Regular articleEvidence for exclusive role in signalling of tumour necrosis factor p55 receptor and a potentiating function of p75 receptor on human endothelial cells
Abstract
The current study was undertaken to investigate the role of TNF-R75 in regulation of E-selectin and ICAM-1 expression by TNF on HUVEC. To this end, we used agonistic anti-TNF-R75 antibodies, being mAb MR2-1 and polyclonal antibodies anti-TNF-R75 (pAb75). The agonistic properties of these antibodies were ascertained by the costimulatory capacity in a T-cell proliferation assay. These anti-TNF-R75 antibodies bound effectively to HUVEC, as evidenced in binding studies using125I-TNF, but they did not induce or enhance E-selectin or ICAM-1 expression as did agonistic anti-TNF-R55 antibodies. In contrast, both MR2-1 and pAb75 inhibited specifically TNF-induced E-selectin and ICAM-1 expression, but not activation by IL-1 or LPS. These results support the hypothesis, that in cells responding to TNF via the signalling pathway of the TNF-R55, the extracellular part of TNF-R75 captures TNF and delivers it to TNF-R55, resulting in an enhanced response to TNF.
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Inducible nitric oxide synthase (iNOS) has been shown to play an important role in the development of liver injury. iNOS deficiency protects mice from hemorrhage/resuscitation as well as from cytokine-mediated liver injury, for example, after administration of concanavalin A (con A). Here we investigated the in vivo effects of tumor necrosis factor (TNF)-α and/or interferon (IFN)-γ, two mediators of con A–induced liver injury, the TNF receptor (TNFR) usage leading to iNOS expression, and its connection with nuclear factor κB (NF-κB) activation. In conclusion, iNOS expression in vivo is dependent on both TNF-α and IFN-γ. Although con A–induced liver injury depends on both TNFR1 and TNFR2, TNF-dependent iNOS expression is mediated exclusively by TNFR1 and requires NF-κB activation. (HEPATOLOGY 2002;36:1061-1069.)
Differential activation of nuclear factor-κB by tumour necrosis factor receptor subtypes. TNFR1 predominates whereas TNFR2 activates transcription poorly
2002, FEBS LettersTumour necrosis factor-α (TNF-α) signals though two receptors, TNFR1 and TNFR2. TNFR1 has a role in cytotoxicity, whereas TNFR2 regulates death responses or proliferation. TNF activates pro-inflammatory transcription factor nuclear factor-κB (NF-κB) by uncertain signalling mechanisms. Here we report the contribution of each TNFR towards the NF-κB activation processes. In human cells expressing endogenous or exogenous TNFR2, in addition to TNFR1, we found both TNFRs capable of activating NF-κB, as measured by IκBα (inhibitor of NF-κB) degradation, electrophoretic mobility shift assay and NF-κB gene reporter assays. TNFR2 activation did not degrade IκBβ. However, TNF-effects on NF-κB activation occurred predominantly through TNFR1, with TNFR2 activating the transcription factor poorly.
TNF receptor subtype signalling: Differences and cellular consequences
2002, Cellular SignallingTumour necrosis factor-α (TNFα) is a multifunctional cytokine belonging to a family of ligands with an associated family of receptor proteins. The pleiotropic actions of TNF range from proliferative responses such as cell growth and differentiation, to inflammatory effects and the mediation of immune responses, to destructive cellular outcomes such as apoptotic and necrotic cell death mechanisms. Activated TNF receptors mediate the association of distinct adaptor proteins that regulate a variety of signalling processes including kinase or phosphatase activation, lipase stimulation, and protease induction. Moreover, the cytokine regulates the activities of transcription factors, heterotrimeric or monomeric G-proteins and calcium ion homeostasis in order to orchestrate its cellular functions. This review addresses the structural basis of TNF signalling, the pathways employed with their cellular consequences, and focuses on the specific role played by each of the two TNF receptor isotypes, TNFR1 and TNFR2.
Endothelial monocyte activating polypeptide-II (EMAP-II) is an inflammatory cytokine known to have a role in neutrophil and macrophage chemotaxis and in apoptosis. It is a tumour-derived cytokine that sensitizes tumour vasculature to the effects of systemic TNF. In order to gain insight into the mechanism by which EMAP-II sensitizes vessels to TNF, we focused on its effects on TNF receptor expression. In human umbilical vein endothelial cells (HUVEC), TNF-R1 mRNA is increased four-fold following incubation with recombinant EMAP-II. Conditioned media from cell lines known to produce high levels of EMAP-II upregulated TNF-R1 but not TNF-R2 by up to twenty-fold compared to media controls and low expressing cell lines; this effect was blocked by anti-EMAP-II antibody. Recombinant EMAP-II upregulated TNF-R1 expression by approximately six-fold. Analysis of HUVEC lysates by ELISA showed increased expression of TNF-R1 within 2 h; TNF-R2 expression was unaffected by recombinant EMAP-II. Finally, immunohistochemistry of human melanomas in vivo showed that TNF-R1 staining is increased on the vessels of tumours known to express high levels of EMAP-II compared to low EMAP-II expressing tumours. These results suggest that EMAP-II upregulates TNF-R1 expression by endothelial cells both in vitro and in vivo. This induction of TNF-R1 expression may be the mechanism by which EMAP-II sensitizes tumour endothelium to the effects of TNF leading to haemorrhagic necrosis.
Activation of Jun N-terminal kinase/stress-activated protein kinase pathway by tumor necrosis factor α leads to intercellular adhesion molecule- 1 expression
1999, Journal of Biological ChemistryTumor necrosis factor α (TNF-α) is a cytokine implicated in the pathogenesis of numerous chronic and acute inflammatory conditions. We have previously shown that mouse Sertoli cells respond to TNF-α by increasing interleukin-6 production and intercellular adhesion molecule-1 (ICAM-1) expression (1). In this cell type TNF-α activates the mitogen-activated protein kinase (MAPK) pathways p42/p44 MAPK, JNK/SAPK, and p38, the last of which is responsible for interleukin-6 production (2). To determine which MAPK signaling pathway is required for TNF-α induction of ICAM-1 expression, we have utilized the protein kinase inhibitor dimethylaminopurine, demonstrating that treatment of Sertoli cells with such compound significantly reduced ICAM-1 expression and JNK/SAPK activation. Moreover, dimethylaminopurine treatment increased the expression of MAPK phosphatase-2, providing a possible mechanism of action of this compound. By using agonist antibodies to p55 and to p75 TNF-α receptors and both human and mouse TNF-α, we demonstrate that both TNF receptors are expressed and that only the p55 receptor is involved in ICAM-1 expression. The p55 receptor activates all of the three pathways, whereas p75 failed to activate any of the MAPKs. Altogether our results demonstrate that TNF-α up-regulates ICAM-1 expression through the activation of the JNK/SAPK transduction pathway mediated by the p55 receptor.
Involvement of protein kinase C-β and ceramide in tumor necrosis factor-α-induced but not Fas-induced apoptosis of human myeloid leukemia cells
1999, Journal of Biological ChemistryThe role of protein kinase C-β (PKC-β) in apoptosis induced by tumor necrosis factor (TNF)-α and anti-Fas monoclonal antibody (mAb) in the human myeloid HL-60 leukemia cell line was studied by using its variant HL-525, which is deficient in PKC-β. In contrast to the parental HL-60 cells, HL-525 is resistant to TNF-α-induced apoptosis but sensitive to anti-Fas mAb-induced apoptosis. Both cell types expressed similar levels of the TNF-receptor I, whereas the Fas receptor was detected only in HL-525 cells. Transfecting the HL-525 cells with an expression vector containing PKC-β reestablished their susceptibility to TNF-α-induced apoptosis. The apoptotic effect of TNF-α in HL-60 and the transfectants was abrogated by fumonisin, an inhibitor of ceramide generation, and by the peptide Ac-YVAD-BoMK, an inhibitor of caspase-1 and -4. Supplementing HL-525 cells with exogenous ceramides bypassed the PKC-β deficiency and induced apoptosis, which was also restrained by the caspase-1 and -4 inhibitor. The apoptotic effect of anti-Fas mAb in HL-525 cells was abrogated by the antioxidantsN -acetylcysteine and glutathione and by the peptide z-DEVD-FMK, an inhibitor of caspase-3 and -7. We suggest that TNF-α-induced apoptosis involves PKC-β and then ceramide and, in turn, caspase-1 and/or -4, whereas anti-Fas mAb-induced apoptosis utilizes reactive oxygen intermediates and, in turn, caspase-3 and/or -7.