Interferon-γ for treatment of severe atopic eczema in two children

Two 4- and 5-year-old children suffering from refractory atopic dermatitis were treated with recombinant interferon- (rIFN-). rIFN- was injected at 50 g subcutaneously three times a week in the first child for 3 weeks, followed by three times 25 g in week 4. In the other child two treatment courses of 4 weeks were given after a break of 2 weeks. Therapy was well tolerated. In child one reductions in eczematous body surface and severity of lesions were observed, while no beneficial effect was seen in the other. Clinical chemistry data remained unchanged. Immunological studies performed in parallel showed a decrease in total serum IgE of 50% in child 1, a decrease in spontaneous in vitro IgE production, an increase in in vitro production of interleukin-6, and a normalization of previously decreased in vitro lymphocyte responses to several mitogens. While marked immunological changes were noted during IFN- treatment, clinical benefits were not encouraging. Diminished IFN- production has been claimed to be a major pathogenic factor in atopic eczema. Our results indicate that the pathogenesis is more complex. Clinically, we were unable to confirm previous observations in adults. Further studies are needed before IFN- can be recommended for therapy of pediatric atopic eczema.Abbreviations IFN- interferon- - IL interleukin     

Synergistic effects of tumour necrosis factor-α and interferon-γ on feline haemopoietic progenitors

Pathogenic mechanisms that underlie feline leukaemia virus subgroup-C (FeLV-C) induced erythroid aplasia are unknown. FeLV-C infection is associated with higher serum levels of interferon- (IFN-) and tumour necrosis factor- (TNF-), which may act synergistically to cause haemopoietic suppression. In the present studies, the synergistic effects of TNF- and IFN- on feline bone marrow progenitors in vitro were evaluated. Bone marrow mononuclear cells from specific-pathogen-free cats were exposed to TNF- (100 and 200 pg/ml) and IFN- (100 or 200 units/ml), alone or in combination, for 2 h before plating for clonal assays of colony forming units. Our results show that TNF- and IFN- in combination caused marked suppression of feline colony forming units-erythroid (CFU-E), burst forming units-erythroid (BFU-E), and colony forming units-fibroblasts (CFU-F), whereas colony forming units-granulocyte/macrophage (CFU-GM) were minimally affected. The same concentrations of TNF- and IFN- alone had minimal effects on CFU-E, BFU-E and CFU-F. These results suggest that TNF- and IFN- may play a significant role in regulating haemopoiesis in cats and may be involved in the pathogenesis of erythroid aplasia in cats infected with feline leukaemia virus.     

Glucocorticoids modulate TGF-beta production by human fetal lung fibroblasts

TGF- is thought to play a central role in pulmonary fibrosis inducing fibroblast differentiation and extracellular matrix synthesis. In human lung fibroblasts, it is still unclear how various TGF- isoforms affect TGF- production and whether glucocorticoids, commonly used agents to treat fibrotic lung disease, modulate these processes. To this end, human fetal lung fibroblasts (HFLF) were cultured with various concentrations of glucocorticoids (budesonide, dexamethasone or hydrocortisone) with and without TGF-1, -2, or -3. Post-culture media were collected for ELISA assays of TGF-1, -2, and -3 . TGF- mRNA was assessed by real time RT-PCR. Smad 2, 3, and 4 and AP-1 complex (c-fos and c-Jun) cellular localization were evaluated by immunostaining. TFG-2 and -3 stimulated TGF-1 production significantly (p < 0.01 relative to control). TGF-1 stimulated TGF-2 production (p < 0.01 relative to control). TGF-3 was undetectable. Glucocorticoids significantly inhibited TGF-1 and TGF-2 production and reduced expression of the up-regulated TGF-1 and TGF-2 mRNA induced by exogenous TGF-1, -2, or -3 (p < 0.01 for each) but had no effect on Smads. Although c-jun-related nuclear staining was not intensified in TGF--stimulated cells, it was reduced by glucocorticoids. Thus, TGF- isoforms may stimulate production of various TGF- isoforms in the lung. Glucocorticoids then may block TGF- production by modulating mRNA levels and c-Jun.     

Interferon β1a Treatment Modulates TH1 Expression in γδ + T Cells from Relapsing-Remitting Multiple Sclerosis Patients

A paradigm exists that multiple sclerosis is causally related to dysregulation of T_H1 inflammatory cytokines and T_H2 antiinflammatory cytokines. The cytokine source(s) that initiate the imbalances are unknown. In this study, , CD4, and CD8 T cell receptor-positive (TCR+) cells were isolated from the blood of 26 definitive relapsing-remitting multiple sclerosis patients prior to interferon -1a (IFN1a) therapy and following 8–10 weeks of this therapy. The bioactivities of interferon (IFN), interleukin 10 (IL10), and interleukin 12 (IL12) were determined. The concentrations of IFN, IL10, and IL12 from each cell type did not change significantly with IFN1a treatment. The IL10 secreted by TCR+ cells strongly correlated with the IL12 secreted by the same TCR+ cells, supporting the paradigm. Furthermore, IFN1a therapy decreased the TCR+ cell secretion of T_H1 cytokines after 8–10 weeks of therapy.     

Synoviocytes in chronic synovitis in situ and cytokine stimulated synovial cells in vitro neo-express α1, α3 and α5 chains of β1 integrins

The expression of the 1 integrins was examined immunohistochemically in synoviocytes from normal synovial membrane and from chronic synovitis of different aetiology and intensity. Normal synoviocytes were 61-positive but lacked 1 through 5. In mild inflammation type A synoviocytes neo-expressed 1, 3, and 5 chains. In severe inflammation both type A and B synoviocytes expressed 3, 4, 5, and 6 chains. The effects of inflammatory cytokines, as single agents or in combination, on the 1 integrin expression in cultured normal synoviocytes was determined by immunocytochemistry and flow cytometry. The 1 chain, while absent in unstimulated synoviocytes, was induced by interleukin-1 (IL-1), tumour necrosis factor- (TNF-), and interferon- (INF-). This effect was enhanced by combining IL-1 and TNF-. Expression of the 3 chain was up-regulated by IL-1 and, more intensely, by IFN-. Transforming growth factor (TGF-) inhibited the up-regulating effect of IL-1 and antagonized the effect of IFN- on 3 chain expression. Expression of the 5 chain was up-regulated significantly by co-stimulation through IL-1 together with TGF- or TNF-. Thus, the 1 integrin profile of cytokine activated synoviocytes in vitro resembled that of synoviocytes in synovitis in situ. These data suggest that IL-1, TNF-, IFN-, and TGF- are likely to be among the effectors regulating 1 integrin expression in synoviocytes in vivo.     

Acute ethanol consumption synergizes with trauma to increase monocyte tumor necrosis factor α production late postinjury

The hypothesis that acute ethanol uptake plus trauma can synergize to increase immunosuppression was tested. We found that, unlike non-alcohol-exposed patients, patients with acute alcohol use prior to trauma have a transient decrease in monocyte tumor necrosis factor (TNF) production during the very early postinjury (0–3 days) period. However, TNF production by these alcoholexposed patients' monocytes (MØ) became hyperelevated late postinjury (>9 days). Consequently, these massively elevated MØ TNF levels can contribute to posttrauma immunosuppression after acute alcohol use. We also demonstrate that normal monocyte activation with the superantigen,Staphylococcus enterotoxin B (SEB), results in a preferential induction of cellassociated MØ TNF production, described as characteristic of immunosuppressed trauma patients. Acutein vitro ethanol treatment down-regulated the elevated TNF production by trauma patients' MØ after either SEB, muramyl-dipeptide (MDP), interferon- plus MDP, or lipopolysaccharide (LPS) stimulation. Both SEB- and LPS-induced TNF mRNA induction was inhibited by acute alcohol treatment in normal MØ, indicating that ethanol can regulate cytokine gene expression. An additional immunosuppressive effect of acute ethanol's stimulation was suggested by its induction of elevated transforming growth factor production in trauma patients' activated MØ.     

γδ T Lymphocytosis Associated with Common Variable Immunodeficiency1

We present the case of a 28-year-old Caucasian female with common variable immunodeficiency (CVID) since age 5 who had a long history of hospitalizations for unexplained fevers and pulmonary infiltrates. The patient developed mild lymphocytosis 7 months prior to our evaluation. Flow cytometry of peripheral blood revealed an expansion of T lymphocytes, mild CD4 T lymphocytopenia, and a reduced CD4/CD8 ratio (0.2). Two subpopulations of T lymphocytes were found (CD3⁺/CD4^–/CD8⁺, 47%; CD3⁺/CD4^–/CD8^–, 53%), the vast majority of which expressed V-1. An infectious cause for the patient's T lymphocytosis could not be found. The sputum was chronically colonized with Staphylococcus aureus, and the organism produced TSST-1 in vitro. A bronchoalveolar lavage (BAL) revealed marked lymphocytosis, but T lymphocytes were not overrepresented in the BAL. Lymphocyte functional studies revealed poor proliferative responses to mitogens and staphylococcal superantigens and diminished cytokine production. V-1 T lymphocytes from the patient's blood were not expanded in vitro in response to staphylococcal superantigens. TCR gene rearrangement studies confirmed the presence of J and J1 clonal rearrangements accounting for only a small subpopulation of the T lymphocytes. These studies were repeated 5 months later and were unchanged. A bone marrow biopsy was negative for leukemia. Hence, the cause of the patient's T lymphocytosis could not be determined despite evaluation for underlying malignancy, occult infection, or superantigen-driven stimulation. The patient ultimately died of progressive respiratory insufficiency. The state of current knowledge regarding T lymphocytosis, decreased production of T lymphocytes, and a low CD4/CD8 ratio in association with CVID is discussed.     

Insulin and C-peptide co-localization in theβ granules of normal human pancreas and insulinomas

Summary It has been shown, by using the immunogold technique, that C-peptide and insulin are co-localized in the mature granules of human pancreatic cells and insulinomas with typical granules. The mean gold bead densities of both C-peptide and insulin were at least twice as high in the normal pancreas when compared with the insulinomas. The mean granule diameter of the insulinoma cells (D=0.30 ±0.12 m) was smaller than that of human pancreatic cells (D=0.45 ±0.15 m). The morphometric data indicate that each of the antigens (C-peptide and insulin) is distributed similarly in the halos and the dense cores of the granules. Thus, no topological segregation of these two antigens occurs within the granules of either normal human pancreas or insulinomas.     

Untersuchungen zur Ultrastruktur lympho-epithelialer Thymustumoren unter besonderer Berücksichtigung der sog. „Emperipolesis“

Summary The ultrastructure of eleven thymomas with lymphocytic predominance, one epitheloid cell thymoma and two normal human thymuses is described with special reference to Emperipolesis. All patients have had myasthenia gravis.The normal human thymus consists of three parts: outer cortex, inner cortex, and medulla. The outer cortex contains mainly lymphoblasts and Metcalf's macrophages within the so-called Clark-packet's. The inner cortex consists mainly lymphocytes and interdigitating reticulum cells, and the medulla of epithelial cells, lymphocytes and Hassall's corpuscles.In all cases of lympho-epithelial thymoma and in normal human thymuses there are enormous interdigitations between epithelial (tumor) cells, lymphocytes and macrophages. The epitheloid cell thymomas also show findings which suggest an epithelial cell interaction. We have not found intact lymphocytes inside the cytoplasm of normal and/or tumor epithelial cells, macrophages or interdigitating reticulum cells.The intracellular existence of intact lymphocytes has been termed Emperipolesis by Humble, Jayne, and Pulvertaft, meaning internal wandering. These investigations indicate that Emperipolesis is not an adequate term for cellular interaction in normal human thymuses and thymomas. A false impression of intraepithelial location of thymic lymphocytes is created by two-dimensional sections of complex thymic structure. These ultrastructural studies revealed damage to lymphocytes only in macrophages with lymphocytolysis within these cells and accumulation of numerous heterophagic vacuoles containing fragments of lymphocytic debris within them.

FrÄulein C. Schürmann danke ich für die gute technische Assistenz, Herrn Priv.-Doz. Dr. med. R. W. Ch. Janzen, Neurologische Klinik der UniversitÄt Hamburg, für die klinischen Daten der Myasthenie-Patienten     

Autogenetic inhibition of γ-motoneurons in the spinal cat uncovered by Dopa injection

Summary The response of identified gastrocnemius -efferents to stretch of the triceps surae muscle was recorded in the acute spinal cat before and after the injection of Dopa. A strong tonic inhibition during static muscle extension was detectable only in those -efferents which had no resting and reflex activity in the spinal cat, but which responded to Dopa injection with an initially high discharge rate. In contrast -motoneurons with background and reflex activity before Dopa, and some previously quiescent units acquiring a low discharge rate after Dopa did not exhibit any signs of a stretch-dependent inhibition. It is concluded that the observed autogenetic inhibition is limited to the static -motoneurons. The responsiveness to stretch-evoked inhibition was found to be correlated with that to antidromic inhibition induced by repetitive stimulation of the ventral root. This could imply that the activation of -motoneurons by muscle stretch leads to a Renshaw inhibition of the -motoneurons. However, this pathway may be only partly responsible for the autogenetic inhibition of -motoneurons in the spinal cat after Dopa, since the inhibitory effects of muscle stretch were more pronounced than those obtained by antidromic activation of motor axons in the ventral root.     

Occult hepatosplenic T-γδ lymphoma

We report on a patient with a rare hepatosplenic T-cell lymphoma ( TCL) presenting clinically with B-symptoms, hepatosplenomegaly and pancytopenia. During the initial stage of the disease the sparse malignant cells could not be detected histologically. Furthermore, their identification was obscured by massive macrophage proliferation with haemophagocytosis in the spleen. Diagnosis was established by detection of a clonal T-cell receptor (TcR) rearrangement and, retrospectively, by demonstration of rare cells expressing an aberrant T-cell phenotype. The findings in this patient emphasize that minimal neoplastic T-cell infiltrates can lead to severe clinical symptoms. Initial biopsy findings may be misinterpreted as benign. TCL may elaborate lymphokines that suppress haematopoiesis, leading to pancytopenia and macrophage proliferation.     

Thymosin β15 expression in tumor cell lines with varying metastatic potential

We previously isolated thymosin 15 from highly metastatic Dunning rat prostatic carcinoma cells. Immunohistochemical study of human prostate cancer specimens revealed a general correlation between Gleason grade and thymosin 15 expression, with high-grade (more malignant) tumors showing increased staining compared to low-grade tumors. To determine whether thymosin 15 may be differentially expressed in cancer cells with different metastatic potential other than in the prostatic carcinoma cells, we examined thymosin 15 mRNA levels in tumor cell lines from different species. We also examined thymosin 15 protein levels in human breast cancer samples. Thymosin 15 was upregulated in the highly metastatic mouse lung and human breast cancer cell lines in comparison to the nonmetastatic counterparts. Immunohistochemical staining showed the evidence of upregulation of thymosin 15 in malignant human breast carcinomas as compared to benign breast tumors. The expression of thymosin b1 5 was correlated with the metastatic potential of the mouse lung carcinoma and human breast carcinoma cells in addition to the prostatic carcinomas. Thymosin 15 may be a useful marker to predict metastatic potential of certain human cancers. © Rapid Science 1998     

Statin treatment and a disease-specific pattern of β-amyloid peptides in Alzheimer’s disease

According to the amyloid cascade hypothesis, sporadic Alzheimers disease (AD) is caused by the production and aggregation of -amyloid (A), and the production of A has recently been linked to the metabolism of cholesterol. We have previously published clinical studies where the effect of statin treatment on A production has been investigated. No effect on A was found, which is in disagreement with cell and animal studies. In the present study we investigated the effect of statin treatment on a disease-specific pattern consisting of a C-terminally-truncated quintet of A peptides. Nineteen patients with AD were treated with simvastatin for 12 months and the quintet of A peptides were analysed in cerebrospinal fluid before and after treatment. Also included was a group of 15 untreated patients with AD. We found that the A peptide pattern at baseline was in agreement with earlier findings; however, we did not find any change in the A peptide pattern after statin treatment. We suggest that clinical studies with extended treatment periods are performed where higher dosages of statins are used. We also believe that the pleiotropic effects of statins should be investigated further in order to elucidate the connection between Alzheimers disease and statin treatment.     

Beta-glucuronidase release from leukocytes in children

Summary The release of -glucuronidase from polymorphonuclear leukocytes (PMNs) is important in the killing of bacteria and in producing tissue damage in acute inflammation. To investigate the effects of various diseases or drugs on degranulation, we studied the kinetics of -glucuronidase release from PMNs exposed to opsonized zymosan. PMNs of children with bacterial infections demonstrated increased degranulation. Within 5, 15, and 30 min the PMNs released 19±3%, 23±3%, and 26±3% of total -glucuronidase compared to 12±2%, 15±2%, and 16±2% of total -glucuronidase of control PMNs. Viral infections induced a significant delay of -glucuronidase release from PMNs. Maintenance therapy of acute lymphoblastic leukemia with 6-mercaptopurine and methotrexate, as well as administration of vincristine, diminished the degranulation. After 5, 15, and 30 min the PMNs released 8±1%, 10±1%, and 11±1%, as well as 6±3%, 8±2%, and 9±2% of total -glucuronidase. This study demonstrated that bacterial infections stimulate -glucuronidase release by PMNs. In contrast, cytostatic drugs inhibit lysosomal enzyme release, increasing the susceptibility to bacterial infections. The total enzyme activities were unchanged.Abbreviations c-AMP cyclic Adenosinemonophosphate - c-GMP cyclic Guanosinemonophosphate - PMNs polymorphonuclear Leukocytes Supported by DFG Ri 275/6-1Dedicated to Prof. Dr. E. Gladtke on the occasion of his 60^th birthday     

γ-Glutamyltranspeptidase-Aktivität im Urin bei Gesunden und Nierenkranken

Zusammenfassung Die -Glutamyltranspeptidase-Aktivität (-GT) in der Niere ist bei chronischen Nephropathien und beim akuten Nierenversagen vermindert. Bei Gesunden wurde im Urin eine -GT-Aktivität von 12,7–37,7 mU/min ermittelt. Serum- und Urinaktivität verhielten sich unabhängig voneinander. Eine erhöhte -GT-Ausscheidung fand sich bei 75% der Nierenkranken mit normaler GFR, aber nur bei 11% mit eingeschränkter GFR.Zwischen -GT-Ausscheidung und Kreatininclearance bestand bei Gesunden (N=68,r=0,67) und Nierenkranken (N=114,r=0,71) eine statistisch hochsignifikante Korrelation. Bei Gesunden wurden deshalb die Toleranzgrenzen für beide Parameter gemeinsam berechnet. Bei Nierenkranken nahm die Enzymausscheidung proportional zur GFR und damit dem funktionsfähig verbleibenden Nierenparenchym ab. Die Relation -GT/C_Kreatinin erlaubt eine pathologisch gesteigerte Enzymabgabe aus den Tubuluszellen bei noch im Normalbereich liegender Gesamtausscheidung zu erkennen.     

Characterization of a macrophage-based system for studying the activiation of latent TGF-β

TGF- has been implicated in scarring and tissue fibrosis. Most cells secrete TGF- as a high molecular weight, latent complex that must be processed to a lower molecular weight, biologically active form. A number of molecules are involved in this activation step including the mannose 6-phosphate/insulin-like growth factor- II receptor, tissue transglutaminase, thrombospondin, plasmin, and others. Here we describe a rapid macrophage-based system for TGF- 1 activation, which could be used for screening potential anti- fibrotic agents. The system employs transformed mouse peritoneal macrophages treated with lipopolysaccharide as a cell line capable of activating latent TGF-. The activation mechanism in our system involves mannose 6-phosphate/insulin-like growth factor-II receptor andtransglutaminase. The activation of latent TGF- in this system can be prevented by the addition of mannose-6-phosphate but not mannose-1-phosphate. In addition, transglutaminase inhibitors, antibodies to thrombospondin, insulin-like growth factor- II in the presence of its binding protein IGFBP-2, but not IGFBP-1, suppressed the activation of TGF-. Anti-inflammatory molecules, such as hydrocortisone, when added to LPS-treated macrophages, inhibited TGF- activation by a mechanism, that may involve downregulation of transglutaminase expression. In summary, this new, rapid and reproducible system allows testing molecules for their ability to inhibit TGF- activation, thus providing a screening method for potential anti-scarring molecules.     

Zur Bedeutung des cyclischen Adenosin-3′:5′-monophosphat als „second messenger“ bei der Säuresekretion des Magens

Zusammenfassung Ob bei einer bestimmten Hormonwirkung cyclisches Adenosin-3:5-monophosphat (cAMP) als intracellulärer Vermittler fungiert, läßt sich nach den von Sutherland entwickelten Kriterien beurteilen; dabei sind die qualitativen, quantitativen und zeitlichen Beziehungen zwischen dem Hormoneffekt auf den intracellulären cAMP-Spiegel und die jeweilige physiologische Antwort zu prüfen. Im Falle der Histamin- bzw. Pentagastrin-stimulierten gastralen Säuresekretion des Frosches (Necturus maculosus) und der Ratte erfüllt das cAMP die Bedingungen eines second messenger. Dagegen ließ sich beim Hund und Menschen eine derartige Funktion des cAMP bisher nicht verifizieren; ob bei diesen Species cyclisches Guanosin-3:5-monophosphat anstelle des cAMP die intracelluläre Vermittlerrolle übernimmt, werden erst noch systematische Untersuchungen erweisen müssen.Die in diesem Zusammenhang zitierten eigenen Arbeiten wurden von der Deutschen Forschungsgemeinschaft, Bad Godesberg, unterstützt.     

Human T-lymphocyte subset production of immune (γ) interferon

Human T, T, and T lymphocyte subpopulations have the capacity to respond to phytohemagglutinin (PHA)in vitro with proliferation and the production of a pH 2 and heat-labile interferon. This occurs both when the subsets are isolated by direct rosetting techniques or by negative selection. Macrophages enhance the production of the interferon by each lymphocyte subset and do not themselves produce interferon in response to products of PHA-activated lymphocyte subsets. Thus our studies indicate that subpopulations of T lymphocytes known to differ with regard to morphology, surface receptors, RNA content, response to corticosteroids and X-irradiation, and other functional capabilities do not differ with regard to their capacity to produce interferon.     

An electrophysiological study of developmental changes in the innervation of the guinea-pig taenia coli

A sucrose-gap technique was used to study the development of neuromuscular transmission in the taenia coli of fetal, 1- to 2-day-old, 3- to 4-week-old and 3-month-old guinea-pigs. In addition, the effects of exogenous, ,-methylene adenosine 5-triphosphate (ATP), noradrenaline, vasoactive intestinal polypeptide (VIP) and carbachol were examined. Taking the response to a single pulse of electrical field stimulation as the index of a developed neuromuscular junction, it was apparent that the non-adrenergic inhibitory system arose before, and matured more quickly than, the cholinergic system. The inhibitory system was present by 8 weeks of gestation in some fetuses, but, in contrast, excitatory cholinergic transmission was not seen until birth. As evidenced by responses to carbachol, ,-methylene ATP and VIP, cholinergic, purinergic and VIP receptors were present on the smooth muscle at the earliest ages studied. No changes in sensitivity to these agents were noted throughout development, although in adults the level of the maximum responses increased.     

Inverse behaviour of "synaptic" ribbon and spherule numbers in the pineal gland of male guinea-pigs exposed to continuous illumination

Summary There is increasing evidence that pineal synaptic ribbons are a heterogeneous population of organelles. In addition to synaptic ribbons (SR) sensu stricto, which consist of an electron-dense rod surrounded by electronlucent vesicles, synaptic spherules (SS) exist, the electrondense core of which is round and much wider than that of the SR. In the guinea-pig SR and SS numbers exhibit an inverse day/night rhythmicity. To gain more insight into the functional significance of SR and SS, guinea-pigs were exposed to continuous illumination for approximately 4 months (LL) and the respective structures in the pineal gland were quantitated under the electron microscope and compared with control animals kept under a lighting regiment of 12 h light and 12 h dark. It was found that SR numbers increase following LL whereas SS numbers decrease. The proximal, intermediate and distal parts of the dumbbell-shaped organ respond differently. The increase in SR numbers is significant in the distal and intermediate regions only, whereas the decrease in SS numbers is significant in the proximal and the intermediate regions only. Within each pineal region analyses of parenchymal subareas measuring 65 m by 65 m exhibit an inverse correlation of SR and SS numbers indicating that there are parenchymal domains in which either SR or SS predominate. Morphometric analyses of a number of pinealocytic parameters reveal minor differences between different pineal regions and that exposure to LL does not strikingly affect the pinealocyte perikarya. By contrast, the numbers of pinealocyte processes increase significantly after LL in the distal and intermediate, but not the proximal region of the pineal gland. These observations suggest structural and functional differences between different parts of the guinea-pig pineal gland.