Interaction of immunoglobulin with actin

Actin can form specific, direct associations with immunoglobulin resulting in soluble complexes or cross-linked matrices. This interaction can be detected by four in vitro assays using purified components: (1) actin enhances the cytophilic activity of guinea pig IgG2; (2) in solutions of low ionic strength, actin and IgG2 co-precipitate: (3) soluble complexes exist in 0.1 M KCl as revealed by the displacement of actin from its expected sedimentation pattern in a gradient of sucrose when in the presence of IgG 1, IgG2, or IgM; (4) immunoglobulin (IgG1, IgG2, BGG)‡: increases the viscosity of F-actin solutions, presumably by crosslinking F-actin filaments. These data suggest that direct interaction of a cytoskeletal protein with a cell surface receptor is possible.     

γ-Hydroxybutyric acid (GHBA) induces pacemaker activity and inhibition of substantia nigra dopamine neurons by activating GABAB-receptors

Summary In the present study the actions of gamma" align="MIDDLE" BORDER="0">-hydroxybutyric acid (GHBA) on dopaminergic neurons in the rat substantia nigra (SN) were pharmacologically analysed utilising extracellular single unit recording techniques. Intravenous administration of GHBA was associated with several effects on the neuronal activity of nigral dopamine (DA) neurons. Low doses (<200 mg/kg) of GHBA produced a slight excitation of the neurons, concomitant with a regularised firing rhythm and lack of burst activity. In higher doses GHBA produced an even higher degree of regularisation but, in contrast to low doses, an inhibition of firing rate. Administration of the GABA_B-receptor agonist baclofen, in all essential respects, mimicked the effect of GHBA on the firing of nigral DA neurons. Both the regularisation of the firing pattern and inhibition of firing rate produced by systemic administration of GHBA were antagonised by the GABA_B-receptor antagonist CGP 35348 (200 mg/kg, i.v.).Our observations show that GHBA affects the firing pattern of nigral DA neurons in doses considerably lower than those required to inhibit the firing rate of the neurons. This action, as well as the decreased firing rate observed after high doses of GHBA, are mediated via activation of GABA_B-receptors. Correspondence to: G. Engberg at the above address     

Decrease of lymphokine (interleukin-2)-activated killer activity in peripheral blood after administration of natural interferon-gamma

Interferon-gamma (IFN-gamma) acts on a large array of different types of cell and has potent immunomodulatory activities besides cytotoxic effects on tumors. In a phase I study, some immunologic parameters of blood mononuclear cells from healthy volunteers who received intramuscular injections of natural human IFN-gamma were analyzed. The percentage of Leu-11a positive cells, natural killer (NK) activity, lymphokine (interleukin-2)-activated killer (LAK) activity and monokine production were measured either in blood mononuclear cells or in purified samples of lymphocytes or monocytes of the donors before and 24 h after IFN-gamma injection. After IFN-gamma injection, the percentage of Leu-11a positive cells and the LAK activity in the blood were significantly reduced, but NK activity and monokine production remained unchanged. These findings suggest that in vivo IFN-gamma acts directly or indirectly on Leu-11a positive cells and reduces LAK activity by changing the recruitment of LAK precursors in the blood.     

Caffeine inhibits cytoplasmic Ca2+ oscillations induced by carbachol and guanosine 5′-O-(3-thiotriphosphate) in hyperpolarized pancreatic ß-cells

The effects of caffeine on cytoplasmic Ca²⁺ oscillations induced by carbachol and guanosine 5-O-(3-thiotriphosphate) (GTP-gamma" align="MIDDLE" BORDER="0">-S) were studied in individual mouse pancreatic ß-cells clamped at a hyperpolarized potential. Addition of 10 mM caffeine did not affect the cytoplasmic Ca²⁺ concentration ([Ca²⁺]₁) in ß-cells exposed to 20 mM glucose and hyperpolarized with diazoxide. Under similar conditions 100 M carbachol induced a typical response with a marked [Ca²⁺]_i peak followed by a lower sustained elevation. Irrespective of whether 10 mM caffeine was present, there were [Ca²⁺]_i transients with frequencies of 1–5/min superimposed on the sustained phase in 50–60% of the cells. In previously non-exposed cells the introduction of 10 mM caffeine caused temporary lowering of the sustained phase with disappearance of the transients. Subsequent omission of caffeine in the continued presence of carbachol caused a marked [Ca²⁺]_i peak followed by reappearance of the [Ca²⁺]_i, transients. However, in cells oscillating in the presence of caffeine its omission caused disappearance of the transients. In this case reintroduction of caffeine restored the transients.In cells kept at –70 mV by a patch pipette containing 100 M GTP-gamma" align="MIDDLE" BORDER="0">-S and 3 mM Mg-ATP there were [Ca²⁺]_i transients with frequencies of 0.5–2.5/min. These transients were sufficiently pronounced to activate repetitively a K⁺ current. Addition of 10 mM caffeine caused disappearance of the [Ca²⁺]_i transients or reduction of their amplitudes and frequencies.The results indicate that caffeine does not activate Ca²⁺-induced Ca²⁺ release in hyperpolarized ß-cells but inhibits the Ca²⁺-mobilizing effect of inositol 1,4,5-trisphosphate. Correspondence to: E. Gylfe at the above address     

An immunohistochemical study of immunological phenomena in minor salivary glands in patients with Sjögren's syndrome

Using different monoclonal antibodies, we performed an immunofluorescent technique on labial salivary glands in order to investigate the immunological phenomena involved in Sjögren's syndrome (SS). An aberrant expression of HLA-DR molecules was detected on cytoplasm of epithelial labial salivary cells in 9 out of 19 (47%) patients, with SS. No such expression was found in 8 patients without SS or in 3 normal controls. HLA-DQ molecules were demonstrated also in two out of ten SS patients without HLA-DR. A lymphocytic infiltration was not correlated with the expression of class II molecules. T cells bearing gamma" align="MIDDLE" BORDER="0"> receptors were not detected. The intracellular adhesion molecules (ICAM-1) and lymphocyte function associated antigen-1 (LFA-1) were not found on epithelial glandular salivary cells of patients and controls. In conclusion, these data suggested that the absence of ICAM-1 and LFA-1 in salivary cells and the absence of infiltrating T cells bearing gamma" align="MIDDLE" BORDER="0"> receptors exclude their immunopathogenetic role in SS; moreover, these data demonstrated that the aberrant expression of HLA class II molecules on epithelial salivary cells of patients with SS is not a phenomenon correlated with the lymphocytic infiltration.     

Interferon gamma and interleukin 10 levels in preimplantation embryo culture media

Purpose The aim of our study is to elucidate whether human oocyteslembryos secrete IFN _{gamma" align="MIDDLE" BORDER="0">} and/or IL-10 and whether the fertilization process depends on the balance between these cytokines.Methods A total of 142 embryo culture media from 24 patients were collected and the cytokine levels were tested with ELISA.Results IFN _{gamma" align="MIDDLE" BORDER="0">} and IL-10 were detectable in 40.1% and 29.6% of culture media respectively. The difference of IFN _{gamma" align="MIDDLE" BORDER="0">} and IL-10 levels in media from fertilized oocytes between day 1 and day 2 are significant (0.46 vs. 1.47 and 34.2 vs. 12.7, respectively). However there was no significant difference between the IFN _{gamma" align="MIDDLE" BORDER="0">} levels of the media from fertilized and nonfertilized oocytes 0.46 vs. 0.85 at day 1 and 1.47 vs. 1.49 at day 2, as well as IL-10 levels 34.2 vs. 30.9 at day 1 and 12.7 vs. 9.58 at day 2 respectively.Conclusions Human preimplantation embryos secrete the cytokines IFN _{gamma" align="MIDDLE" BORDER="0">} and IL-10. No effect of these cytokines on fertilization process could be shown.Presented at the IXth World Congress on In Vitro Fertilization and Alternate Assisted Reproduction, April 3–7, 1995, Vienna, Austria.     

Molecular Human Reproduction: Evidence for {gamma}-aminobutyric acid specific binding sites on human spermatozoa

The possible presence of gamma.gif" ALT="{gamma}" BORDER="0">-aminobutyric acid (GABA) specificbinding sites on human spermatozoa was investigated. Swim-uppreparations of human spermatozoa were incubated with radiolabelledGABA in the presence of unlabelled GABA, alternatively displacersof GABA_A/B receptors and GABA transport proteins. The resultsindicate that GABA specific binding sites are present on thesurface of human spermatozoa, and that these binding sites possiblyindicate the presence of GABA transport proteins. Furthermore,GABA at different concentrations was added to swim-up preparationsof human spermatozoa. Possible effects of GABA on sperm motility,hyperactivation and acrosome reaction were explored. No significantdifferences were observed between treated groups and controlsconcerning motility parameters and hyperactivation. Incubationwith GABA did not cause any increase in spontaneous acrosomereaction. However, spermatozoa treated with the calcium ionophoreA-23187 showed a small but significantly increased ability toundergo the acrosome reaction following preincubation in 10–⁴M GABA (P < 0.05).     

The selective P2X purinoceptor agonist, β,γ-methylene-L-adenosine 5′-triphosphate,discriminates between smooth muscle and neuronal P2X purinoceptors

The effects of the putative selective P_2X purinoceptor agonist, ,gamma" align="MIDDLE" BORDER="0">-methylene-l-adenosine 5-triphosphate (gamma" align="MIDDLE" BORDER="0">me-l-ATP), were determined at rat neuronal and smooth muscle P_2X purinoceptors.gamma" align="MIDDLE" BORDER="0">Me-l-ATP had no effect on the extracellularly recorded membrane potential of the rat isolated vagus nerve preparation at concentrations up to 300 M. In contrast, the archetypal P_2X purinoceptor agonist, , methylene ATP (meATP;1–100 M), produced concentration-related depolarisation responses with a mean EC₅₀ value of 10.8 M. The depolarising effects of meATP were not attenuated by gamma" align="MIDDLE" BORDER="0">me-l-ATP (100 M). In voltage clamp experiments on single nodose ganglion neurones, ATP (100 M), but not gamma" align="MIDDLE" BORDER="0">me-l.-ATP (1–300 M), evoked rapid ( < 20 ms onset) inward currents when applied using a concentration-clamp method. In receptor binding studies to rat brain membranes, gamma" align="MIDDLE" BORDER="0">me-d-ATP and meATP competed with high affinity for [³H]Lx meATP binding sites, with mean pIC₅₀ values of 7.7 and 8.3, respectively. However, gamma" align="MIDDLE" BORDER="0">me-l-ATP possessed low affinity for these sites and competed only at concentrations in excess of 10 M (mean pIC₅₀ value 4.1).In prostatic segments of the rat vas deferens, gamma" align="MIDDLE" BORDER="0">me-l-ATP (1–100 M) and meATP (0.3–100 M) each produced concentration-related contractile responses with mean EC₅₀ values of 17.1 and 3.6 M, respectively. gamma" align="MIDDLE" BORDER="0">Me-l-ATP (1–10 M) evoked fast inward currents in freshly dispersed vas deferens smooth muscle cells, indicative of an action at ligand-gated ion channels. Binding sites in vas deferens membranes labelled using 1 nM [³H]meATP exhibited high affinity for gamma" align="MIDDLE" BORDER="0">me-l-ATP, meATP and gamma" align="MIDDLE" BORDER="0">me-d-ATP with mean PIC₅₀ values of 7.7, 8.4 and 7.3, respectively.These results indicate that gamma" align="MIDDLE" BORDER="0">me-l-ATP exhibits neither agonist nor antagonist properties at P_2X purinoceptors on rat vagal neurones and possesses only very low affinity for [³H]meATP binding sites in rat brain. In contrast, gamma" align="MIDDLE" BORDER="0">me-l-ATP is a potent, high affinity agonist at smooth muscle P_2X purinoceptors of the rat vas deferens. This selective agonist action of gamma" align="MIDDLE" BORDER="0">me-l-ATP suggests that P_2X purinoceptors in smooth muscle and neurones are different and represent distinct P_2X purinoceptor subtypes.     

Analysis of T-cell receptor expressing lymphocytes infiltrating squamous cell carcinomas of the upper aerodigestive tract

T-lymphocytes expressing T -cell receptors (TCRs) of the gamma" align="MIDDLE" BORDER="0">/ type have been suggested to play an important role in mucosal defense against infection and neoplastic transformation. In this study, an immunohistochemical investigation was performed on the distribution of / and gamma" align="MIDDLE" BORDER="0">/ TCRs among tumor-infiltrating lymphocytes. Thirteen patients with squamous cell carcinomas of the upper aerodigestive tract were studied, using monoclonal antibodies and an avidin-biotin-peroxidase technique. Most of the T-cells had an / TCR. Only 1.6% of the T-cells within the cancer tissue and 1.2% of the T-cells in the parenchyma adjacent to the cancer tissue expressed gamma" align="MIDDLE" BORDER="0">/gd TCRs. These results are consistent with the results of similar studies in bronchial and breast carcinomas. Biopsies from normal oral mucosa in nine healthy individuals showed that 1.3% of the T -cells within the epithelium and 1.0% of those in the lamina propria adjacent to the epithelium expressed gamma" align="MIDDLE" BORDER="0">/ TCRs. Quantitatively the results do not support the theory that gamma" align="MIDDLE" BORDER="0">/ T-cells play an important role in the immunological response against cancer tissue in the mucosa of the upper aerodigestive tract. The functional role of these cells in the mucosa and in response to carcinomas is, however, still uncertain.