bluuebunny/crossref_metadata_2025_split huggingface dataset:
Direct Action through the Sertoli Cells Is Essential for Androgen Stimulation of Spermatogenesis
Authors: O'Shaughnessy P. J. , Verhoeven G. , De Gendt K. , Monteiro A. , Abel M. H. 
Published: January 2014

ABSTRACT
Androgens act to stimulate spermatogenesis through androgen receptors (ARs) on the Sertoli cells and peritubular myoid cells. Specific ablation of the AR in either cell type will cause a severe disruption of spermatogenesis. To determine whether androgens can stimulate spermatogenesis through direct action on the peritubular myoid cells alone or whether action on the Sertoli cells is essential, we crossed hypogonadal (hpg) mice that lack gonadotrophins and intratesticular androgen with mice lacking ARs either ubiquitously (ARKO) or specifically on the Sertoli cells (SCARKO). These hpg.ARKO and hpg.SCARKO mice were treated with testosterone (T) or dihydrotestosterone (DHT) for 7 d and testicular morphology and cell numbers assessed. Androgen treatment did not affect Sertoli cell numbers in any animal group. Both T and DHT increased numbers of spermatogonia and spermatocytes in hpg mice, but DHT has no effect on germ cell numbers in hpg.SCARKO and hpg.ARKO mice. T increased germ cell numbers in hpg.SCARKO and hpg.ARKO mice, but this was associated with stimulation of FSH release. Results show that androgen stimulation of spermatogenesis requires direct androgen action on the Sertoli cells.
Identification of Adipocyte Genes Regulated by Caloric Intake
Authors: Franck Niclas , Gummesson Anders , Jernås Margareta , Glad Camilla , Svensson Per-Arne , Guillot Gilles , Rudemo Mats , Nyström Fredrik H , Carlsson Lena M. S. , Olsson Bob 
Published: January 2014

Context
Changes in energy intake have marked and rapid effects on metabolic functions, and some of these effects may be due to changes in adipocyte gene expression that precede alterations in body weight.

Objective
The aim of the study was to identify adipocyte genes regulated by changes in caloric intake independent of alterations in body weight.

Research Design and Methods
Obese subjects given a very low-caloric diet followed by gradual reintroduction of ordinary food and healthy subjects subjected to overfeeding were investigated. Adipose tissue biopsies were taken at multiple time-points, and gene expression was measured by DNA microarray. Genes regulated in the obese subjects undergoing caloric restriction followed by refeeding were identified using two-way ANOVA corrected with Bonferroni. From these, genes regulated by caloric restriction and oppositely during the weight-stable refeeding phase were identified in the obese subjects. The genes that were also regulated, in the same direction as the refeeding phase, in the healthy subjects after overfeeding were defined as being regulated by caloric intake. Results were confirmed using real-time PCR or immunoassay.

Results
Using a significance level of P &lt; 0.05 for all comparisons, 52 genes were down-regulated, and 50 were up-regulated by caloric restriction and regulated in the opposite direction by refeeding and overfeeding. Among these were genes involved in lipogenesis (ACLY, ACACA, FASN, SCD), control of protein synthesis (4EBP1, 4EBP2), β-oxidation (CPT1B), and insulin resistance (PEDF, SPARC).

Conclusions
Metabolic genes involved in lipogenesis, protein synthesis, and insulin resistance are central in the transcriptional response of adipocytes to changes in caloric intake.
Altered Autophagy in Human Adipose Tissues in Obesity
Authors: Kovsan Julia , Blüher Matthias , Tarnovscki Tanya , Klöting Nora , Kirshtein Boris , Madar Liron , Shai Iris , Golan Rachel , Harman-Boehm Ilana , Schön Michael R. , Greenberg Andrew S. , Elazar Zvulun , Bashan Nava , Rudich Assaf 
Published: January 2014

Context
Autophagy is a housekeeping mechanism, involved in metabolic regulation and stress response, shown recently to regulate lipid droplets biogenesis/breakdown and adipose tissue phenotype.

Objective
We hypothesized that in human obesity autophagy may be altered in adipose tissue in a fat depot and distribution-dependent manner.

Setting and Patients
Paired omental (Om) and sc adipose tissue samples were used from obese and nonobese (n = 65, cohort 1); lean, sc-obese and intraabdominally obese (n = 196, cohort 2); severely obese persons without diabetes or obesity-associated morbidity, matched for being insulin sensitive or resistant (n = 60, cohort 3).

Results
Protein and mRNA levels of the autophagy genes Atg5, LC3A, and LC3B were increased in Om compared with sc, more pronounced among obese persons, particularly if with intraabdominal fat accumulation. Both adipocytes and stromal-vascular cells contribute to the expression of autophagy genes. The increased number of autophagosomes and elevated autophagic flux assessed in fat explants incubated with lysosomal inhibitors were observed in obesity, particularly in Om. The degree of visceral adiposity and adipocyte hypertrophy accounted for approximately 50% of the variance in Atg5 mRNA levels by multivariate regression analysis, whereas age, sex, measures of insulin sensitivity, inflammation, and adipose tissue stress were excluded from the model. Moreover, in cohort 3, the autophagy marker genes were increased in those who were insulin resistant compared with insulin sensitive, particularly in Om.

Conclusions
Autophagy is up-regulated in adipose tissue of obese persons, especially in Om, correlating with the degree of obesity, visceral fat distribution, and adipocyte hypertrophy. This may precede the occurrence of obesity-associated morbidity.
Expanding the Phenotype and Genotype of Female GnRH Deficiency
Authors: Shaw Natalie D. , Seminara Stephanie B. , Welt Corrine K. , Au Margaret G. , Plummer Lacey , Hughes Virginia A. , Dwyer Andrew A. , Martin Kathryn A. , Quinton Richard , Mericq Veronica , Merino Paulina M. , Gusella James F. , Crowley William F. , Pitteloud Nelly , Hall Janet E. 
Published: January 2014

Abstract

Context:
GnRH deficiency is a rare genetic disorder of absent or partial pubertal development. The clinical and genetic characteristics of GnRH-deficient women have not been well-described.

Objective:
To determine the phenotypic and genotypic spectrum of a large series of GnRH-deficient women.
Design, Setting, and Subjects: Retrospective study of 248 females with GnRH deficiency evaluated at an academic medical center between 1980 and 2010.

Main Outcome Measures:
Clinical presentation, baseline endogenous GnRH secretory activity, and DNA sequence variants in 11 genes associated with GnRH deficiency.

Results:
Eighty-eight percent had undergone pubarche, 51% had spontaneous thelarche, and 10% had 1-2 menses. Women with spontaneous thelarche were more likely to demonstrate normal pubarche (P = 0.04). In 27% of women, neuroendocrine studies demonstrated evidence of some endogenous GnRH secretory activity. Thirty-six percent (a large excess relative to controls) harbored a rare sequence variant in a gene associated with GnRH deficiency (87% heterozygous and 13% biallelic), with variants in FGFR1 (15%), GNRHR (6.6%), and PROKR2 (6.6%) being most prevalent. One woman had a biallelic variant in the X-linked gene, KAL1, and nine women had heterozygous variants.

Conclusions:
The clinical presentation of female GnRH deficiency varies from primary amenorrhea and absence of any secondary sexual characteristics to spontaneous breast development and occasional menses. In this cohort, rare sequence variants were present in all of the known genes associated with GnRH deficiency, including the novel identification of GnRH-deficient women with KAL1 variants. The pathogenic mechanism through which KAL1 variants disrupt female reproductive development requires further investigation.
Induction of Carbonic Anhydrase II Expression in Osteoclast Progenitors Requires Physical Contact with Stromal Cells*
Authors: Biskobing Diane M. , Fan Dongjie , Fan Xian , Rubin Janet 
Published: January 2014

Abstract
Carbonic anhydrase II (CA II) expression is vital to normal osteoclast function. We and others have previously reported induction of CA II messenger RNA (mRNA) expression by 1,25(OH)2D3 in myelomonocytic cells and marrow culture. However, since 1,25(OH)2D3 stimulates osteoclast differentiation as well, we wished to separate direct effects of 1,25(OH)2D3 on the CA II gene from the differentiating effects of the hormone. Using primary murine mixed marrow cultures, we measured CA II mRNA expression by RT-PCR. 10 nm 1,25(OH)2D3 dose dependently induced expression of CA II mRNA (4.12 ± 0.68-fold) at day 4 in culture compared with control with an ED50 of 0.25 nm. When nonadherent marrow cells containing osteoclast progenitors were depleted of stromal cells and exposed to 10 nm 1,25(OH)2D3, CA II mRNA expression was decreased by more than 60%. Coculture of progenitors with ST-2 stromal cells for 3 days with 10 nm 1,25(OH)2D3 stimulated CA II expression by 22 ± 3.6-fold. 1,25(OH)2D3 stimulated CA II mRNA expression in progenitors separated from ST-2 cells by transwells was insignificant demonstrating that the two cell types must be in physical contact. PTH also stimulated CA II mRNA expression (4.91 ± 0.01-fold) to a similar degree as seen with 1,25(OH)2D3 treatment. These results demonstrate that induction of CA II in osteoclast progenitors requires their physical communication with stromal cells and is inseparable from the osteoclast differentiation process.
A Role for Phospholipase C Activity in GLUT4-Mediated Glucose Transport*
Authors: Van Epps-Fung Mark , Gupta Kiran , Hardy Robert W. , Wells Alan 
Published: January 2014

AbstractOverexpression of surrogate receptors [epidermal growth factor (EGF) receptor (EGFR) and platelet-derived growth factor receptor] in adipocytes has demonstrated that multiple signaling pathways may lead to GLUT4-mediated glucose uptake. These implicated pathways function independently of IRS-1 phosphorylation and PI3-kinase activation. In addition, we previously demonstrated that EGFR tyrosyl autophosphorylation is required to stimulate GLUT4-mediated glucose transport in 3T3-L1 adipocytes. This observation suggests that signaling molecules that are dependent on EGFR auto-phosphorylation, such as phospholipase C (PLC), may lie in the signaling pathway to glucose transport. As PLC has been implicated in glucose transport by several clinical and basic mechanistic studies, we investigated whether EGFR signaling may promote glucose transport via modulation of PLC activity. Activation of EGFR overexpressing 3T3-L1 adipocytes leads to a 3.4 ± 1.2-fold stimulation of PLC activity over basal levels vs. only 1.06 ± 0.01-fold stimulation by insulin. Pharmacological inhibition of PLC by 50 μm U73122 reduced phosphoinositide accumulation by 79.2 ± 16.9% and resulted in a concomitant 56.0 ± 12.7% decrease in EGF-induced glucose transport. This inhibition of glucose transport by U73122 was specific, because the inactive congener, U73343, failed to block EGF-induced glucose transport. Despite the low levels of insulin-induced PLC activity, insulin-stimulated glucose transport activity was similarly inhibited by U73122 (55.9 ± 13.1% inhibition). Inhibition of PLC activation did not impair either EGF- or insulin-induced activation of glycogen synthase or incorporation of glucose into lipid, supporting the hypothesis that both EGF- and insulin-induced glucose disposal can be independent of GLUT4-mediated glucose transport. The diminution of glucose transport secondary to inhibition of PLC activity was reflected by a decrease in GLUT4 translocation to the plasma membrane upon either EGF or insulin stimulation. These results are consistent with either a permissive or an active role for PLC activity in the translocation of GLUT4 to the plasma membrane.
Insulin-Like Growth Factor I (IGF-I) Receptor Overexpression Abolishes the IGF Requirement for Differentiation and Induces a Ligand-Dependent Transformed Phenotype in C2 Inducible Myoblasts*
Authors: Navarro Magali , Barenton Bruno , Garandel Veronique , Schnekenburger Juergen , Bernardi Henri 
Published: January 2014

AbstractInsulin-like growth factors (IGFs) stimulate both proliferation and differentiation of myogenic cell lines, and these actions are mostly mediated through the type I IGF receptor (type I IGF-R). To further investigate the role of this receptor in phenotypic characteristics of C2 murine myoblasts, we overexpressed the human type I IGF-R in the inducible clone of C2 cells, which requires IGFs in the differentiation medium to undergo terminal differentiation.Inducible myoblasts were transfected with either the eukaryotic expression vector pNTK or pNTK containing the human type I IGF-R complementary DNA, and we isolated two clones named Ind-Neo and Ind-R, respectively. Binding and autophosphorylation experiments indicate that Ind-R cells express about 10 times as much type I IGF-R compared with Ind-Neo control cells and that the transfected type I IGF-R is functional in Ind-R cells.We show that overexpression of the human type I IGF-R makes inducible myoblasts able to differentiate spontaneously, as assessed by expression of the myogenic transcription factors MyoD and myogenin, detection of the muscle-specific protein troponin T, and myotube formation. Moreover, when exposed to IGF-I, Ind-R cells lose contact inhibition, grow in the presence of a low level of growth factors and form colonies in soft agar, which is characteristic of a ligand-dependent transformed phenotype.It emerges from this study that 1) the type I IGF-R is strongly involved in the phenotypic differences between inducible and permissive cells with respect to the differentiation program; and 2) overexpression causes this receptor to act as a ligand-dependent transforming protein in muscle cells. We suggest that type I IGF-R abundance and level of activation may determine the efficiency of the autocrine mode of action of IGFs and discriminate their biological functions.
The Type 2 and Type 3 Iodothyronine Deiodinases Play Important Roles in Coordinating Development in Rana catesbeiana Tadpoles*
Authors: Becker Kathryn B. , Stephens Kristen C. , Davey Jennifer C. , Schneider Mark J. , Galton Valerie Anne 
Published: January 2014

AbstractIn developing Rana catesbeiana tadpoles, the timing of the thyroid hormone (TH)-dependent metamorphic responses varies markedly among tissues. Yet at any one time these tissues are exposed to the same plasma concentration of TH, suggesting that TH action is regulated in part at the level of the peripheral tissues. A major factor in TH action is the intracellular level of the active TH, T3. This level is dependent not only on the plasma concentration of TH (mostly T4) but also on the intracellular activities of the type 2 5′-deiodinase (D2) and the type 3 5-deiodinase (D3), which are responsible, respectively, for generating and degrading T3. (D1 is not present in this species.) To determine whether differential expression of D2 and D3 among tissues could be a significant factor in the coordination of metamorphic events, the ontogenic profiles of the two enzyme activities and corresponding messenger RNA levels in most tissues of R. catesbeiana tadpoles have been documented. The profiles of D2 expression in tail, hindlimb, forelimb, intestine, skin, and eye differed markedly at both activity and messenger RNA levels, but it was notable that expression was invariably highest in a given tissue at the time of its major metamorphic change. D2 expression was very low in brain and heart and did not vary during development. D2 was not expressed in liver, kidney, or red blood cells. With the exception of red blood cells, D3 expression was detected in all tissues studied. Furthermore, it was evident that in tissues that expressed both deiodinase genes, the two expression profiles were comparable, indicating a potential for tight control of intracellular T3 levels.Direct evidence of the importance of the intracellular conversion of T4 to T3 for TH-dependent metamorphic events was obtained in tadpoles in which endogenous TH synthesis was blocked with methimazole, and the activities of D2 and D3 were inhibited by iopanoic acid. This treatment inhibited metamorphosis. The inhibition could be overcome by the concomitant administration of replacement levels of T3, but not T4.These results strongly support the view that coordinated development in amphibia depends in part on the tissue-specific expression patterns of the D2 and D3 genes, which ensure that the requisite level of intracellular T3 is attained in a given tissue, regardless of the current level of circulating TH, at the appropriate stage of metamorphosis.
Immunization of Male Bonnet Monkeys (M. radiata) with a Recombinant FSH Receptor Preparation Affects Testicular Function and Fertility
Authors: Moudgal N. R. , Sairam M. R. , Krishnamurthy H. N. , Sridhar Surekha , Krishnamurthy H. , Khan H. 
Published: January 2014

Abstract
Immunization of proven fertile adult male monkeys (n = 3) with a recombinant FSH receptor protein preparation (oFSHR-P) (representing amino acids 1–134 of the extracellular domain of the receptor Mr∼ 15KDa) resulted in production of receptor blocking antibodies. The ability of the antibody to bind a particulate FSH receptor preparation and receptors in intact granulosa cells was markedly (by 30–80%) inhibited by FSH. Serum T levels and LH receptor function following immunization remained unchanged. The immunized monkeys showed a 50% reduction (p&lt; 0.001) in transformation of spermatogonia (2C) to primary spermatocytes (4C) as determined by flow cytometry and the 4C:2C ratio showed a correlative change (R 0.81, p&lt; 0.0007) with reduction in fertility index (sperm counts X motility score). Breeding studies indicated that monkeys became infertile between 242–368 days of immunization when the fertility index was in the range of 123 ± 76 to 354 ± 42 (compared to a value of 1602 ± 384 on day 0). As the effects observed are near identical to that seen following immunization with FSH it is suggestive that oFSHR-P can substitute for FSH in the development of a contraceptive vaccine.
Ligand-Independent GLUT4 Translocation Induced by Guanosine 5′-O-(3-Thiotriphosphate) Involves Tyrosine Phosphorylation*
Authors: Haruta Tetsuro , Morris Aaron J. , Vollenweider Peter , Nelson James G. , Rose David W. , Mueckler Michael , Olefsky Jerrold M. 
Published: January 2014

AbstractTo delineate the signaling pathway leading to glucose transport protein (GLUT4) translocation, we examined the effect of microinjection of the nonhydrolyzable GTP analog, guanosine 5′-O-(3-thiotriphosphate) (GTPγS), into 3T3-L1 adipocytes. Thirty minutes after the injection of 5 mm GTPγS, 40% of injected cells displayed surface GLUT4 staining indicative of GLUT4 translocation compared with 55% for insulin-treated cells and 10% in control IgG-injected cells. Treatment of the cells with the phosphatidylinositol 3-kinase inhibitor wortmannin or coinjection of GST-p85 SH2 fusion protein had no effect on GTPγS-mediated GLUT4 translocation. On the other hand, coinjection of antiphosphotyrosine antibodies (PY20) blocked GTPγS-induced GLUT4 translocation by 65%. Furthermore, microinjection of GTPγS led to the appearance of tyrosine-phosphorylated proteins around the periphery of the plasma membrane, as observed by immunostaining with PY20. Treatment of the cells with insulin caused a similar phosphotyrosine-staining pattern. Electroporation of GTPγS stimulated 2-deoxy-d-glucose transport to 70% of the extent of insulin stimulation. In addition, immunoblotting with phosphotyrosine antibodies after electroporation of GTPγS revealed increased tyrosine phosphorylation of several proteins, including 70- to 80-kDa and 120- to 130-kDa species. These results suggest that GTPγS acts upon a signaling pathway either downstream of or parallel to activation of phosphatidylinositol 3-kinase and that this pathway involves tyrosine-phosphorylated protein(s).
Vasoactive Intestinal Peptide Is an Important Endocrine Regulatory Factor of Fetal Rat Testicular Steroidogenesis*
Authors: El-Gehani Faraj , Tena-Sempere Manuel , Huhtaniemi Ilpo 
Published: January 2014

AbstractThis study elaborates our recent preliminary finding that vasoactive intestinal peptide (VIP) has a specific stimulatory effect on fetal rat Leydig cells. We examined the dose-response relationship for the effect of VIP on cAMP and testosterone production by dispersed fetal Leydig cells isolated from rat testes on embryonic day (E) 18.5. Further, we used RT-PCR to examine the expression of the VIP gene in fetal brain and testes and that of the VIP receptor genes in fetal testes and used RIA to measure VIP in testes and plasma during the fetal period. VIP stimulated fetal testicular cAMP production at a dose of 10−9 mol/liter, whereas a dose as low as 10−12 mol/liter stimulated testosterone production. This suggests that VIP at low doses may stimulate testosterone production using second messenger pathways other than cAMP. RT-PCR analysis could not reveal either VIP messenger RNA (mRNA) in fetal tissues or VIP1 receptor mRNA in the fetal or newborn testes, whereas VIP2 receptor mRNA was detected in fetal testes as early as E15.5. Northern hybridization analysis showed that the level of expression of VIP2 receptor mRNA is very low in fetal and neonatal testes and increases with age. The testicular VIP content was unmeasurable by our RIA method (i.e. &lt;1 fmol/testis), whereas the circulating level of VIP was 82.9 ± 1.1 pmol/liter on E17.5 and decreased with advancing fetal age.In conclusion, our results suggest that VIP from an extratesticular source, possibly from the maternal compartment, may regulate fetal testicular steroidogenesis through type 2 receptors as early as E15.5. These findings may be of physiological significance, because the onset of fetal testicular steroidogenesis occurs at an age (E15.5–19.5) before the onset of pituitary LH secretion.
Clathrin Gene Expression Is Androgen Regulated in the Prostate*
Authors: Prescott James L. , Tindall Donald J. 
Published: January 2014

AbstractAndrogens are required for the development and function of the prostate. In a normal human prostate, androgens control the synthesis of proteins such as prostate-specific antigen and human glandular kallikrein. The prostate secretes these proteins as well as a number of other compounds to form the prostatic fluid. Using differential display PCR to detect novel androgen-regulated genes, clathrin heavy chain expression was identified as potentially being up-regulated by androgens in the prostate cancer cell line LNCaP. We report here that the clathrin heavy chain and light chain genes are regulated by androgens. Clathrin heavy chain messenger RNA was up-regulated by androgens in a concentration- and time-specific manner in the LNCaP cell line. Translation of clathrin heavy chain messenger RNA was stimulated by androgens. Steady state levels of clathrin light chains a and b were up-regulated in the presence of androgen in LNCaP cells. Clathrin gene expression was examined in normal rat prostates, and similar results were found. Clathrin heavy chain protein levels in the rat prostate are also affected by the androgen status of the animal. We hypothesize that clathrin may be involved in the exocytosis of androgen-regulated secretory proteins such as prostate-specific antigen and human glandular kallikrein.