scholarly journals Modulation of Progesterone Receptor Isoform Expression in Pregnant Human Myometrium

2017 ◽  
Vol 2017 ◽  
pp. 1-17 ◽  
Author(s):  
Marina Ilicic ◽  
Tamas Zakar ◽  
Jonathan W. Paul
Keyword(s):  
Progesterone Receptor ◽  
Ex Situ ◽  
Human Myometrium ◽  
Mrna Levels ◽  
Protein Acetylation ◽  
Nonhistone Protein ◽  
Serum Supplementation ◽  
Isoform Expression ◽  
Induced Changes

Background. Regulation of myometrial progesterone receptor (PR) expression is an unresolved issue central to understanding the mechanism of functional progesterone withdrawal and initiation of labor in women. Objectives. To determine whether pregnant human myometrium undergoes culture-induced changes in PR isoform expression ex situ and, further, to determine if conditions approaching the in vivo environment stabilise PR isoform expression in culture. Methods. Term nonlaboring human myometrial tissues were cultured under specific conditions: serum supplementation, steroids, stretch, cAMP, PMA, PGF2α, NF-κB inhibitors, or TSA. Following 48 h culture, PR-T, PR-A, and PR-B mRNA levels were determined using qRT-PCR. PR-A/PR-B ratios were calculated. Results. PR-T and PR-A expression and the PR-A/PR-B ratio significantly increased in culture. Steroids prevented the culture-induced increase in PR-T and PR-A expression. Stretch blocked the effects of steroids on PR-T and PR-A expression. PMA further increased the PR-A/PR-B ratio, while TSA blocked culture-induced increases of PR-A expression and the PR-A/PR-B ratio. Conclusion. Human myometrial tissue in culture undergoes changes in PR gene expression consistent with transition toward a laboring phenotype. TSA maintained the nonlaboring PR isoform expression pattern. This suggests that preserving histone and/or nonhistone protein acetylation is critical for maintaining the progesterone dependent quiescent phenotype of human myometrium in culture.

scholarly journals Methanogens and Methanogenesis in the Rumens and Ceca of Lambs Fed Two Different High-Grain-Content Diets

2012 ◽  
Vol 79 (6) ◽  
pp. 1777-1786 ◽  
Author(s):  
M. Popova ◽  
D. P. Morgavi ◽  
C. Martin
Keyword(s):  
Metabolic Activity ◽  
Methane Production ◽  
Direct Evidence ◽  
Species Level ◽  
Ex Situ ◽  
Content Type ◽  
Copy Numbers ◽  
Dietary Starch ◽  
Induced Changes

ABSTRACTThe amount and nature of dietary starch are known to influence the extent and site of feed digestion in ruminants. However, how starch degradability may affect methanogenesis and methanogens along the ruminant's digestive tract is poorly understood. This study examined the diversity and metabolic activity of methanogens in the rumen and cecum of lambs receiving wheat or corn high-grain-content diets. Methane productionin vivoandex situwas also monitored.In vivodaily methane emissions (CH4g/day) were 36% (P< 0.05) lower in corn-fed lambs than in wheat-fed lambs.Ex situmethane production (μmol/h) was 4-fold higher for ruminal contents than for cecal contents (P< 0.01), while methanogens were 10-fold higher in the rumen than in the cecum (mcrAcopy numbers;P< 0.01). Clone library analysis indicated thatMethanobrevibacterwas the dominant genus in both sites. Diet induced changes at the species level, as theMethanobrevibacter millerae-M. gottschalkii-M. smithiiclade represented 78% of the sequences from the rumen of wheat-fed lambs and just about 52% of the sequences from the rumen of the corn-fed lambs. Diet did not affectmcrAexpression in the rumen. In the cecum, however, expression was 4-fold and 2-fold lower than in the rumen for wheat- and corn-fed lambs, respectively. Though we had no direct evidence for compensation of reduced rumen methane production with higher cecum methanogenesis, the ecology of methanogens in the cecum should be better considered.

scholarly journals Culture-Induced Changes in Blood—Brain Barrier Transcriptome: Implications for Amino-Acid Transporters in vivo

2009 ◽  
Vol 29 (9) ◽  
pp. 1491-1502 ◽  
Author(s):  
Ruth Lyck ◽  
Nadine Ruderisch ◽  
Anton G Moll ◽  
Oliver Steiner ◽  
Clemens D Cohen ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Blood Brain Barrier ◽  
Brain Barrier ◽  
Amino Acid Transporters ◽  
Mrna Levels ◽  
Blood Brain ◽  
Barrier Formation ◽  
Platelet Endothelial Cell Adhesion ◽  
Induced Changes

Tight homeostatic control of brain amino acids (AA) depends on transport by solute carrier family proteins expressed by the blood—brain barrier (BBB) microvascular endothelial cells (BMEC). To characterize the mouse BMEC transcriptome and probe culture-induced changes, microarray analyses of platelet endothelial cell adhesion molecule-1-positive (PECAM1+) endothelial cells (ppMBMECs) were compared with primary MBMECs (pMBMEC) cultured in the presence or absence of glial cells and with b.End5 endothelioma cell line. Selected cell marker and AA transporter mRNA levels were further verified by reverse transcription real-time PCR. Regardless of glial coculture, expression of a large subset of genes was strongly altered by a brief culture step. This is consistent with the known dependence of BMECs on in vivo interactions to maintain physiologic functions, for example, tight barrier formation, and their consequent dedifferentiation in culture. Seven ( 4F2hc, Lat1, Taut, Snat3, Snat5, Xpct, and Cat1) of nine AA transporter mRNAs highly expressed in freshly isolated ppMBMECs were strongly downregulated for all cultures and two ( Snat2 and Eaat3) were variably regulated. In contrast, five AA transporter mRNAs with low expression in ppMBMECs, including y+ Lat2, xCT, and Snat1, were upregulated by culture. We hypothesized that the AA transporters highly expressed in ppMBMECs and downregulated in culture have a major in vivo function for BBB transendothelial transport.

Cytokine-induced changes in chromatin structure and in vivo footprints in the inducible NOS promoter

2001 ◽  
Vol 280 (3) ◽  
pp. L390-L399 ◽  
Author(s):  
Jane K. Mellott ◽  
Harry S. Nick ◽  
Michael F. Waters ◽  
Timothy R. Billiar ◽  
David A. Geller ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Protein Interactions ◽  
Molecular Mechanisms ◽  
Cell Types ◽  
Specific Pattern ◽  
Mrna Levels ◽  
Inos Gene ◽  
In Vivo Footprinting ◽  
Induced Changes

Transcription of the human inducible nitric oxide synthase ( iNOS) gene is regulated by inflammatory cytokines in a tissue-specific manner. To determine whether differences in cytokine-induced mRNA levels between pulmonary epithelial cells (A549) and hepatic biliary epithelial cells (AKN-1) result from different protein or DNA regulatory mechanisms, we identified cytokine-induced changes in DNase I-hypersensitive (HS) sites in 13 kb of the iNOS 5′-flanking region. Data showed both constitutive and inducible HS sites in an overlapping yet cell type-specific pattern. Using in vivo footprinting and ligation-mediated PCR to detect potential DNA or protein interactions, we examined one promoter region near −5 kb containing both constitutive and cytokine-induced HS sites. In both cell types, three in vivo footprints were present in both control and cytokine-treated cells, and each mapped within a constitutive HS site. The remaining footprint appeared only in response to cytokine treatment and mapped to an inducible HS site. These studies, performed on chromatin in situ, identify a portion of the molecular mechanisms regulating transcription of the human iNOS gene in both lung- and liver-derived epithelial cells.

scholarly journals Transendothelial movement of adiponectin is restricted by glucocorticoids

2017 ◽  
Vol 234 (2) ◽  
pp. 101-114 ◽  
Author(s):  
Thanh Q Dang ◽  
Nanyoung Yoon ◽  
Helen Chasiotis ◽  
Emily C Dunford ◽  
Qilong Feng ◽  
...  
Keyword(s):  
Critical Role ◽  
Endothelial Permeability ◽  
The Body ◽  
Diabetic Rat ◽  
Mrna Levels ◽  
Endothelial Monolayers ◽  
Basolateral Side ◽  
Induced Changes

Altered permeability of the endothelial barrier in a variety of tissues has implications both in disease pathogenesis and treatment. Glucocorticoids are potent mediators of endothelial permeability, and this forms the basis for their heavily prescribed use as medications to treat ocular disease. However, the effect of glucocorticoids on endothelial barriers elsewhere in the body is less well studied. Here, we investigated glucocorticoid-mediated changes in endothelial flux of Adiponectin (Ad), a hormone with a critical role in diabetes. First, we used monolayers of endothelial cells in vitro and found that the glucocorticoid dexamethasone increased transendothelial electrical resistance and reduced permeability of polyethylene glycol (PEG, molecular weight 4000 Da). Dexamethasone reduced flux of Ad from the apical to basolateral side, measured both by ELISA and Western blotting. We then examined a diabetic rat model induced by treatment with exogenous corticosterone, which was characterized by glucose intolerance and hyperinsulinemia. There was no change in circulating Ad but less Ad protein in skeletal muscle homogenates, despite slightly higher mRNA levels, in diabetic vs control muscles. Dexamethasone-induced changes in Ad flux across endothelial monolayers were associated with alterations in the abundance of select claudin tight junction (TJ) proteins. shRNA-mediated knockdown of one such gene, claudin-7, in HUVEC resulted in decreased TEER and increased adiponectin flux, confirming the functional significance of Dex-induced changes in its expression. In conclusion, our study identifies glucocorticoid-mediated reductions in flux of Ad across endothelial monolayers in vivo and in vitro. This suggests that impaired Ad action in target tissues, as a consequence of reduced transendothelial flux, may contribute to the glucocorticoid-induced diabetic phenotype.

scholarly journals TBX2, a Novel Regulator of Labour

Medicina ◽  
2021 ◽  
Vol 57 (6) ◽  
pp. 515
Author(s):  
Febilla Fernando ◽  
Geertruda J.M. Veenboer ◽  
Martijn A. Oudijk ◽  
Marlies A.M. Kampman ◽  
Karst Y. Heida ◽  
...  
Keyword(s):  
Preterm Labour ◽  
Therapeutic Interventions ◽  
Human Myometrium ◽  
Mrna Levels ◽  
Myometrial Cells ◽  
Cytokines And Chemokines ◽  
Term Labour ◽  
Upstream Regulators

Background and Objectives: Therapeutic interventions targeting molecular factors involved in the transition from uterine quiescence to overt labour are not substantially reducing the rate of spontaneous preterm labour. The identification of novel rational therapeutic targets are essential to prevent the most common cause of neonatal mortality. Based on our previous work showing that Tbx2 (T-Box transcription factor 2) is a putative upstream regulator preceding progesterone withdrawal in mouse myometrium, we now investigate the role of TBX2 in human myometrium. Materials and Methods: RNA microarray analysis of (A) preterm human myometrium samples and (B) myometrial cells overexpressing TBX2 in vitro, combined with subsequent analysis of the two publicly available datasets of (C) Chan et al. and (D) Sharp et al. The effect of TBX2 overexpression on cytokines/chemokines secreted to the myometrium cell culture medium were determined by Luminex assay. Results: Analysis shows that overexpression of TBX2 in myometrial cells results in downregulation of TNFα- and interferon signalling. This downregulation is consistent with the decreased expression of cytokines and chemokines of which a subset has been previously associated with the inflammatory pathways relevant for human labour. In contrast, CXCL5 (C-X-C motif chemokine ligand 5), CCL21 and IL-6 (Interleukin 6), previously reported in relation to parturition, do not seem to be under TBX2 control. The combined bioinformatical analysis of the four mRNA datasets identifies a subset of upstream regulators common to both preterm and term labour under control of TBX2. Surprisingly, TBX2 mRNA levels are increased in preterm contractile myometrium. Conclusions: We identified a subset of upstream regulators common to both preterm and term labour that are activated in labour and repressed by TBX2. The increased TBX2 mRNA expression in myometrium collected during a preterm caesarean section while in spontaneous preterm labour compared to tissue harvested during iatrogenic preterm delivery does not fit the bioinformatical model. We can only explain this by speculating that the in vivo activity of TBX2 in human myometrium depends not only on the TBX2 expression levels but also on levels of the accessory proteins necessary for TBX2 activity.

scholarly journals Changes in Gene Expression during G-CSF-Induced Emergency Granulopoiesis in Humans

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 2202-2202
Author(s):  
Corinna Cavan Pedersen ◽  
Rehannah Borup ◽  
Anne Fischer-Nielsen ◽  
Helena Mora-Jensen ◽  
Anna Fossum ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mirna Expression ◽  
Conflicts Of Interest ◽  
Expression Patterns ◽  
Critical Role ◽  
Severe Infection ◽  
Mrna Levels ◽  
Healthy Individuals ◽  
Induced Changes

Abstract Emergency granulopoiesis refers to the increased production of neutrophils in bone marrow and their release into circulation induced by severe infection. Several studies point to a critical role for granulocyte colony-stimulating factor (G-CSF) as the main mediator of emergency granulopoiesis. However, the consequences of G-CSF stimulation on the transcriptome of neutrophils and their precursors have not yet been elucidated in humans. Here, we investigate the changes in mRNA and miRNA expression in successive stages of neutrophil development following in vivo administration of G-CSF in humans, mimicking emergency granulopoiesis. Blood samples were collected from healthy individuals after five days of G-CSF administration. Neutrophil precursors were sorted into discrete stages of maturation by flow cytometry and extracted RNA was subjected to microarray analysis. mRNA levels were compared to previously published expression levels in corresponding populations of neutrophil precursors isolated from bone marrow of untreated, healthy individuals. miRNA expression was investigated in the most mature cell population to determine G-CSF-induced changes in circulating neutrophils. G-CSF substantially affected mRNA and miRNA expression patterns, demonstrating significant impact on neutrophil development and function. 1110 mRNAs were differentially expressed more than 2-fold with G-CSF while the treatment induced changes in the levels of 73 miRNAs in the mature population. In addition, G-CSF treatment reduced the levels of four out of five measured granule proteins in mature neutrophils including hCAP-18, which was completely deficient in neutrophils from G-CSF-treated donors. Cell cycle analysis pointed towards an induced proliferative capacity of myelocytes. These results indicate that multiple biological processes are altered in order to satisfy the increased demand for neutrophils during G-CSF-induced emergency granulopoiesis. Disclosures No relevant conflicts of interest to declare.

scholarly journals Differential expression and regulation of progesterone receptor isoforms in rat and mouse pituitary cells and LβT2 gonadotropes

2006 ◽  
Vol 190 (3) ◽  
pp. 837-846 ◽  
Author(s):  
Judith L Turgeon ◽  
Dennis W Waring
Keyword(s):  
Progesterone Receptor ◽  
Mrna Level ◽  
Primary Cultures ◽  
Ovariectomized Rats ◽  
Pituitary Cells ◽  
Mrna Levels ◽  
Mouse Cells ◽  
Isoform Expression ◽  
Mouse Pituitary ◽  
Lh Secretion

Manipulation of endogenous progesterone receptor (PR) does not produce equivalent physiological effects in mouse and rat pituitary cells. To test whether this may be due in part to difference in PR isoform expression, we examined hormonally regulated pituitary PR-A and PR-B mRNA levels using quantitative real-time PCR. The LβT2 mouse gonadotrope line or pituitary cells from adult, ovariectomized rats or mice were cultured with or without 0.2 nM 17β-estradiol (E2) for 3 days. PR-A was the predominant form expressed for all groups. For mouse cells, E2 led to an increase in both isoforms without a change in the A:B ratio; for rat cells, the PR-B response to E2 was more robust resulting in a decrease in the A:B ratio. Exposure of E2-treated pituitary cells to 200 nM progesterone for 6 h decreased both PR-A and PR-B levels in rat cells, but had no effect on PR isoform expression in mouse cells even when exposure was extended to 12 h. The low level of PR expression found in LβT2 gonadotropes was unaffected by E2, alone or with progesterone. The weak PR expression and lack of responsiveness of LβT2 cells cannot be explained by a male phenotype as was shown by the more than tenfold higher PR mRNA level in primary cultures of male mouse pituitary cells, which responded to E2 stimulation with a proportional increase in PR isoforms similar to female cells. Functionally, E2-stimulated changes in PR mRNA isoform ratios in rat, mouse or LβT2 cells correlated with the degree of progesterone augmentation of GnRH-stimulated LH secretion in these models. These results are consistent with the hypothesis that robust GnRH priming and progesterone augmentation of LH secretion in the rat compared to these events in the mouse are a consequence, in part, of differences in the E2-modulated ratio of PR isoforms.

CD36-dependent fatty acid uptake regulates expression of peroxisome proliferator activated receptors

2005 ◽  
Vol 33 (1) ◽  
pp. 311-315 ◽  
Author(s):  
V.A. Drover ◽  
N.A. Abumrad
Keyword(s):  
Plasma Lipids ◽  
Fatty Acid Uptake ◽  
Peroxisome Proliferator ◽  
Acid Uptake ◽  
Mrna Levels ◽  
Peroxisome Proliferator Activated Receptor ◽  
Peroxisome Proliferator Activated Receptors ◽  
Induced Changes ◽  
Deficient Mice

CD36 is an important regulator of lipid metabolism in vivo due to its role in the facilitated uptake of long-chain FAs (fatty acids). CD36-deficient mice display reduced TAG (triacylglycerol) in muscle, but elevated hepatic TAG. Also, insulin sensitivity is enhanced peripherally, while it appears impaired in the liver [Goudriaan, Dahlmans, Teusink, Ouwens, Febbraio, Maassen, Romijn, Havekes, and Voshol (2003) J. Lipid. Res. 44, 2270–2277; and Hajri, Han, Bonen and Abumrad (2002) J. Clin. Invest. 109, 1381–1389]. Tissues such as muscle, which normally express high levels of CD36, shift to high glucose utilization in CD36 deficiency [Hajri, Han, Bonen and Abumrad (2002) J. Clin. Invest. 109, 1381–1389], so we hypothesized that this shift must involve adaptive changes in the PPAR (peroxisome-proliferator-activated receptor) transcription factors which regulate FA metabolism. To test this, we examined mRNA levels for the three PPAR isoforms in tissues of WT (wild-type) and CD36-deficient mice following the administration of saline, glucose or olive oil by intragastric gavage. Compared with WT mice, CD36-null mice had 5–10-fold increased PPAR mRNA in adipose tissue in the basal state, and did not exhibit diet-induced changes. Correlations between adipose PPAR mRNA abundance and plasma lipids were observed in WT mice, but not in CD36-null mice. The opposite was true for hepatic PPAR mRNA levels, which correlated with plasma FA, TAG and/or glucose only in CD36-null mice. No significant differences were observed in PPAR mRNA levels in the intestine, where CD36 does not impact on FA uptake. The data suggest that CD36 and the PPARs are components of the FA-sensing machinery to respond to changes in FA flux in a tissue-specific manner.

Effect of chronic energy deprivation on cardiac thyroid hormone receptor and myosin isoform expression

1994 ◽  
Vol 266 (2) ◽  
pp. E254-E260 ◽  
Author(s):  
S. J. Swoap ◽  
F. Haddad ◽  
P. Bodell ◽  
K. M. Baldwin
Keyword(s):  
Thyroid Hormone ◽  
Thyroid Hormone Receptor ◽  
Binding Capacity ◽  
Relative Content ◽  
Female Rats ◽  
Mrna Levels ◽  
Isoform Expression ◽  
Energy Deprivation

Thyroid hormone (3,5,3'-triiodothyronine; T3) and its receptor (TR) play an important regulatory role for in vivo and in vitro cardiac myosin heavy chain (MHC) isoform gene expression by activating the alpha- and inhibiting the beta-MHC genes. Previous studies have shown that chronic energy deprivation (CED; 50% of normal caloric intake) in the rat impacts cardiac MHC protein expression and hemodynamic parameters in a pattern typically seen with hypothyroidism; yet, unlike hypothyroidism, circulating T3 levels are not depressed. Therefore, the goal of this study was to determine if the altered MHC isoform expression observed in CED is associated with altered TR expression, both at the mRNA and protein levels. Female rats weighing approximately 250 g were allocated into two groups, designated as normal control (NC) and CED. After 5 wk, the relative content of alpha-MHC protein and mRNA levels decreased in CED ventricles by 20% (P < 0.05). In contrast, the relative content of both beta-MHC protein and mRNA levels increased five- to sixfold in CED (P < 0.05). Although there were no changes in TR mRNA levels relative to 18S rRNA in CED, the total number of nuclear TRs decreased 3.5-fold in the CED group (P < 0.05), from a maximum binding capacity of 840 +/- 130 fmol/mg DNA in NC to 241 +/- 118 fmol/mg DNA in CED, with no change in the affinity of the receptor.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Evaluation of [18F]F-DPA as a target for TSPO in head and neck cancer under normal conditions and after radiotherapy

2020 ◽  
Author(s):  
Sanni Tuominen ◽  
Thomas Keller ◽  
Nataliia Petruk ◽  
Francisco López-Picón ◽  
Dominik Eichin ◽  
...  
Keyword(s):  
Head And Neck ◽  
Inflammatory Responses ◽  
Ex Vivo ◽  
Mrna Levels ◽  
Malignant Tumours ◽  
Pre Treatment ◽  
Induced Changes ◽  
Tspo Expression

Abstract Background Many malignant tumours have increased TSPO expression, which has been related to a poor prognosis. TSPO-PET tracers have not comprehensively been evaluated in peripherally located tumours. This study aimed to evaluate whether N,N-diethyl-2-(2-(4-([18F]fluoro)phenyl)-5,7-dimethylpyrazolo[1,5-a]pyrimidin-3-yl)acetamide ([18F]F-DPA) can reflect radiotherapy (RT)-induced changes in TSPO activity in head and neck squamous cell carcinoma (HNSCC). Methods RT was used to induce inflammatory responses in HNSCC xenografts and cells. [18F]F-DPA uptake was measured in vivo in non-irradiated and irradiated tumours, followed by ex vivo biodistribution, autoradiography, and radiometabolite analysis. In vitro studies were performed in parental and TSPO-silenced (TSPO siRNA) cells. TSPO protein and mRNA expression, as well as tumour-associated macrophages (TAMs), were also assessed. Results In vivo imaging and ex vivo measurement revealed significantly higher [18F]F-DPA uptake in irradiated, compared to non-irradiated tumours. In vitro labelling studies with cells confirmed this finding, whereas no effect of RT on [18F]F-DPA uptake was detected in TSPO siRNA cells. Radiometabolite analysis showed that the amount of unchanged [18F]F-DPA in tumours was 95%, also after irradiation. PK11195 pre-treatment reduced the tumour-to-blood ratio of [18F]F-DPA by 73% in xenografts and by 88% in cells. TSPO protein and mRNA levels increased after RT, but were highly variable. The proportion of M1/M2 TAMs decreased after RT, whereas the proportion of monocytes and migratory monocytes/macrophages increased. Conclusions [18F]F-DPA can detect changes in TSPO expression levels after RT in HNSCC, which does not seem to reflect inflammation. Further studies are however needed to clarify the physiological mechanisms regulated by TSPO after RT.

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