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T Bartfai

Publications and source records attributed to T Bartfai.

At least 55 records · Page 3Linked to original sources
PubMed2000

Activation of neutral sphingomyelinase by IL-1beta requires the type 1 interleukin 1 receptor.

The cytokine interleukin 1beta (IL-1beta) plays an important role in host defence reactions and neuro-immune interactions but it is still not clear which of the two interleukin 1 receptor subtypes is coupled to activation of neutral sphingomyelinase (nSMase) by IL-1beta. To investigate involvement of neutral sphingomyelinase (nSMase) in central IL-1beta effects we used P(2)fractions of brain cerebral cortex from wild-type mice and mice deficient in the type 1 IL-1 receptor. IL-1beta (human, recombinant) was shown to activate, in a dose-dependent manner, nSMase in the P(2)brain fraction of the wild-type mice while in the knock-out mice the stimulatory effect of IL-1beta on nSMase was absent. In the presence of an IL-1 receptor antagonist (IL-1ra), IL-1beta did not activate nSMase either in the cortex of wild-type or knock-out mice. These data suggest that nSMase, a key enzyme of the sphingomyelin signal transduction pathway, might be involved in IL-1beta signalling in the brain and that activation of the enzyme requires the IL-1 receptor type 1.

Animals↗
PubMed2000

Renal cytokine responses in acute Escherichia coli pyelonephritis in IL-6-deficient mice.

The aim of this study was to investigate the influence of IL-6 on mortality, bacterial growth and cytokine expression in experimental acute pyelonephritis. Female IL-6-deficient mice and their wild-type counterparts, 8-10 weeks old, were infected with Escherichia coli CFT 073 or injected with NaCl 0.9% (w/v) via the urethra and thereafter obstructed for 6 h. Animals were killed at 48 h, 6 days or 8 weeks and cytokine and bacterial renal levels were assessed at each time point. We found that IL-6-deficient mice had increased mortality and extensive renal bacterial growth on day 6, compared with wild-type mice (P < 0.05) and the histopathological changes were generally more severe and widespread in the IL-6-deficient mice. Peak mRNA expression of IL-1beta, IL-4, IL-10, IL-12 and interferon-gamma (IFN-gamma) occurred 48 h after infection in both IL-6 knock out and wild-type mice. Transforming growth factor-beta (TGF-beta) levels also peaked at 48 h in E. coli-infected wild-type mice, while in the IL-6-deficient strain both TGF-beta mRNA and protein levels were significantly lower at 48 h than wild-type levels (P < 0.0008 and P < 0.03, respectively) and remained stationary throughout the study period. Animals injected with NaCl 0.9% (w/v) displayed a similar decrease in TGF-beta expression (P < 0.02). When splenocytes from the IL-6-deficient mice were incubated with murine recombinant IL-6, TGF-beta levels increased to those of wild-type mice. No increase was observed when splenocytes from wild-type mice were incubated with the same doses of rIL-6. We therefore conclude that IL-6 plays an important role in bacterial clearance and directly influences the TGF-beta levels in experimental acute pyelonephritis. We also demonstrate that urethral obstruction per se induces an increase in TGF-beta the magnitude of which is decreased in IL-6-deficient mice.

Animals↗
PubMed2000

Role of third intracellular loop of galanin receptor type 1 in signal transduction.

To determine the domains essential for G-protein coupling of the human galanin receptor type 1 (GalR1), we have used both GalR1 mutants and synthetic receptor-derived peptides in(125)I-galanin and [(35)S]-GTPgammaS binding studies. Replacement of potential phosphorylation sites by Leu in the third intracellular loop (IC3) of GalR1 did not affect K(D)values for the receptor. Peptides derived form the IC3 loop, and especially the N-terminal part of it were able to increase the rate of [(35)S]-GTPgammaS binding to the trimeric Gialpha1beta1gamma2, but not to Gsalphabeta1gamma2, whereas the peptides corresponding to the IC1 and IC2 loops had no such effect. IC3 loop peptides also inhibited the binding of(125)I-galanin to GalR1 in membranes from Rin m5F cells. Our results suggest that the IC3 loop of GalR1, especially its N-terminal part, defines the coupling of the receptor to the Gialpha1beta1gamma2 protein and consequently, to the signal transduction cascade.

Amino Acid Sequence↗
PubMed1999

Leukocyte recruitment in the cerebrospinal fluid of mice with experimental meningitis is inhibited by an antibody to junctional adhesion molecule (JAM).

The mechanisms that govern leukocyte transmigration through the endothelium are not yet fully defined. Junctional adhesion molecule (JAM) is a newly cloned member of the immunoglobulin superfamily which is selectively concentrated at tight junctions of endothelial and epithelial cells. A blocking monoclonal antibody (BV11 mAb) directed to JAM was able to inhibit monocyte transmigration through endothelial cells in in vitro and in vivo chemotaxis assays. In this study, we report that BV11 administration was able to attenuate cytokine-induced meningitis in mice. The intravenous injection of BV11 mAb significantly inhibited leukocyte accumulation in the cerebrospinal fluid and infiltration in the brain parenchyma. Blood-brain barrier permeability was also reduced by the mAb. We conclude that JAM may be a new target in limiting the inflammatory response that accompanies meningitis.

Animals↗
PubMed1999

Fever: links with an ancient receptor.

Recent advances have provided evidence that prostaglandin E2 mediates the generation of fever in response to interleukin-1 or lipopolysaccharide and have reinforced the similarities of signaling downstream of these two pyrogens.

Animals↗
PubMed1999

Intrathecal galanin alleviates allodynia-like behaviour in rats after partial peripheral nerve injury.

We have previously suggested that the neuropeptides galanin and galanin message-associated peptide (GMAP) may have an inhibitory role in spinal nociception. The present study examined the effects of intrathecal (i.t.) administration of these two peptides on allodynia-like behaviours in response to mechanical and cold stimulation in rats after photochemically induced ischaemic peripheral nerve injury. I.t. galanin significantly alleviated the mechanical- and cold-allodynia-like behaviours in nerve injured rats, and was not associated with motor impairment or sedation. I.t. GMAP relieved mechanical allodynia much less than galanin. I.t. M-35, a high-affinity galanin receptor antagonist, did not significantly alter the response of the rats to mechanical or cold stimulation. At 1 or 2 weeks postinjury, around 15% of dorsal root ganglion (DRG) neuron profiles showed galanin-like immunoreactivity. These profiles were mostly small sized. Although the number of galanin positive cells was thus increased in the DRG in the present model, the increase was substantially less than after complete sciatic nerve section, as previously shown. The present results showed that spinal administration of galanin inhibited some abnormal pain-like behaviours in rats after partial peripheral nerve injury. These results further support an inhibitory function for galanin in nociception. However, endogenous galanin may not play a significant role in suppressing nociceptive input after partial ischaemic peripheral nerve injury, as the upregulation of galanin is moderate.

Animals↗
PubMed1999

Acute-phase responses in transgenic mice with CNS overexpression of IL-1 receptor antagonist.

The interleukin-1 (IL-1) receptor antagonist (IL-1ra) is an endogenous antagonist that blocks the effects of the proinflammatory cytokines IL-1alpha and IL-1beta by occupying the type I IL-1 receptor. Here we describe transgenic mice with astrocyte-directed overexpression of the human secreted IL-1ra (hsIL-1ra) under the control of the murine glial fibrillary acidic protein (GFAP) promoter. Two GFAP-hsIL-1ra strains have been generated and characterized further: GILRA2 and GILRA4. These strains show a brain-specific expression of the hsIL-1ra at the mRNA and protein levels. The hsIL-1ra protein was approximated to approximately 50 ng/brain in cytosolic fractions of whole brain homogenates, with no differences between male and female mice or between the two strains. Furthermore, the protein is secreted, inasmuch as the concentration of hsIL-1ra in the cerebrospinal fluid was 13 (GILRA2) to 28 (GILRA4) times higher in the transgenic mice than in the control animals. To characterize the transgenic phenotype, GILRA mice and nontransgenic controls were injected with recombinant human IL-1beta (central injection) or lipopolysaccharide (LPS, peripheral injection). The febrile response elicited by IL-1beta (50 ng/mouse icv) was abolished in hsIL-1ra-overexpressing animals, suggesting that the central IL-1 receptors were occupied by antagonist. The peripheral LPS injection (25 micrograms/kg ip) triggered a fever in overexpressing and control animals. Moreover, no differences were found in LPS-induced (100 and 1,000 micrograms/kg ip; 1 and 6 h after injection) IL-1beta and IL-6 serum levels between GILRA and wild-type mice. On the basis of these results, we suggest that binding of central IL-1 to central IL-1 receptors is not important in LPS-induced fever or LPS-induced IL-1beta and IL-6 plasma levels.

Acute-Phase Reaction↗
PubMed1998

Mutagenesis study on human galanin receptor GalR1 reveals domains involved in ligand binding.

Many receptor mutants were generated and several NH2-terminally modified galanin analogs synthesized to define the regions of hGalR1 involved in galanin binding. Ligand binding properties and functionality of mutant receptors were evaluated. The His264Ala and Phe282Ala receptor mutants, although deficient in binding in the concentration range of galanin used, remained functional albeit at least 20-fold less efficient than the wild-type receptor in the inhibition of stimulated cAMP production. Hence, His264 and Phe282 of hGalR1 are directly involved in galanin binding. NH2-terminal carboxylic acid analogs of galanin (1-16) have a very low affinity for the wild-type receptor, but substantially increased affinity for the Glu271Lys-hGalR1, suggesting that the NH2-terminus of galanin binds to the receptor near the transmembrane (TM) VI. Based on these findings and computer-aided molecular modeling, we propose a binding site model for the hGalR1 receptor (possibly also for other galanin receptor subtypes): galanin binds with its NH2-terminus to the pocket between TM III and TM VI, Trp2 of galanin interacts with His264 of the receptor, and Tyr9 is involved in an aromatic-aromatic type of interaction with Phe282 of ECIII of GalR1.

Amino Acid Sequence↗
PubMed1998

NMR study of the conformation and localization of porcine galanin in SDS micelles. Comparison with an inactive analog and a galanin receptor antagonist.

Galanin is a 29/30-residue neuro-endocrine peptide which performs its many important physiological functions via a membrane-bound receptor. By using two-dimensional proton NMR spectroscopy, complete relaxation matrix analysis, and simulated annealing, the conformation of porcine galanin was determined in a membrane-mimicking solvent containing sodium dodecyl sulfate (SDS) micelles. The final family of calculated structures displays three well-defined beta- or gamma-turn regions, comprising residues 1-5, 7-10, and 24-27, but has otherwise a random conformation. The receptor-interacting N-terminal part, residues 1-5, was found to be best defined with a backbone RMSD value of 0.12 A. The mode of association between galanin and the SDS micelle was determined by observing the broadening effect on proton resonances, when spin-labeled 5- and 12-doxyl stearate molecules were added. It was concluded that galanin is located close to the surface of the micelle with two regions, residues 6-9 and 24-29, as well as two single residues, 18 and 21, reaching out into the aqueous solvent. Additional NMR studies were carried out on an inactive analogue, Ala2-galanin, and an antagonist M40. The results show that the proton resonances of galanin and M40 have identical chemical shifts in the N-terminal receptor-interacting region, indicating similar solution structures in this region. For Ala2-galanin, the same region displays a spectral heterogeneity with chemical shifts clearly different from the other two peptides, indicative of different secondary structures. These results may provide a structural background for the antagonist activity of M40 and the hormonal inactivity of Ala2-galanin, as compared to galanin.

Alanine↗
PubMed1998

Interleukin-1 receptor accessory protein interacts with the type II interleukin-1 receptor.

Stably transfected HEK-293 cells express on their surface the murine type II IL-1 receptor (mIL-1RII) as demonstrated by FACS analysis using the mAb 4E2, however binding of [125I]-hrIL-1beta to these cells is nearly absent. Saturable high affinity binding of [125I]-hrIL-1beta is observed when the murine IL-1 receptor accessory protein (mIL-1RAcP) is coexpressed with mIL-1RII. Binding of [125I]-hrIL-1beta to mIL-1RII-mIL-1RAcP complex can be inhibited either with antibodies to mIL-1RII (mAb 4E2), or by antibodies to mIL-1RAcP (mAb 4C5). The number of high affinity binding sites in cells stably transfected with the cDNA for mIL-1RII is dependent on the dose of cDNA for mIL-1RAcP used to transfect the cells. The high affinity complex between mIL-1RII and mIL-1RAcP is not preformed by interaction between the intracellular domains of these two transmembrane proteins, rather it appears to require the extracellular portions of mIL-1RII and mIL-1RAcP and the presence of a ligand. We suggest that in addition to its earlier described decoy receptor role, IL-1RII may modulate the responsiveness of cells to IL-1 by binding the IL-1RAcP in unproductive/non-signalling complexes and thus reducing the number of signalling IL-1RI-IL-1RAcP-agonist complexes when IL-1 is bound.

Animals↗
PubMed1998

Human/mouse interleukin-1 receptor/receptor accessory protein interactions in IL-1beta-induced NFkappaB activation.

We examined whether functional heterologous complexes between human IL-1RI (hIL-1RI) and murine IL-1R accessory protein (mIL-1RAcP) can be formed, utilizing human fibroblast HEK 293 cells and murine fibroblast C127 cells, nontransfected or stably transfected with hIL-1RI (C127-hIL-1RI), respectively. In non-transfected C127 cells, IL-1beta signalled through the mIL-1RI-mIL-1RAcP complex and activated NFkappaB p50/p65 heterodimers. In C127-hIL-1RI cells, IL-1beta signalled through the hIL-1RI and activated both p65/p65 and p50/p65 NFkappaB complexes, where only the activation of NFkappaB p65/p65 was dependent on mIL-1RAcP. Thus, clearly both homologous and heterologous IL-1RI-IL-1RAcP interactions support NFkappaB translocation, but with differences in signalling pattern.

Animals↗
PubMed1998

The interleukin-1 system: receptors, ligands, and ICE in the brain and their involvement in the fever response.

The ligands and the receptors of the interleukin 1 (IL-1) system constitute a highly inducible set of proteins whose expression in infection and inflammation is of key importance in the host defense. The IL-1 system participates in the stimulation of the immune system, the neuroendocrine system, and the neuroimmune system. The major soluble and secreted agonist of the system, IL-1 beta, has been studied by mutational and transgenic approaches. Furthermore, involvement of the signal-transducing type I IL-1 receptor (IL-1RI), in fever and other responses, has been studied by null mutation technique. We describe the inducible expression of the two agonists, IL-1 alpha (31 kDa and 17 kDa) and IL-1 beta (17 kDa) and of the IL-1 receptor subtypes IL-1RI and IL-1RII in the brain and in the adrenals (as well as in the pituitary cell line AtT20). We also describe an additional member of the IL-1 family: the IL-1 receptor antagonist (IL-1ra), an endogenous antagonist to IL-1 alpha and IL-1 beta. Furthermore, the IL-1 beta-converting enzyme (ICE) and its differential regulation and expression in brain and adrenals is also discussed. Fever is a systemic response to intraperitoneal (i.p.) or intracerebroventricular (i.c.v.) injection of IL-1 alpha or IL-1 beta. IL-1 beta-induced fever can be blocked by IL-1ra pretreatment. The fever response seems to be mediated via the IL-1RI as inferred from studies with receptor subtype-specific mutants of IL-1 beta and from studies in IL-1RI knock-out (IL-1RI KO) mice. IL-1 beta knock-out mice showed a hyperresponsive fever to both IL-1 agonists, IL-1 alpha and IL-1 beta, as well as to LPS.

Animals↗
PubMed1998

Interleukin 1 (IL-1) type I receptors mediate activation of rat hypothalamus-pituitary-adrenal axis and interleukin 6 production as shown by receptor type selective deletion mutants of IL-1beta.

The cytokine interleukin 1 (IL-1) plays an important role in the activation of the hypothalamus-pituary-adrenal (HPA)-axis and interleukin 6 (IL-6) production during infection or inflammation. Which of the interleukin-1 receptor types mediates these effects is not known. To investigate this issue a pharmacological approach was chosen by using recently developed IL-1 receptor type selective ligands. Rats were given one of various doses of recombinant human IL-1beta (rhIL-1beta; 1 and 10 microg/kg) and of several IL-1beta mutants (DeltaSND, DeltaQGE and DeltaI; 1, 10 and 100 microg/kg), that differ in their affinities for the IL-1 type I receptor but have similar affinities for the IL-1 type II receptor. One hour after intravenous administration of rhIL-1beta or IL-1beta mutants, plasma levels of ACTH, corticosterone (cort) and IL-6 were measured. Doses of 1 and 10 microg/kg rhIL-1beta markedly elevated plasma levels of ACTH, cort and IL-6. However, 10-100-fold higher doses of IL-1beta mutants DeltaSND and DeltaQGE and at least 100-fold higher doses of DeltaI have to be administered to increase plasma levels of ACTH, cort and IL-6. The potency differences correlate with their respective affinity for the type I receptor but not with that of the IL-1 type II receptor. It is concluded that IL-1beta induced ACTH, cort and IL-6 production is mediated by interleukin 1 type I receptors.

Adrenocorticotropic Hormone↗
PubMed1998

Identification of the spinal degradation products and inhibition of adenylate cyclase by recombinant rat galanin message-associated peptide.

In rat preprogalanin, galanin is C-terminally flanked by a 60 amino acid long peptide: galanin message-associated peptide (GMAP). GMAP sequences in different species show high degree of homology, suggesting a biological role. However, the study of the physiological and pharmacological actions of this peptide have been hampered by lack of availability of this large peptide, its fragments and well-characterized antibodies to GMAP. This study report the production of GMAP in Escherichia coli and the use of the recombinant peptide to define its degradation products in the spinal cord. The GMAP fragments formed upon incubation of GMAP with membranes of lumbar spinal cord were identified by sequencing and were also produced by solid phase synthesis for studies on second messenger systems. Furthermore, the study demonstrates that GMAP inhibits forskolin stimulated adenylate cyclase activity in a concentration dependent manner, while GMAP and its synthetic fragments did not affect cGMP level.

Adenylyl Cyclase Inhibitors↗
PubMed1998

Purification of a galanin degrading 70 kDa metallo-peptidase from bovine spinal cord.

Galanin is a 29/30 amino acids long neuroendocrine peptide, acting as an inhibitory modulator in the spinal cord. Several studies show that galanin is involved in control of pain threshold and acts synergistically with morphine, hence, inhibition of galanin degradation may be a pharmaceutical target for treatment of pain. In this study we have designed an 11 amino acids long substrate based on the first 10 N-terminal amino acids of galanin (this part contains the major pharmacophores of galanin), with a N-terminal fluorescent marker, anthranilic acid, and a C-terminal internal fluorescence quencher, 3-nitrotyrosine, coupled to the epsilon-amino group of the linker Lys11. Using this substrate to detect galanin degradation, we have purified a membrane bound galanin inactivating metallo-peptidase from bovine spinal cord. This enzyme, cleaving galanin between Trp2 and Thr3, is a novel 70 kDa, Zn2+ dependent metallo-peptidase.

Animals↗