scholarly journals Divergent expression patterns of pituitary gonadotropin subunit and GnRH receptor genes to continuous GnRH in vitro and in vivo

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Marija M. Janjic ◽  
Rafael M. Prévide ◽  
Patrick A. Fletcher ◽  
Arthur Sherman ◽  
Kosara Smiljanic ◽  
...  
Keyword(s):  
Expression Profiles ◽  
Expression Patterns ◽  
Reproductive Function ◽  
Gnrh Receptor ◽  
Continuous Treatment ◽  
Pituitary Cells ◽  
Cell Content ◽  
Major Factors

AbstractContinuous, as opposed to pulsatile, delivery of hypothalamic gonadotropin-releasing hormone (GnRH) leads to a marked decrease in secretion of pituitary gonadotropins LH and FSH and impairment of reproductive function. Here we studied the expression profile of gonadotropin subunit and GnRH receptor genes in rat pituitary in vitro and in vivo to clarify their expression profiles in the absence and continuous presence of GnRH. Culturing of pituitary cells in GnRH-free conditions downregulated Fshb, Cga, and Gnrhr expression, whereas continuous treatment with GnRH agonists upregulated Cga expression progressively and Gnrhr and Fshb expression transiently, accompanied by a prolonged blockade of Fshb but not Gnrhr expression. In contrast, Lhb expression was relatively insensitive to loss of endogenous GnRH and continuous treatment with GnRH, probably reflecting the status of Egr1 and Nr5a1 expression. Similar patterns of responses were observed in vivo after administration of a GnRH agonist. However, continuous treatment with GnRH stimulated LH secretion in vitro and in vivo, leading to decrease in LH cell content despite high basal Lhb expression. These data suggest that blockade of Fshb expression and depletion of the LH secretory pool are two major factors accounting for weakening of the gonadotroph secretory function during continuous GnRH treatment.

Chronic Lymphocytic Leukemia-Associated Macrophages: Not Just Nurse Cells

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 46-46
Author(s):  
Loic Ysebaert ◽  
Mary Poupot ◽  
Yovan Sanchez-Ruiz ◽  
Camille Laurent ◽  
Guy Laurent ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Conditioned Medium ◽  
Apoptotic Cell ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Lymphocytic Leukemia ◽  
Functional Interpretation

Abstract Abstract 46 Introduction: CLL cells interact with many accessory cells in an environment mimicking that of normal mature B cells. Role of antigen, cytokines, adhesion pathways are critical for many aspects in the disease course (proliferation/survival, migration or homing, drug resistance, and presumably relapse). Nurse-like cells (NLC) belong to a monocytic-derived, bystander population among CLL lymph node and spleen stromal cells. Aim: To investigate the nature, functions, and location of NLC within CLL microenvironment. Methods: Gene expression profiles (GEP) from in vitro expanded NLC from patients (n=10) were produced and compared to those from normal CD14+ monocytes, M1-polarized macrophages, M2-polarized macrophages and tumor-associated macrophages (produced in the lab or downloaded from GEO datasets). Principal Component Analysis was used to categorize these five populations of cells and in-house-built GSEA software was used for functional interpretation of their relevant gene lists. Protein expression patterns were validated with multi-analyte ELISArray kits, proteome profiler arrays, flow cytometry (FC) or immunohistochemistry (IHC). Results: New insights into the physiopathological role of NLC in CLL are suggested from five lines of evidence: 1/a Òmonocytic gene signatureÓ (i.e. a set of 549 genes) is shared by the NLC and the monocyte subtypes. The genes over-represented in NLC vs normal monocytes pinpointed positive modulation of apoptotic cell clearance (scavenger, mannose and complement receptors, LXRalpha), lipid metabolism (Apolipoprotein E, PPAR signaling), extracellular matrix-receptor interactions (integrins, SPARC, Matrix MetalloProteinases) and actin cytoskeleton remodeling. 2/unsupervised clustering show that NLC represent an M2-skewed, TAM-like cell population. They down-regulate mRNA and proteins for classic M1 inflammatory markers (e.g. IL-1, IL-6, IL-12, COX2) while increase secretion of TGFbeta, IL-10, CCL17 and CCL22 soluble factors. 3/these and previously published observations suggest that B-CLL-to-NLC interactions may orchestrate immunosuppression in this disease. PBMCs from Òwatch and waitÓ CLL patients (all stage A/Rai 0, mutated IgVH, low risk cytogenetics profile) or healthy donors were stimulated with anti-CD3/CD28 beads + IL-2, either in standard RPMI+10% FCS or in conditioned medium (CM, after 14d CLL-NLC co-culture in vitro) and their proliferation/phenotype were compared after 2 weeks. Significant expansion of T cells with Treg (CD4+CD25+FoxP3+) phenotype was observed only from CLL PBMCs grown in conditioned medium (mean % Treg: 2.85 vs 3.05 in CM for normal PBMCs, and 1.54 vs 15.9 in CM for CLL PBMCs, P< 0.05). 4/although NLC make immune synapses with live B-CLL, they do not phagocytose them. Over-expression of CD47 (ÒdonÕt eat meÓ signal) by B-CLL cells (mfi= 3490 vs 2581 on normal cells, P< 0.05, n=18) may provide them with a protective signal against NLC. 5/from our GEP, flow cytometric and IHC analyses, we propose CD163 (classic M2 marker) as a reliable tool to identify NLC in vivo. Although in vitro, CLL cells can pervert healthy donor monocytes into NLC, only CLL-derived NLC are truly CD14+ CD163+. In vivo, CD163 staining reveals putative NLC in CLL lymph nodes(LN)/spleen sections but not in bone marrow. In LN from all patients, NLC reside in the subcapsular areas and line vessel structures, suggesting a role in CLL cells trafficking. Most interestingly, NLC infiltrate pseudofollicles structures only in a subset of cases. We will present updated IHC and clinical presentation correlation studies. Conclusions: Our results suggest that the role of NLC in CLL might be broader than initially thought. Beside of nursing and conferring drug resistance, NLC may also be crucial in the setting of immunosuppression, of CLL cells recruitment, and should thus be considered as therapeutic targets. Disclosures: Off Label Use: GA101 is not currently approved for CLL treatment.

scholarly journals Phosphorylation and Stabilization of HURP by Aurora-A: Implication of HURP as a Transforming Target of Aurora-A

2005 ◽  
Vol 25 (14) ◽  
pp. 5789-5800 ◽  
Author(s):  
Chang-Tze Ricky Yu ◽  
Jung-Mao Hsu ◽  
Yuan-Chii Gladys Lee ◽  
Ann-Ping Tsou ◽  
Chen-Kung Chou ◽  
...  
Keyword(s):  
Expression Profiles ◽  
Expression Patterns ◽  
Ectopic Expression ◽  
Gene Expression Profiles ◽  
Aurora A ◽  
Initial Attempt ◽  
Stable Transfectants ◽  
Low Serum

ABSTRACT Aurora-A, a mitotic serine/threonine kinase with oncogene characteristics, has recently drawn intense attention because of its association with the development of human cancers and its relationship with mitotic progression. Using the gene expression profiles of Aurora-A as a template to search for and compare transcriptome expression profiles in publicly accessible microarray data sets, we identified HURP (encodes hepatoma upregulated protein) as one of the best Aurora-A-correlated genes. Empirical validation indicates that HURP has several characteristics in common with Aurora-A. These two genes have similar expression patterns in hepatocellular carcinoma, liver regeneration after partial hepatectomy, and cell cycle progression and across a variety of tissues and cell lines. Moreover, Aurora-A phosphorylated HURP in vitro and in vivo. Ectopic expression of either the catalytically inactive form of Aurora-A or the HURP-4P mutant, in which the Aurora-A phosphorylation sites were replaced with Ala, resulted in HURP instability and complex disassembly. In addition, HURP-wild-type stable transfectants were capable of growing in low-serum environments whereas HURP-4P grew poorly under low-serum conditions and failed to proliferate. These studies together support the view that the ability to integrate evidence derived from microarray studies into biochemical analyses may ultimately augment our predictive power when analyzing the potential role of poorly characterized proteins. While this combined approach was simply an initial attempt to answer a range of complex biological questions, our findings do suggest that HURP is a potential oncogenic target of Aurora-A.

scholarly journals Developmental and molecular correlates of bovine preimplantation embryos

Reproduction ◽  
2006 ◽  
Vol 131 (5) ◽  
pp. 895-904 ◽  
Author(s):  
Hakan Sagirkaya ◽  
Muge Misirlioglu ◽  
Abdullah Kaya ◽  
Neal L First ◽  
John J Parrish ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methyltransferase ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Blastocyst Stage ◽  
Culture Conditions ◽  
Preimplantation Embryos ◽  
Developmental Potential

Expression of embryonic genes is altered in different culture conditions, which influence developmental potential both during preimplantation and fetal development. The objective of this study was to define the effects of culture conditions on: bovine embryonic development to blastocyst stage, blastocyst cell number, apoptosis and expression patterns of a panel of developmentally important genes. Bovine embryos were culturedin vitroin three culture media containing amino acids, namely potassium simplex optimization medium (KSOMaa), Charles Rosenkrans 1 (CR1aa) and synthetic oviductal fluid (SOFaa). Apoptosis in blastocysts was determined by TUNEL assay and expression profiles of developmentally important genes were assayed by real-time PCR.In vivo-produced bovine blastocysts were used as controls for experiments determining gene expression patterns. While the cleavage rates did not differ, embryos cultured in SOFaa had higher rates of development to blastocyst stage (P< 0.05). Mean cell numbers and percentages of apoptotic cells per blastocyst did not differ among the groups. Expression of the heat shock protein 70 (Hsp70) gene was significantly up-regulated in both CR1aa and KSOMaa when compared with SOFaa (P< 0.001). DNA methyltransferase 3a (Dnmt3a) expression was higher in embryos cultured in CR1aa than in those cultured in SOFaa (P< 0.001). Expression of interferon tau (IF-τ) and insulin-like growth factor II receptor (Igf-2r) genes was significantly up-regulated in KSOMaa when compared with CR1aa (P< 0.001). Gene expression did not differ betweenin vivo-derived blastocysts and theirin vitro-derived counterparts. In conclusion, SOFaa supports higher development to blastocyst stage than KSOMaa and CR1aa, and the culture conditions influence gene expression.

scholarly journals Liver X Receptors Regulate Adrenal Steroidogenesis and Hypothalamic-Pituitary-Adrenal Feedback

2007 ◽  
Vol 21 (1) ◽  
pp. 126-137 ◽  
Author(s):  
Maria Nilsson ◽  
Thomas M. Stulnig ◽  
Chin-Yo Lin ◽  
Ai Li Yeo ◽  
Peter Nowotny ◽  
...  
Keyword(s):  
Glucose Metabolism ◽  
Hormone Receptors ◽  
Expression Profiles ◽  
Hormone Secretion ◽  
Pituitary Cells ◽  
Hormone Production ◽  
Hypothalamic Pituitary Adrenal ◽  
Adrenal Steroid

Abstract The nuclear hormone receptors liver X receptor α (LXRα) (NR1H3) and LXRβ (NR1H2) are established regulators of cholesterol, lipid, and glucose metabolism and are attractive drug targets for the treatment of diabetes and cardiovascular disease. Adrenal steroid hormones including glucocorticoids and mineralocorticoids are known to interfere with glucose metabolism, insulin signaling, and blood pressure regulation. Here we present genome-wide expression profiles of LXR-responsive genes in both the adrenal and the pituitary gland. LXR activation in cultured adrenal cells inhibited expression of multiple steroidogenic genes and consequently decreased adrenal steroid hormone production. In addition, LXR agonist treatment elevated ACTH mRNA expression and hormone secretion from pituitary cells both in vitro and in vivo. Reduced expression of the glucocortioid-activating enzyme 11β-hydroxysteroid dehydrogenase 1 in pituitary cells upon LXR activation suggests blunting of the negative feedback of glucocorticoids by LXRs. In conclusion, LXRs independently interfere with the hypothalamic-pituitary-adrenal axis regulation at the level of the pituitary and the adrenal gland.

Inhibin, a member of a new peptide family

1989 ◽  
Vol 1 (1) ◽  
pp. 1 ◽  
Author(s):  
HG Burger
Keyword(s):  
Feedback System ◽  
Suppressive Effect ◽  
Commercial Application ◽  
Pituitary Cells ◽  
Cell Content ◽  
Peptide Family ◽  
Normal Menstrual Cycle ◽  
Loop Feedback

There is considerable experimental evidence that follicle stimulating hormone (FSH) and luteinizing hormone (LH) are regulated by separate mechanisms in some circumstances. Part of this differential regulation involves the gonadal factor inhibin, which preferentially affects FSH. A sensitive and specific bioassay based on suppression of FSH cell content in dispersed cultured pituitary cells was used to monitor the purification of inhibin to homogeneity. The two subunits were cloned and the full amino acid sequence of the molecule established. Much evidence has been gathered to support the hypothesis that FSH and inhibin form a classic endocrine closed-loop feedback system in which FSH stimulates inhibin secretion both in vivo and in vitro and inhibin in turn exerts a significant suppressive effect on FSH secretion. The establishment of an inhibin radioimmunoassay has allowed the description of its concentrations in various physiological states such as in the normal menstrual cycle, during puberty and in pregnancy. Inhibin levels were shown to be within the normal range in the polycystic ovarian syndrome. A potential commercial application of inhibin is as a vaccine to increase fertility in domestic animals.

Effect of chronic exposure to a gonadotrophin-releasing hormone agonist on in vitro responsiveness of ovine pituitary cells to inhibin, activin and oestradiol

1994 ◽  
Vol 140 (3) ◽  
pp. 483-493 ◽  
Author(s):  
S Muttukrishna ◽  
P G Knight
Keyword(s):  
Gnrh Agonist ◽  
Inhibitory Effect ◽  
Pituitary Cells ◽  
Cell Content ◽  
Releasing Hormone ◽  
Basal Release ◽  
Lh Release ◽  
Gonadotrophin Releasing Hormone

Abstract To investigate the extent to which the direct actions of inhibin, activin and oestradiol on pituitary output of FSH and LH are dependent on the presence of functional gonadotrophin-releasing hormone (GnRH) receptors, we have compared the effects of these agents on cultured ovine pituitary cells derived from control and GnRH agonist-suppressed ewes. Chronic treatment with GnRH agonist reduced plasma LH and FSH levels (P<0·01) and abolished GnRH-induced release of LH and FSH both in vivo and in vitro. As expected, basal LH release and LH cell content in vitro were drastically reduced in GnRH agonist-suppressed cells (P<0·001). However, basal FSH release and FSH cell content were approximately twofold higher than in control cells (P<0·001). Irrespective of whether the cells had been desensitized to GnRH, inhibin and oestradiol were both found to suppress basal FSH release and FSH cell content in a dose-dependent fashion (P<0·001). Although inhibin had no effect on basal release of LH from control cells, it markedly enhanced GnRH-induced release (P<0·001). In contrast, inhibin increased (P<0·001) basal LH release from GnRH agonist-suppressed cells (which were unresponsive to the GnRH challenge). Inhibin had no overall effect on total LH content/well for either control or GnRH agonist-suppressed cells. Treatment with oestradiol, on the other hand, reduced total LH content/well, an effect which was more pronounced with GnRH agonist-suppressed cells (−44%; P<0·001) than with control cells (−14%, P<0·01). Whereas in control cells activin had no significant effect on any aspect of FSH production examined, in GnRH agonist-treated cells activin enhanced basal FSH release, residual cell content and total FSH content/well (P<0·001). Altering GnRH receptor status also modified the LH response to activin. With control cells activin increased basal release (P<0·001), decreased GnRH-induced release (P<0·001) and increased total LH content/well (P<0·001). With GnRH agonist-treated cells, however, activin had a uniform inhibitory effect on each aspect of LH production examined (P<0·001 in each case). It was concluded that desensitization of ovine gonadotrophs to GnRH by chronic agonist treatment results in a paradoxical enhancement of FSH output in vitro but has little effect on the responsiveness of the cells (in terms of gonadotrophin release and content) to either inhibin or oestradiol. In contrast, GnRH agonist treatment leads to qualitative changes in cellular reponsiveness to activin. Journal of Endocrinology (1994) 140, 483–493

scholarly journals Effects of different oocyte retrieval and in vitro maturation systems on bovine embryo development and quality

Zygote ◽  
2014 ◽  
Vol 23 (3) ◽  
pp. 367-377 ◽  
Author(s):  
Sandra Milena Bernal ◽  
Julia Heinzmann ◽  
Doris Herrmann ◽  
Bernd Timmermann ◽  
Ulrich Baulain ◽  
...  
Keyword(s):  
Gene Expression ◽  
Embryo Development ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Developmental Competence ◽  
Bovine Embryo ◽  
Global Methylation ◽  
Oocyte Developmental Competence

SummaryCyclic adenosine monophosphate (cAMP) modulators have been used to avoid spontaneous oocyte maturation and concomitantly improve oocyte developmental competence. The current work evaluated the effects of the addition of cAMP modulators forskolin, 3-isobutyl-1-methylxanthine (IBMX) and cilostamide during in vitro maturation on the quality and yields of blastocysts. The following experimental groups were evaluated: (i) slicing or (ii) aspiration and maturation in tissue culture medium (TCM)199 for 24 h (TCM24slicing and TCM24aspiration, respectively), (iii) aspiration and maturation in the presence of cAMP modulators for 30 h (cAMP30aspiration) and in vivo-produced blastocysts. In vitro-matured oocytes were fertilized and presumptive zygotes were cultured in vitro to assess embryo development. Cleavage, blastocyst formation, blastocyst cell number, mRNA abundance of selected genes and global methylation profiles were evaluated. Blastocyst rate/zygotes for the TCM24aspiration protocol was improved (32.2 ± 2.1%) compared with TCM24slicing and cAMP30aspiration (23.4 ± 1.2% and 23.3 ± 2.0%, respectively, P<0.05). No statistical differences were found for blastocyst cell numbers. The mRNA expression for the EGR1 gene was down-regulated eight-fold in blastocysts that had been produced in vitro compared with their in vivo counterparts. Gene expression profiles for IGF2R, SLC2A8, COX2, DNMT3B and PCK2 did not differ among experimental groups. Bovine testis satellite I and Bos taurus alpha satellite methylation profiles from cAMP30aspiration protocol-derived blastocysts were similar to patterns that were observed in their in vivo equivalents (P > 0.05), while those from the other groups were significantly elevated. It is concluded that retrieval, collection systems and addition of cAMP modulators can affect oocyte developmental competence, which is reflected not only in blastocyst rates but also in global DNA methylation and gene expression patterns.

scholarly journals miR-31 Functions as a Negative Regulator of Lymphatic Vascular Lineage-Specific Differentiation In Vitro and Vascular Development In Vivo

2010 ◽  
Vol 30 (14) ◽  
pp. 3620-3634 ◽  
Author(s):  
Deena M. Leslie Pedrioli ◽  
Terhi Karpanen ◽  
Vasilios Dabouras ◽  
Giorgia Jurisic ◽  
Glenn van de Hoek ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Vascular System ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Vascular Development ◽  
Negative Regulator ◽  
Tissue Fluid ◽  
Specific Differentiation

ABSTRACT The lymphatic vascular system maintains tissue fluid homeostasis, helps mediate afferent immune responses, and promotes cancer metastasis. To address the role microRNAs (miRNAs) play in the development and function of the lymphatic vascular system, we defined the in vitro miRNA expression profiles of primary human lymphatic endothelial cells (LECs) and blood vascular endothelial cells (BVECs) and identified four BVEC signature and two LEC signature miRNAs. Their vascular lineage-specific expression patterns were confirmed in vivo by quantitative real-time PCR and in situ hybridization. Functional characterization of the BVEC signature miRNA miR-31 identified a novel BVEC-specific posttranscriptional regulatory mechanism that inhibits the expression of lymphatic lineage-specific transcripts in vitro. We demonstrate that suppression of lymphatic differentiation is partially mediated via direct repression of PROX1, a transcription factor that functions as a master regulator of lymphatic lineage-specific differentiation. Finally, in vivo studies of Xenopus and zebrafish demonstrated that gain of miR-31 function impaired venous sprouting and lymphatic vascular development, thus highlighting the importance of miR-31 as a negative regulator of lymphatic development. Collectively, our findings identify miR-31 is a potent regulator of vascular lineage-specific differentiation and development in vertebrates.

scholarly journals Regulation of SpeB in Streptococcus pyogenes by pH and NaCl: a Model for In Vivo Gene Expression

2006 ◽  
Vol 188 (2) ◽  
pp. 399-408 ◽  
Author(s):  
Jennifer A. Loughman ◽  
Michael Caparon
Keyword(s):  
Gene Expression ◽  
Streptococcus Pyogenes ◽  
Growth Phase ◽  
Soft Tissues ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Transcriptional Response ◽  
Culture Conditions

ABSTRACT For a pathogen such as Streptococcus pyogenes, ecological success is determined by its ability to sense the environment and mount an appropriate adaptive transcriptional response. Thus, determining conditions for analyses of gene expression in vitro that are representative of the in vivo environment is critical for understanding the contributions of transcriptional response pathways to pathogenesis. In this study, we determined that the gene encoding the SpeB cysteine protease is up-regulated over the course of infection in a murine soft-tissue model. Conditions were identified, including growth phase, acidic pH, and an NaCl concentration of <0.1 M, that were required for expression of speB in vitro. Analysis of global expression profiles in response to these conditions in vitro identified a set of coregulated genes whose expression patterns showed a significant correlation with that of speB when examined during infection of murine soft tissues. This analysis revealed that a culture medium that promotes high levels of SpeB expression in vitro produced an expression profile that showed significant correlation to the profile observed in vivo. Taken together, these studies establish culture conditions that mimic in vivo expression patterns; that growth phase, pH, and NaCl may mimic relevant cues sensed by S. pyogenes during infection; and that identification of other environmental cues that alter expression of speB in vitro may provide insight into the signals that direct global patterns of gene expression in vivo.

Cryotolerance and global gene-expression patterns of Bos taurus indicus and Bos taurus taurus in vitro- and in vivo-produced blastocysts

2014 ◽  
Vol 26 (8) ◽  
pp. 1129 ◽  
Author(s):  
Mateus J. Sudano ◽  
Ester S. Caixeta ◽  
Daniela M. Paschoal ◽  
Alicio Martins ◽  
Rui Machado ◽  
...  
Keyword(s):  
Gene Expression ◽  
Lipid Metabolism ◽  
Bos Taurus ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Global Gene Expression ◽  
Embryo Viability ◽  
Bos Taurus Indicus

In a 2 × 2 factorial experimental design, embryo development, cryotolerance and global gene expression of Nellore (Bos taurus indicus) and Simmental (Bos taurus taurus) blastocysts produced in vitro (IVP) and in vivo (multiple ovulation derived embryo, MODE) were assessed. Blastocyst production was higher in Nellore than in Simmental (47.7 ± 2.0% vs 27.0 ± 2.0%) cows. The total numbers of ova or embryos recovered (5.5 ± 0.9 vs 3.7 ± 0.8) and transferable embryos (3.8 ± 1.0 vs 2.3 ± 0.8) per cow were not different between breeds. Simmental and MODE (34.6% and 38.5%, n = 75 and 70) blastocysts had higher survival rates after cryopreservation compared with Nellore and IVP (20.2% and 18.1%, n = 89 and 94) embryos, respectively. Differences between transcriptomes were addressed by principal-component analysis, which indicated that gene expression was affected by subspecies (158 genes), origin (532 genes) and interaction between both subspecies and origin (53 genes). Several functional processes and pathways relevant to lipid metabolism and embryo viability involving differentially expressed genes were identified. The lipid metabolism-related genes were upregulated in Simmental (AUH and ELOVL6) and IVP (ACSL3 and ACSL6) blastocysts. The expression profiles of genes related to mitochondrial metabolism (ATP5B), oxidative stress (GPX4), apoptosis (DAD1, DAP, PRDX2), heat shock (HSPA5), pregnancy (IFNT2, PAG2) and cell differentiation (KRT18) varied between experimental groups.

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