TNF Signaling in Peritoneal Mesothelial Cells: Pivotal Role of CflipL

2017 ◽  
Vol 37 (3) ◽  
pp. 250-258
Author(s):  
Willie M. Lüdemann ◽  
Danijela Heide ◽  
Lars Kihm ◽  
Martin Zeier ◽  
Peter Scheurich ◽  
...  
Keyword(s):  
Cell Surface ◽  
Signaling Pathway ◽  
Molecular Mechanisms ◽  
Mesothelial Cells ◽  
Cell Surface Expression ◽  
Peritoneal Membrane ◽  
Peritoneal Mesothelial Cells ◽  
Healthy Donors ◽  
Ultrafiltration Failure ◽  
Human Peritoneal Mesothelial Cells

BackgroundPeritoneal dialysis (PD) coincides with high concentrations of proinflammatory cytokines, such as tumor necrosis factor (TNF), in the peritoneal cavity. During treatment, chronic inflammatory processes lead to damage of the peritoneal membrane and a subsequent ultrafiltration failure. Human peritoneal mesothelial cells (HPMCs) play a central role as mediators and targets of PD-related inflammatory changes. Although TNF Receptor 1 (TNFR1) is expressed in high numbers on the cells, TNF-induced apoptosis is inhibited. Here, the underlying molecular mechanisms of TNFR1 signaling in HPMCs are investigated.MethodsHuman peritoneal mesothelial cells were isolated from the omentum of healthy donors and the dialysis solution of PD patients. Flow cytometry was applied to determine cell surface expression of TNFR1 on HPMCS from healthy donors in absence or presence of TNF or PD fluid (PDF) and were compared to TNFR1 expression on cells from PD patients. To investigate TNFR1- mediated signaling, HPMCs were treated with PDF or TNF, and expression patterns of proteins involved in the TNFR1 signaling pathway were assessed by western blot.ResultsIncubation with PDF led to a significant up-regulation of TNFR1 on the cell surface correlating with elevated TNFR1 numbers on HPMCs from PD patients. Investigations of under-lying molecular mechanisms of TNFR1 signaling showed that PDF affects TNFR1 signaling at the proapoptotic signaling pathway by upregulation of IκBα and downregulation of cFLIPL. In contrast, TNF exclusively induces the activation of NFκB by an increase of phosphorylated IκBα.ConclusionsNovel and relevant insights into the mechanisms of TNFR1-mediated signaling in HPMCs with an impact on our understanding of PD-associated damage of the peritoneal membrane are shown.

Expression of Aquaporin-1 in Human Peritoneal Mesothelial Cells and Its Upregulation by GlucoseIn Vitro

2001 ◽  
Vol 12 (5) ◽  
pp. 1036-1045 ◽  
Author(s):  
KAR NENG LAI ◽  
FU KEUNG LI ◽  
HAO YUI LAN ◽  
SYDNEY TANG ◽  
ANITA W. L. TSANG ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Peritoneal Dialysis ◽  
Mesothelial Cells ◽  
Dependent Manner ◽  
Peritoneal Membrane ◽  
Dialysis Adequacy ◽  
Peritoneal Mesothelial Cells ◽  
Aqp1 Expression ◽  
Human Peritoneal Mesothelial Cells ◽  
Osmotic Agents

Abstract. Aquaporin (AQP) is a family of water channels that are highly selective for the passage of water and occasionally glycerol. In previous studies, only AQP1 was found in human peritoneal endothelial cells in both control subjects and patients on peritoneal dialysis. As human peritoneal mesothelial cells (HPMC) play an important role in dialysis adequacy and fluid balance in continuous ambulatory peritoneal dialysis patients, this study examined whether AQP1 is present in HPMC. It was found that AQP1 mRNA and protein are present in HPMC constitutively. The localization of AQP1 protein in peritoneal mesothelial cells was confirmed by double immunohistochemical staining of the mesothelial lining of human peritoneal membrane. More important, the expression of AQP1 in HPMC is not constitutive and the transcription and biosynthesis of AQP1 in HPMC is inducible by osmotic agents such as glucose and mannitol. There was significant enhancement of AQP1 biosynthesis upon exposure to glucose in a time- and dose-dependent manner (P< 0.0001). Similar findings were observed in the AQP1 biosynthesis by an endothelial cell line, EA.hy 926. Of particular interest, the upregulation in AQP1 mRNA or biosynthesis in mesothelial cells was always significantly higher than that of endothelial cells when the experiments were conducted under identical settings (P< 0.001). AQP1 expression in HPMC was demonstrated for the first time. Osmotic agents upregulate both mRNA and protein expression of this aquaporin. The role of AQP1 in HPMC in maintaining the ultrafiltration of the peritoneal membrane is potentially of clinical interest.

Production and Regulation of Matrix Metalloproteinases and Their Inhibitors by Human Peritoneal Mesothelial Cells

2000 ◽  
Vol 20 (5) ◽  
pp. 524-533 ◽  
Author(s):  
John Martin ◽  
Susan Yung ◽  
Rachel L. Robson ◽  
Robert Steadman ◽  
Malcolm Davies
Keyword(s):  
Matrix Metalloproteinases ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Mesothelial Cells ◽  
Tissue Inhibitors Of Metalloproteinases ◽  
Apical Surface ◽  
Peritoneal Membrane ◽  
Peritoneal Mesothelial Cells ◽  
The Matrix ◽  
Human Peritoneal Mesothelial Cells

Objective Human peritoneal mesothelial cells (HPMC) are likely to be involved in maintenance of the peritoneal membrane. We determined whether these cells were able to synthesize the matrix degrading enzymes, matrix metalloproteinases (MMPs), likely to be responsible for the breakdown of this membrane, and whether this secretion could be modulated by cytokines involved in the inflammatory response. Design MMP activity in conditioned medium of growth-arrested HPMC was measured by zymography. Cultures were incubated in the presence and absence of the cytokines transforming growth factor-beta (TGFβ) and interleukin (IL)-1β in order to determine the effects of these cytokines on this process. The mRNA for these MMPs, together with that of their specific inhibitors, tissue inhibitors of metalloproteinases (TIMPs), was also examined by reverse transcriptase polymerase chain reaction (RT-PCR). Results HPMC were shown to constitutively secrete the metalloproteinases MMP-2 and MMP-3 in vitro. In response to the proinflammatory cytokine IL-1β, the protein and mRNA for MMP-9 was induced, while secretion of MMP-2 was unaltered. Similarly, the mRNA for MMP-3 was also increased relative to actin following the addition of IL-1β. TGFβ was shown to slightly induce the secretion of MMP-2 together with the mRNA for TIMP I, TIMP II, and, to a greater extent, TIMP III. Used peritoneal dialysate was also shown to induce MMP-9 secretion, and this effect was blocked by the co-incubation of IL-1 receptor antagonist. The secretion of enzyme activity was shown to be from the apical surface of the cells. Conclusion HPMC have the ability to control the accumulation of extracellular matrix by secreting the matrix degrading molecules MMP-2, MMP-3, and MMP-9. In addition, the secretion of these enzymes, together with that of their inhibitors (TIMPs) is regulated by the cytokines IL-1β and TGFβ. This process is likely to be important in both the normal maintenance of the integrity of the peritoneal membrane and in the changes that occur following prolonged peritoneal dialysis.

scholarly journals Activation of General Control Nonderepressible-2 Kinase Ameliorates Glucotoxicity in Human Peritoneal Mesothelial Cells, Preserves Their Integrity, and Prevents Mesothelial to Mesenchymal Transition

Biomolecules ◽  
2019 ◽  
Vol 9 (12) ◽  
pp. 832
Author(s):  
Theodoros Eleftheriadis ◽  
Georgios Pissas ◽  
Georgia Antoniadi ◽  
Evdokia Nikolaou ◽  
Spyridon Golfinopoulos ◽  
...  
Keyword(s):  
High Glucose ◽  
Cell Apoptosis ◽  
Glucose Transporter ◽  
Mesothelial Cells ◽  
General Control ◽  
Mesenchymal Transition ◽  
Peritoneal Mesothelial Cells ◽  
Ultrafiltration Failure ◽  
Human Peritoneal Mesothelial Cells ◽  
Tgf Β1

Along with infections, ultrafiltration failure due to the toxicity of glucose-containing peritoneal dialysis (PD) solutions is the Achilles’ heel of PD method. Triggered by the protective effect of general control nonderepressible-2 (GCN-2) kinase activation against high-glucose conditions in other cell types, we evaluated whether the same occurs in human peritoneal mesothelial cells. We activated GCN-2 kinase with halofuginone or tryptophanol, and assessed the impact of this intervention on glucose transporter-1, glucose transporter-3, and sodium-glucose cotransporter-1, glucose influx, reactive oxygen species (ROS), and the events that result in glucotoxicity. These involve the inhibition of glyceraldehyde 3-phosphate dehydrogenase and the diversion of upstream glycolytic products to the aldose pathway (assessed by D-sorbitol), the lipid synthesis pathway (assessed by protein kinase C activity), the hexosamine pathway (determined by O-linked β-N-acetyl glucosamine-modified proteins), and the advanced glycation end products generation pathway (assessed by methylglyoxal). Then, we examined the production of the profibrotic transforming growth factor-β1 (TGF-β1), the pro-inflammatory interleukin-8 (IL-8). Cell apoptosis was assessed by cleaved caspase-3, and mesothelial to mesenchymal transition (MMT) was evaluated by α-smooth muscle actin protein. High-glucose conditions increased glucose transporters, glucose influx, ROS, all the high-glucose-induced harmful pathways, TGF-β1 and IL-8, cell apoptosis, and MMT. Halofuginone and tryptophanol inhibited all of the above high glucose-induced alterations, indicating that activation of GCN-2 kinase ameliorates glucotoxicity in human peritoneal mesothelial cells, preserves their integrity, and prevents MMT. Whether such a strategy could be applied in the clinic to avoid ultrafiltration failure in PD patients remains to be investigated.

Klotho is a novel therapeutic target in peritoneal fibrosis via Wnt signaling inhibition

2020 ◽  
Vol 35 (5) ◽  
pp. 773-781
Author(s):  
Hiroyuki Kadoya ◽  
Minoru Satoh ◽  
Yuko Nishi ◽  
Megumi Kondo ◽  
Yoshihisa Wada ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Therapeutic Target ◽  
Molecular Mechanisms ◽  
Elongation Factor ◽  
Mesothelial Cells ◽  
Peritoneal Fibrosis ◽  
Peritoneal Mesothelial Cells ◽  
Pathway Activation ◽  
Klotho Protein

Abstract Background Long-term exposure to bioincompatible peritoneal dialysate causes the loss of mesothelial cells and accumulation of matrix proteins, leading to an increase in the thickness of the submesothelial layer, thereby limiting the long-term effectiveness of peritoneal dialysis (PD). However, the detailed molecular mechanisms underlying the process of peritoneal fibrosis have not been clearly elucidated. Wnt/β-catenin signaling pathway activation has been suggested to play a pivotal role in the development of organ fibrosis. Moreover, Klotho protein can regulate Wnt/β-catenin signaling. We examined the role of Klotho protein in reducing peritoneal fibrosis by inhibiting Wnt/β-catenin signaling. Methods The β-catenin-activated transgenic (BAT) driving expression of nuclear β-galactosidase reporter transgenic (BAT-LacZ) mice, the alpha-Klotho gene under control of human elongation factor 1 alpha promoter [Klotho transgenic (KLTG) and C57BL/6 background] and C57BL/6 mice [wild-type (WT)] were used. The mice received daily intraperitoneal (i.p.) injections of 4.25% glucose with lactate (PD solution) or saline as a control for 4 weeks. Other mice received daily i.p. injections of the same volume of saline (normal control). Results After exposure to PD, Wnt signal activation was observed on the peritoneal mesothelial cells in WT-PD mice. The peritoneal fibrosis was also accelerated in WT-PD mice. The protein expression of β-catenin and Wnt-inducible genes were also remarkably increased in WT-PD mice. On the other hand, KLTG-PD mice attenuated activation of Wnt/β-catenin signaling after exposure to PD and ameliorated the progression of peritoneal fibrosis. Conclusions Overexpression of Klotho protein protects the peritoneal membrane through attenuation of the Wnt/β-catenin signaling pathway. The availability of recombinant Klotho protein would provide a novel potential therapeutic target in peritoneal fibrosis.

scholarly journals LncRNA GAS5 Competitively Combined With miR-21 Regulates PTEN and Influences EMT of Peritoneal Mesothelial Cells via Wnt/β-Catenin Signaling Pathway

2021 ◽  
Vol 12 ◽  
Author(s):  
Yi Fan ◽  
Xingxu Zhao ◽  
Jianfei Ma ◽  
Lina Yang
Keyword(s):  
Signaling Pathway ◽  
Molecular Mechanisms ◽  
Epithelial Mesenchymal Transition ◽  
Specific Binding ◽  
Mesothelial Cells ◽  
Pten Expression ◽  
Pten Gene ◽  
Mesenchymal Transition ◽  
Peritoneal Mesothelial Cells ◽  
Lncrna Gas5

ObjectiveEpithelial-mesenchymal transition (EMT) is an important factor leading to peritoneal fibrosis (PF) in end-stage renal disease (ESRD) patients. The current research aimed to evaluate the effect of long non-coding RNA growth arrest-specific 5 (lncRNA GAS5) in human peritoneal mesothelial cells (HPMCs) EMT and explore the potential molecular mechanisms.Materials and MethodsHPMCs were cultured under control conditions or with high glucose (HG). The cells were then treated with lncRNA GAS5, lncRNA GAS5 siRNA, with or without miR-21 inhibitor and PTEN transfection. Expression of lncRNA GAS5, miR-21, α-SMA, Vimentin, E-cadherin, phosphatase and tensin homolog deleted on chromosome ten (PTEN), Wnt3a, and β-catenin were measured by real time PCR and Western blotting. Bioinformatics analyses were used to test the specific binding sites between the 3′ UTR of the PTEN gene, miR-21, and lncRNA GAS5. Rescue experiments were performed to confirm the lncRNA GAS5/miR-21/PTEN axis in HPMC EMT.ResultsWe found that HG-induced EMT decreased lncRNA GAS5 and that overexpression of lncRNA GAS5 can attenuate EMT in HPMCs. In addition, lncRNA GAS5 regulated HG-induced EMT through miR-21/PTEN. Cotransfection of miR-21 inhibitors remarkably increased PTEN expression and attenuated EMT in lncRNA GAS5 knockdown HPMCs. Moreover, rescue experiments showed that overexpression of PTEN attenuated the EMT effects of lncRNA GAS5 siRNA in HPMCs. We also confirmed that the Wnt/β-catenin pathway was stimulated in lncRNA GAS5/miR-21/PTEN-mediated EMT.ConclusionOur research showed that lncRNA GAS5 competitively combined with miR-21 to regulate PTEN expression and influence EMT of HPMCs via the Wnt/β-catenin signaling pathway. This study provides novel evidence that lncRNA GAS5 may be a potential therapeutic target for HPMC EMT.

scholarly journals Physical dissociation of the TCR-CD3 complex accompanies receptor ligation.

1995 ◽  
Vol 182 (6) ◽  
pp. 1997-2006 ◽  
Author(s):  
H Kishimoto ◽  
R T Kubo ◽  
H Yorifuji ◽  
T Nakayama ◽  
Y Asano ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Cell Surface ◽  
Molecular Mechanisms ◽  
Signal Transduction Pathway ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Protein Levels ◽  
Before And After ◽  
Physical Dissociation ◽  
Receptor Ligation

Recent studies indicate that there may be functional uncoupling of the TCR-CD3 complex and suggest that the TCR-CD3 complex is composed of two parallel signal-transducing units, one made of gamma delta epsilon chains and the other of zeta chains. To elucidate the molecular mechanisms that may explain the functional uncoupling of TCR and CD3, we have analyzed their expression by using flow cytometry as well as immunochemical means both before and after stimulation with anti-TCR-beta, anti-CD3 epsilon, anti-CD2, staphylococcal enterotoxin B, and ionomycin. We present evidence that TCR physically dissociates from CD3 after stimulation of the TCR-CD3 complex. Stimulation with anti-CD3 resulted in down-modulation of TCR within 45 min whereas CD3 epsilon was still expressed on the cell surface as detected by flow cytometry. However, the cell surface expression of TCR and CD3 was not affected when cells were stimulated with anti-TCR-beta under the same conditions. In the case of anti-CD3 treatment of T cells, the TCR down-modulation appeared to be due to the internalization of TCR, as determined by immunoelectron microscopy. Immunochemical analysis of cells after stimulation with either anti-TCR or anti-CD3 mAbs revealed that the overall protein levels of TCR and CD3 were similar. More interestingly, the dissociation of the TCR-CD3 complex was observed with both treatments and occurred in a manner that the TCR and the associated TCR-zeta chain dissociated as a unit from CD3. These results provide the first report of physical dissociation of TCR and CD3 after stimulation through the TCR-CD3 complex. The results also suggest that the signal transduction pathway triggered by TCR may differ from that induced by CD3.

Sign in / Sign up

Export Citation Format

Share Document