234 MORPHOLOGICAL CHARACTERIZATION OF TESTES FROM INSULIN RECEPTOR SUBSTRATE (IRS) 2-DEFICIENT MICE

2011 ◽  
Vol 23 (1) ◽  
pp. 216
Author(s):  
R. J. Griffeth ◽  
D. J. Burks
Keyword(s):  
Insulin Receptor ◽  
Sertoli Cells ◽  
Insulin Signalling ◽  
Seminiferous Epithelium ◽  
Structural Defects ◽  
Seminiferous Tubules ◽  
Testicular Development ◽  
Insulin Receptor Substrate ◽  
Morphological Criteria ◽  
Deficient Mice

In mammals, reproduction is controlled by the hypothalamic-pituitary-gonadal axis but it is also affected by changes in energy homeostasis and metabolism. Insulin exhibits pleiotropic effects that are tissue and development dependent. In humans, sperm from males with diabetes exhibit severe structural defects and lower motility and concentration. Insulin receptor substrate 2 (IRS2) is a key mediator in the insulin/insulin-like growth factor-1 signalling pathway. Deletion of IRS2 in mice leads to diabetes and causes female infertility and impaired male fertility. The objective of this study was to determine if the absence of IRS2 in male mice causes a testicular phenotype. Mice were categorized into 3 groups based on genotyping and blood glucose concentrations: wildtype (WT), knockout normal (KOn), or knockout diabetic (KOd); n = 6 mice per group. Testes weighed significantly less in KOn (107.5 ± 10.4 mg) and KOd (112.9 ± 11.1 mg) mice, which corresponds to a reduction in testes size of 46 and 43%, respectively, compared with WT (197.8 ± 29.9 mg; P < 0.001). To determine the difference between the size of the testes in KO and WT mice, the diameter of the seminiferous tubules and the length of the seminiferous epithelium were measured. In both KO groups, both parameters were significantly reduced (P < 0.001). The diameter of the seminiferous tubules of KOn (123.0 ± 12.6 μm) and KOd (137.1 ± 13.1 μm) was reduced by 42 and 35%, respectively, compared with WT (210.5 ± 16.5 μm). The length of the seminiferous epithelium of the KOn (44.0 ± 5.5 μm) and KOd (41.5 ± 6.9 μm) was reduced by 43 and 45%, respectively, compared with WT (76.6 ± 7.25 μm). Because the number of Sertoli cells determines the size of the testes and the number of germ cells that can be supported during spermatogenesis, Sertoli cells were counted. The number of Sertoli cells per cross section of seminiferous tubule was significantly lower in KO v. WT mice (KOn 13.7 ± 2.5, KOd 15 ± 2.8, WT 24.1 ± 2.5; P < 0.001), corresponding to a 43% reduction in KOn and a 38% reduction in KOd. To determine how the differences in morphology affect spermatogenesis, epididymal sperm were counted. As expected, there were fewer sperm in KO mice than WT mice (KOn 3.80 × 106, KOd 8.58 × 106, WT 1.34 × 107). After in vitro capacitation, sperm from all groups were motile; however, sperm from WT mice displayed a more characteristic motility pattern. In summary, there was a reduction in all morphological criteria studied in the testes of KO mice compared with WT. Although IRS2-deficient mice are initially able to breed, when they become severely diabetic, they are infertile. It appears that reductions in testicular morphology and declining numbers of sperm are likely causes of the reduced fertility. The data suggest that insulin signalling in the testes is important for normal testicular development and function and that metabolic abnormalities such as diabetes lead to reduced fertility.

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.

Effect of thyroid hormone on the pre- and post-natal development of the rat testis

1991 ◽  
Vol 129 (1) ◽  
pp. 35-NP ◽  
Author(s):  
S. Francavilla ◽  
G. Cordeschi ◽  
G. Properzi ◽  
L. Di Cicco ◽  
E. A. Jannini ◽  
...  
Keyword(s):  
Germ Cells ◽  
Sertoli Cells ◽  
Serum Levels ◽  
Seminiferous Tubules ◽  
Testicular Development ◽  
Postnatal Life ◽  
Fetal Life ◽  
Newborn Rats ◽  
Total Thyroxine ◽  
Delayed Maturation

ABSTRACT The relationship between thyroid function and testicular development in the rat was investigated. Hypothyroidism was induced during fetal or postnatal life by adding methimazole (MMI) to the drinking water of pregnant or lactating mothers. A group of newborn rats was treated with MMI and i.p. injections of l-tri-iodothyronine (l-T3). Hypothyroidism was shown by the reduced serum levels of total T3 and of total thyroxine (T4) in pregnant mothers and in pubertal rats. Testes were studied using light microscopy at 18 and 21 days post coitum or during puberty (21, 35 and 50 days after birth); serum levels of gonadotrophins were also evaluated in pubertal rats. Hypothyroidism had no effect on testicular development during fetal life and when induced in newborn rats it was associated at puberty with reduced serum levels of FSH and LH and with delayed maturation of the testis compared with control rats. The delay in maturation consisted of a reduction in the diameter of seminiferous tubules, and a reduction in the number of germ cells per tubule; this was associated with increased degeneration and arrested maturation of germ cells. In addition, Sertoli cells demonstrated retarded development, as indicated by a delay in the appearance of cytoplasmic lipids and in the development of a tubule lumen. Hormonal and morphological abnormalities were absent in rats treated with MMI plus l-T3. In conclusion, hypothyroidism occurring soon after birth caused reduced levels of gonadotrophins in the serum and a delay in pubertal spermatogenesis, possibly due to retarded differentiation of the Sertoli cells. Journal of Endocrinology (1991) 129, 35–42

scholarly journals A novel regulation of IRS1 (insulin receptor substrate-1) expression following short term insulin administration

2005 ◽  
Vol 392 (2) ◽  
pp. 345-352 ◽  
Author(s):  
Antonio J. Ruiz-Alcaraz ◽  
Hui-Kang Liu ◽  
Daniel J. Cuthbertson ◽  
Edward J. Mcmanus ◽  
Simeen Akhtar ◽  
...  
Keyword(s):  
Insulin Receptor ◽  
Insulin Signalling ◽  
Insulin Injection ◽  
Underlying Mechanism ◽  
Type Ii Diabetes Mellitus ◽  
Insulin Receptor Substrate ◽  
Insulin Receptor Substrate 1 ◽  
Insulin Receptor Tyrosine Kinase ◽  
Cell Culture Systems ◽  
Or Gene

Reduced insulin-mediated glucose transport in skeletal muscle is a hallmark of the pathophysiology of T2DM (Type II diabetes mellitus). Impaired intracellular insulin signalling is implicated as a key underlying mechanism. Attention has focused on early signalling events such as defective tyrosine phosphorylation of IRS1 (insulin receptor substrate-1), a major target for the insulin receptor tyrosine kinase. This is required for normal induction of signalling pathways key to many of the metabolic actions of insulin. Conversely, increased serine/threonine phosphorylation of IRS1 following prolonged insulin exposure (or in obesity) reduces signalling capacity, partly by stimulating IRS1 degradation. We now show that IRS1 levels in human muscle are actually increased 3-fold following 1 h of hyperinsulinaemic euglycaemia. Similarly, transient induction of IRS1 (3-fold) in the liver or muscle of rodents occurs following feeding or insulin injection respectively. The induction by insulin is also observed in cell culture systems, although to a lesser degree, and is not due to reduced proteasomal targeting, increased protein synthesis or gene transcription. Elucidation of the mechanism by which insulin promotes IRS1 stability will permit characterization of the importance of this novel signalling event in insulin regulation of liver and muscle function. Impairment of this process would reduce IRS1 signalling capacity, thereby contributing to the development of hyperinsulinaemia/insulin resistance prior to the appearance of T2DM.

scholarly journals 4PS/Insulin Receptor Substrate (IRS)-2 Is the Alternative Substrate of the Insulin Receptor in IRS-1-deficient Mice

1995 ◽  
Vol 270 (42) ◽  
pp. 24670-24673 ◽  
Author(s):  
Mary-Elizabeth Patti ◽  
Xiao-Jian Sun ◽  
Jens C. Bruening ◽  
Eiichi Araki ◽  
Myra A. Lipes ◽  
...  
Keyword(s):  
Insulin Receptor ◽  
Insulin Receptor Substrate ◽  
Alternative Substrate ◽  
Deficient Mice

scholarly journals Cyclic Expression of Endothelin-converting Enzyme-1 Mediates the Functional Regulation of Seminiferous Tubule Contraction

1999 ◽  
Vol 145 (5) ◽  
pp. 1027-1038 ◽  
Author(s):  
Antonella Tripiciano ◽  
Carmelina Peluso ◽  
Anna Rita Morena ◽  
Fioretta Palombi ◽  
Mario Stefanini ◽  
...  
Keyword(s):  
Sertoli Cell ◽  
Sertoli Cells ◽  
Seminiferous Tubule ◽  
Seminiferous Epithelium ◽  
Seminiferous Tubules ◽  
Converting Enzyme ◽  
Myoid Cells ◽  
Endothelin 1 ◽  
Endothelin Converting Enzyme ◽  
Cyclic Expression

The potent smooth muscle agonist endothelin-1 (ET-1) is involved in the local control of seminiferous tubule contractility, which results in the forward propulsion of tubular fluid and spermatozoa, through its action on peritubular myoid cells. ET-1, known to be produced in the seminiferous epithelium by Sertoli cells, is derived from the inactive intermediate big endothelin-1 (big ET-1) through a specific cleavage operated by the endothelin-converting enzyme (ECE), a membrane-bound metalloprotease with ectoenzymatic activity. The data presented suggest that the timing of seminiferous tubule contractility is controlled locally by the cyclic interplay between different cell types. We have studied the expression of ECE by Sertoli cells and used myoid cell cultures and seminiferous tubule explants to monitor the biological activity of the enzymatic reaction product. Northern blot analysis showed that ECE-1 (and not ECE-2) is specifically expressed in Sertoli cells; competitive enzyme immunoassay of ET production showed that Sertoli cell monolayers are capable of cleaving big ET-1, an activity inhibited by the ECE inhibitor phosphoramidon. Microfluorimetric analysis of intracellular calcium mobilization in single cells showed that myoid cells do not respond to big endothelin, nor to Sertoli cell plain medium, but to the medium conditioned by Sertoli cells in the presence of big ET-1, resulting in cell contraction and desensitization to further ET-1 stimulation; in situ hybridization analysis shows regional differences in ECE expression, suggesting that pulsatile production of endothelin by Sertoli cells (at specific “stages” of the seminiferous epithelium) may regulate the cyclicity of tubular contraction; when viewed in a scanning electron microscope, segments of seminiferous tubules containing the specific stages characterized by high expression of ECE were observed to contract in response to big ET-1, whereas stages with low ECE expression remained virtually unaffected. These data indicate that endothelin-mediated spatiotemporal control of rhythmic tubular contractility might be operated by Sertoli cells through the cyclic expression of ECE-1, which is, in turn, dependent upon the timing of spermatogenesis.

scholarly journals Autologous transplantation of spermatogonial stem cells restores fertility in congenitally infertile mice

2020 ◽  
Vol 117 (14) ◽  
pp. 7837-7844
Author(s):  
Mito Kanatsu-Shinohara ◽  
Narumi Ogonuki ◽  
Shogo Matoba ◽  
Atsuo Ogura ◽  
Takashi Shinohara
Keyword(s):  
Stem Cells ◽  
Sertoli Cells ◽  
Expression Patterns ◽  
Autologous Transplantation ◽  
Spermatogonial Stem Cells ◽  
Congenital Defects ◽  
Seminiferous Tubules ◽  
Special Environment ◽  
Deficient Mice

The blood–testis barrier (BTB) is thought to be indispensable for spermatogenesis because it creates a special environment for meiosis and protects haploid cells from the immune system. The BTB divides the seminiferous tubules into the adluminal and basal compartments. Spermatogonial stem cells (SSCs) have a unique ability to transmigrate from the adluminal compartment to the basal compartment through the BTB upon transplantation into the seminiferous tubule. Here, we analyzed the role ofCldn11, a major component of the BTB, in spermatogenesis using spermatogonial transplantation.Cldn11-deficient mice are infertile due to the cessation of spermatogenesis at the spermatocyte stage.Cldn11-deficient SSCs failed to colonize wild-type testes efficiently, andCldn11-deficient SSCs that underwent double depletion ofCldn3andCldn5showed minimal colonization, suggesting that claudins on SSCs are necessary for transmigration. However,Cldn11-deficient Sertoli cells increased SSC homing efficiency by >3-fold, suggesting that CLDN11 in Sertoli cells inhibits transmigration of SSCs through the BTB. In contrast to endogenous SSCs in intactCldn11-deficient testes, those from WT orCldn11-deficient testes regenerated sperm inCldn11-deficient testes. The success of this autologous transplantation appears to depend on removal of endogenous germ cells for recipient preparation, which reprogrammed claudin expression patterns in Sertoli cells. Consistent with this idea, in vivo depletion ofCldn3/5regenerated endogenous spermatogenesis inCldn11-deficient mice. Thus, coordinated claudin expression in both SSCs and Sertoli cells expression is necessary for SSC homing and regeneration of spermatogenesis, and autologous stem cell transplantation can rescue congenital defects of a self-renewing tissue.

scholarly journals Axl Promotes Zika Virus Entry and Modulates the Antiviral State of Human Sertoli Cells

mBio ◽  
2019 ◽  
Vol 10 (4) ◽  
Author(s):  
Daniel P. Strange ◽  
Boonyanudh Jiyarom ◽  
Nima Pourhabibi Zarandi ◽  
Xuping Xie ◽  
Coleman Baker ◽  
...  
Keyword(s):  
Sertoli Cells ◽  
Zika Virus ◽  
Leydig Cells ◽  
Virus Entry ◽  
Seminiferous Epithelium ◽  
Seminiferous Tubules ◽  
Type I ◽  
Cell Type ◽  
Zikv Infection ◽  
Antiviral State

ABSTRACT Zika virus (ZIKV) is unique among mosquito-borne flaviviruses in its ability to be sexually transmitted. Persistent ZIKV infection in the testes, which are immune privileged organs, long after peripheral clearance suggests involvement of immunosuppressive pathways; however, the underlying mechanisms remain undetermined. We recently demonstrated that ZIKV infects human Sertoli cells (SC), the major cell type of the seminiferous epithelium responsible for maintaining the immune privileged compartment of seminiferous tubules. Recent reports have identified the TAM (Tyro3, Axl, Mer) receptor tyrosine kinase Axl as an entry receptor and/or immune modulator for ZIKV in a cell type-specific manner. Interestingly, the seminiferous epithelium exhibits high basal expression of the Axl receptor where it is involved in clearance of apoptotic germ cells and immunosuppression. Here, we show that Axl was highly expressed in SC compared to Leydig cells (LC) that correlated with robust ZIKV infection of SC, but not LC. Further, neutralization of Axl receptor and its ligand Gas6 strongly attenuated virus entry in SC. However, inhibition of Axl kinase did not affect ZIKV entry but instead led to decreased protein levels of suppressor of cytokine signaling 1 (SOCS1) and SOCS3, increased expression of interferon-stimulated genes (ISGs), and reduced ZIKV replication. Similarly, treatment of multicellular human testicular organoids with an Axl kinase inhibitor attenuated ZIKV replication and increased ISG expression. Together, our data demonstrate that Axl promotes ZIKV entry and negatively regulates the antiviral state of SC to augment ZIKV infection of the testes and provides new insights into testis antiviral immunity and ZIKV persistence. IMPORTANCE Recent Zika virus (ZIKV) outbreaks have identified sexual transmission as a new route of disease spread not reported for other flaviviruses. ZIKV crosses the blood-testis barrier and establishes infection in seminiferous tubules, the site for spermatozoa development. Currently, there are no therapies to treat ZIKV infection, and the immune mechanisms underlying testicular persistence are unclear. We found that multiple human testicular cell types, except Leydig cells, support ZIKV infection. Axl receptor, which plays a pivotal role in maintaining the immunosuppressive milieu of the testis, is highly expressed in Sertoli cells and augments ZIKV infection by promoting virus entry and negatively regulating the antiviral state. By using testicular organoids, we further describe the antiviral role of Axl inhibition. The significance of our research lies in defining cross talk between Axl and type I interferon signaling as an essential mechanism of immune control that can inform therapeutic efforts to clear ZIKV from the testis.

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