INFLUENCE OF GERM CELLS UPON TRANSFERRIN SECRETION BY RAT SERTOLI CELLS in vitro

1988 ◽  
Vol 118 (3) ◽  
pp. R13-R16 ◽  
Author(s):  
B. LE MAGUERESSE ◽  
C. PINEAU ◽  
F. GUILLOU ◽  
B. JEGOU
Keyword(s):  
Sertoli Cell ◽  
Cell Cultures ◽  
Germ Cells ◽  
Sertoli Cells ◽  
Seminiferous Tubules ◽  
Adult Rats ◽  
Hypotonic Treatment ◽  
Old Rats ◽  
Pachytene Spermatocytes

ABSTRACT Indirect approach (hypotonic treatment) and direct approaches (co-cultures and conditioned media) were used in order to investigate the effects of germ cells from adult rats upon transferrin secretion by Sertoli cell cultures prepared from 20-day-old rats. Removal of germ cells contaminating the Sertoli cell cultures resulted in a significant decrease in transferrin secretion whereas the addition of crude germ cell preparations or of enriched preparations of pachytene spermatocytes, early spermatids and of liver epithelial cells (LEC) markedly stimulated this parameter. Furthermore, spent media of pachytene spermatocytes and of early spermatids, but not of LEC, also stimulated transferrin production. It is concluded that germ cells normally located within the adluminal compartment of the seminiferous tubules may be capable of controlling their own supply of iron via their influence upon transferrin secretion by the Sertoli cells.

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

Development ◽  
1994 ◽  
Vol 120 (7) ◽  
pp. 1759-1766 ◽  
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.

148. HORMONALLY REGULATED miRNAS TARGET THE TUBULOBULBAR COMPLEX IN THE TESTIS

2010 ◽  
Vol 22 (9) ◽  
pp. 66
Author(s):  
P. K. Nicholls ◽  
P. G. Stanton ◽  
K. L. Walton ◽  
R. I. McLachlan ◽  
L. O'Donnell ◽  
...  
Keyword(s):  
Sertoli Cell ◽  
Sertoli Cells ◽  
Hormonal Regulation ◽  
Focal Adhesions ◽  
Intercellular Junctions ◽  
Protein Translation ◽  
Seminiferous Epithelium ◽  
Seminiferous Tubules ◽  
Micro Rnas

Spermatogenesis is absolutely dependent on follicle stimulating hormone (FSH) and androgens; acute suppression of these hormones inhibits germ cell development and thus sperm production. The removal of intercellular junctions and release of spermatids by the Sertoli cell, a process known as spermiation, is particularly sensitive to acute hormone suppression(1). To define the molecular mechanisms that mediate FSH and androgen effects in the testis, we investigated the expression and hormonal regulation of micro-RNAs (miRNA), small non-coding RNAs that regulate protein translation and modify cellular responses. By array analysis, we identified 23 miRNAs that were upregulated >2-fold in stage VIII seminiferous tubules following hormone suppression, and in vitro in primary Sertoli cells. We subsequently validated the expression and hormonal regulation of several miRNAs, including miR-23b, -30d and -690 by quantitative PCR in primary Sertoli cells. Bioinformatic analysis of potential targets of hormonally-suppressed miRNAs identified genes associated with Focal adhesions (54 genes, P = –ln(17.97)) and the Regulation of the actin cytoskeleton (52 genes, P = –ln(10.16)), processes known to be intimately associated with adhesion of spermatids to Sertoli cells(2, 3). Furthermore, this analysis identified numerous components of the testicular tubulobulbar complex (TBC) as being targets of hormonally sensitive miRNAs. The TBC is a podosome-like structure between Sertoli and adjacent spermatids in the testis, which internalises intact inter-cellular junctions by endocytotic mechanisms prior to spermiation(4). We then demonstrate the hormonal regulation of predicted miRNA target proteins, and validate novel inhibitory miRNA interactions with Pten, nWASP, Eps15 and Picalm by luciferase knockdown in vitro. We hypothesise that hormonally suppressed miRNAs inhibit TBC function, and subsequently, endocytosis of intercellular junctions. In conclusion, we have demonstrated that hormonal suppression in the testis stimulates the expression of a subset of Sertoli cell miRNAs that are likely regulators of cell adhesion protein networks involved in spermiation. (1) Saito K, O’Donnell L, McLachlan RI, Robertson DM 2000 Spermiation failure is a major contributor to early spermatogenic suppression caused by hormone withdrawal in adult rats. Endocrinology 141: 2779–2.(2) O’Donnell L, Stanton PG, Bartles JR, Robertson DM 2000 Sertoli cell ectoplasmic specializations in the seminiferous epithelium of the testosterone-suppressed adult rat. Biol Reprod 63: 99–108.(3) Beardsley A, Robertson DM, O’Donnell L 2006 A complex containing alpha6beta1-integrin and phosphorylated focal adhesion kinase between Sertoli cells and elongated spermatids during spermatid release from the seminiferous epithelium. J Endocrinol 190(3): 759–70.(4) Young JS, Guttman JA, Vaid KS, Vogl AW 2009 Tubulobulbar complexes are intercellular podosome-like structures that internalize intact intercellular junctions during epithelial remodeling events in the rat testis. Biol Reprod 80: 162–74.

236. Effects of long term recombinant rat follicle-stimulating hormone replacement on the restoration of spermatogenesis after chronic suppression of gonadotrophins in adult rats

2008 ◽  
Vol 20 (9) ◽  
pp. 36
Author(s):  
S. M. Ruwanpura ◽  
P. G. Stanton ◽  
D. M. Robertson ◽  
R. I. McLachlan ◽  
Y. Makanji ◽  
...  
Keyword(s):  
Sertoli Cell ◽  
Germ Cells ◽  
Follicle Stimulating Hormone ◽  
Inhibin B ◽  
Testis Weight ◽  
Adult Rats ◽  
Early Pachytene ◽  
Pachytene Spermatocytes ◽  
Stimulating Hormone

Follicle stimulating hormone (FSH) in short-term rat studies supports spermatogenesis at multiple levels, notably spermatogonial development. The role of FSH in supporting full spermatogenesis in rats is still in question as long-term studies have not been possible due the development of neutralising antibodies to heterologous FSH preparations. This study sought to assess the effects of a homologous recombinant rat FSH (rr-FSH) preparation on the long-term restoration of spermatogenesis. Adult rats were GnRH-immunised (GnRH-im) for 12 weeks then, administered an anti-androgen; flutamide (flut), alone or together with rr-FSH (8µg/rat/daily) for 56 days (1 spermatogenic cycle). Germ and Sertoli cell numbers were quantified using an optical disector stereological method. Testis weight, serum FSH and inhibin B and Sertoli cell nuclear volume were significantly reduced to 15%, 13%, 25% and 57% of controls respectively, following GnRH-im+flut treatment. GnRH-im+flut treatment reduced A/I spermatogonial, type B spermatogonial+preleptotene, leptotene+zygotene and early pachytene spermatocyte numbers to 28%, 68%, 50% and 19% (P < 0.001) of controls respectively, with later germ cells rarely observed. After FSH treatment, no significant affect on testis weight, serum FSH and inhibin B or Sertoli cell number were observed. However, rr-FSH treatment significantly increased numbers of A/I spermatogonia, leptotene+zygotene and early pachytene spermatocytes from 28 = >42%, 50 = >69% and 19 = >27% of controls, respectively, while no differences were observed in later germ cell types. rr-FSH also increased (P < 0.05) the volume of Sertoli cell nuclei from 57 = >66% of control. In conclusion, FSH is unable to support full rat spermatogenesis; however, FSH can partially support germ cells notably spermatogonia through to early pachytene spermatocytes, despite the absence of androgenic support.

scholarly journals Pachytene spermatocytes in co-culture inhibit rat Sertoli cell synthesis of inhibin beta B-subunit and inhibin B but not the inhibin alpha-subunit

2002 ◽  
Vol 172 (3) ◽  
pp. 565-574 ◽  
Author(s):  
RJ Clifton ◽  
L O'Donnell ◽  
DM Robertson
Keyword(s):  
Mrna Expression ◽  
Sertoli Cell ◽  
Sertoli Cells ◽  
Inhibin B ◽  
Alpha Subunit ◽  
Subunit Mrna ◽  
B Subunit ◽  
Subunit Expression ◽  
Pachytene Spermatocytes

This study investigates the effects of spermatogenic germ cells on inhibin alpha-subunit and beta B-subunit expression, and inhibin alpha-subunit and inhibin B production by rat Sertoli cells in vitro. Sertoli cells isolated from 19-day-old rats were cultured for 48 h at 32 degrees C, in the presence or absence of FSH (2.3-2350 mIU/ml), and in the presence of pachytene spermatocytes, round spermatids or cytoplasts of elongated spermatids purified from adult rat testis by elutriation and density gradient separation. Sertoli cell secretion of inhibin alpha-subunit and inhibin B, as measured by immunoassay, was dose-dependently stimulated by FSH (maximal stimulation 13- and 2-fold, respectively). Round spermatids or cytoplasts co-cultured with Sertoli cells had no effect on basal or FSH-induced secretion of inhibin alpha-subunit or inhibin B. When Sertoli cells were co-cultured with pachytene spermatocytes, inhibin alpha-subunit secretion was unaltered, while inhibin B secretion was suppressed in a cell concentration-dependent manner to reach a maximal suppression of 45% compared with Sertoli cells alone (P<0.01). A similar suppression in inhibin B was still observed (64% of Sertoli cells alone) when the pachytene spermatocytes were separated from Sertoli cells by a 0.45 microm pore membrane barrier in bicameral chambers. Pachytene spermatocytes also suppressed FSH-induced inhibin B levels in Sertoli cell co-cultures and this suppression was attributed to a decrease in basal inhibin B production rather than a change in FSH responsiveness. Quantitation of Sertoli cell inhibin alpha- and beta B-subunit mRNA by quantitative (real-time) PCR demonstrated that pachytene spermatocytes did not alter Sertoli cell alpha-subunit mRNA expression, but significantly (P<0.01) suppressed basal and FSH-induced beta B-subunit mRNA expression to a similar degree to that seen with inhibin B protein levels. It is concluded that pachytene spermatocytes in vitro suppress Sertoli cell inhibin B secretion via factor-mediated suppression of inhibin beta B-subunit expression. These findings support the hypothesis that specific germ cell types can influence inhibin B secretion by the testis independent of FSH regulation.

scholarly journals Glial cell-line-derived neurotropic factor and its receptors are expressed by germinal and somatic cells of the rat testis

2006 ◽  
Vol 190 (1) ◽  
pp. 59-71 ◽  
Author(s):  
Sophie Fouchécourt ◽  
Murielle Godet ◽  
Odile Sabido ◽  
Philippe Durand
Keyword(s):  
Cell Line ◽  
Glial Cell ◽  
Sertoli Cells ◽  
Small Population ◽  
Seminiferous Tubules ◽  
Rat Testis ◽  
Old Rats ◽  
Neurotropic Factor ◽  
Pachytene Spermatocytes ◽  
Factor Α

Glial cell-line-derived neurotropic factor (GDNF) and its receptors glial cell-line-derived neurotropic factor α (GFR1α) and rearranged during transformation (RET) have been localized in the rat testis during postnatal development. The three mRNAs, and GDNF and GFR1α proteins were detected in testis extracts from 1- to 90-day-old rats by reverse transcriptase PCR and Western blotting respectively. The three mRNAs were present in Sertoli cells from 20- and 55-day-old rats, pachytene spermatocytes (PS), and round spermatids (RS). The GDNF and GFR1α proteins were detected in PS, RS, and Sertoli cells. GDNF and GFR1α were also detected using flow cytometry in spermatogonia and preleptotene spermatocytes, and in secondary spermatocytes. The localization of GDNF and GFR1α in germ and Sertoli cells was confirmed by immunocytochemistry. The hypothesis that GDNF may control DNA synthesis of Sertoli cells and/or spermatogonia in the immature rat was addressed using cultures of seminiferous tubules from 7- to 8-day-old rats. Addition of GDNF for 48 h resulted in a twofold decrease in the percentage of spermatogonia able to duplicate DNA, whereas Sertoli cells were not affected. These results are consistent with a role of GDNF in inhibiting the S-phase entrance of a large subset of differentiated type A spermatogonia, together with an enhancing effect of the factor on a small population of undifferentiated (stem cells) spermatogonia. Moreover, the wide temporal and spatial expression of GDNF and its receptors in the rat testis suggest that it might act at several stages of spermatogenesis.

scholarly journals Establishment and applications of male germ cell and Sertoli cell lines

Reproduction ◽  
2016 ◽  
Vol 152 (2) ◽  
pp. R31-R40 ◽  
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.

A combination of insulin-like growth factor I (IGF-I) and FSH promotes proliferation of prepubertal bovine Sertoli cells isolated and cultured in vitro

2017 ◽  
Vol 29 (8) ◽  
pp. 1635 ◽  
Author(s):  
A. Dance ◽  
J. Kastelic ◽  
J. Thundathil
Keyword(s):  
Growth Factor ◽  
Sertoli Cell ◽  
Sertoli Cells ◽  
Seminiferous Tubules ◽  
Insulin Like Growth Factor ◽  
Factor I ◽  
Bull Calves ◽  
Igf I ◽  
Growth Factor I

Beef and dairy bull calves fed a low-nutrition diet during early life had decreased concentrations of circulating insulin-like growth factor I (IGF-I), delayed increases in testosterone, smaller testes and delayed puberty compared with those fed high-nutrition diets. Although IGF-1 has important roles in Sertoli cell function in rats and mice, this has not been well documented in bulls. The objectives of this study were to: (1) isolate Sertoli cells from bull calves at 8 weeks of age, (2) culture them in vitro and (3) determine the effects of IGF-I, FSH and a combination of both hormones on cell proliferation. For Sertoli cell isolation, minced testicular tissues were treated with collagenase followed by trypsin and hyaluronidase to digest seminiferous tubules and release Sertoli cells. In this study, Sertoli cells were successfully isolated from 8-week-old Holstein bull calves (n = 4) and these cells were cultured for up to 8 days. A combination of IGF-I and FSH increased proliferation (~18%) and therefore cell number (1.5-fold) of prepubertal bovine Sertoli cells in culture, providing clear evidence that IGF-I has a similar role in bovine Sertoli cells as reported in rodents.

Sertoli cell replacement in explanted mouse testis tissue supporting host spermatogenesis

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.

Effects of FSH and testosterone on highly purified rat Sertoli cells: inhibin α-subunit mRNA expression and inhibin secretion are enhanced by FSH but not by testosterone

1989 ◽  
Vol 122 (3) ◽  
pp. 757-762 ◽  
Author(s):  
A. M. W. Toebosch ◽  
D. M. Robertson ◽  
I. A. Klaij ◽  
F. H. de Jong ◽  
J. A. Grootegoed
Keyword(s):  
Mrna Expression ◽  
Effective Dose ◽  
Sertoli Cell ◽  
Germ Cells ◽  
Sertoli Cells ◽  
Osmotic Shock ◽  
Shock Treatment ◽  
Α Subunit ◽  
Subunit Mrna ◽  
Old Rats

ABSTRACT The effects of FSH and testosterone on inhibin mRNA expression and inhibin production by highly purified Sertoli cell preparations were examined. Sertoli cells were isolated from testes of 22-day-old rats by sequential trypsin, collagenase and hyaluronidase treatments, with subsequent osmotic shock treatment on day 3 of culture. Contamination by peritubular and germ cells was <0·5 and 1–3% respectively. Intracellular and secreted inhibin levels were measured by radioimmunoassay, using Sertoli cells which were incubated for 24 h in the absence or presence of FSH and testosterone from days 4 to 5 of culture. FSH stimulated the cellular inhibin content and the secreted inhibin level by four- and sevenfold respectively, with a half-maximal effective dose of 5–50 ng/ml. Under the present incubation conditions, testosterone (1 μmol/l) had no effect on immunoreactive inhibin levels in either the presence or absence of FSH. Similarly, the expression of inhibin α-subunit mRNA was increased following FSH stimulation, whereas testosterone had no effect. The expression of inhibin βB-subunit mRNAs was not influenced by FSH or testosterone. It is concluded that highly purified Sertoli cell preparations, with a very low number of peritubular or germ cells, are fully responsive to FSH with respect to inhibin mRNA expression and inhibin production. Journal of Endocrinology (1989) 122, 757–762

METABOLIC CHANGES IN TESTICULAR CELLS FROM RATS AFTER LONG-TERM EXPOSURE TO 37 °C IN VIVO OR IN VITRO

1980 ◽  
Vol 85 (3) ◽  
pp. 471-479 ◽  
Author(s):  
F. F. G. ROMMERTS ◽  
F. H. DE JONG ◽  
J. A. GROOTEGOED ◽  
H. J. VAN DER MOLEN
Keyword(s):  
Sertoli Cells ◽  
Leydig Cells ◽  
Biochemical Properties ◽  
Effect Of Temperature ◽  
Seminiferous Tubules ◽  
Experimental Conditions ◽  
Testicular Cells ◽  
Old Rats

Biochemical properties of isolated Leydig cells, Sertoli cells and spermatocytes from rat testes have been investigated after in-vivo or in-vitro exposure of these cells to abdominal temperature (37 °C). The rate of production of testosterone and pregnenolone by isolated Leydig cells from cryptorchid and normal testes from mature rats was not different. Production of pregnenolone by mitochondria prepared from cryptorchid testes was 6·7 times higher than production by mitochondria from normal testes. Sertoli cells prepared from immature rats and incubated in vitro at 32 or 37 °C showed, on day 1 of the culture period, an initial twofold increase in the secretion of androgen-binding protein which was absent after 6 days in culture. In contrast, incorporation of [3H]leucine into secreted proteins was stimulated twofold on day 1 as well as by day 6 of culture. Secretion of oestradiol was increased 30-fold by day 6 when compared with the level found on day 1 when cells had been cultured at 37 °C and the increased secretion of oestradiol was maintained for approximately 2 days when the temperature of incubation was decreased to 32 °C Spermatocytes isolated from seminiferous tubules incubated for 20 h at 37 °C were active in the synthesis of RNA. No degeneration of these cells was observed in testes of 25-day-old rats 5 days after experimental cryptorchidism, whereas under similar conditions massive degeneration of spermatocytes was shown in the testes of mature rats. These results suggest that the effects of temperature on the different testicular cells greatly depend on the experimental conditions used to study the effect of temperature.

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