Activation of nuclear factor of activated T cells 5 in the peritoneal membrane of uremic patients

2015 ◽  
Vol 308 (11) ◽  
pp. F1247-F1258 ◽  
Author(s):  
Daniel Kitterer ◽  
Joerg Latus ◽  
Christoph Ulmer ◽  
Peter Fritz ◽  
Dagmar Biegger ◽  
...  
Keyword(s):  
T Cells ◽  
Mrna Expression ◽  
Mrna Levels ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Significant Difference ◽  
Ccl2 Mrna ◽  
Uremic Patients ◽  
High Concentrations

Peritoneal inflammation and fibrosis are responses to the uremic milieu and exposure to hyperosmolar dialysis fluids in patients on peritoneal dialysis. Cells respond to high osmolarity via the transcription factor nuclear factor of activated T cells (NFAT5). In the present study, the response of human peritoneal fibroblasts to glucose was analyzed in vitro. Expression levels of NFAT5 and chemokine (C-C motif) ligand (CCL2) mRNA were quantified in peritoneal biopsies of five nonuremic control patients, five uremic patients before PD (pPD), and eight patients on PD (oPD) using real-time PCR. Biopsies from 5 control patients, 25 pPD patients, and 25 oPD patients were investigated using immunohistochemistry to detect the expression of NFAT5, CCL2, NF-κB p50, NF-κB p65, and CD68. High glucose concentrations led to an early, dose-dependent induction of NFAT5 mRNA in human peritoneal fibroblasts. CCL2 mRNA expression was upregulated by high concentrations of glucose after 6 h, but, most notably, a concentration-dependent induction of CCL2 was present after 96 h. In human peritoneal biopsies, NFAT5 mRNA levels were increased in uremic patients compared with nonuremic control patients. No significant difference was found between the pPD group and oPD group. CCL2 mRNA expression was higher in the oPD group. Immunohistochemistry analysis was consistent with the results of mRNA analysis. CD68-positive cells were significantly increased in the oPD group. In conclusion, uremia results in NFAT5 induction, which might promote early changes of the peritoneum. Upregulation of NFAT5 in PD patients is associated with NFκB induction, potentially resulting in the recruitment of macrophages.

scholarly journals Nuclear factor of activated T cells 2 is required for osteoclast differentiation and function in vitro but not in vivo

2018 ◽  
Vol 119 (11) ◽  
pp. 9334-9345 ◽  
Author(s):  
Jungeun Yu ◽  
Stefano Zanotti ◽  
Lauren Schilling ◽  
Ernesto Canalis
Keyword(s):  
T Cells ◽  
Osteoclast Differentiation ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
And Function

scholarly journals Physiological tonicity improves human chondrogenic marker expression through nuclear factor of activated T-cells 5 in vitro

2010 ◽  
Vol 12 (3) ◽  
pp. R100 ◽  
Author(s):  
Anna E van der Windt ◽  
Esther Haak ◽  
Ruud HJ Das ◽  
Nicole Kops ◽  
Tim JM Welting ◽  
...  
Keyword(s):  
T Cells ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Chondrogenic Marker ◽  
Marker Expression

scholarly journals Regulation of Myosin Heavy Chain Expression during Rat Skeletal Muscle Development In Vitro

2001 ◽  
Vol 12 (5) ◽  
pp. 1499-1508 ◽  
Author(s):  
Carol E. Torgan ◽  
Mathew P. Daniels
Keyword(s):  
Skeletal Muscle ◽  
T Cells ◽  
Cyclosporin A ◽  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Fiber Formation ◽  
Immunofluorescent Staining ◽  
Nuclear Factor ◽  
Activated T Cells

Signals that determine fast- and slow-twitch phenotypes of skeletal muscle fibers are thought to stem from depolarization, with concomitant contraction and activation of calcium-dependent pathways. We examined the roles of contraction and activation of calcineurin (CN) in regulation of slow and fast myosin heavy chain (MHC) protein expression during muscle fiber formation in vitro. Myotubes formed from embryonic day 21 rat myoblasts contracted spontaneously, and ∼10% expressed slow MHC after 12 d in culture, as seen by immunofluorescent staining. Transfection with a constitutively active form of calcineurin (CN*) increased slow MHC by 2.5-fold as determined by Western blot. This effect was attenuated 35% by treatment with tetrodotoxin and 90% by administration of the selective inhibitor of CN, cyclosporin A. Conversely, cyclosporin A alone increased fast MHC by twofold. Cotransfection with VIVIT, a peptide that selectively inhibits calcineurin-induced activation of the nuclear factor of activated T-cells, blocked the effect of CN* on slow MHC by 70% but had no effect on fast MHC. The results suggest that contractile activity-dependent expression of slow MHC is mediated largely through the CN–nuclear factor of activated T-cells pathway, whereas suppression of fast MHC expression may be independent of nuclear factor of activated T-cells.

scholarly journals Brain Endothelial miR-146a Negatively Modulates T-Cell Adhesion through Repressing Multiple Targets to Inhibit NF-κB Activation

2015 ◽  
Vol 35 (3) ◽  
pp. 412-423 ◽  
Author(s):  
Dongsheng Wu ◽  
Camilla Cerutti ◽  
Miguel A Lopez-Ramirez ◽  
Gareth Pryce ◽  
Josh King-Robson ◽  
...  
Keyword(s):  
T Cells ◽  
Endothelial Cells ◽  
Nuclear Translocation ◽  
Leukocyte Adhesion ◽  
Cell Types ◽  
Nuclear Factor ◽  
Autoimmune Encephalomyelitis ◽  
Activated T Cells ◽  
Cytokine Activation

Pro-inflammatory cytokine-induced activation of nuclear factor, NF-κB has an important role in leukocyte adhesion to, and subsequent migration across, brain endothelial cells (BECs), which is crucial for the development of neuroinflammatory disorders such as multiple sclerosis (MS). In contrast, microRNA-146a (miR-146a) has emerged as an anti-inflammatory molecule by inhibiting NF-κB activity in various cell types, but its effect in BECs during neuroinflammation remains to be evaluated. Here, we show that miR-146a was upregulated in microvessels of MS-active lesions and the spinal cord of mice with experimental autoimmune encephalomyelitis. In vitro, TNFα and IFNγ treatment of human cerebral microvascular endothelial cells (hCMEC/D3) led to upregulation of miR-146a. Brain endothelial overexpression of miR-146a diminished, whereas knockdown of miR-146a augmented cytokine-stimulated adhesion of T cells to hCMEC/D3 cells, nuclear translocation of NF-κB, and expression of adhesion molecules in hCMEC/D3 cells. Furthermore, brain endothelial miR-146a modulates NF-κB activity upon cytokine activation through targeting two novel signaling transducers, RhoA and nuclear factor of activated T cells 5, as well as molecules previously identified, IL-1 receptor-associated kinase 1, and TNF receptor-associated factor 6. We propose brain endothelial miR-146a as an endogenous NF-κB inhibitor in BECs associated with decreased leukocyte adhesion during neuroinflammation.

Abstract 439: Biomechanical Stimulation Promotes Translocation of Nuclear Factor of Activated T-cells 5 Into The Nucleus of Vascular Smooth Muscle Cells

2015 ◽  
Vol 35 (suppl_1) ◽  
Author(s):  
Maren Hödebeck ◽  
Markus Hecker ◽  
Thomas Korff
Keyword(s):  
T Cells ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Mrna Expression ◽  
Vascular Smooth Muscle Cells ◽  
Splice Variant ◽  
Nuclear Translocation ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Biomechanical Stimulation

Chronic alterations in the biomechanical stimulation of vascular smooth muscle cells (VSMC) as experienced during hypertension lead to changes in VSMC phenotype and function and further enable the structural remodeling of the vessel wall. In this context, we recently reported that an increase in wall stress or biomechanical stretch is sufficient to activate nuclear factor of activated T-cells 5 (NFAT5). This transcription factor promotes the expression of gene products such as tenascin-C and κ-actin, both involved in VSMC migration. Based on these findings, we hypothesized that biomechanical stretch elicits NFAT5 mRNA expression and induces biochemical modifications of NFAT5 on the post-translational level, a prerequisite for its entry into the nucleus and transcriptional activity. To scrutinize this hypothesis, human arterial VSMC were exposed to biomechanical stretch (13%, 0.5 Hz) and subjected to detailed mRNA expression analyses. While a ~3-fold reduction in NFAT5 splice variant 1 (isoform A) mRNA expression was observed in stretch-stimulated VSMC as compared to the static controls (n=3, p<0.05), splice variant 3 (isoform C) mRNA levels were induced ~1.8-fold (n=3, p<0.05). Overexpression of corresponding Flag-tagged NFAT5 proteins in VSMC and subsequent immunofluorescence as well as biochemical analyses revealed that isoform A was primarily located in the cytoplasm of static and stretch-stimulated VSMC while isoform C was preferentially localized in the nucleus under baseline conditions and further accumulated in the nucleus upon biomechanical stimulation (n=3, p<0.05). Nuclear translocation of isoform C was amplified for phosphorylation-deficient mutants generated by exchanging serine to alanine at position 1197 even under static culture conditions while a phosphomimetic mutation at this residue (serine to glutamate) inhibited NFAT5c nuclear translocation (n=3, p<0.05). Collectively, our findings indicate that exposure of VSMC to biomechanical stretch triggers the expression of NFAT5 isoform C and controls its entry into the nucleus via phosphorylation at S-1197. Current investigations are focusing on the impact of NFAT5 on hypertensive remodeling utilizing inducible smooth muscle cell-specific NFAT5-deficient mice.

249 EXPRESSION OF mRNA ENCODING EPIDERMAL GROWTH FACTOR-LIKE FACTORS IN BOVINE CUMULUS CELLS DURING IN VITRO MATURATION: EFFECTS OF TIME AND FOLLICLE-STIMULATING HORMONE

2011 ◽  
Vol 23 (1) ◽  
pp. 222
Author(s):  
E. S. Caixeta ◽  
M. F. Machado ◽  
P. Ripamonte ◽  
P. F. Lima ◽  
A. C. S. Castilho ◽  
...  
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Mrna Expression ◽  
In Vitro Maturation ◽  
Cumulus Cells ◽  
Mrna Abundance ◽  
Mrna Levels ◽  
Significant Difference ◽  
Epidermal Growth

Epidermal growth factor (EGF)-like family members [amphiregulin (AREG), epiregulin (EREG), and betacellulin (BTC)] have been shown to be important regulators of cumulus–oocyte complex (COC) maturation, particularly cumulus expansion. The aim of this study was to determine the temporal expression patterns of mRNA encoding EGF-like growth factors in bovine cumulus cells (CC) during COC in vitro maturation and to assess the effects of grading doses of FSH on EGF-like mRNA expression in CC. Immature COC (grades 1 and 2) were obtained from 2- to 8-mm follicles from abattoir ovaries. In the first experiment, CC were separated from 20 COC and frozen before (immature group) or after COC culture for 4, 8, 12, 16, and 20 h with (10 ng mL–1) or without FSH. In the second experiment, pools containing 20 COC were matured for 12 h with grading doses of FSH (0, 0.1, 1, 10, and 100 ng mL–1). After culture, CC were mechanically separated and stored at –80°C. Total RNA was extracted using RNeasy® (Qiagen, Valencia, CA, USA), and 100 ng of RNA was reverse transcribed. Expression of target genes was assessed by real-time PCR and normalized by Cyclophilin (CYC-A). Relative quantification of mRNA abundance was determined by the Pfaffl equation. Effects of time of culture and FSH treatment were tested by ANOVA, and groups were compared by Tukey-Kramer honestly significant difference test. Nonparametric analysis was used when data were not normally distributed. Differences were considered significant when P < 0.05. In the presence of FSH, AREG and EREG mRNA abundance was increased at 4 h of culture, whereas in the absence of FSH, AREG but not EREG mRNA levels were increased by 4 h of culture. The addition of FSH stimulated AREG mRNA expression from 4 to 16 h of culture. In contrast, BTC mRNA was more expressed in immature CC, decreased after 4 h of culture with FSH, and did not vary during maturation in the absence of FSH. In the dose–response experiment, AREG and EREG mRNA expression was stimulated by FSH starting from 10 ng mL–1 and did not increase from 10 ng mL–1 to 100 ng mL–1. Again in contrast, BTC mRNA expression was inhibited by FSH at 100 ng mL–1. In conclusion, the present data suggest that FSH differently regulates the expression of EGF-like factors during bovine COC maturation, although AREG and EREG are stimulated, BTC is inhibited by FSH. This work was supported by FAPESP.

A central role for Fos in human B- and T-cell NFAT (nuclear factor of activated T cells): an acidic region is required for in vitro assembly

1994 ◽  
Vol 14 (10) ◽  
pp. 6886-6895
Author(s):  
N R Yaseen ◽  
J Park ◽  
T Kerppola ◽  
T Curran ◽  
S Sharma
Keyword(s):  
Amino Acids ◽  
T Cells ◽  
T Cell ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Nuclear Extracts ◽  
In Vitro Reconstitution ◽  
Direct Binding ◽  
Acidic Region

Nuclear factor of activated T cells (NFAT) is a multicomponent transcription factor that contains Fos and Jun family proteins in addition to a constitutively expressed factor(s). It is important for the production of interleukin 2 (IL-2) by T cells and is also expressed in B cells. Here we show that NFAT complexes in B- and T-cell nuclear extracts can be supershifted prominently with Fos antibodies and to a variable extent with Jun family protein antibodies. Fos and Jun proteins appear to participate in NFAT complexes as heterodimers, since efficient in vitro reconstitution of NFAT in unstimulated B- or T-cell nuclear extracts required both Fos and Jun. Using Fos and Jun deletion derivatives, we found that an acidic Fos region (amino acids 118 to 138), outside the DNA binding and dimerization domains, was necessary for the in vitro reconstitution of the NFAT complex in both B- and T-lymphocyte extracts although it was not required for binding to an AP-1 site. Fos-Jun heterodimers exhibited low-affinity direct binding to the NFAT site in the absence of nuclear extracts. This binding also required the Fos acidic region, amino acids 118 to 138. Mutating a variant AP-1 site in the NFAT oligonucleotide abolished both direct binding of Fos-Jun heterodimers and in vitro reconstitution of NFAT. These results demonstrate a central role of Fos in NFAT complex formation in both B and T lymphocytes and show that NFAT assembly involves direct binding of Fos-Jun heterodimers to a variant AP-1 site within the human NFAT recognition site.

Osmolarity determines the in vitro chondrogenic differentiation capacity of progenitor cells via nuclear factor of activated T-cells 5

Bone ◽  
2013 ◽  
Vol 53 (1) ◽  
pp. 94-102 ◽  
Author(s):  
Marjolein M.J. Caron ◽  
Anna E. van der Windt ◽  
Pieter J. Emans ◽  
Lodewijk W. van Rhijn ◽  
Holger Jahr ◽  
...  
Keyword(s):  
T Cells ◽  
Progenitor Cells ◽  
Chondrogenic Differentiation ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Differentiation Capacity

Berberine Suppresses RANKL-Induced Osteoclast Differentiation by Inhibiting c-Fos and NFATc1 Expression

2019 ◽  
Vol 47 (02) ◽  
pp. 439-455 ◽  
Author(s):  
Sang-Yong Han ◽  
Yun-Kyung Kim
Keyword(s):  
Bone Loss ◽  
Osteoclast Differentiation ◽  
Osteoclast Formation ◽  
Bone Diseases ◽  
Mrna Levels ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Murine Bone Marrow

Osteoporosis is a common disorder of bone remodeling, marked by excessive osteoclast formation. Recent studies indicated that berberine (BBR) is a potential natural drug for the treatment of various bone diseases. However, it still needs to be further studied for the treatment of osteoporosis. The current study investigated the inhibitory effects of BBR on receptor activator of nuclear factor-[Formula: see text]B ligand (RANKL)-induced osteoclast differentiation in vitro and in vivo. Cell-based assays were performed using osteoclasts generated in cultures of murine bone marrow-derived macrophages (BMMs) treated with RANKL and M-CSF. The effects of BBR on in vivo lipopolysaccharide (LPS)-mediated bone loss were evaluated using ICR mice. BBR significantly inhibited TRAP-positive osteoclast formation induced by RANKL. BBR also inhibited RANKL-induced Akt, p38 and ERK phosphorylation and I[Formula: see text]B degradation, and suppressed RANKL-induced expression of c-Fos and nuclear factor of activated T cells c1 (NFATc1), which is a key transcription factors for osteoclast formation. BBR reduced the mRNA levels of osteoclast markers, including TRAP, osteoclast-associated receptor (OSCAR), cathepsin K, and ATPase H[Formula: see text] transporting V0 subunit d2 (ATP6v0d2). Moreover, BBR prevented LPS-mediated bone loss in vivo. We suggest BBR as a natural compound that can be a potential therapeutic agent for osteoclast-related bone diseases.

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