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Biomedical subjects

Sara Fuchs

Publications and source records attributed to Sara Fuchs.

At least 19 recordsLinked to original sources
PubMed2006

Intravenous immunoglobulin suppresses experimental myasthenia gravis: immunological mechanisms.

Intravenous immunoglobulin (IVIG) administration has been beneficially used in the treatment of several autoimmune disorders including myasthenia gravis (MG), although its mechanism of action is still not clear. To study the optimal conditions of IVIG treatment and delineate its mechanism of action we established a suitable model in rat experimental autoimmune MG (EAMG). We show that IVIG has a suppressive effect on the clinical symptoms of ongoing EAMG that is associated with decreased AChR-specific cellular and humoral immune reactivity. Costimulatory factors and cytokine profile analyses suggest that IVIG immunomodulation in EAMG involves suppression of B and Th1-type T cell responses with no generation of T-regulatory cells. Our data contribute to the understanding of the immunological mechanisms underlying IVIG treatment in MG and in other autoimmune disorders.

Animals↗
PubMed2006

Low levels of alpha7-nicotinic acetylcholine receptor mRNA on peripheral blood lymphocytes in schizophrenia and its association with illness severity.

BACKGROUND: There is evidence that the nicotinic alpha7-acetylcholine receptor (alpha7-AChR) is involved in the pathophysiology of schizophrenia. Several neurotransmitter receptors, including alpha7-AChR, have been demonstrated on peripheral blood lymphocytes (PBL) and it has been suggested that these peripheral receptors may reflect corresponding brain receptors. OBJECTIVE: In this study we compare alpha7 mRNA expression in PBL between schizophrenia patients and control individuals in order to determine whether any correlation exists between alpha7 mRNA expression in PBL and severity of schizophrenia. In addition, the isoforms of alpha7-AChR expressed are identified. METHOD: Peripheral venous blood samples were collected from individuals with schizophrenia (n = 44) and from healthy subjects (n = 16). Symptomatology and illness severity were assessed using standard clinical psychiatric evaluation scales. RNA was prepared from isolated lymphocytes and alpha7 mRNA was measured by RT-PCR. RESULTS: We observed a significantly lower level of alpha7 mRNA on PBLs of schizophrenia patients in comparison with healthy controls (p < 0.00). A tendency to a negative correlation was noted between the CGI score, reflecting illness severity, and the alpha7-subunit gene expression. CONCLUSION: Observations confirm that the alpha7 mRNA in PBL represents the duplicated alpha7-AChR gene rather than the classic alpha7-AChR gene. Our study observations further substantiate the involvement of alpha7-AChR in the pathophysiology of schizophrenia and, while preliminary, indicate that the alpha7-AChR may be expressed and be readily measured in the peripheral blood circulation.

Actins↗
PubMed2005

Overexpression of phosphodiesterases in experimental autoimmune myasthenia gravis: suppression of disease by a phosphodiesterase inhibitor.

Myasthenia gravis (MG) and experimental autoimmune MG (EAMG) are T cell-dependent antibody-mediated autoimmune disorders, in which the nicotinic acetylcholine receptor (AChR) is the major autoantigen. DNA microarray analysis revealed increased levels of several phosphodiesterase (PDE) subtypes in lymph node cells (LNC) and muscles of EAMG rats compared with healthy controls. Quantitative real-time PCR analysis indicated that EAMG is characterized by an increase of PDE subtypes 1, 3, 4, and 7 in LNC and of PDE subtypes 2, 3, 4, and 7 in muscles. Pentoxifylline (PTX), a general PDE inhibitor, inhibited the progression of EAMG when treatment started at either the acute or chronic stages of disease. This suppression was associated with down-regulation of humoral and cellular AChR-specific responses, as well as down-regulation of PDE4, TNF-alpha, IL-18, IL-12, and IL-10 in LNC and of PDEs 1, 4, 7, and TNF-alpha in muscles. The expression of Foxp3, a transcription factor essential for CD4+CD25+ regulatory T cell function, was increased in splenocytes although the number of these cells remained unchanged. PTX also reduced the expression of the endopeptidase cathepsin-l, a marker of muscle damage, in EAMG muscles. This study demonstrates the involvement of PDE regulation in EAMG pathogenesis and suggests that PDE inhibitors may be considered for immunotherapy of MG.

Animals↗
PubMed2005

Overexpression of IFN-induced protein 10 and its receptor CXCR3 in myasthenia gravis.

Myasthenia gravis (MG) and its animal model, experimental autoimmune MG (EAMG), are autoimmune disorders in which the acetylcholine receptor (AChR) is the major autoantigen. Microarray technology was used to identify new potential drug targets for treatment of myasthenia that would reduce the need for the currently used nonspecific immunosuppression. The chemokine IFN-gamma-inducible protein 10 (IP-10; CXCL10), a CXC chemokine, and its receptor, CXCR3, were found to be overexpressed in lymph node cells of EAMG rats. Quantitative real-time PCR confirmed these findings and revealed up-regulated mRNA levels of another chemoattractant that activates CXCR3, monokine induced by IFN-gamma (Mig; CXCL9). TNF-alpha and IL-1beta, which act synergistically with IFN-gamma to induce IP-10, were also up-regulated. These up-regulations were observed in immune response effector cells, namely, lymph node cells, and in the target organ of the autoimmune attack, the muscle of myasthenic rats, and were significantly reduced after suppression of EAMG by mucosal tolerance induction with an AChR fragment. The relevance of IP-10/CXCR3 signaling in myasthenia was validated by similar observations in MG patients. A significant increase in IP-10 and CXCR3 mRNA levels in both thymus and muscle was observed in myasthenic patients compared with age-matched controls. CXCR3 expression in PBMC of MG patients was markedly increased in CD4(+), but not in CD8(+), T cells or in CD19(+) B cells. Our results demonstrate a positive association of IP-10/CXCR3 signaling with the pathogenesis of EAMG in rats as well as in human MG patients.

Adolescent↗
PubMed2005

Vaccines against myasthenia gravis.

Myasthenia gravis (MG) is an autoimmune disease mediated by antibodies to nicotinic acetylcholine receptor (AChR) interfering with the neuromuscular transmission. Experimental autoimmune MG serves as an excellent animal model to study possible therapeutic modalities for MG. This review will focus on the different ways to turn off the autoimmune response to AChR, which results in suppression of myasthenia. This paper will describe the use of fragments or peptides derived from the AChR, antigen-presenting cells and anti-T cell receptor antibodies, and will discuss the underlying mechanisms of action. Finally, the authors propose new promising therapeutic prospects, including treatment based on the modulation of regulatory T cells, which have recently been found to be functionally defective in MG patients.

Animals↗
PubMed2004

Dopaminergic regulation of immune cells via D3 dopamine receptor: a pathway mediated by activated T cells.

Neuro-immune interactions enable mutual regulation of the nervous and immune systems. To date, evidence exists for manipulations of immune cells by neurotransmitters in the periphery. In this study, we suggest the existence of a pathway by which the brain affects immune cells. The pathway we describe here is mediated by dopamine receptors expressed on activated T cells, termed blasts. Blasts can cross the blood brain barrier regardless of antigen specificity and can therefore encounter neurotransmitters in the brain. We show that blasts have a unique response to dopaminergic activation, which has no counterpart in resting T cells. Dopaminergic activation of blasts induces a Th1 bias in their cytokine profile and causes changes in surface marker expression. We further suggest that these changes can subsequently be transferred to peripheral T cells. We have tested this pathway in two in vivo systems: in rats exogenously administered with L-dopa, and in schizophrenia, which is characterized by a central nervous system-restricted increase in dopamine. In both models, peripheral T cells exhibit similar features to those of dopaminergically activated blasts. The existence of such a pathway by which the brain can regulate immune cells opens a conceptually new direction in neuro-immune interactions.

Animals↗
PubMed2004

Comparative phenotypic resolution of spontaneous, D2-like and D1-like agonist-induced orofacial movement topographies in congenic mutants with dopamine D2 vs. D3 receptor "knockout".

Using a novel system, the role of D2-like dopamine receptors in distinct topographies of orofacial movement was assessed in mutant mice with congenic D2 vs. D3 receptor knockout, and compared with findings in D1A mutants. Under spontaneous conditions, D2 mutants evidenced increased vertical jaw movements and unaltered horizontal jaw movements, with reductions in tongue protrusions and incisor chattering; in D3 mutants, only incisor chattering was reduced. Given previous evidence that D1A mutants show reduced horizontal but not vertical jaw movements, this indicates that apparent oppositional D1-like:D2-like interactions in the regulation of composited jaw movements may in fact reflect the independent actions of D2 receptors to inhibit vertical jaw movements and of D1A receptors to facilitate horizontal jaw movements. Effects of the D2-like agonist RU 24213 to exert greater reduction in horizontal than in vertical jaw movements were not altered prominently in either D2 or D3 mutants. The D1-like agonists A 68930 and SK&F 83959 induced vertical jaw movements, tongue protrusions, and incisor chattering; induction of tongue protrusions by A 68930 was reduced in D2 mutants. D2 receptors exert topographically specific regulation of orofacial movements in a manner distinct from their D1A counterparts, while D3 receptors exert only minor regulation of such movements.

Animals↗
PubMed2004

Immunosuppression of rat myasthenia gravis by oral administration of a syngeneic acetylcholine receptor fragment.

A syngeneic rat recombinant fragment of the extracellular domain of the acetylcholine receptor (AChR) alpha-subunit (Ralpha1-205), administered orally, suppresses ongoing experimental autoimmune myasthenia gravis (EAMG) in rats. The underlying mechanism is a shift from Th1 to Th2 regulation as evidenced by downregulated mRNA expression levels of IFN-gamma and TNF-alpha, upregulated IL-10, changes in anti-AChR IgG isotypes and diminished Th1 signaling via CD28/CTLA-4:B7. Unlike the xenogeneic fragment, the syngeneic Ralpha1-205 does not induce elevation in TGF-beta and elicitation of autoregulatory cells. The ability to suppress EAMG by a non-immunogenic syngeneic fragment of AChR is encouraging and may in the future be applied for the treatment of myasthenia gravis in humans.

Animals↗
PubMed2003

The alpha7 nicotinic acetylcholine receptor in schizophrenia: decreased mRNA levels in peripheral blood lymphocytes.

Recent studies have suggested that the alpha7 nicotinic acetylcholine receptor (alpha7 AChR) may play a role in the pathogenesis of schizophrenia. In search for peripheral biological markers for schizophrenia we have investigated alpha7 mRNA levels in peripheral blood lymphocytes (PBLs) of schizophrenic patients and healthy controls. Peripheral blood samples were collected from medicated and non-medicated (drug naive) schizophrenic patients as well as from healthy (non-mentally ill) smokers and non-smokers. RNA was prepared from isolated lymphocytes. Polymerase chain reaction products specific for human alpha7 AChR were quantified by densitometry using Scion image-analysis (shared NIH software). We observed a significant decrease of alpha7 mRNA levels on PBLs of schizophrenic patients compared with controls. The decrease in alpha7 mRNA levels was not a result of medication management, because non-medicated schizophrenic patients displayed the same level of reduction in alpha7 mRNA as did patients receiving medication. In addition, we exclude the possibility that the observed decrease in alpha7 mRNA levels resulted from nicotine consumption in smoking, because healthy smokers exhibited the same levels of alpha7 mRNA as non-smokers. We propose that alpha7 AChR may be involved in the pathophysiology of the disease and may serve as a reliable peripheral biological marker in schizophrenia.

Adolescent↗
PubMed2003

A monoclonal antibody specific for rat IL-18BP and its application in determining serum IL-18BP.

IL-18 binding protein (IL-18BP) is a soluble inhibitory receptor for IL-18, a proinflammatory cytokine that plays a major role in IL-12-driven Th1 cell differentiation. IL-18BP is presumed to play a vital regulatory function in IL-18-mediated immune responses, although its precise role in immunomodulation in various diseases is not well understood. In the present study, we have generated and characterized a mouse monoclonal antibody (2A6H11), specific for rat IL-18BP. Its binding epitope is within amino acids 29-60 on IL-18BP and it does not interfere with the function of IL-18BP. Using this monoclonal antibody, we have developed an ELISA for determining IL-18BP levels. We observed elevated levels of IL-18BP in sera of rats with experimental autoimmune myasthenia gravis (EAMG), in comparison with the levels in healthy rats. Thus, this monoclonal antibody can serve as a valuable tool to elucidate the role of IL-18BP in EAMG and in other rat models for autoimmune diseases.

Amino Acid Sequence↗
PubMed2003

Experimental autoimmune myasthenia gravis in mice expressing human immunoglobulin loci.

Antibodies (Abs) specifically directed against the muscular acetylcholine receptor (AChR) mediate the pathogenesis of myasthenia gravis (MG). The animal model experimental autoimmune MG (EAMG) can be induced by passive transfer or by active immunization of anti-AChR Abs. We report a new EAMG mouse model that generates human anti-AChR Abs upon immunization with Torpedo AChR (tAChR). Mice transgenic for human mu, gamma1, and kappa germ line genes (HuMAb-Mice) were immunized with tAChR. Serum titers of anti-tAChR Abs were in the nanomolar range, and anti-rodent AChR Abs were in picomolar range. Some HuMAb-Mice had signs of muscle weakness, clearly indicating their susceptibility to EAMG. Human Ab-mouse AChR complexes were found at the neuromuscular junction, while AChR loss was up to 65%. Spleen and lymph nodes were used for producing hybridomas. Of the anti-tAChR monoclonal Ab-producing hybridomas, 2% had cross-reactivity with rodent AChR and none with human AChR. Immunization with a fusion protein, Trx-Halpha1-210, displaying the human main immunogenic region did not result in EAMG or the generation of human anti-human AChR monoclonal Abs. These experiments show that the HuMAb-Mouse represents a suitable model to generate and study the effects of human anti-AChR Abs in vivo.

Animals↗
PubMed2003

Breakage of tolerance to hidden cytoplasmic epitopes of the acetylcholine receptor in experimental autoimmune myasthenia gravis.

The acetylcholine receptor (AChR) is the major autoantigen in the antibody-mediated disease myasthenia gravis (MG) and its animal model experimental autoimmune myasthenia gravis (EAMG). This study demonstrates that rats immunized with a recombinant fragment corresponding to the normally exposed extracellular region of the rat AChR alpha-subunit first develop antibodies to the injected extracellular portion only, but later develop antibodies to intracellular cytoplasmic epitopes of AChR. The presence of autoantibodies to intracellular epitopes seems to be correlated with development of clinical signs of disease. We propose that a similar process of epitope spreading may take place in the natural course of myasthenia.

Animals↗
PubMed2003

Design and synthesis of peptides that bind alpha-bungarotoxin with high affinity and mimic the three-dimensional structure of the binding-site of acetylcholine receptor.

Alpha-bungarotoxin (alpha-BTX) is a highly toxic snake neurotoxin that binds to acetylcholine receptor (AChR) at the neuromuscular junction, and is a potent inhibitor of this receptor. In the following we review multi-phase research of the design, synthesis and structure analysis of peptides that bind alpha-BTX and inhibit its binding to AChR. Structure-based design concomitant with biological information of the alpha-BTX/AChR system yielded 13-mer peptides that bind to alpha-BTX with high affinity and are potent inhibitors of alpha-BTX binding to AChR (IC(50) of 2 nM). X-Ray and NMR spectroscopy reveal that the high-affinity peptides fold into an anti-parallel beta-hairpin structure when bound to alpha-BTX. The structures of the bound peptides and the homologous loop of acetylcholine binding protein, a soluble analog of AChR, are remarkably similar. Their superposition indicates that the toxin wraps around the binding-site loop, and in addition, binds tightly at the interface of two of the receptor subunits and blocks access of acetylcholine to its binding site. The procedure described in this article may serve as a paradigm for obtaining high-affinity peptides in biochemical systems that contain a ligand and a receptor molecule.

Animals↗
PubMed2003

The binding site of acetylcholine receptor: from synthetic peptides to solution and crystal structure.

Our group has been employing short synthetic peptides, encompassing sequences from the acetylcholine receptor (AChR) alpha-subunit for the analysis of the binding site of the AChR. A 13-mer peptide mimotope, with similar structural motifs to the AChR binding region, was selected by alpha-bungarotoxin (alpha-BTX) from a phage-display peptide library. The solution structure of a complex between this library-lead peptide and alpha-BTX was solved by NMR spectroscopy. On the basis of this NMR study and on structure-function analysis of the AChR binding site, and in order to obtain peptides with higher affinity to alpha-BTX, additional peptides resulting from systematic residue replacement in the lead peptide were designed and characterized. Of these, four peptides, designated high-affinity peptides (HAPs), homologous to the binding region of the AChR, inhibited the binding of alpha-BTX to the AChR with an IC(50) of 2 nM. The solution and crystal structures of complexes of alpha-BTX with HAP were solved, demonstrating that the HAP fits snugly to alpha-BTX and adopts a beta-hairpin conformation. The X-ray structures of the bound HAP and the homologous loop of the acetylcholine binding protein (AChBP) are remarkably similar. Their superposition results in a model indicating that alpha-BTX wraps around the receptor binding-site loop and, in addition, binds tightly at the interface of two of the receptor subunits, where it inserts a finger into the ligand-binding site. Our proposed model explains the strong antagonistic activity of alpha-BTX and accommodates much of the biochemical data on the mode of interaction of alpha-BTX with the AChR.

Animals↗
PubMed2003

Suppression of myasthenia gravis by antigen-specific mucosal tolerance and modulation of cytokines and costimulatory factors.

We have shown that mucosal administration of recombinant fragments corresponding to the human acetylcholine receptor (AChR) alpha subunit suppresses chronic ongoing experimental autoimmune myasthenia gravis (EAMG) in rats. Treated animals exhibit a Th1 to Th2/Th3 shift in their cytokine profile and downregulation of costimulatory factors. However, application of a xenogeneic recombinant fragment may have limitations when considered as a possible approach for the treatment of MG in humans. We therefore tested the potential of a syngeneic fragment and of long synthetic peptides to suppress EAMG. We found that a syngeneic fragment corresponding to the extracellular region of the rat AChR alpha subunit was as effective as the formerly described human xenogeneic fragment in suppressing ongoing EAMG. This is encouraging in view of the potential use of mucosally administered recombinant AChR fragments for the treatment of MG in humans. However, in severely affected individuals, this antigen-specific approach may need to be supported by direct modulation of cytokines and costimulatory factors known to be involved in the pathogenesis of EAMG. To test the potential of this approach, myasthenic rats were injected by antibodies either to the proinflammatory cytokine IL-18 or to the costimulatory factor CD40L. These treatments act via different mechanisms, but both lead to the alleviation of clinical symptoms even when given at the chronic phase of EAMG. We suggest that antagonists to key cytokines and/or costimulatory factors be used to augment antigen-specific treatments of myasthenia such as mucosal administration of AChR recombinant fragments.

Animals↗
PubMed2002

betagamma Dimers mediate synergy of dopamine D2 and adenosine A2 receptor-stimulated PKA signaling and regulate ethanol consumption.

Dopamine release is activated by ethanol and addicting drugs, but molecular mechanisms linking dopaminergic signaling to neuronal responses and drinking behavior are poorly understood. We report that dopamine-D2 receptors induce PKA Calpha translocation and increase CRE-regulated gene expression. Ethanol also activates PKA signaling. Subthreshold concentrations of the D2 agonist NPA and ethanol, without effect alone, together cause synergistic PKA translocation and CRE-mediated gene transcription. D2 or adenosine A2 receptor blockade, pertussis toxin, Rp-cAMPS, or overexpression of dominant-negative peptides that sequester betagamma dimers prevent synergy. Importantly, overexpression of a betagamma inhibitor peptide in the nucleus accumbens strikingly reduces sustained alcohol consumption. We propose that synergy of D2 and A2 confers ethanol hypersensitivity and that betagamma dimers are required for voluntary drinking.

Adenoviridae↗
PubMed2002

Phenotypic, ethologically based resolution of spontaneous and D(2)-like vs D(1)-like agonist-induced behavioural topography in mice with congenic D(3) dopamine receptor "knockout".

Uncertainty as to the functional role of the D(3) dopamine receptor, due primarily to a paucity of selective agonists or antagonists, is being addressed in mice with targeted gene deletion ("knockout") thereof. This study describes, for the first time, the phenotype of congenic D(3)-null mice. Initially, 129/Sv x C57BL/6 D(3)-null mice were backcrossed 14 times onto C57BL/6; they were then assessed using an ethologically based approach which resolves all topographies of behaviour within the mouse repertoire. The ethogram of D(3)-null mice, on comparison with wildtypes, was characterised by no alteration in any topography of behaviour over an initial period of exploration; subsequent assessment over several hours revealed only increased rearing among females due to delayed habituation. Low doses of the selective D(2)-like agonist RU 24213 (0.016-0.25 mg/kg) inhibited topographies of exploratory behaviour; this effect was diminished in D(3)-null mice only when investigated following prolonged habituation, and then only for certain topographies of behaviour, primarily sniffing and rearing. High doses of RU 24213 (0.1-12.5 mg/kg) induced stereotyped sniffing and "ponderous" locomotion, while the selective D(1)-like agonist SK&F 83959 (0.016-2.0 mg/kg) promoted characteristic grooming syntax; these effects did not differ materially between the genotypes. When examined topographically on an essentially congenic C57BL/6 background (<0.005% 129/Sv), the resultant phenotype indicated essential conservation of the mouse ethogram, high-dose D(2)-like stimulatory effects, and D(1)-like stimulatory effects in the absence of D(3) receptors. A role for D(3) receptors in inhibitory processes appeared topographically circumscribed and only when baseline levels of behaviour were low.

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗