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1.
Biol Reprod ; 98(4): 491-500, 2018 04 01.
Article in English | MEDLINE | ID: mdl-29365049

ABSTRACT

Human female reproductive aging features declining ovarian follicle reserve and oocyte quality, and rising levels of circulating follicle-stimulating hormone (FSH). We determined the effects of elevated FSH on oocyte-embryo development in mature mice exhibiting premature infertility caused by progressively rising transgenic human FSH (TgFSH) levels. Oocyte-embryo developmental competence and quality were examined using oocyte maturation and aneuploidy rates, biomarkers of oocyte quality, and reciprocal embryo transfers assessed for implantation and pregnancy. In vitro maturation suggested that TgFSH exposure only hindered oocyte developmental competence in old females, as significantly more oocytes from ≥12-month-old TgFSH females remained at germinal vesicle stage compared with age-matched control oocytes. Aneuploidy rates were equivalent in oocytes from aging TgFSH compared with wildtype females. Cumulus cell expression levels of candidate biomarker Inhba, Egfr, and Rgs2 transcripts were elevated in associated aneuploid vs euploid oocytes from both TgFSH and wildtype females. In vivo, embryos transferred from subfertile 6-month-old TgFSH females to wildtype recipients yielded normal implantation rates and more pups born compared with controls. Transfer of wildtype embryos rescued the fertility of 6-month-old TgFSH-recipient females, although pup birth weight was reduced in TgFSH vs wildtype recipients. Our current findings show that elevated FSH had minimal disruption of either embryo developmental capacity or uterine function when examined in isolation, and the subfertility of TgFSH female mice was not caused by altered oocyte aneuploidy or quality.


Subject(s)
Embryonic Development/physiology , Follicle Stimulating Hormone/genetics , Infertility, Female/genetics , Oocytes/metabolism , Uterus/metabolism , Animals , Cumulus Cells/metabolism , ErbB Receptors/genetics , ErbB Receptors/metabolism , Female , Follicle Stimulating Hormone/metabolism , Infertility, Female/metabolism , Inhibin-beta Subunits/genetics , Inhibin-beta Subunits/metabolism , Mice , Mice, Transgenic , Pregnancy , RGS Proteins/genetics , RGS Proteins/metabolism
2.
Horm Cancer ; 7(5-6): 316-326, 2016 12.
Article in English | MEDLINE | ID: mdl-27506975

ABSTRACT

Phosphatase and tensin homologue (PTEN) is a known tumour suppressor. To explore the role of Pten in ovarian tumorigenesis, we used transgenic (Tg) SOX2. Cre and AMH. Cre mouse models to direct global Pten haploinsufficiency (Pten +/-) or ovary-specific granulosa cell (GC) Pten disruption (Pten GC ). Pten mutant models were combined with progressively rising Tg-follicle-stimulating hormone (TgFSH) levels to study the tumorigenic potential of combined genetic/endocrine modification in vivo. Global Pten +/- mice exhibited grossly detectable tumours in multiple organs including uterine and mammary tissue and displayed reduced survival. Despite extra-ovarian tumorigenesis, Pten +/- females had no detectable ovarian tumours, although elevated corpus luteum numbers increased ovary size and estrous cycling was altered. Combined TgFSH/Pten +/- mice also had no ovarian tumours, but early survival was reduced in the presence of TgFSH. Ovary-specific Pten GC  ± TgFSH females exhibited no detectable ovarian or uterine tumours, and corpus luteum numbers and estrous cycling remained unchanged. The non-tumorigenic ovarian phenotypes in Pten +/- and Pten GC  ± TgFSH mice support the proposal that multi-hit genetic mutations (including ovarian and extra-ovarian tissue) initiate ovarian tumours. Our findings suggest that elevated FSH may reduce early cancer survival; however, the ovary remains remarkably resistant to Pten-induced tumorigenic changes even in the presence of uterine and reproductive cancers.


Subject(s)
Follicle Stimulating Hormone/metabolism , Granulosa Cells/metabolism , Ovarian Neoplasms/pathology , Ovarian Neoplasms/veterinary , PTEN Phosphohydrolase/genetics , SOXB1 Transcription Factors/genetics , Animals , Female , Mammary Neoplasms, Animal/genetics , Mice , Mice, Transgenic , Mutation , Organ Size , Ovarian Neoplasms/genetics , Ovarian Neoplasms/metabolism , PTEN Phosphohydrolase/metabolism , Survival Analysis , Uterine Neoplasms/genetics
3.
Am J Physiol Endocrinol Metab ; 311(2): E396-404, 2016 08 01.
Article in English | MEDLINE | ID: mdl-27354237

ABSTRACT

Recently, we created a unique gain-of-function mouse model with Sertoli cell-specific transgenic androgen receptor expression (TgSCAR) showing that SCAR activity controls the synchronized postnatal development of somatic Sertoli and Leydig cells and meiotic-postmeiotic germ cells. Moderate TgSCAR (TgSCAR(m)) expression reduced testis size but had no effect on male fertility. Here, we reveal that higher TgSCAR expression (TgSCAR(H)) causes male infertility. Higher SCAR activity, shown by upregulated AR-dependent transcripts (Rhox5, Spinw1), resulted in smaller adult TgSCAR(H) testes (50% of normal) despite normal or elevated circulating and intratesticular testosterone levels. Unlike fertile TgSCAR(m) males, testes of adult TgSCAR(H) males exhibited focal regions of interstitial hypertrophy featuring immature adult Leydig cells and higher intratesticular dihydrotestosterone and 5α-androstane 3α,17ß-diol levels that are normally associated with pubertal development. Mature TgSCAR(H) testes also exhibited markedly reduced Sertoli cell numbers (70%), although meiotic and postmeiotic germ cell/Sertoli cell ratios were twofold higher than normal, suggesting that elevated TgSCAR activity supports excessive spermatogenic development. Concurrent with the higher germ cell load of TgSCAR(H) Sertoli cells were increased levels of apoptotic germ cells in TgSCAR(H) relative to TgSCAR(m) testes. In addition, TgSCAR(H) testes displayed unique morphological degeneration that featured accumulated cellular and spermatozoa clusters in dilated channels of rete testes, consistent with reduced epididymal sperm numbers. Our findings reveal for the first time that excessive Sertoli cell AR activity in mature testes can reach a level that disturbs Sertoli/germ cell homeostasis, impacts focal Leydig cell function, reduces sperm output, and disrupts male fertility.


Subject(s)
Benzamides/metabolism , Fertility/genetics , Infertility, Male/genetics , Piperidines/metabolism , Receptors, Androgen/genetics , Sertoli Cells/metabolism , Androstane-3,17-diol/metabolism , Animals , Dihydrotestosterone/metabolism , Epididymis , Homeodomain Proteins/genetics , Male , Meiosis , Mice, Transgenic , Proteinase Inhibitory Proteins, Secretory/genetics , Proteins/genetics , Real-Time Polymerase Chain Reaction , Rete Testis/pathology , Spermatogenesis , Spermatozoa , Testis , Transcription Factors/genetics
4.
Horm Cancer ; 6(4): 142-52, 2015 Aug.
Article in English | MEDLINE | ID: mdl-25943777

ABSTRACT

BRCA1 mutations are associated with ovarian cancer. Previous studies reported that murine granulosa cell (GC) Brca1 loss caused ovarian-uterine tumors resembling serous cystadenomas, but the pathogenesis of these tumors may have been confounded by ectopic Brca1 expression and altered estrous cycling. We have used Tg.AMH.Cre conferring proven ovarian and GC-specific Cre activity to selectively target Brca1 disruption, denoted Brca1(GC-/-). Furthermore, ovary-specific Brca1(GC-/-) was combined with global Trp53 haploinsufficiency (Trp53(+/-)) and transgenic follicle-stimulating hormone (Tg.FSH) overexpression as a multi-hit strategy to investigate additional genetic and hormonal ovarian tumorigenesis mechanisms. However, 12-month-old Brca1(GC-/-) mice had no detectable ovarian or uterine tumors. Brca1(GC-/-) mice had significantly increased ovary weights, follicles exhibiting more pyknotic granulosa cells, and fewer corpora lutea with regular estrous cycling compared to controls. Isolated Brca1(GC-/-) mutation lengthened the estrous cycle and proestrus stage; however, ovarian cystadenomas were not observed, even when Brca1(GC-/-) was combined with Trp53(+/-) and overexpressed Tg.FSH. Our Brca1(GC-/-) models reveal that specific intra-follicular Brca1 loss alone, or combined with cancer-promoting genetic (Trp53 loss) and endocrine (high serum FSH) changes, was not sufficient to cause ovarian tumors. Our findings show that the ovary is remarkably resistant to oncogenesis, and support the emerging view of an extragonadal, multi-hit origin for ovarian tumorigenesis.


Subject(s)
BRCA1 Protein/genetics , Follicle Stimulating Hormone/genetics , Haploinsufficiency , Ovarian Neoplasms/pathology , Tumor Suppressor Protein p53/genetics , Animals , Cystadenoma/genetics , Cystadenoma/pathology , Estradiol/metabolism , Female , Follicle Stimulating Hormone/metabolism , Granulosa Cells/metabolism , Mice , Mice, Transgenic , Ovarian Neoplasms/genetics , Ovary/pathology , Uterus/pathology
5.
Endocrinology ; 155(3): 1120-30, 2014 Mar.
Article in English | MEDLINE | ID: mdl-24424066

ABSTRACT

We determined the functional role of the Sertoli cell glucocorticoid receptor (GR) in vivo using a transgenic Cre-loxP approach to conditionally disrupt GR expression. Sertoli cell GR knockout (SCGRKO) was shown by absent Sertoli cell-specific GR immunolocalization and reduced levels of glucocorticoid-responsive Stc1 and Tsc22d3 mRNA in SCGRKO relative to control testes. Adult SCGRKO testes exhibited distinct morphological changes, including reduced seminiferous tubular lumen formation, decreased total Sertoli cell numbers, and parallel reductions in meiotic spermatocyte and postmeiotic spermatid numbers. Conversely, tubular diameter was increased and testis size was normal in SCGRKO males. Decreased serum FSH and testicular Fshr mRNA levels were consistent with reduced Sertoli cell number. Adult SCGRKO testes also displayed atypical germ cells and interstitial focal accumulations of hypertrophic lipid-laden, immature-like Leydig cells. Circulating LH, and testicular Lhr mRNA, testosterone, dihydrotestosterone, and 3α/3ß-diol levels were all reduced in mature SCGRKO mice, whereas serum testosterone and dihydrotestosterone levels remained normal. Moreover, Sertoli cell GR disruption caused differential changes to steroidogenic enzyme transcripts, with down-regulated testicular Cyp11a1 contrasting with up-regulated Hsd17b3 expression. Reduced SCGRKO testicular expression of Hsd11b2, encoding an enzyme for corticosterone inactivation, supports a dynamic coupling between Hsd11b and androgen production. Our novel SCGRKO model has revealed that Sertoli cell-mediated GR actions support normal testicular function. Sertoli cell GR is required to maintain normal testicular Sertoli/germ cell numbers and circulating gonadotropin levels, as well as optimal Leydig cell maturation and steroidogenesis, providing new insight into gluocorticoid-mediated impact on male reproduction.


Subject(s)
Gene Expression Regulation , Receptors, Glucocorticoid/genetics , Receptors, Glucocorticoid/physiology , Sertoli Cells/metabolism , Testis/growth & development , 11-beta-Hydroxysteroid Dehydrogenase Type 2/metabolism , 17-Hydroxysteroid Dehydrogenases/metabolism , Animals , Cholesterol Side-Chain Cleavage Enzyme/metabolism , Gene Expression Regulation, Enzymologic , Glucocorticoids/metabolism , Leydig Cells/cytology , Leydig Cells/metabolism , Luteinizing Hormone/metabolism , Male , Mice , Mice, Knockout , Sertoli Cells/cytology , Spermatogenesis , Testis/metabolism , Transgenes , Up-Regulation
6.
Am J Physiol Endocrinol Metab ; 305(6): E717-26, 2013 Sep 15.
Article in English | MEDLINE | ID: mdl-23880317

ABSTRACT

Homozygous androgen receptor (AR)-knockout (ARKO) female mice are subfertile due to both intra- and extraovarian (neuroendocrine) defects as defined by ovary transplantation. Using ARKO mice, this study set out to reveal the precise AR-regulated pathways required for optimal androgen-regulated ovulation and fertility. ARKO females exhibit deficient neuroendocrine negative feedback, with a reduced serum luteinizing hormone (LH) response to ovariectomy (OVX) (P < 0.01). Positive feedback is also altered as intact ARKO females, at late proestrus, exhibit an often mistimed endogenous ovulatory LH surge. Furthermore, at late proestrus, intact ARKO females display diminished preovulatory serum estradiol (E2; P < 0.01) and LH (P < 0.05) surge levels and reduced Kiss1 mRNA expression in the anteroventral periventricular nucleus (P < 0.01) compared with controls. However, this reduced ovulatory LH response in intact ARKO females can be rescued by OVX and E2 priming or treatment with endogenous GnRH. These findings reveal that AR regulates the negative feedback response to E2, E2-positive feedback is compromised in ARKO mice, and AR-regulated negative and positive steroidal feedback pathways impact on intrahypothalamic control of the kisspeptin/GnRH/LH cascade. In addition, intraovarian AR-regulated pathways controlling antral to preovulatory follicle dynamics are disrupted because adult ARKO ovaries collected at proestrus have small antral follicles with reduced oocyte/follicle diameter ratios (P < 0.01) and increased proportions of unhealthy large antral follicles (P < 0.05) compared with controls. As a consequence of aberrant follicular growth patterns, proestrus ARKO ovaries also exhibit fewer preovulatory follicle (P < 0.05) and corpora lutea numbers (P < 0.01). However, embryo development to the blastocyst stage is unchanged in ARKO females, and hence, the subfertility is a consequence of reduced ovulations and not altered embryo quality. These findings reveal that the AR has a functional role in neuroendocrine regulation and timing of the ovulatory LH surge as well as antral/preovulatory follicle development.


Subject(s)
Hypothalamus/metabolism , Infertility, Female/metabolism , Ovary/metabolism , Ovulation/metabolism , Receptors, Androgen/metabolism , Animals , Corpus Luteum/metabolism , Estradiol/blood , Estrous Cycle/blood , Estrous Cycle/genetics , Estrous Cycle/metabolism , Female , Hypothalamus/physiopathology , Infertility, Female/genetics , Infertility, Female/physiopathology , Kisspeptins/genetics , Kisspeptins/metabolism , Luteinizing Hormone/blood , Luteinizing Hormone/metabolism , Mice , Mice, Knockout , Ovarian Follicle/metabolism , Ovary/physiopathology , Ovulation/blood , Ovulation/genetics , Receptors, Androgen/genetics
7.
Endocrinology ; 154(9): 3410-22, 2013 Sep.
Article in English | MEDLINE | ID: mdl-23766127

ABSTRACT

We recently created a mouse model displaying precocious Sertoli cell (SC) and spermatogenic development induced by SC-specific transgenic androgen receptor expression (TgSCAR). Here we reveal that TgSCAR regulates the development, function, and absolute number of Leydig cells (LCs). Total fetal and adult type LC numbers were reduced in postnatal and adult TgSCAR vs control testes, despite normal circulating LH levels. Normal LC to SC ratios found in TgSCAR testes indicate that SC androgen receptor (SCAR)-mediated activity confers a quorum-dependent relationship between total SC and LC numbers. TgSCAR enhanced LC differentiation, shown by elevated ratios of advanced to immature LC types, and reduced LC proliferation in postnatal TgSCAR vs control testes. Postnatal TgSCAR testes displayed up-regulated expression of coupled ligand-receptor transcripts (Amh-Amhr2, Dhh-Ptch1, Pdgfa-Pdgfra) for potential SCAR-stimulated paracrine pathways, which may coordinate LC differentiation. Neonatal TgSCAR testes displayed normal T and dihydrotestosterone levels despite differential changes to steroidogenic gene expression, with down-regulated Star, Cyp11a1, and Cyp17a1 expression contrasting with up-regulated Hsd3b1, Hsd17b3, and Srd5a1 expression. TgSCAR males also displayed elevated postnatal and normal adult serum testosterone levels, despite reduced LC numbers. Enhanced adult-type LC steroidogenic output was revealed by increased pubertal testicular T, dihydrotestosterone, 3α-diol and 3ß-diol levels per LC and up-regulated steroidogenic gene (Nr5a1, Lhr, Cyp11a1, Cyp17a1, Hsd3b6, Srd5a1) expression in pubertal or adult TgSCAR vs control males, suggesting regulatory mechanisms maintain androgen levels independently of absolute LC numbers. Our unique gain-of-function TgSCAR model has revealed that SCAR activity controls temporal LC differentiation, steroidogenic function, and population size.


Subject(s)
Cell Differentiation , Leydig Cells/cytology , Receptors, Androgen/metabolism , Sertoli Cells/metabolism , Testis/cytology , Testosterone Congeners/metabolism , Animals , Animals, Newborn , Biomarkers/metabolism , Cell Count , Hemizygote , Isoenzymes/biosynthesis , Isoenzymes/genetics , Isoenzymes/metabolism , Leydig Cells/metabolism , Ligands , Male , Mice , Mice, Transgenic , Patched Receptors , Patched-1 Receptor , Receptor, Platelet-Derived Growth Factor alpha/biosynthesis , Receptor, Platelet-Derived Growth Factor alpha/genetics , Receptor, Platelet-Derived Growth Factor alpha/metabolism , Receptors, Androgen/biosynthesis , Receptors, Androgen/genetics , Receptors, Cell Surface/biosynthesis , Receptors, Cell Surface/genetics , Receptors, Cell Surface/metabolism , Receptors, Peptide/biosynthesis , Receptors, Peptide/genetics , Receptors, Peptide/metabolism , Receptors, Transforming Growth Factor beta/biosynthesis , Receptors, Transforming Growth Factor beta/genetics , Receptors, Transforming Growth Factor beta/metabolism , Sertoli Cells/cytology , Sexual Development , Testis/growth & development , Testis/metabolism , Testosterone Congeners/blood , Up-Regulation
8.
Asian J Androl ; 15(1): 87-8, 2013 Jan.
Article in English | MEDLINE | ID: mdl-23223029
9.
Mol Endocrinol ; 27(1): 12-24, 2013 Jan.
Article in English | MEDLINE | ID: mdl-23160479

ABSTRACT

Sertoli cell (SC) androgen receptor (AR) activity is vital for spermatogenesis. We created a unique gain-of-function transgenic (Tg) mouse model to determine the temporal role of SCAR expression in testicular development. The SC-specific rat Abpa promoter directed human Tg AR [Tg SC-specific AR (TgSCAR)] expression, providing strong premature postnatal AR immunolocalized to SC nuclei. Independent Tg lines revealed that TgSCAR dose dependently reduced postnatal and mature testis size (to 60% normal), whereas androgen-dependent mature seminal vesicle weights and serum testosterone levels remained normal. Total SC numbers were reduced in developing and mature TgSCAR testes, despite normal or higher Fshr mRNA and circulating FSH levels. Postnatal TgSCAR testes exhibited elevated levels of AR-regulated Rhox5 and Spinlw1 transcripts, and precocious SC function was demonstrated by early seminiferous tubular lumen formation and up-regulated expression of crucial SC tight-junction (Cldn11 and Tjp1) and phagocytic (Elmo1) transcripts. Early postnatal Amh expression was elevated but declined to normal levels in peripubertal-pubertal TgSCAR vs. control testes, indicating differential age-related regulation featuring AR-independent Amh down-regulation. TgSCAR induced premature postnatal spermatogenic development, shown by increased levels of meiotic (Dmc1 and Spo11) and postmeiotic (Capza3 and Prm1) germ cell transcripts, elevated meiotic-postmeiotic germ:Sertoli cell ratios, and accelerated spermatid development. Meiotic germ:Sertoli cell ratios were further increased in adult TgSCAR mice, indicating predominant SCAR-mediated control of meiotic development. However, postmeiotic germ:Sertoli cell ratios declined below normal. Our unique TgSCAR paradigm reveals that atypical SC-specific temporal AR expression provides a direct molecular mechanism for induction of precocious testicular development, leading to reduced adult testis size and decreased postmeiotic development.


Subject(s)
Receptors, Androgen/metabolism , Sertoli Cells/metabolism , Spermatogenesis , Animals , Female , Follicle Stimulating Hormone/blood , Gene Expression , Gene Expression Regulation, Developmental , Humans , Male , Mice , Mice, Transgenic , Organ Size , Rats , Receptors, Androgen/genetics , Seminiferous Tubules/anatomy & histology , Seminiferous Tubules/growth & development , Spermatocytes/metabolism , Spermatogonia/metabolism , Testis/cytology , Testis/growth & development , Testosterone/blood
10.
Biol Reprod ; 87(6): 151, 2012 Jun.
Article in English | MEDLINE | ID: mdl-23115271

ABSTRACT

Ovarian granulosa cells display strong androgen receptor (AR) expression, suggesting a functional role for direct AR-mediated actions within developing mammalian follicles. By crossing AR-floxed and anti-Müllerian hormone (AMH)-Cre recombinase mice, we generated granulosa cell-specific androgen receptor knockout mice (GCARKO). Cre expression, assessed by lacZ activity, localized to 70%-100% of granulosa cells in most preantral to antral follicles, allowing for selected evaluation of granulosa cell AR-dependent actions during follicle development. Relative to wild-type (WT) females, GCARKO females were subfertile, producing a 24% reduction in the number of litters (P < 0.05) over 6 mo and an age-dependent decrease in total number of pups born, evident from 6 mo of age (P < 0.05). Follicle dynamics were altered in GCARKO ovaries at 3 mo of age, with a significant reduction in large preantral and small antral follicle numbers compared to WT ovaries (P < 0.05). Global premature follicle depletion was not observed, but increased follicular atresia was evident in GCARKO ovaries at 6 mo of age, with an 81% increase in unhealthy follicles and zona pellucida remnants (P < 0.01). Cumulus cell expansion was decreased (P < 0.01) and oocyte viability was diminished in GCARKO females, with a significant reduction in the percentage of oocytes fertilized after natural mating and, thus, in the rate of progression to the two-cell embryo stage (P < 0.05). In addition, compared with age-matched WT females, 6-mo-old GCARKO females exhibited significantly prolonged estrous cycles (P ≤ 0.05), suggesting altered hypothalamic-pituitary-gonadal feedback signaling. In conclusion, our findings revealed that selective loss of granulosa cell AR actions during preantral and antral stages of development leads to a premature reduction in female fecundity through reduced follicle health and oocyte viability.


Subject(s)
Granulosa Cells/metabolism , Infertility, Female/metabolism , Oogenesis , Receptors, Androgen/metabolism , Signal Transduction , Animals , Anti-Mullerian Hormone/genetics , Anti-Mullerian Hormone/metabolism , Cell Survival , Crosses, Genetic , Cumulus Cells/metabolism , Cumulus Cells/pathology , Estrous Cycle/metabolism , Female , Fertilization , Follicular Atresia/metabolism , Granulosa Cells/pathology , Heterozygote , Infertility, Female/etiology , Infertility, Female/pathology , Mice , Mice, Knockout , Mice, Transgenic , Ovarian Follicle/metabolism , Ovarian Follicle/pathology , Receptors, Androgen/deficiency , Receptors, Androgen/genetics , Recombinant Proteins/biosynthesis , Recombinant Proteins/metabolism , Recombinases/genetics , Recombinases/metabolism , Zona Pellucida/metabolism , Zona Pellucida/pathology
11.
Biol Reprod ; 87(2): 38, 2012 Aug.
Article in English | MEDLINE | ID: mdl-22623623

ABSTRACT

Sertoli cell tight junctions (TJs) form at puberty as a major component of the blood-testis barrier (BTB), which is essential for spermatogenesis. This study characterized the hormonal induction of functional Sertoli cell TJ formation in vivo using the gonadotropin-deficient hypogonadal (hpg) mouse that displays prepubertal spermatogenic arrest. Androgen actions were determined in hpg mice treated for 2 or 10 days with dihydrotestosterone (DHT). Follicle-stimulating hormone (FSH) actions were studied in hpg mice expressing transgenic human FSH (hpg+tgFSH) with or without DHT treatment. TJ formation was examined by mRNA expression and immunolocalization of TJ proteins claudin-3 and claudin-11, and barrier functionality was examined by biotin tracer permeability. Immunolocalization of claudin-3 and claudin-11 was extensive at wild-type (wt) Sertoli cell TJs, which functionally excluded permeability tracer. In contrast, seminiferous tubules of hpg testes lacked claudin-3, but claudin-11 protein was present in adluminal regions of Sertoli cells. Biotin tracer permeated throughout these tubules, demonstrating dysfunctional TJs. In hpg+tgFSH testes, claudin-3 was generally absent, but claudin-11 had redistributed basally toward the TJs, where function was variable. In hpg testes, DHT treatment stimulated the redistribution of claudin-11 protein toward the basal region of Sertoli cells by Day 2, increased Cldn3 and Cldn11 mRNA expression, then induced the formation of functional TJs containing both proteins by Day 10. In hpg+tgFSH testes, TJ protein redistribution was accelerated and functional TJs formed by Day 2 of DHT treatment. We conclude that androgen stimulates initial Sertoli cell TJ formation and function in mice, whereas FSH activity is insufficient alone, but augments androgen-induced TJ function.


Subject(s)
Androgens/physiology , Follicle Stimulating Hormone/physiology , Sertoli Cells/physiology , Tight Junctions/physiology , Animals , Connexins/metabolism , Dihydrotestosterone , Disease Models, Animal , Humans , Hypogonadism , Male , Mice , Mice, Transgenic , Organ Size , RNA, Messenger/metabolism , Rats
12.
Biol Reprod ; 86(5): 149, 1-12, 2012 May.
Article in English | MEDLINE | ID: mdl-22337333

ABSTRACT

Polycystic ovary syndrome (PCOS) is the most frequent female endocrine disorder, affecting 5%-10% of women, causing infertility due to dysfunctional follicular maturation and ovulation, distinctive multicystic ovaries and hyperandrogenism, together with metabolic abnormalities including obesity, hyperinsulinism, an increased risk of type 2 diabetes, and cardiovascular disease. The etiology of PCOS is unclear, and decisive clinical studies are limited by ethical and logistic constraints. Consequently treatment is palliative rather than curative and focuses on symptomatic approaches. Hence, a suitable animal model could provide a valuable means with which to study the pathogenesis of the characteristic reproductive and metabolic abnormalities and thereby identify novel and more effective treatments. So far there is no consensus on the best experimental animal model, which should ideally reproduce the key features associated with human PCOS. The prenatally androgenized rhesus monkey displays many characteristics of the human condition, including hyperandrogenism, anovulation, polycystic ovaries, increased adiposity, and insulin insensitivity. However, the high cost of nonhuman primate studies limits the practical utility of these large-animal models. Rodent models, on the other hand, are inexpensive, provide well-characterized and stable genetic backgrounds readily accessible for targeted genetic manipulation, and shorter reproductive life spans and generation times. Recent rodent models display both reproductive and metabolic disturbances associated with human PCOS. This review aimed to evaluate the rodent models reported to identify the advantages and disadvantages of the distinct rodent models used to investigate this complex endocrine disorder.


Subject(s)
Disease Models, Animal , Polycystic Ovary Syndrome/metabolism , Androgens/adverse effects , Animals , Aromatase Inhibitors/adverse effects , Estrogens/adverse effects , Female , Humans , Leptin/biosynthesis , Leptin/genetics , Luteinizing Hormone/biosynthesis , Mice , Mutation , Obesity/genetics , Obesity/metabolism , Plasminogen Activator Inhibitor 1/biosynthesis , Plasminogen Activator Inhibitor 1/genetics , Polycystic Ovary Syndrome/chemically induced , Polycystic Ovary Syndrome/genetics , Progestins/antagonists & inhibitors , Rats , Receptors, Leptin/biosynthesis , Receptors, Leptin/genetics
14.
Proc Natl Acad Sci U S A ; 107(52): 22629-34, 2010 Dec 28.
Article in English | MEDLINE | ID: mdl-21149714

ABSTRACT

Elevated follicle-stimulating hormone (FSH) activity is proposed to directly cause bone loss independent of estradiol deficiency in aging women. Using transgenic female mice expressing human FSH (TgFSH), we now reveal that TgFSH dose-dependently increased bone mass, markedly elevating tibial and vertebral trabecular bone volume. Furthermore, TgFSH stimulated a striking accrual of bone mass in hypogonadal mice lacking endogenous FSH and luteinizing hormone (LH) function, showing that FSH-induced bone mass occurred independently of background LH or estradiol levels. Higher TgFSH levels increased osteoblast surfaces in trabecular bone and stimulated de novo bone formation, filling marrow spaces with woven rather than lamellar bone, reflective of a strong anabolic stimulus. Trabecular bone volume correlated positively with ovarian-derived serum inhibin A or testosterone levels in TgFSH mice, and ovariectomy abolished TgFSH-induced bone formation, proving that FSH effects on bone require an ovary-dependent pathway. No detectable FSH receptor mRNA in mouse bone or cultured osteoblasts or osteoclasts indicated that FSH did not directly stimulate bone. Therefore, contrary to proposed FSH-induced bone loss, our findings demonstrate that FSH has dose-dependent anabolic effects on bone via an ovary-dependent mechanism, which is independent of LH activity, and does not involve direct FSH actions on bone cells.


Subject(s)
Bone Density/physiology , Bone and Bones/metabolism , Follicle Stimulating Hormone/physiology , Animals , Bone and Bones/cytology , Cell Line , Cells, Cultured , Estradiol/blood , Estradiol/metabolism , Female , Follicle Stimulating Hormone/blood , Follicle Stimulating Hormone/genetics , Humans , Inhibins/blood , Inhibins/metabolism , Luteinizing Hormone/blood , Male , Mice , Mice, Inbred C3H , Mice, Inbred C57BL , Mice, Transgenic , Osteoblasts/cytology , Osteoblasts/metabolism , Osteocalcin/genetics , Osteoclasts/cytology , Osteoclasts/metabolism , Ovariectomy , Ovary/metabolism , Peptide Fragments/blood , Procollagen/blood , Receptors, FSH/genetics , Reverse Transcriptase Polymerase Chain Reaction , Testosterone/blood , Testosterone/metabolism
15.
Endocrinology ; 151(6): 2800-10, 2010 Jun.
Article in English | MEDLINE | ID: mdl-20410197

ABSTRACT

Both testosterone and its nonaromatizable metabolite dihydrotestosterone (DHT) induce spermatogenesis in gonadotropin-deficient hpg mice. Surprisingly, because aromatization is not required, estradiol (E2) also induces spermatogenesis and increases circulating FSH in hpg mice, but the mechanism remains unclear. We studied E2-induced spermatogenesis in hpg mice on an estrogen receptor (ER)-alpha (hpg/alphaERKO) or ERbeta (hpg/betaERKO) knockout or wild-type ER (hpg/WT) background treated with subdermal E2 or DHT implants for 6 wk. In hpg/WT and hpg/betaERKO, but not hpg/alphaERKO mice, E2 increased testis and epididymal weight, whereas DHT-induced increases were unaffected by ERalpha or ERbeta inactivation. E2 but not DHT treatment increased serum FSH (but not LH) in hpg/WT and hpg/betaERKO but not hpg/alphaERKO hpg mice. DHT or E2 alone increased (premeiotic) spermatogonia and (meiotic) spermatocytes without significant change in Sertoli cell numbers. DHT alone increased postmeiotic spermatids, regardless of ER presence, compared with variable ERalpha-dependent E2 postmeiotic responses. An ERalpha-mediated effect was confirmed by treating hpg mice for 6 wk by subdermal selective ER-alpha (16alpha-LE(2)) or ERbeta (8beta-VE(2)) agonist implants. ERalpha (but not ERbeta) agonist increased testis and epididymal weight, Sertoli cell, spermatogonia, meiotic, and postmeiotic germ cell numbers. Only ERalpha agonist markedly increased serum FSH, whereas either agonist induced small rises in serum LH. Administration of ERalpha agonist or E2 in the presence of functional ERalpha induced prominent gene expression of specific Sertoli (Eppin, Rhox5) and Leydig cell (Cyp11a1, Hsd3b1) markers. We conclude that E2-induced spermatogenesis in hpg mice involves an ERalpha-dependent neuroendocrine mechanism increasing blood FSH and Sertoli cell function.


Subject(s)
Estradiol/pharmacology , Estrogen Receptor alpha/metabolism , Estrogen Receptor beta/metabolism , Follicle Stimulating Hormone/blood , Neurosecretory Systems/drug effects , Neurosecretory Systems/metabolism , Spermatogenesis/drug effects , Animals , Dihydrotestosterone/pharmacology , Estrogen Receptor alpha/agonists , Estrogen Receptor alpha/genetics , Estrogen Receptor beta/agonists , Estrogen Receptor beta/genetics , Female , Gonadotropins/deficiency , Gonadotropins/genetics , Leydig Cells/drug effects , Leydig Cells/metabolism , Male , Mice , Mice, Knockout , Organ Size/drug effects , Seminal Vesicles/drug effects , Seminal Vesicles/metabolism , Spermatogenesis/genetics , Testis/drug effects , Testis/metabolism
16.
Endocrinology ; 150(10): 4755-65, 2009 Oct.
Article in English | MEDLINE | ID: mdl-19574395

ABSTRACT

We examined the biological importance of Sertoli cell androgen receptor (AR) genomic interaction, using a Cre-loxP approach to selectively disrupt the AR DNA-binding domain (AR-DBD). Sertoli cell (SC)-specific transgenic Abpa or AMH promoters targeted Cre-mediated inframe excision of mouse Ar exon-3, encoding the AR-DBD second zinc-finger (ZF2), generating SC-specific mutant AR(DeltaZF2) lines designated Abp.SCAR(DeltaZF2) and AMH.SCAR(DeltaZF2), respectively. Both SCAR(DeltaZF2) lines produced infertile males exhibiting spermatogenic arrest, despite normal SC numbers and immunolocalized SC nuclear AR. Adult homozygous TgCre((+/+)) SCAR(DeltaZF2) or double-TgCre((+/-)) Abp/AMH.SCAR(DeltaZF2) males displayed equivalent small testes 30% of normal size, representing maximal Cre-loxP-disruption of Sertoli AR function. Hemizygous TgCre((+/-)) vs. homozygous TgCre((+/+)) Abp.SCAR(DeltaZF2) testes were larger (47% normal size) with more postmeiotic development, indicating dose-dependent Cre-mediated disruption of SC-specific AR-DBD activity. SCAR(DeltaZF2) males exhibited adult Leydig cell hypertrophy but normal serum testosterone levels. Sertoli cell-specific Rhox5 and Spinlw1 transcription, regulated by divergent or classical androgen-response elements, respectively, were both decreased in postnatal SCAR(DeltaZF2) vs. control testes, demonstrating SC-specific AR-DBD function as early as postnatal d 5. However, Rhox5 expression declined dose-dependently, whereas Spinlw1 expression increased, in adult TgCre((+/-)) and TgCre((+/+)) SCAR(DeltaZF2) testes, revealing differential temporal control for distinct AR-regulated transcripts. Androgen-repressed Ngfr was not up-regulated in SCAR(DeltaZF2) testes, suggesting maintenance of a nonclassical mechanism independent of AR-DBD. Thus, our unique SCAR(DeltaZF2) paradigm provided dose-dependent Cre-mediated disruption of testicular development and gene expression revealing that the AR-DBD is essential for SC function and postmeiotic spermatogenesis. Nongenomic or AR-DBD-independent pathways appear secondary or play no major independent role in SC function.


Subject(s)
Receptors, Androgen/metabolism , Sertoli Cells/metabolism , Spermatogenesis , Animals , Cell Adhesion , DNA/metabolism , DNA-Binding Proteins/metabolism , Gene Expression , Immunohistochemistry , Male , Mice , Mice, Transgenic , Organ Size , Phenotype , Protein Interaction Domains and Motifs , Testis/anatomy & histology , Transgenes
17.
Am J Physiol Endocrinol Metab ; 296(5): E1022-8, 2009 May.
Article in English | MEDLINE | ID: mdl-19293333

ABSTRACT

We have characterized the in vivo actions of human wild-type FSH receptor (FSHR) overexpressed in Sertoli cells of transgenic (Tg) mice (TgFSHRwt) compared with transgenic overexpression of the human activated mutant FSHR*D567G (TgFSHR*D567G). Testicular TgFSHRwt expression significantly elevated specific FSH binding (>2-fold, P < 0.01) relative to nontransgenic testes, similar to increased FSH binding in TgFSHR*D567G testes. Isolated TgFSHRwt Sertoli cells exhibited higher FSH-stimulated cAMP levels compared with non-Tg or TgFSHR*D567G cells but did not display the elevated FSH-independent basal cAMP levels found in TgFSHR*D567G Sertoli cells. Furthermore, Sertoli cell overexpression of TgFSHR*D567G but not TgFSHRwt allowed promiscuous cAMP responses to human chorionic gonadotropin (300 IU/ml) and TSH (30 mIU/ml), demonstrating increased constitutive signaling and altered glycoprotein hormone specificity via the intracellular D567G substitution rather than FSHR overexpression. Despite elevating Sertoli cell FSH sensitivity, overexpression of TgFSHRwt had no detectable effect upon normal testis function and did not stimulate Sertoli and germ cell development in testes of gonadotropin-deficient hypogonadal (hpg) mice, in contrast to the increased meiotic and postmeiotic germ cell development in TgFSHR*D567G hpg testes. Increased steroidogenic potential of TgFSHR*D567G hpg testes was demonstrated by elevated Cyp11a1 and Star expression, which was not detected in TgFSHRwt hpg testes. Androgen-regulated and Sertoli cell-specific Rhox5 gene expression was increased in TgFSHR*D567G but not TgFSHRwt hpg testes, providing evidence of elevated LH-independent androgen activity due to mutant FSHR*D567G. Hence, transgenic FSHR overexpression in Sertoli cells revealed that the D567G mutation confers autonomous signaling and steroidogenic activity in vivo as well as promiscuous glycoprotein hormone receptor activation, independently of FSHR overexpression alone.


Subject(s)
Receptors, FSH/genetics , Sertoli Cells/metabolism , Testis/metabolism , Animals , Cholesterol Side-Chain Cleavage Enzyme/metabolism , Cyclic AMP/metabolism , Follicle Stimulating Hormone/metabolism , Histocytochemistry , Humans , Male , Mice , Mice, Transgenic , Mutation , Organ Size/physiology , Phosphoproteins/metabolism , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , Receptors, FSH/biosynthesis , Receptors, FSH/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Spermatogenesis/physiology , Testis/cytology
18.
Reprod Fertil Dev ; 20(8): 861-70, 2008.
Article in English | MEDLINE | ID: mdl-19007549

ABSTRACT

Spermatogenesis requires androgen but, paradoxically, oestradiol (E2) treatment stimulates spermatogenic development in gonadotrophin- and androgen-deficient hypogonadal (hpg) mice. The mechanisms of E2-induced spermatogenesis were investigated by determining intratesticular E2 levels and testis cell populations in E2-treated hpg male mice, and E2 spermatogenic actions were determined in androgen receptor-knockout (ARKO) mice. Despite increased serum E2 concentrations (150-300 pmol L(-1)), intratesticular E2 concentrations declined fivefold (P < 0.001) in E2-treated v. untreated hpg male mice. Serum FSH reached 40% of normal and total testicular numbers of known FSH-responsive Sertoli, spermatogonia and meiotic spermatocyte populations were significantly (P < 0.001) elevated 1.7-, 4- and 13-fold, respectively. However, E2 administration also increased androgen-dependent pachytene spermatocytes and post-meiotic spermatids to levels comparable with testosterone-treated hpg testes. Selective investigation of androgen receptor involvement used E2-treated ARKO mice, which were found to exhibit increased (1.6-fold; P < 0.05) intratesticular E2 concentrations and suppression of the elevated serum gonadotrophins, although FSH remained twofold higher than normal. However, testis size and total Sertoli, spermatogonia and spermatocyte numbers were not increased in E2-treated ARKO male mice. Therefore, E2-stimulated murine spermatogenic development occurs with markedly suppressed and not elevated intratesticular E2 levels and displays an absolute requirement for functional androgen receptors. We propose that this paradoxical E2 spermatogenic response is explained by predominantly extratesticular E2 actions, increasing FSH to combine with residual androgen activity in hpg testes to stimulate pre- to post-meiotic development.


Subject(s)
Estradiol/pharmacology , Receptors, Androgen/physiology , Spermatogenesis/drug effects , Testis/drug effects , Animals , Estradiol/metabolism , Follicle Stimulating Hormone/blood , Hypogonadism/physiopathology , Male , Mice , Mice, Knockout , Organ Size/drug effects , Receptors, Androgen/genetics , Sertoli Cells/cytology , Sertoli Cells/drug effects , Signal Transduction/physiology , Spermatogenesis/physiology , Spermatozoa/cytology , Spermatozoa/drug effects , Testis/cytology , Testis/metabolism , Testosterone/metabolism
19.
Endocrinology ; 148(9): 4432-9, 2007 Sep.
Article in English | MEDLINE | ID: mdl-17540727

ABSTRACT

Rising serum FSH levels is one of the earliest signs of human female reproductive aging. Whether or not elevated FSH remains a passive reflection of a diminishing ovarian follicle pool or actively contributes to declining female fertility with age has not been established. We therefore investigated female reproduction in mice expressing progressively rising serum levels of transgenic human FSH (Tg-FSH, 2.5-10 IU/liter) independently of follicle depletion. We show that serum LH and estradiol levels and uterine size remained normal in Tg-FSH females, whereas ovarian weight and corpora lutea number were significantly increased up to 1.3- and 5-fold, respectively. Furthermore, the monotrophic FSH rise produced a striking biphasic effect on female fertility. Tg-FSH females less than 22 wk old delivered increased litter sizes, then beyond 23 wk, litter sizes decreased rapidly culminating in premature infertility despite continued ovary follicle development, and increased ovulation and uterine embryo implantation sites as well as normal serum levels of anti-Mullerian hormone, a marker of ovarian follicle reserve. We found that rising circulating Tg-FSH produced premature infertility by increasing embryo-fetal resorption and parturition failure with age. Thus, our Tg-FSH mice present a novel paradigm to investigate selective contributions of elevated FSH to age-related female infertility, which revealed that rising FSH levels, despite no exhaustion of ovarian reserve, actively accelerates female reproductive aging primarily by postimplantation reduction of embryo-fetal survival.


Subject(s)
Aging/physiology , Fertility/physiology , Follicle Stimulating Hormone/physiology , Reproduction/physiology , Animals , Female , Follicle Stimulating Hormone/blood , Humans , Insulin/genetics , Luteinizing Hormone/blood , Mice , Mice, Transgenic , Ovarian Follicle/drug effects , Ovarian Follicle/physiology , Ovary/physiology , Polymerase Chain Reaction , Promoter Regions, Genetic , Rats
20.
Endocrinology ; 148(5): 2264-72, 2007 May.
Article in English | MEDLINE | ID: mdl-17317769

ABSTRACT

Prostate development and maturation requires stromal-epithelial interactions and androgen action via the androgen receptor (AR) within these compartments. However, the specific roles of epithelial and stromal AR in postnatal prostate differentiation are unclear. We used Cre-LoxP technology to determine the prostate phenotype in mice with epithelial-selective genetic inactivation of the AR leaving the stromal AR functionally intact. We find that prostate development abolished in mice globally lacking a functional AR can be rescued by restricting the AR knockout to the postnatal prostate epithelium. We show that, at 8 wk of age, prostate epithelial AR knockout (PEARKO) mice exhibit prostate development with normal branching morphogenesis but lobe-specific decrease in prostate weight and hindered structural and functional differentiation of the mature prostate epithelium. No change was observed in PEARKO testis weight or serum testosterone compared with littermate controls. The most striking change was increased proliferation and abnormal lesions of epithelial cells predominantly in the anterior lobe of PEARKO mice. These findings highlight the vital role of stromal AR in postnatal prostate growth and structural differentiation and emphasize the requirement of epithelial AR in maintaining functional differentiation and restraining proliferation of epithelial cells in a lobe-specific manner. This unique PEARKO mouse provides a new paradigm with which to define the molecular mechanisms of the androgen signaling in mature prostate lobes in vivo and provides insight into the identification of better targets for treatment of prostate cancer and hyperplasia.


Subject(s)
Epithelial Cells/pathology , Prostate/growth & development , Prostate/pathology , Receptors, Androgen/genetics , Receptors, Androgen/metabolism , Animals , Cell Differentiation/physiology , Cell Division/physiology , Epithelial Cells/physiology , Gene Expression Regulation, Developmental/physiology , Integrases/genetics , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Organ Size , Phenotype , Prostate/physiology , RNA, Messenger/metabolism , Stromal Cells/metabolism , Stromal Cells/physiology , Transgenes
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