The catecholamine cytokine balance: interaction between the brain and the immune system.

Cytokines are involved both in various immune reactions and in controlling certain events in the central nervous system (CNS). In our earlier studies, it was shown that monoamine neurotransmitters, released in stress situations, represent a tonic sympathetic control on cytokine production and on the balance of proinflammatory/anti-inflammatory cytokines. Basic and clinical studies have provided evidence that the biophase level of monoamines, determined by the balance of their release and uptake, is involved in the pathophysiology and treatment of depression, while inflammatory mediators might also have a role in its etiology. In this work, we studied the role of changes in norepinephrine (NE) level on the lipopolysaccharide (LPS) evoked tumor necrosis factor (TNF)-alpha and interleukin (IL)-10 response both in the plasma and in the hippocampus of mice. We demonstrated that the LPS induced TNF-alpha response is in direct correlation with the biophase level of NE, as it is significantly higher when the release of NE of vesicular origin was completely inhibited in an animal model of depression (reserpine treatment) and it is significantly lower in the case of increasing biophase levels of NE by genetic (NET-KO) or chemical (desipramine) disruption of NE reuptake. IL-10 was changed inversely to TNF-alpha levels only in the desipramine-treated animals. Our results showed that depression is related both to changes in peripheral and in hippocampal inflammatory cytokine production and to monoamine neurotransmitter levels. Since several anti-inflammatory drugs also have antidepressant effects, we hypothesized that antidepressants are also able to modulate the LPS-induced inflammatory response, which might contribute to their antidepressant effect.

Platelet-activating factor evokes Ca2+ transients after the blockade of ryanodine receptor by dantrolene in RAW 264.7 macrophages.

In the present study we studied platelet-activating factor (PAF)-, and ATP-induced increases in intracellular Ca2+ concentration ([Ca2+]i) using RAW 264.7 macrophages filled with fura-2/AM and imaged with fluorescence video microscopy. We found that the prevalence of detectable [Ca2+]i responses to PAF application was significantly higher in the presence of dantrolene. Dantrolene itself significantly decreased basal [Ca2+]i of macrophages compared to control cases after a 20-min incubation period. In the dantrolene-treated cells even the peak [Ca2+]i in response to PAF (as an average of all cells) was below the baseline of control suggesting that decreased [Ca2+]i plays a permissive role in the Ca2+ rise induced by PAF in macrophages. In contrast to the effect of PAF, neither the amplitude of response to ATP nor the frequency of responding cells changed significantly during dantrolene treatment in our experiments. These cells were able to respond to a standard immune stimulus as well: lipopolysaccharide (LPS) was able to increase [Ca2+]i. Our data indicate that the effectiveness of PAF to increase [Ca2+]i in RAW 264.7 macrophages depends on the resting [Ca2+]i. It has also been shown in this study that PAF and ATP differently regulate Ca2+ homeostasis in macrophages during inflammatory response and therefore they possibly differently modulate cytokine production by macrophages.

Enhanced tumor necrosis factor-alpha-specific and decreased interleukin-10-specific immune responses to LPS during the third trimester of pregnancy in mice.

It is increasingly apparent that there is a bidirectional interaction between the maternal immune system and the reproductive system during pregnancy. Pregnancy is associated with a suppression of maternal specific immune responses, which process underlies the protection of fetal tissues expressing paternally inherited alloantigens. However, recent evidence indicates that the suppression of specific, lymphocyte-mediated immune responses during pregnancy is accompanied by activation of the non-specific arm of the maternal immune response. In the present study, we have investigated the effect of pregnancy on the non-specific immune response induced by bacterial lipopolysaccharide (LPS, endotoxin) in mice. Pregnancy enhanced the LPS-induced production of proinflammatory cytokines, including tumor necrosis factor-alpha, interleukin (IL)-6, and interferon-gamma. On the other hand, LPS-induced levels of the anti-inflammatory cytokine IL-10 were suppressed in pregnant mice. These alterations in cytokine production correlated with an increased susceptibility for endotoxemic mortality in the pregnant mice. Although adrenergic receptors are important regulators of cytokine production in non-pregnant mice, the alpha(2)- and the beta-adrenoceptor-mediated modulation of cytokine production ceases to operate during pregnancy associated with severe endotoxemia. These data may explain how excessive activation of the non-specific immune responses during pregnancy can contribute to the increased severity of some maternal diseases, including septic shock, and can be an important pathophysiological factor in disseminated intravascular coagulation or preeclampsia.

Cytokines and the central nervous system.

Cytokines are involved both in the immune response and in controlling various events in the central nervous system, that is, they are equally immunoregulators and modulators of neural functions and neuronal survival. On the other hand, cytokine production is under the tonic control of the peripheral and the central nervous system and the cytokine balance can be modulated by the action of neurotransmitters released from nonsynaptic varicosities [131]. The neuroimmune interactions are therefore bidirectional-cytokines and other products of the immune cells can modulate the action, differentiation, and survival of neuronal cells, while the neurotransmitter and neuropeptide release play a pivotal role in influencing the immune response. Cytokines and their receptors are constitutively expressed by and act on neurons in the central nervous system, in both its normal and its pathological state, but cytokine overexpression in the brain is an important factor in the pathogenesis of neurotoxic and neurodegenerative disorders. Accordingly, it can be accepted that the peripheral and central cytokine compartments appear to be integrated, and their effects might synergize or inhibit each other; however, it should always be taken into account that they are spatiotemporally differentially regulated. New concepts are reviewed in the regulation of relations between cytokine balance and neurodegeneration, including intracellular receptor-receptor, cell-cell, and systemic neuroimmune interactions that promote the further elucidation of the complexities and cascade of the possible interactions between cytokines and the central nervous system.

Synthesis and biological activity of the structural analogues of (-)-cabenegrin A-I.

A series of phenylbutene and butanol derivatives (6a-j, 12, 13, 15, 17, 24b,c, 26, 27a,b) were prepared from the readily available resorcinol derivatives 2a-f and 7-hydroxy-chroman (18). The products were tested for inhibitory activity on the LPS-induced TNF-alpha production in the plasma in comparison with that of cabenegrin A-I (1a).

Streptozotocin-induced diabetes alters the oligomerization pattern of acetylcholinesterase in rat skeletal muscle.

AIMS/HYPOTHESIS: Diabetes mellitus has a serious effect on most of the properties of skeletal muscles. Changes in neuromuscular transmission are also involved in propagating the disease. METHODS: In our experiments, acetylcholinesterase was extracted from the fast extensor digitorum longus and slow soleus muscles of control, non-treated 6-week-diabetic and insulin-treated diabetic rats. The extracts were applied to velocity sedimentation and acetylcholinesterase activity was determined. RESULTS: We observed considerable differences in the distribution of individual acetylcholinesterase molecular forms in diabetic fast muscles. This included a 59% decline in G4 content together with a fivefold increase in A8 and a 53 % increase in A12 activity resulting in a shift of acetylcholinesterase profile characteristically towards slow muscles. These alterations were partly reversed by insulin treatment. CONCLUSION/INTERPRETATION: In slow muscles diabetes caused an increase in G4 activity without affecting the sedimentation profile. Decline in G4 content in fast muscles could contribute to enhanced desensitization of acetylcholine receptors in diabetes.

Opposite role of alpha2- and beta-adrenoceptors in the modulation of interleukin-10 production in endotoxaemic mice.

Our aim was to investigate the role of adrenoceptors in the modulation of in vivo interleukin-10 (IL-10) production in lipopolysaccharide (LPS)-treated mice. The effect of different adrenergic drugs on plasma concentration of IL-10 was measured by ELISA 90 min after LPS injection. Our results confirmed the involvement of beta-adrenoceptors since the beta-agonist isoproterenol significantly increased the IL-10 production in response to LPS stimulation, whereas the beta-antagonists propranolol decreased it. In contrast, the alpha2-agonists UK-14304, clonidine and xylazine significantly decreased the IL-10 plasma level, whereas the alpha2-antagonists CH-38083, prazosine and WB-4101 increased it. Our results provide the first in vivo evidence that, in addition to beta-adrenoceptors; alpha-adrenoceptors play also a very important role in the regulation of IL-10 production under endotoxaemic conditions.

Differential involvement of sympathetic nervous system and immune system in the modulation of TNF-alpha production by alpha2- and beta-adrenoceptors in mice.

In the present study, the regulation of tumor necrosis factor-alpha (TNF-alpha) production by alpha2- and beta-adrenoceptors located on noradrenergic nerve terminals and on macrophages was studied in endotoxaemic mice. We found that reduction of the sympathetic outflow by reserpine dramatically increased the lipopolysaccharide (LPS)-induced TNF-alpha production, demonstrating that the release of endogenous noradrenaline (NA), controlled by presynaptic alpha2-adrenoceptors, was a determinant factor in this model. By using alpha2- and beta-adrenergic drugs (clonidine, CH-38083, isoproterenol, propranolol) we provided the first in vivo evidence that, beside the dominance of neuronal alpha2- and macrophage beta-adrenoceptors, the alpha2-adrenoceptors on macrophages were also involved in the modulation of LPS-induced TNF-alpha production. Since adrenergic drugs are widely used in the clinical practice, our findings may have therapeutical implications.

Contribution of differently localized alpha 2- and beta-adrenoceptors in the modulation of TNF-alpha and IL-10 production in endotoxemic mice.

Evidence is presented that the immune response to endotoxemia is under tonic control of the sympathetic nervous system. Adrenergic agents may influence the immune response both directly through alpha- and beta-adrenergic receptors expressed by immunologically competent cells and indirectly via alteration of the endogenous NA level by influencing the activity of release-regulating presynaptic alpha 2-adrenoceptors located on the sympathetic nerve terminals. In the immunomodulatory effect of NA/adrenergic drugs, their action on beta-adrenoceptors was dominant, but the considerable role of alpha-adrenoceptors on macrophages was also demonstrated. According to our findings, regulation of the ascending wing of the inflammatory response, that is, TNF-alpha production, is more sensitive to the adrenoceptor effect, whereas modulation of its deregulation by IL-10 production also involves some other determining factors.

The polymorphic human TLX-B/CD46/MCP system and its implications in transplantation and reproduction.

TLX antigens have been found on most peripheral blood cells, trophoblasts, seminal vesicle cells and sperms. These antigens seem to be associated with the membrane cofactor protein (MCP) and the CD46 antigen. Alloantibodies to TLX antigens with Fc tau RII-blocking features were obtained by transfusion of leucocytes or platelets. Preliminary population studies revealed that alloantibodies to TLX/CD46/MCP recognize four overlapping specificities. The terminology TLX-B was introduced with specificities TLX-B1, B2, B3, B4 and frequencies obtained in the population were: 38%, 46%, 42% and 26%, respectively. Family studies showed an independent segregation of the TLX and HLA alleles. At the cellular protein on trophoblast, the alloantibody detected a glycoprotein of 66-67 kDa molecular mass, which may correspond to the alpha chain of the TLX/CD46/MCP isotypes. A direct association of the alloantibody with Fc tau RII could be excluded thus its FcR blocking feature is probably based on an indirect functional effect. After transfusion and in pregnancy the induction of TLX alloantibody production depended on the mismatching in the TLX/CD46/MCP phenotypes. Probable associations were revealed in the case of recurrent habitual abortion between the lack of Fc tau R blocking antibody production and the matched TLX specificities of the couples. After transfusion, TLX alloantibody production with Fc tau R and MLR blocking function was induced only when the recipient was lacking the TLX specificities expressed on the donor cells. Suppression of MLR was found only when TLX specificity in sera corresponded to the TLX specificity of the effector cell. The immunopathological importance of these findings in transplantation and reproductive medicine has yet to be clarified.

Differences in the isoelectric focusing patterns of serum and cerebrospinal fluid transthyretin.

Transthyretin isolated by polyacrylamide gel electrophoresis from human serum and cerebrospinal fluid, dissociated into its subunits, was subjected to isoelectric focusing in polyacrylamide gels containing 8 mole/L urea. The isoelectric focusing multi-component patterns of serum and cerebrospinal fluid transthyretin differ in a characteristic way, having only one main protein zone in common. Double diffusion immunotest and immunoblotting revealed the immunological identity of serum and cerebrospinal fluid transthyretin and of the main components separated by isoelectric focusing. The different isoelectric focusing zones can be consistently explained when they are ascribed to structurally identical transthyretin subunits associated with different ligands specifically occurring in either serum or cerebrospinal fluid. Only the protein zone located at the same pI in serum and cerebrospinal fluid transthyretin patterns may be assigned to ligand-free subunits. Thus, the typical differences in isoelectric focusing patterns may point to different carrier functions of transthyretin in serum and cerebrospinal fluid.

Molecular characterization of beta-thalassemia in Hungary.

We have identified seven different beta-thalassemia mutations and one delta beta-thalassemia determinant (the Sicilian type) in 32 members of 17 Hungarian families. The most common mutation is the IVS-I-1 (G-->A) change; its high frequency is comparable to that observed in neighboring Czechoslovakia. Additional mutations are of Mediterranean origin. One rare mutation (initiation codon ATG-->GTG) was identified as an independent mutation because of the absence of known polymorphisms in the beta-globin gene. One new frameshift at codon 51 (-C) was observed in a single individual; hematological data were as expected for a beta zero-thalassemia heterozygosity.

[Study of the origin of hyperglycemia in acute myocardial infarct]. / A heveny myocardialis infarctust kísérö hyperglykaemia eredetének vizsgálata.

In order to clarify the origin of hyperglycaemia, blood glucose, glycated haemoglobin (GHb) and protein-corrected serum fructosamine (SFA) values were simultaneously determined at admission of 65 patients with acute myocardial infarction while oral glucose tolerance test was performed later at discharge. In 29 patients no alterations in carbohydrate metabolism were found (blood glucose: 5.2 +/- 0.1 mmol/l, GHb: 4.4 +/- 0.1%, SFA: 2.20 +/- 0.08 mmol/l) while in 9 patients diabetes was already recorded in the medical history (blood glucose: 11.5 +/- 1.1 mmol/l, GHb: 7.9 +/- 0.9%, SFA: 3.36 +/- 0.31 mmol/l, p < 0.001). Undiagnosed diabetes was documented in 8 patients (blood glucose: 11.8 +/- 1.3 mmol/l, GHb: 7.3 +/- 0.6%, SFA: 3.51 +/- 0.24 mmol/l) while stress-hyperglycaemia was found in 19 patients (blood glucose: 8.4 +/- 0.3 mmol/l, GHb: 4.5 +/- 0.1%, SFA: 2.55 +/- 0.17 mmol/l). Undiagnosed diabetes could be recorded in one seventh while stress-hyperglycaemia could be found in one third of non-diabetic patients with acute myocardial infarction. Due to overlapping values SFA is not suitable to distinguish between stress-hyperglycaemia and undiagnosed diabetes in patients with acute myocardial infarction.

[Genetic traits in the area of Bodrogköz]. / Genetikai jellegek vizsgálata a Bodrogközben.

In 1984 a late malaria endemic area, called Bodrogköz was studied. This was a reexamination of the population genetic work performed by Walter, Nemeskéri. In six villages of Bodrogköz 328 persons were tested for AB0, Rh blood groups, haptoglobins, haemoglobin concentration, haematocrit, erythrocyte amount, the MCV, the MCH and the G-6-PD were analyzed. The quantitative determination of HbF and HbA2, red cell osmotic resistance and thalassemia were measured as well. Thalassemia heterozygote carriers and an increased level of HbF were revealed. The frequency of G-6-PD deficiency was 0.39%. In Bodrogköz the frequencies of AB0, Rh and haptoglobin types were similar in the present and all previous studies. The background of this similarity might be the genetic similarity between two following generations. On the basis of these facts, the Hb0 Arab and partially DNA work we suggested an alternative hypothesis that these mutant genes got into Bodrogköz by the rather later migration than with ancient Hungarian people during the period of conquest of Hungary.

TCR gamma/delta bearing lymphocytes in peripheral blood of allogenic bone marrow transplanted patients.

The presence of two distinct T-cell receptors (TCR) alpha/beta and gamma/delta dimers as well as of the activated T cells was analysed in peripheral blood mononuclear cells from seventeen recipients of allogeneic bone marrow transplants for leukemia and for severe aplastic anemia. Nine of seventeen recipients expressed an elevated percentage of T cells bearing TCR gamma/delta receptors in their peripheral blood. Seven out of nine cases having elevated gamma/delta positive cells showed chronic graft-versus-host (GVH) disease; one patient was treated with Cyclosporin A, and one patient was asymptomatic. In the twelve patients with GVH or other clinical symptoms, activated T cells (CD3+/HLA-DR+) were elevated indicating an autoreactive or alloreactive cell population. Our results confirmed earlier in vitro data showing that TCR-gamma/delta-bearing lymphocytes may be an activated T-cell population, and this T cell subset might be involved in mediating GVH disease, or in prolonging immunodeficiency after transplantation.

Plasma desorption mass spectrometry of haemoglobin tryptic peptides for the characterization of a Hungarian alpha-chain variant.

S-Aminoethylated-alpha A and -beta A globin tryptic peptides separated by reversed-phase high-performance liquid chromatography have been analysed by plasma desorption mass spectrometry. Almost all the expected alpha A and beta A tryptic fragments were tentatively assigned relative to the known globin chain sequences based on the molecular weight obtained by plasma desorption mass spectrometric analysis of the purified peptides. The application of plasma desorption mass spectrometry for structure elucidation of a haemoglobin alpha-chain variant revealed the first case of Hb Hasharon in Hungary.

Decrease in the carbamylcholine-induced chemotaxis of monocytes in myasthenia gravis.

The carbamylcholine-induced chemotaxis of monocytes was decreased in patients with myasthenia gravis, whereas no change was found in the C5a-induced locomotion of these cells compared with that of the normal controls. The decrease in the chemotaxis induced by carbamylcholine correlated with the severity of clinical symptoms. The beneficial effect of thymectomy was also reflected in the improvement of chemotaxis. The method is simple, not expensive and could be used in the diagnosis of myasthenia gravis.

[T gamma-lymphoproliferative disease]. / T gamma-lymphoproliferatív betegség.

The case of a rare type of T-cell malignant lymphomas, clinically with a relatively favorable course, a T gamma-lymphoproliferative disease (T gamma-cell chronic lymphocytic leukaemia) was presented. The cytomorphological, cytochemical, immuncytochemical and cytogenetical markers and functional tests of the peripheral blood lymphocytes from the patient were tested. The leukaemic cells with light- and electron microscopy showed the so called LGL (large granular lymphocyte) morphology with multifocal reactions of acid hydrolase enzymes. These cells also expressed IgG-Fc-receptor, CD 8 monoclonal antibody positivity and a monoclonally rearranged T-cell receptor gen expression. Functionally the patient's lymphocytes developed a blastic response to the T-cell mitogen Concanavalin A (ConA), they suppressed the immunoglobulin production of B-lymphocytes in co-cultures and had a normal NK-activity but decreased ADCC values. The patient was diagnosed by blood, bone marrow and lymph node examination and does not need any therapy. This case was published because of it's diagnostic, immunological and prognostical interests.

Locus assignment of human alpha-globin structural mutants by selective enzymatic amplification of alpha 1 and alpha 2-globin cDNAs.

We have used the powerful methodology of DNA enzymatic amplification in order to assign human alpha-globin structural mutants to one of the two highly homologous alpha-globin genes. Selectively amplified alpha 1 and alpha 2-globin cDNAs were dot-blotted and further hybridized to synthetic oligonucleotides encompassing either the normal or the mutated sequences. The generated signals corresponded specifically to one of the two alpha-globin genes. Using this approach the alpha-globin structural mutants J-Buda and G-Pest were found to be encoded by the alpha 2 and the alpha 1-globin genes, respectively. Furthermore, the exact nucleotide changes were determined. We propose this technique to serve as a simple and definitive method for assigning alpha-globin structural mutants.