 8 weeks. We found that VEGF promoted C18-4 proliferation in vitro, indicating its role in germ cell survival. HE staining revealed that seminiferous tubules were reconstituted and male germ cells from spermatogonia to spermatids could be observed in testis-like tissues 8 weeks after grafting. A few advantaged male germ cells, including spermatocytes and spermatids, were found in SU5416-treated grafts. Moreover, VEGF enhanced the expression of genes specific for male germ cells and vascularization in 8-week grafts, whereas SU5416 decreased the expression of these genes. SU5416-treated grafts had a lower expression of MVH and CD31, indicating that blockade of VEGF/VEGFR2 signaling reduces the efficiency of seminiferous tubule reconstitution. Collectively, these data suggest that VEGF/VEGFR2 signaling regulates germ cell proliferation and promotes testicular regeneration via direct action on germ cells and the enhancement of vascularization.

Download Full-text

Comparison of the cellular composition and steroidogenic properties of preparations of interstitial cells isolated from immature and mature rat testis

Journal of Endocrinology ◽

10.1677/joe.0.1120361 ◽

1987 ◽

Vol 112 (3) ◽

pp. 361-NP ◽

Cited By ~ 10

Author(s):

A. P. N. Themmen ◽

R. Molenaar ◽

W. J. Visser ◽

J. F. Jongkind ◽

F. F. G. Rommerts ◽

...

Keyword(s):

Leydig Cells ◽

Interstitial Cells ◽

Cellular Composition ◽

Percoll Gradient ◽

Dibutyryl Cyclic Amp ◽

Steroid Production ◽

Immature Rat ◽

Collagenase Treatment ◽

Ficoll Gradient ◽

Immature Rats

ABSTRACT The morphological and steroidogenic properties of preparations of interstitial cells isolated by collagenase treatment from testes of immature and mature rats have been compared. After additional purification on a Ficoll gradient, 80% of cells from mature rat testes were found to be Leydig cells; 20% were macrophages. Forty to sixty per cent of collagenase-dispersed cells isolated from immature rats were Leydig cells, 37% were mesenchymal cells and there were no macrophages. A preparation in which 90% cells were Leydig cells could be obtained from immature testes after further purification by centrifugation through a Percoll gradient. The distribution of steroidogenic enzymes through the gradient corresponded to the distribution of LH-dependent steroid production. The results indicate that the steroidogenic activity per Leydig cell from mature rats is fourfold greater than the activity in immature rat Leydig cells in control incubations or after stimulation with LH, dibutyryl cyclic AMP or in the presence of 22R-hydroxycholesterol. J. Endocr. (1987) 112, 361–366

Download Full-text

288 THE EXPRESSION PATTERN OF ACETYLATED ALPHA-TUBULIN IS CONSERVED IN PORCINE AND MURINE SPERMATOGONIAL STEM CELLS

Reproduction Fertility and Development ◽

10.1071/rdv20n1ab288 ◽

2008 ◽

Vol 20 (1) ◽

pp. 223

Author(s):

J. Luo ◽

S. Megee ◽

I. Dobrinski

Keyword(s):

Stem Cells ◽

Basement Membrane ◽

Expression Pattern ◽

Germ Cells ◽

Sertoli Cells ◽

Spermatogonial Stem Cells ◽

Seminiferous Tubules ◽

Histone Deacetylase 6 ◽

Pgp 9.5

During mammalian spermatogenesis, spermatogonial stem cells (SSCs) reside in the stem cell niche on the basement membrane where they undergo self-renewing divisions. Differentiating daughter cells are located progressively more toward the tubular lumen where they ultimately form spermatozoa. The mechanisms responsible for maintenance of SSCs at the basement membrane are unclear. Microtubules consisting of α/β-tubulin heterodimers are associated with many cellular functions. Reversible acetylation of α-tubulin at Lys40 has been implicated in regulating microtubule stability and function. Acetylation of α-tubulin is abundant in stable microtubules but absent from dynamic cellular structures. Deacetylation of α-tubulin is controlled by histone deacetylase 6 which is predominantly expressed in mouse testis. Here, we tested the hypothesis that differential acetylation of α-tubulin might be involved in maintenance of SSCs. Immunohistochemistry for acetylated α-tubulin (Ac-α-Tu) and the spermatogonia specific proteins PGP 9.5, DAZL, and PLZF were used to characterize the expression pattern of Ac-α-Tu in porcine and murine germ cells at different stages of testis development. In immature boar testes, Ac-α-Tu was present exclusively in gonocytes but not in other testicular cells at 1 week of age, and in a subset of spermatogonia at 10 weeks of age. At this age, spermatogonia are migrating to the basement membrane of the seminiferous tubules, and Ac-α-Tu appeared to be polarized toward the basement membrane. In immature mouse testes, Ac-α-Tu was present in germ cells and Sertoli cells at 6 days of age, whereas at 2 weeks of age, Ac-α-Tu expression was stronger in spermatogonia co-expressing PGP 9.5 and in spermatocytes than in Sertoli cells or PGP 9.5-negative spermatogonia. In adult boar and mouse testes, Ac-α-Tu was detected in a few single or paired spermatogonia expressing PGP 9.5 localized on the basement membrane as well as in spermatocytes, spermatids, and spermatozoa. Spermatogonia with high levels of Ac-α-Tu expressed PLZF but did not express DAZL, suggesting that only undifferentiated spermatogonia maintain a high level of Ac-α-Tu. When seminiferous tubules from 1-week-old and adult boar testes were maintained in vitro for 1–2 days, high levels of Ac-α-Tu were detected in single or paired round spermatogonia with a large nucleus, compared to low levels in elongated paired and aligned spermatogonia. The unique expression pattern of Ac-α-Tu in undifferentiated germ cells during postnatal development appears to be conserved in mammalian testes. Since Ac-α-Tu is a component of long-lived stable microtubules and reducing acetylation of α-tubulin enhances cell motility, these results suggest that stabilization of microtubules might contribute to the maintenance of spermatogonial stem cells. This work was supported by 1R01 RR 17359-05.

Download Full-text