Down-regulation of tumor necrosis factor α activity by acute ethanol treatment in human peripheral blood monocytes

As the most commonly used drug that can modulate both metabolic and immune pathways, ethanol is evaluated in this report as a regulator of tumor necrosis factor (TNF) production in human peripheral blood monocytes (M) in combination with a variety of stimuli. While acute ethanol treatment did not induce TNF in M, it was a potent down-regulator of M TNF production whether induced by the combination of interferon- plus muramyl dipeptide (MDP) (P<0.001), lipopolysaccharide (LPS) alone (P<0.01), or interferon- plus LPS. Down-regulation of M TNF by ethanol was dose dependent and statistically significant in the biologically relevant, 25–150 mM, ethanol concentration range. We also demonstrate that these ethanol concentrations did not affect M viability. TNF down-regulation by ethanol was most effective when ethanol was administered 4 hr prior to MDP stimulation; however, it was also effective—though to a lesser extent—if it was added at the time of MDP stimulation. Furthermore, ethanol also down-regulated TNF production of thein vivo preactivated M of trauma patients, which produce hyperelevated levels of TNF. We have previously shown that the majority of posttrauma elevated M TNF is produced by the M subpopulation expressing high-affinity type I Fc receptors (FcRI). When the FcRI cross-linking-stimulated M subpopulation was treated with acute ethanol, TNF production was suppressed again both inin vivo preactivated M of trauma patients and in M of normal controls. In experiments utilizing cyclooxygenase inhibitor, we also demonstrate that ethanol has a direct, prostaglandin E₂-independent, effect on M TNF production. These results demonstrate that acute ethanol exposure has the potential to down-regulate M production of TNF significantly regardless of the TNF-inducing stimulus. Decreased capacity of M to produce TNF might, therefore, contribute to the immunological and metabolic abnormalities described after ethanol uptake.     

Differential effects of interleukin-1α and β on the arachidonic acid cascade in human synovial cells and chondrocytes in culture

The effects of interleukin-1 and were tested on the [³H]-arachidonic acid release and the prostaglandin synthesis by human cultured synovial cells and chondrocytes. Both forms of interleukin-1 stimulated the arachidonic acid release but interleukin-1 was more potent than IL-1. Human synovial cells and chondrocytes synthesized three types of prostaglandins upon stimulation with interleukin-1 or : prostaglandin E₂, F₂ and 6-keto-prostaglandin F₁. Regarding the synthesis of these prostaglandins, IL-1 was again more potent than IL-1. A comparison between interleukin-1-stimulated synovial cells and chondrocytes revealed neither significant quantitative nor qualitative differences in both the arachidonic acid release and the prostaglandin synthesis.     

Immunocytochemical characterization of porcine zona pellucida during follicular development

Sections of bovine ovaries fixed in Bouin's fluid or methanol-acetic acid and embedded in paraffin were incubated with chicken polyclonal antibodies to HPLC-purified zona glycoproteins ZP3 and ZP3. Oocytes of primordial follicles as well as of primary follicles showed weak labelling with anti-ZP3 and anti-ZP3. No immunostaining could be observed in the follicle cells. The ZP of primary follicles displayed distinct immunoreactivity for both ZP3 and ZP3. In secondary follicles, distinct labelling with anti-ZP3 and weak labelling with anti-ZP3 could be seen in the oocyte. The ZP showed immunoreactivity with antibodies to ZP3 and ZP3. Both antibodies labelled single follicle cells. In tertiary follicles, the oocytes were weakly labelled with anti-ZP3 and anti-ZP3. Some granulosa cells showed staining for ZP3 and ZP3. The ZP displayed strong immunoreactivity for ZP3 and ZP3. Cells of the corona radiata were strongly immunopositive for ZP3 and ZP3. Similar histotopography of immunoreactive cells could be seen in preovulatory follicles. The characteristic pattern observed for the distribution of ZP3 and ZP3 strongly suggests that in the porcine ovary both the oocyte and the follicle cells contribute to the synthesis of the ZP, perhaps in sequence.     

β-Subunits co-determine the sensitivity of rat neuronal nicotinic receptors to antagonists

We have investigated the effect of 4 ganglionic cholinergic antagonists (hexamethonium, mecamylamine, pentolinium, trimetaphan) on rat 32 and 34 neuronal nicotinic acetylcholine receptors (nAChRs) expressed in Xenopus oocytes. Current responses were elicited by fast application of acetylcholine on voltage-clamped oocytes (holding potential Vinh = -80mV). Concentration-inhibition curves were used to get estimates of IC₅₀, the antagonist concentration yielding 50% reduction of the peak current. The K_B's of the antagonists were calculated using estimates of the apparent K_D of acetylcholine. The order of affinity of the antagonists was similar for both receptor subtypes: mecamylamine pentolinium > hexamethonium > trimetaphan. However, 34 neuronal nAChRs were 9 to 22 times more sensitive to each of the 4 antagonists than 32 receptors. These results further underline the importance of the -subunit as co-determinant of the functional properties of neuronal nAChRs.     

GM1-gangliosidosis: Chromosome 3 assignment of theβ-galactosidase-A gene (β GAL A )

The structural gene (GAL_A) coding for lysosomal -galactosidase- A (EC 3.2.1.23) has been assigned to human chromosome 3 using man-mouse somatic cell hybrids. Human -galactosidase-A was identified in cell hybrids with a species-specific antiserum to human liver -galactosidase-A. The antiserum precipitates -galactosidase-A from human tissues, cultured cells, and cell hybrids, and recognizes cross-reacting material from a patient with G_M1 gangliosidosis. We have analyzed 90 primary man-mouse hybrids derived from 12 separate fusion experiments utilizing cells from 9 individuals. Enzyme segregation analysis excluded all chromosomes for GAL_A assignment except chromosome 3. Concordant segregation of chromosomes and enzymes in 16 cell hybrids demonstrated assignment of GAL_A to chromosome 3; all other chromosomes were excluded. The evidence suggests that G_M1 gangliosidosis is a consequence of mutation at this GAL_A locus on chromosome 3.     

Parathyroid hormone enhances calcium current in snail neurones — Simulation of the effect by phorbol esters

Effects of parathyroid hormone substance (PTH) on the voltage-activated calcium current (I _Ca) were studied on intracellularly perfused neurones of the snail, Helix pomatia, under voltage-clamp conditions. Application of 0.1 nM PTH produced a marked potentiation of the current. The effect developed slowly (60–70 min) and remained after removal of PTH. Potentiation could be observed in most neurones, but varied considerably from cell to cell; in some neurones I _Ca was increased 2- to 3-fold. Addition of ethylenebis(oxonitrilo)tetraacetate (EGTA, 10 mM) to, or removal of adenosine 5-triphosphate (ATP, 2 mM) from the intracellular perfusing solution resulted in a suppression or attenuation of the potentiating effect. The effect could be reproduced by the synthetic 1–34 amino acid fragment of PTH. Extracellularly applied protein kinase-C (PK-C) activator phorbol ester phorbol 12-myristate 13-acetate (PMA, 0.1–10 M) produced a similar slow increase in I _Ca (up to 1.5- to 2-fold), while its inactive analogue (4-phorbol ester) had no effect on I_Ca. The effects of PTH and PMA were not additive. PK-C inhibitors [1-(5-isoquinoline-sulphonyl)-2-methylpiperazine hydrochloride] (H-7, 100 M) and staurosporine (100 M) as well as calcium channel antagonists Cd²⁺, verapamil, nifedipine and nimodipine depressed the effect of PTH. The chloride channel blocker 4,4-diisothiocyanato-stilbene-2,2-disulphonic acid (DIDS, 1 mM) did not affect the potentiating action of PTH. Activation of the adelylate cyclase system also potentiated I _Ca in some neurones, but this effect had a different time course and was additive to the effect of PTH. A conclusion is made that activation of PK-C may mediate the slowly developing enhancement of I _Ca by PTH.     

TRP channels: a TR(I)P through a world of multifunctional cation channels

The transient receptor potential (TRP) family of ion channels comprises more than 50 cation-permeable channels expressed from yeast to man. On the basis of structural homology, the TRP family can be subdivided in to seven main subfamilies: the TRPC (Canonical) group, the TRPV (Vanilloid) group, the TRPM (Melastatin) group, the TRPP (Polycystin), the TRPML (Mucolipin), the TRPA (Ankyrin) and the TRPN (NOMP) family. The cloning and characterization of members of this cation channel family has exploded during recent years, leading to a plethora of data concerning TRPs in a variety of cell types, tissues and species. This paper briefly reviews the TRP superfamily and the basic properties of its many members as a readers guide in this Special Issue. Hopefully, a better understanding of TRP channel physiology will provide important insight into the relationship between TRP channel dysfunction and human diseases.     

Isoenzymes of γ-glutamyl transpeptidase in liver diseases

Summary A new method for the separation of isoenzymes of-glutamyl-transpeptidase is described, using electrophoresis on acetate cellulose gel and a developing solution composed by-glutamyl-naphthylamide, and a colored diazonium compound.The method permits the separation of up to four different isoenzymes, which we called-GT₁,-GT₂,-GT₃,-GT₄, the first two showing an electrophoretic migration similar to that of ₁- and ₂-globulins and the other two to that of-globulins.The present technique has proved its usefulness in detecting isoenzymes in serum with values of total-glutamyl-transpeptidase higher than 80 U/L.The application of this method in 52 patients with different types of biliary obstruction and hepatocellular damage has shown that it provides new possibilities in differential diagnosis.     

Application of the polymerase chain reaction to study the M protein(-like) gene family in beta-hemolytic streptococci

Evaluation of homologous regions of published M protein (emm) gene sequences from group A streptococci (GAS; Streptococcus pyogenes) was used to design three primer pairs for polymerase chain reaction (PCR) and three oligonucleotide probe sequences internal to the amplified products. One set of primers and corresponding probe should detect and lead to amplification of emm(-like) genes of virtually every type (all M), another (SOR^-M) should only amplify emm(-like) genes from GAS negative for serum opacity reaction (SOR) and the third (SOR⁺M) should expand only emm(-like) genes from SOR⁺ GAS. Using the all M primer pair for PCR on the genomic DNA from GAS of 29 different M types as well as from a group C and a group G streptococcal isolate, DNA fragments within the expected size range were amplified in every assay. All PCR products reacted with the all M probe. Related sequences were not detected in genomic DNA of an S. agalactiae and an Enterococcus faecalis isolate. Applying the SOR^-M and SOR⁺M primers to identical assays led to mutually exclusive amplification products. The SOR⁺M and SOR⁺M probes hybridized only to their corresponding products. Exceptions to this exclusivity were the SOR⁺ GAS of M types 3, 8, 27, 34, 42, 67, and 69, which consistently reacted only with the SOR⁺M primer/probe set. Analysis of sequence data from the amplified emm(-like) 2, 3, 18, and 19 genes revealed interesting specific features such as conserved gaps in the C-terminal sequence regions from SOR⁺ and the exceptional SOR^- GAS strains. These data indicate the existence of a subgroup of strains among SOR^- GAS and may advance our understanding of phylogenetic relationship between different serotypes of GAS.     

A comparative immunohistological study of cerebellar enolases

Summary A comparative immunohistological study of the neurone-specific enolase and enolase, demonstrates the exclusive neuronal localization of enolase and its absence from glial cells. In contrast, enolase is located in astroglial cells. The validity of enolase as a neuronal marker and enolase as an astrocytic marker, is confirmed both by a double labelling technique, using antibodies to and to revealed with fluorescence or peroxidase in the same tissue sections, and by immunoelectronmicroscopy.     

Immunglobuline γG, γA und γM im Serum von Schwangeren

Zusammenfassung Der Gehalt an Immunglobulinen G, A und M im Serum schwangerer, gesunder Frauen wurde in Abhängigkeit vom Alter der Gravidität geprüft und verglichen mit den Werten aus Seren eines Kollektivs weiblicher Blutspender gleicher Altersstufe. Es wurde festgestellt, daß der Serumspiegel an G stark absinkt, der Gehalt an A eine sinkende Tendenz zeigt und der M-Spiegel unbeeinflußt bleibt.

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Summary Serum of pregnant healthy women was investigated for its percentage of Ig G, Ig A and Ig M dependent from the stage of pregnancy. These measurements were compared with results from a group of female blood-donors of the same age. It was concluded, that the serumcontent of G decreases very distinctly, the Ig A percentage is somewhat diminished whereas the M content shows no differences between the two groups.

     

Chromaticity of synaptic inputs to H1 horizontal cells in carp retina: analysis by voltage-clamp and spectral adaptation

Summary Cone photoreceptor inputs to H1 horizontal cells (H1 HCs) in carp retina were studied by measuring light-modulated currents (I_L) to monochromatic stimuli (460, 533, 688 nm) under a voltage-clamp condition. By using double-barrelled micro-electrodes H1 HCs were voltage-clamped whilst perfusing with dopamine to uncouple the cells. The I_L of the H1 HCs driven by each cone input was segregated by selective chromatic adaptation, and differences in the kinetics of the I_L of the H1 HCs were revealed. Thus, all together, three types of I_L were observed: (1) a fast outward current to the long-wavelength stimulus; (2) a slow outward current to the middle-wavelength stimulus; and (3) a delayed inward current that followed the peak of slow outward current to the short-wavelength stimulus. The reversal potentials of the three currents were estimated to be at least 20 mV more positive than the dark resting potential by extrapolation of the I_L-V curve. These observations are consistent with the idea that the H1 HCs receive sign-inverting, conductance decreasing synaptic input(s) from at least one other cone mechanism, in addition to the main conventional EPSP type synaptic input from red-sensitive cones.     

ω-Grammotoxin blocks action-potential-induced Ca2+ influx and whole-cell Ca2+ current in rat dorsal-root ganglion neurons

Field-potential stimulation of rat dorsal-root ganglion (DRG) neurons evoked action-potential-mediated transient increases in intracellular free calcium concentration ([Ca²⁺]_i) as measured by indo-1-based microfluorimetry. Field-potential-evoked [Ca²⁺]_i transients were abolished by tetrodotoxin, and their dependence on stimulus intensity exhibited an abrupt threshold. -Conotoxin GVIA (-CgTx, 100 nM) inhibited action-potential-mediated Ca²⁺ influx by 79%, while nitrendipine (1 M) had little effect. -Grammotoxin SIA (-GsTx, 267 nM), a peptide toxin purified from the venom of the tarantula spider, Grammostola spatulata, blocked action-potential-mediated Ca²⁺ influx as effectively as did -CgTx, suggesting that -GsTx blocks N-type Ca²⁺ channels. In contrast to block by -CgTx, the block produced by -GsTx reversed upon washout of the peptide. -GsTx (270 nM) blocked 80%, and -CgTx (1 M) blocked 64%, of whole-cell Ca²⁺ current (I _Ca) elicited by step depolarization to 0 mV from a holding potential of –80 mV. -GsTx completely occluded inhibition of I _Ca by -CgTx. However, when applied after -CgTx, -GsTx produced an additional inhibition of 27%, indicating that -GsTx also blocked a non-N-type Ca²⁺ channel. BayK8644 (1 M) elicited an increase in I _Ca in the presence of maximally effective concentrations of -GsTx, suggesting that -GsTx does not block L-type channels. Thus, -GsTx displays a selectivity for Ca²⁺ channel subtypes which should prove useful for studying Ca²⁺ channels and Ca²⁺-channel-mediated processes.     

Examination of a purified A′ influenza virus preparation by potentiometry

Summary The potentiometric titration of a purified influenza A virus preparation revealed 100.7×10^–4 M base and 68.8 × 10^–4 M acid-binding capacity per g. of virus protein N. The titration curve was characterized by the following fourpK values:pK ₁ = 3.37;pK ₂ = 4.50;pK ₃ = 6.37, andpK ₄ = 9.75. The isoionic point was at pH 5.43.Attempt was made to identify the dissociating groups and it was found that the carboxylic groups (pK ₁ andpK ₂) may either be glutamyl or aspartyl groups, while the cationic groups are probably the imidazolium of histidine (pK ₃) and the -amino residues of lysine (pK ₄).Inaotivation of the hemagglutinating activity of the virus preparation by mild treatment with formaldehyde at pH 8.0 resulted in a simultaneous disappearence of the -amino groups of lysine (pK ₄). The same treatment at pH 9.0 resulted in the loss of all the cationic groups previously demonstrable.The possible role of the stable positive charges on the surface of the virus at physiological pH is discussed from the point of view of the physico-chemistry of the hemagglutination.     

The effect of high altitude on the protein composition of human blood

Summary Twelve healthy young individuals had the protein composition of their blood determined in Stalinabad (850 m) and during their sojourn in the mountain of East Pamir between the months of May and October, 1958, at the altitude of 4200 m.During the first month of sojourn at high altitude, the total concentration of protein went up, while towards the end of the 4 month period it dropped somewhat, but still remained above the initial level. The relative and the absolute albumin content in the blood serum dropped immediately after the ascent and remained low for a month after descent from 4200 m to 850 m. The figures for ₁-, - and -globulins rose immediately after ascent. During the 4 month stay at high altitude, the ₁- -globulins went back to normal. During the first month after descent, figures for ₁-, - and -globulins were higher than the initial levels; ₂-globulin went up only after descent. Oncotic pressure of blood rose during the first month after ascent, then returned to normal at the expense of increased concentration of the globulin fraction, which compensates for the decreased albumin content in blood serum.(Presented by Active Member AMN SSSR S. R. Severin) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 50, No. 10, pp. 78–82, October, 1960.     

Über die Beziehungen zwischen dem Druck und der Stromstärke in der Arteria pulmonalis sowie der Leistung des rechten Ventrikels beim „unbeeinflußten Kreislauf“ der Katze und im oligämischen Schock

Zusammenfassung Die Druck-Stromstärkebeziehung in der Arteria pulmonalis und die Leistung des rechten Herzens wurden beim unbeeinflußten Kreislauf und im oligämischen Schock gemessen, sowie der Strömungswiderstand und der relative Gefäßquerschnitt berechnet. Die an Katzen durchgeführten Untersuchungen ergaben folgende Ergebnisse:1. Die Druckstromstärkekurve entspricht bei beiden untersuchten Kreislaufzuständen einer Geraden. Die Kurve des oligämischen Schocks ist gegenüber der des unbeeinflußten Kreislaufs im Koordinatensystem nach links zu kleineren Stromstärken und niedrigeren Druckwerten verschoben. Die Beziehung zwischen der wirksamen Druckdifferenz (mittlerer arterieller Druck — linker Vorhofdruck) folgt ebenfalls einer Geraden.2. Im oligämischen Schock ist die Leistung des rechten Herzens herabgesetzt (Einschränkung der Druckentwicklung und des Minutenvolumens bei erhöhtem rechten Vorhofdruck).3. Der Strömungswiderstand ist im oligämischen Schock kleiner als beim unbeeinflußten Kreislaufzustand. Er nimmt mit steigendem Druck ab. Der relative Gefäßquerschnitt verhält sich gegensinnig.4. Die Pulsfrequenz nimmt mit steigendem Blutvolumen in der Mehrzahl der Versuche ab. Sie ist im oligämischen Schock deutlich niedriger als beim unbeeinflußten Kreislauf.Mit 3 Textabbildungen     

Glomerulonephritis as late manifestation of severe αl-antitrypsin deficiency

An association of ₁-antitrypsin deficiency with glomerulonephritis is rare and has so far been observed only in children or young adults. We report a 63-year-old man with severe ₁-antitrypsin deficiency associated with pulmonary emphysema, cirrhosis of the liver, and mesangioproliferative glomerulonephritis with nephrotic syndrome. Following initial presentation with nephropathy, further work-up revealed ₁-antitrypsin deficiency of proteinase inhibitor Z. In the absence of glomerular ₁-antitrypsin deposits the relationship between renal disease and ₁-antitrypsin deficiency remains unclear. ₁-Antitrypsin deficiency should be considered in adults with abnormal renal function and chronic liver disease.Abbreviations A1AT ₁-antitrypsin - Pi proteinase inhibitor     

Investigation of the “valency” of antibodies by means of azoproteins

Summary The valency of antibodies was studied by the method of exhaustion of antisera against mono-and diazoproteins, and subsequent cross reactions both with the antibodies left over in the supernatant fluid of the serum and with the precipitating and nonprecipitating antibodies isolated from the precipitate.It was proved that the antibodies interact with the antigens as multivalent compounds.The valency determined with regard to the azoproteins is dependent upon the number of groups introduced.Thus, bivalent antibodies correspond to monoazoproteins and trivalent ones to diazoproteins.The valency of antibodies is, evidently, determined by the structural similarity of the heterologous and the immunizing antigens as well as by the less complete specific conformity between the individual structural peculiarities of the antigen and its antibody.From the Tashkent Pharmaceutical Institute (Director-Docent M. A. Azizov)Presented by Active Member AMN SSSR N. N. Zhukov-Verezhnikov     

Cardiac Purkinje fibres

Summary The influence of intracellular calcium concentration [Ca²⁺] _i on the steady state membrane currentsi was studied in a range of clamp potentials between –20 and –100 mV. Injection of CaCl₂ or Ca-EGTA (pCa 6) increasedi whereas injection of K-EGTA diminished it. The changes i were attributed to a change in steady state potassium conductance, g_K, by four arguments: i was restricted to potentials negative to –20 mV and depended on clamp potential in an inward rectifying manner. i displayed a reversal potential, E_rev, which followed log [K⁺]₀ with 60 mV for a tenfold change. Since E_rev obtained during Ca injection agreed with E_rev observed during EGTA injection the potassium driving force had to be constant. Wheng _K was blocked by superfusion with 20 mM Cesium neither CaCl₂ nor K-EGTA injection modifiedi .Supported by SFB 38, Membranforschung, project G2     

Distribution of renal integrin receptors and their ligands in congenital nephrotic syndrome of the finnish type

The aim of this study was to examine the distribution of 1 and v integrins (Ints) and some of their ligands in the kidneys of patients with congenital nephrotic syndrome of the Finnish type (CNF) and in controls using indirect immunofluorescence with monoclonal antibodies. The mesangial reactivity of Int 1 and Int 1 subunits was more variable and an increased glomerular reactivity with Int 3 and Int-6 antibodies was found in CNF kidneys than in controls. Int 2 subunit was either completely missing from or found in significantly lesser amounts in CNF kidney glomeruli. The immunoreactivity for Int v was more variable, fainter and also more granular in CNF samples than in control kidneys. The glomerular reactivity for Int 5 was more diffuse and weaker, and in sclerotic Bowman's capsules more intense in CNF kidneys than in controls. Immunoreactivity for Int 6 was restricted and was comparable in extent in CNF and control kidneys. Of the extracellular matrix components studied, the expression of EDAFn, EDBFn, OncFn, Ln 2 chain, Ln 1 chain and tenascin was increased. This is also seen in several glomerular diseases with inflammation and sclerosis. Immunoreactivity for vitronectin was decreased. Several differences were found in the intensity or location of the immunostaining for the 1 and v Ints and their ligands in CNF kidneys compared with controls, which have not been found in any other proteinuric disease. Disturbed Int expression pattern in CNF may specifically reflect the disturbance of glomerular function caused by the primary defect in this disease.