Defective signal transduction by the CD2 molecule in immature T-cell receptor/CD3- thymocytes. The CD2 accessory molecule mediates an activation pathway in mature T cells, transducing signals similar to those observed following stimulation of the T-cell receptor/CD3 (TCR/CD3) complex. CD2 is also one of the earliest cell surface markers to appear during thymic ontogeny and has been proposed to be a stimulatory pathway for immature thymocytes that have not yet expressed TCRs on their surface (TCR/CD3-). To examine this hypothesis highly purified TCR/CD3- human thymocytes were stimulated using mitogenic combinations of anti-CD2 monoclonal antibodies or individual biotinylated anti-CD2 monoclonal antibodies crosslinked with avidin. TCR/CD3+ thymocytes responded readily to either stimulus as determined by anti-phosphotyrosine immunoblotting, and the pattern of tyrosine phosphorylated substrates was similar to that of mature T cells. In contrast, TCR/CD3- thymocytes responded weakly and with a distinct substrate pattern. In addition, the altered signal transduced by CD2 in TCR/CD3- thymocytes did not lead to a rise in intracellular calcium, failed to induce interleukin 2 receptor expression, and did not serve as a comitogen with phorbol ester or interleukin 2, functions that were all intact in TCR/CD3+ thymocytes. Failure of TCR/CD3- thymocytes to respond to CD2 stimulation was not due to an intrinsic defect in these cells as they responded normally to phorbol ester plus calcium ionophore. In TCR/CD3- thymocytes, CD2 stimulation also failed to affect steady-state mRNA levels of the recombination-activating genes RAG1 and RAG2, whereas in TCR/CD3+ cells activation of the CD2 pathway terminated their expression. Together, these data support the concept that CD2 engagement does not deliver a stimulus to TCR/CD3- thymocytes and suggests that this molecule may not directly participate in the earliest stages of thymic development.


Evidence that the deep keratin filament systems of the Xenopus embryo act to ensure normal gastrulation. To study the role of keratin filaments in Xenopus development, fertilized eggs were injected with anti-keratin monoclonal antibodies. The anti-keratin monoclonal antibodies AE1 and AE3 induce abnormal gastrulation; in the most severely affected embryos gastrulation fails completely. In contrast, embryos injected with the anti-keratin antibody 1h5 develop normally. Immunocytochemical data indicate that injected 1h5 binds to the dense superficial keratin filament system of the embryo but not to the deeper keratin filament networks of ectodermal and subectodermal cells. Injected AE1 and AE3 do not bind to the superficial keratin system but appear to interact preferentially with the deep keratin filament systems of the embryo. We conclude that the superficial keratin filament system is not involved in the process of gastrulation per se but may protect the embryo from mechanical damage. On the other hand, our results suggest that the integrity of the deeper keratin filament systems is required for the mechanical integration of the morphogenetic movements that underlie gastrulation in Xenopus.


Escherichia coli MutY protein has both N-glycosylase and apurinic/apyrimidinic endonuclease activities on A.C and A.G mispairs. In Escherichia coli the mutY (or micA)-dependent DNA mismatch repair pathway can convert A degrees G and A degrees C mismatches to C.G and G.C base pairs, respectively, through a short repair-tract mechanism. The MutY protein has been purified to near homogeneity from an E. coli overproducer strain. Purified MutY has been shown to contain both N-glycosylase and 3' apurinic/apyrimidinic (AP) endonuclease activities. The N-glycosylase removes the mispaired adenines of A degrees G and A degrees C mismatches, and the AP endonuclease acts on the first phosphodiester bond 3' to the AP sites. The N-glycosylase and the nicking (combined N-glycosylase and AP endonuclease) activities copurified through multiple chromatographic steps without a change in relative specific activities. Furthermore, both N-glycosylase and AP endonuclease activities can be recovered by renaturation of a single polypeptide band from an SDS/polyacrylamide gel. Renaturation required the presence of iron and sulfide. These findings suggest that the MutY protein, like endonuclease III, is an iron-sulfur protein. DNA fragments with A degrees C mismatches were 20-fold less active than DNA with A degrees G mispairs as a substrate for purified MutY.


Studies on antigen specifity of immunoreactive arabinogalactan proteins extracted from Baptisia tinctoria and Echinacea purpurea. In a series of experiments the cross-reactivity of antibodies raised against arabinogalactan proteins from Baptisia tinctoria and Echinacea purpurea was studied in order to prove the antigen specificity of the extracted glycoproteins/polysaccharides. Using the antigen-antibody reaction in a competitive ELISA it was evident that antibodies against glycoproteins from Baptisia tinctoria were specific because none of the other antigens like those from Echinacea purpurea, Thuja occidentalis, arabinogalactan from larch, LPS from E. coli 055:B5, and from Salmonella typhimurium were able to inhibit the antigen-antibody reaction. The same results were obtained from ELISA experiments with Echinacea purpurea. From these studies it was concluded that the antigenic regions of immunoreactive proteins from both medicinal plants show structural differences.


Calcium-dependent increase in tyrosine kinase activity stimulated by angiotensin II. The cellular effects of numerous hormones and neurotransmitters, including the vasoactive agents angiotensin II (AngII) and [Arg8]vasopressin, are mediated in part by protein-serine threonine kinases activated by increase of cytosolic Ca2+ concentration. In this study, we have tested the ability of Ca(2+)-mobilizing agents to activate cellular tyrosine kinases. Treatment of intact GN4 liver epithelial cells with AngII rapidly (less than or equal to 15 sec) increased tyrosine kinase activity measured either in unfractionated cell lysates or in anti-phosphotyrosine immune complexes from detergent-solubilized cells. Increased phosphorylation of the exogenous substrate poly(Glu80Tyr20) (3- to 4-fold over control) by immunoprecipitated kinases closely paralleled the time- and dose-dependence of the appearance of tyrosine phosphoproteins in intact cells. This effect of AngII was mimicked by thapsigargin, a Ca(2+)-elevating tumor promoter. The ability of AngII, but not epidermal growth factor, to increase tyrosine kinase activity was blocked in cells loaded with the Ca2+ chelator bis-(O-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid. Dephosphorylation of immunoprecipitated proteins by tyrosine phosphatase treatment was accompanied by a 60-70% loss in in vitro kinase activity, suggesting that the AngII-sensitive kinase(s) are activated by phosphorylation in intact cells. These findings demonstrate a link between two widely occurring signaling pathways, the tyrosine kinases and the Ca2+ second-messenger system, and suggest the possible involvement of Ca(2+)-activated tyrosine kinases in the endocrine actions of AngII and [Arg8]vasopressin.


[The body--a symbol of the unconscious?]. Based on Lacan's differentiation between the real, the imaginary and the symbolic it is shown that Moser's connection between psychoanalysis and therapy is almost exclusively concentrated on the body in the sphere of the imaginary. According to Bittner, when the body is regarded symbolically, it represents a highly emotional "transcription" of the unconscious.


[Hepatitis C infection and liver cell carcinoma]. Epidemiological data disclose that the incidence of hepatocellular carcinoma (HCC) is increasing world-wide, whereas the number of cases positive for HBV-marker has remained almost stable, at least in Japan. Data from Europe show positivity of antibodies against hepatitis C virus (anti-HCV) in 72% of HBsAg negative cases with HCC and in 28% of patients positive for HBsAg. Nearly 90% of HBsAg negative patients with HCC showed a histology of cirrhosis or chronic active hepatitis in the noncancerous liver. Almost every third patient had a history of blood transfusion. These results suggest an increasing incidence of HCV - associated HCC's, as it already has been shown for patients suffering from chronic HBV infection.


[Growth factors and hematopoiesis. Physiopathology and clinical applications]. Recently biotechnologic progress has, through the technique of the recombinant DNA, allowed a low cost production of large amount of several growth factors. Such a large availability has made possible to either carry out deeper investigations on the physiopathology of the hemopoietic regulation and perform new therapeutic approaches under different pathologic conditions. The most interesting acquisition concerning the biology of hemopoiesis to which such investigations have addressed us is the inadequacy of the protocols adopted till now. Such protocols considered only a simple vision of an elective action of a given growth factor during an exact maturation period of a determined cell colony. On the contrary, it was possible to point out a close network of inter-relationship among the different factors, which sometime impedes a clear distinction for each single factor, between actions of competence and progression in the cell maturation phenomena. However, the present uncertainty pertaining to the regulation of hemopoiesis has not impede the performance of clinical trials with positive findings in several pathologic conditions. The administration of recombinant erythropoietin has for example allowed to intervene in a resolutive way on the anemia in uremic subjects, and seems giving satisfactory results also in subjects with non renal origin anemic conditions. Satisfactory results were also obtained through the use of the Granulocyte-Macrophage CSF and of the Granulocyte-CSF, which by preventing neutropenia have allowed the performance of more adequate chemotherapeutic protocols in neoplastic subjects. New interesting perspectives are now coming for the use of Interleukin-3 in the treatment of the aplastic anaemia.


Modulation of the dimerization of a transcriptional antiterminator protein by phosphorylation. The transcriptional antiterminator protein BglG inhibits transcription termination of the bgl operon in Escherichia coli when it is in the nonphosphorylated state. The BglG protein is now shown to exist in two configurations, an active, dimeric nonphosphorylated form and an inactive, monomeric phosphorylated form. The migration of BglG on native polyacrylamide gels was consistent with it existing as a dimer when nonphosphorylated and as a monomer when phosphorylated. Only the nonphosphorylated dimer was found to bind to the target RNA. When the dimerization domain of the lambda repressor was replaced with BglG, the resulting chimera behaved like an intact lambda repressor in its ability to repress lambda gene expression, which suggests that BglG dimerizes in vivo. Repression by the lambda-BglG hybrid was significantly reduced by BglF, the BglG kinase, an effect that was relieved by conditions that stimulate dephosphorylation of BglG by BglF. These results suggest that the phosphorylation and the dephosphorylation of BglG regulate its activity by controlling its dimeric state.


Salvage of infarcted myocardium by angiogenic action of basic fibroblast growth factor. Coronary collateral vessels reduce damage to ischemic myocardium after coronary obstruction. Factors that stimulate collateral formation are expected to have ameliorating effects on myocardial infarction. In a canine experimental myocardial infarct model, intracoronary injection of basic fibroblast growth factor (bFGF) improved cardiac systolic function and reduced infarct size. Treatment with bFGF increased the number of arterioles and capillaries in the infarct. Thus, the angiogenic action of bFGF might lead to a reduction in infarct size. The application of bFGF might bring about a therapeutic modality for the salvage of infarcted myocardium.


Control by asparagine residues of calcium permeability and magnesium blockade in the NMDA receptor. The N-methyl-D-aspartate (NMDA) receptor forms a cation-selective channel with a high calcium permeability and sensitivity to channel block by extracellular magnesium. These properties, which are believed to be important for the induction of long-term changes in synaptic strength, are imparted by asparagine residues in a putative channel-forming segment of the protein, transmembrane 2 (TM2). In the NR1 subunit, replacement of this asparagine by a glutamine residue decreases calcium permeability of the channel and slightly reduces magnesium block. The same substitution in NR2 subunits strongly reduces magnesium block and increases the magnesium permeability but barely affects calcium permeability. These asparagines are in a position homologous to the site in the TM2 region (Q/R site) of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors that is occupied by either glutamine (Q) or arginine (R) and that controls divalent cation permeability of the AMPA receptor channel. Hence AMPA and NMDA receptor channels contain common structural motifs in their TM2 segments that are responsible for some of their ion selectivity and conductance properties.


Malignant transformation by a mutant of the IFN-inducible dsRNA-dependent protein kinase. The double-stranded RNA-dependent protein kinase (dsRNA-PK) is thought to be a key mediator of the antiviral and antiproliferative effects of interferons (IFNs). Studies examining the physiological function of the kinase suggest that it participates in cell growth and differentiation by regulating protein synthesis. Autophosphorylation and consequent activation of dsRNA-PK in vitro and in vivo result in phosphorylation of the alpha subunit of eukaryotic initiation factor-2 (eIF-2) and inhibition of protein synthesis. Expression of a functionally defective mutant of human dsRNA-PK in NIH 3T3 cells resulted in malignant transformation, suggesting that dsRNA-PK may function as a suppressor of cell proliferation and tumorigenesis.


Regulation by ATP and ADP of CFTR chloride channels that contain mutant nucleotide-binding domains. Regulation of the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel is unusual in that phosphorylated channels require cytosolic adenosine triphosphate (ATP) to open. The CFTR contains two regions predicted to be nucleotide-binding domains (NBDs); site-directed mutations in each NBD have now been shown to alter the relation between ATP concentration and channel activity, which indicates that ATP stimulates the channel by direct interaction with both NBDs. The two NBDs are not, however, functionally equivalent: adenosine diphosphate (ADP) competitively inhibited the channel by interacting with NBD2 but not by interacting with NBD1. Four cystic fibrosis-associated mutations in the NBDs reduced absolute chloride channel activity, and one mutation also decreased the potency with which ATP stimulates channel activity. Dysfunction of ATP-dependent stimulation through the NBDs may be the basis for defective CFTR chloride channel activity in some cystic fibrosis patients.


Pharmacological manipulation of peripheral resistance during distal vascular reconstruction. A randomised placebo-controlled trial was conducted to investigate the effects of iloprost, a stable prostacyclin analogue, on peripheral resistance, during femoro-crural arterial bypass. 3000 ng of iloprost, infused into the graft, produced an immediate drop in peripheral resistance by a mean (range) of 43% (4-80%; p less than 0.01, Wilcoxon). Decreased peripheral resistance persisted to 20 minutes. Graft flow during the same period increased by 59.8% (-7 to 294%; p less than 0.01) and this increase persisted after the operation. Iloprost produces an immediate decrease in peripheral resistance associated with a prolonged increase in graft blood flow. This may reduce graft failure in the early postoperative period.


[Effects of mepartricin on the growth of hypertrophic prostatic tissue in primary culture. Creation of the experimental model and preliminary results]. The aim of this study was to standardise a method for the in vitro culture of hypertrophic prostatic tissue, to assay the morphological type of isolated cells, to evaluate the degree of proliferation and to identify their differentiated iter. In addition, the effects of mepartricina on growth was also studied obtaining preliminary results. Of a total of 40 biopsies used in this experiment, 30 were placed directly in culture using Freshney's method, whereas the remaining 10 were used in a method involving the primary culture of the dispersed cells obtained from enzymatically disaggregated tissue. In vitro proliferation was analysed using optic microscopy, histological and histochemical techniques, and a scanning electron microscope. Cellular kinetics were also studied by bromodeoxyuridine marking. Using these tests it was found that it is possible to obtain the development and growth in this form of culture of fibroblastic-type colonies of epithelial origin characterised by different morphologies and growth curves. In addition, cells from the epithelioid colonies are characterised by a high level of proliferative activity and a low degree of differentiation. Mepartricina appears, on the other hand, to inhibit the in vitro growth of hypertrophic prostatic tissue.


Dimethylnitrosamine (DMN)-induced IL-1 beta, TNF-alpha, and IL-6 inflammatory cytokine expression. Altered immune functions have been demonstrated in mice following exposure to dimethylnitrosamine (DMN). In particular, changes in cell-mediated immune responses resulted from chronic DMN exposure in vivo. Since cytokines are potent immunoregulatory peptides, experiments were performed to determine whether DMN exposure results in the induction of serum-borne inflammatory cytokines. Animals were exposed to either vehicle (PBS) or DMN (5.0 mg/kg) every 24 hr for 14 days. Serum and liver samples were obtained from individual mice at 0, 1, 2, 3, 6, 12, and 24 hr following the first exposure, with additional samples collected every 24 hr preceding the daily DMN exposure. Sera were then analyzed for IL-1 beta, IL-3, IL-6, CSF-1, GM-CSF, and TNF-alpha activities using either biological or immunological assays. In addition, liver total cellular RNA was probed for the induction of IL-1 beta transcripts using the solution hybridization/RNase protection assay. IL-1 beta, IL-6, and TNF-alpha serum activities were observed within 2 hr of DMN exposure and returned to vehicle control levels by 3 days even though DMN exposure was maintained. Chronic expression of cytokine activity (after 72 hr) was only observed for GM-CSF. A rapid induction of IL-1 beta transcripts (within 1 hr) in both vehicle and DMN-treated animals was observed by solution hybridization. However, by 3 hr postexposure, transcript levels decreased in the vehicle-treated animals while remaining elevated in the DMN-treated animals for 6 hr. These results demonstrated that DMN exposure in vivo induced: (1) the expression of serum-borne cytokine activities, and (2) IL-1 beta transcription in liver tissue.


Neutralization of myotoxic phospholipases A2 from the venom of the snake Bothrops asper by monoclonal antibodies. The neutralization of two myotoxic phospholipases A2 from the venom of Bothrops asper, myotoxins I and II, by two murine monoclonal antibodies is reported. The monoclonal antibodies, MAb-3 and Mab-4, recognize different epitopes of the toxin. Both antibodies completely neutralized myotoxic activity of myotoxin I and crude venom. MAb-3 also completely neutralized myotoxic activity of myotoxin II, a lysine-49 phospholipase A2 isoform, whereas MAb-4 neutralized this toxin only partially. MAb-3 neutralized myotoxin II at a molar ratio of 1:1, showing a higher efficiency than affinity-purified polyclonal antibodies. A dissociation of myotoxic and enzymatic activities of myotoxin I was observed with both monoclonal antibodies.


Finding new genes in the nervous system by cDNA analysis. There are several methods to isolate cDNAs that do not require prior biochemical characterization of their protein products, including isolation of new members of particular protein families based on sequences of known members, selection of specific clones by differential screening, and shotgun analysis of a particular cDNA population. This third approach will produce a huge catalogue of cDNA sequences and anatomical data that will be useful to investigators with diverse interests. Recent methodological advances allow cDNAs to be isolated and sequenced rapidly. These methods, coupled with fast and reliable techniques for in situ hybridization histochemistry, should permit cDNA catalogues to be assembled efficiently.


The nerve growth factor family of receptors. The neurotrophins, of which nerve growth factor (NGF) is the best known example, support the survival and differentiation of chick embryo sensory neurons at extremely low concentrations, 10(-12) M or less. These same neurons display two different classes of neurotrophin receptors with dissociation constants of 10(-11) M and 10(-9) M, respectively, implying that only low occupancy of the higher affinity receptor is required to mediate the biological actions of the neurotrophins. Two structurally unrelated receptors have now been characterized for NGF, and one of them, p75NGFR, serves as a receptor for all the known neurotrophins. This is the receptor with a dissociation constant of 10(-9) M. The second NGF receptor is a member of the trk family of tyrosine kinase receptors, p140trkA. Other members, p145trkB and p145trkC, are receptors for brain-derived neurtrophic factor (BDNF) and neurotrophin-4 (NT-4) and neurotrophin-3 (NT-3), respectively, when assayed in fibroblasts. The specificity of neurotrophin binding to these receptors appears to be much higher in neurons than in the non-neuronal cells. The receptor p140trkA has many of the properties of the higher affinity class of NGF receptors, and is able to mediate survival and differentiation of the PC12 cell line, and cell growth and transformation in fibroblast cells. On the other hand, expression of p75NGFR in several types of cells displaying p140trkA induces a component of higher affinity NGF binding not seen in its absence. Since it is unlikely that p75NGFR and p140trkA interact at the level of the receptors, the crosstalk between receptors probably occurs through their signal transduction mechanisms.(ABSTRACT TRUNCATED AT 250 WORDS)


Vive la différence!

Vive la différence! Hormonal effects are increasingly recognized as important influences on neuronal function and, ultimately, on animal behavior. Such 'higher' behavioral effects are well studied, particularly in relation to sexually dimorphic behaviors. Yet, somewhat surprisingly, a significant proportion of more basic neuroscience research papers fail to specify the sex of the subjects used. In this brief article Karen Berkley argues that knowledge of, and controlling for, the sex of research animals is important. In addition, if females are used, their reproductive-cycle status could provide a deliberate strategy to investigate the effects of gonadal steroid hormones on biological functions.


Protein kinase C modulation of NMDA currents: an important link for LTP induction. A brief high frequency tetanic stimulation of afferent fibers induces a long-term potentiation (LTP) of synaptic transmission, which is manifested by an increase in the size of the synaptic response elicited by low frequency stimulation of the same synapse. LTP persists for several hours in vitro and up to several weeks in vivo, and is at present the most extensively studied form of activity-dependent synaptic plasticity. This article focuses on the relationship between two key elements in the induction of LTP--the N-methyl-D-aspartate (NMDA) subtype of glutamate receptor and the Ca(2+)-phospholipid-dependent protein kinase C (PKC). In view of several recent findings that describe a direct positive modulation of NMDA currents by PKC, we suggest that PKC activity may, in fact, determine the threshold of LTP induction. Enhanced kinase activity may underlie the central role of the NMDA receptor--channel complex in neuronal plasticity.


Individual differences in the cognitive and neurobiological consequences of normal aging. Defining the neural basis of age-related cognitive dysfunction is a major goal of current research on aging. Compelling evidence from laboratory animals and humans indicates that aging does not inevitably lead to cognitive decline. Conducting neurobiological investigations in subjects that have previously undergone behavioral characterization has therefore emerged as a promising strategy for identifying those alterations in brain structure and function that are specifically associated with age-related cognitive impairment.


Voltage-gated ion channels in Schwann cells and glia. In the past decade, particularly its last half, it has become clear that the satellite cells of the nervous system (oligodendrocytes and astrocytes in the CNS, Schwann cells in the PNS) are liberally endowed with a vast array of voltage-gated ion channels. These original observations of the various channel types have been followed by the elucidation in considerable detail of the electrophysiological characteristics of the channels. Virtually all of the types of voltage-gated channel found in neurons have now also been found in the various populations of satellite cells. In spite of this, the intriguing questions concerning the physiological function of these channels remain to be fully elucidated.


Distribution and functional significance of the P-type, voltage-dependent Ca2+ channels in the mammalian central nervous system. In addition to the three types of voltage-dependent calcium channels presently recognized in the CNS, the L-, the T- and the N-types, a fourth distinct type known as the P-type channel has recently been described. This channel, initially recognized in Purkinje cells (and thus the name), is not blocked by dihydropyridines or by omega-conotoxin (GVIA), but is blocked by native funnel-web spider venom and by a polyamine (FTX) extracted from such venom. In addition, a synthetic polyamine (sFTX) has been produced that also specifically blocks P-channels in brain slices and at the neuromuscular junction, and blocks presynaptic Ca2+ currents in other vertebrate and invertebrate forms, as well as channels expressed in Xenopus oocytes following CNS mRNA injections. Using sFTX to form an affinity gel, a protein was isolated and reconstituted into lipid bilayers where it manifests single-channel properties that are electrophysiologically and pharmacologically similar to those of the native P-channels. Rabbits immunized with the isolated protein produced a polyclonal antibody that gave a positive western blot with the purified P-channel protein and generated a reaction product at specific sites in the CNS that agree with the physiological distribution of P-channel activity.


Structural basis of voltage-gated K+ channel pharmacology. Major advances have been made in understanding the domains and amino acid sidechains important for the function of voltage-gated K+ channels, by combining recombinant DNA techniques with pharmacological and electrophysiological approaches. As explained in this review by Olaf Pongs, the results of these experiments have enabled description of a detailed model of the K+ channel pore structure and provide an exciting picture of how open-channel blockers occlude the pore of K+ channels. Since the pore is a highly conserved structure among voltage-gated K+ channels, there are only limited possibilities for open K+ channel blockers to distinguish between the many distinct voltage-gated K+ channels, which have diverse kinetic and conductance properties.


Distinctive pattern of infection and replication of HIV1 strains in blood-derived macrophages. The macrophage-tropic virus HIV1-PAR, isolated from cerebrospinal fluid of HIV1-seropositive man, induced cytopathic effect accompanied by different magnitude of the virus production in blood-derived macrophages (BDM) obtained from different donors. HIV1-PAR-specific RNA was detected by in situ hybridization in 15 and 66% of BDM producing low and high levels of virus, respectively. In contrast with HIV1-PAR, infection of BDM with two laboratory strains adapted to T-cell lines, HIV1-LAV prototype and HIV1-NDK, a Zairian virus that is highly cytopathic for T-lymphocytes, resulted in a low production of HIV1 p24gag in culture fluid. Expression of HIV1-LAV and HIV1-NDK RNA was detected by in situ hybridization in a maximum of 1% of macrophages. Only HIV1-NDK, and not HIV1-LAV, induced ultrastructural alterations in BDM. In contrast with a striking difference in the production of macrophage-tropic and T-lymphotropic viruses, no significant differences were found in the proportion of macrophages containing retrotranscribed genomes of HIV1-. HIV1 DNA was detected by in situ hybridization in 93, 100, and 80% of macrophages infected with HIV1-PAR, HIV1-LAV, and HIV1-NDK, respectively. A higher level of HIV1 DNA was detected by polymerase chain reaction in the BDM infected with HIV1-PAR than in that infected with HIV1-LAV and HIV1-NDK. The results indicate that both macrophage-tropic as well as T-lymphotropic viruses can enter and retrotranscribe their genomes in a vast majority of macrophages.


Amino acid residues of the human immunodeficiency virus type I gp120 critical for the binding of rat and human neutralizing antibodies that block the gp120-sCD4 interaction. We have characterized the discontinuous epitopes recognized by two rat and three human neutralizing monoclonal antibodies (mAb) by examining the effect of single amino acid changes in conserved residues of gp120 on mAb recognition. A human mAb derived from an infected individual, 448D, and two rat mAbs, 39.13g and 39.3b, respectively, derived by immunization with native recombinant gp120, recognize similar epitopes. Recognition of the envelope glycoproteins by these mAbs was affected by changes in gp120 amino acid residues 88, 113, 117, 257, 368, or 370. The gp120 amino acids 257, 368, and 370 have previously been reported to be important for CD4 binding, which is consistent with the ability of these mAbs to block the gp120-CD4 interaction. Residues 88, 113, and 117 are not thought to be important for CD4 binding, suggesting that the antibody epitopes overlap, but are distinct from, the CD4 binding region. We also found that some alterations in gp120 residues 88, 117, 368, or 421 reduced the ability of polyclonal sera from HIV-1-infected individuals to inhibit the interaction of the mutant gp120 glycoproteins with soluble CD4. Thus, changes in the HIV-1 gp120 glycoprotein that minimally affect the receptor binding may allow escape from neutralizing antibodies directed against the CD4 binding region.


Viral spliced RNA are produced, encapsidated and reverse transcribed during in vivo woodchuck hepatitis virus infection. By the use of reverse transcription followed by polymerase chain reaction (RT-PCR), we have identified one shorter than full-length, pregenomic viral RNA species in liver samples of woodchucks chronically infected with the woodchuck hepatitis virus (WHV). The spliced WHV RNA of about 2.4 kb in length was cloned and partially sequenced. The splicing donor and acceptor sites of this novel RNA are located, respectively, 130 nucleotides downstream of the ATG initiation codon of the core gene and 21 nucleotides upstream of the initiation codon of the pre-S2 surface gene. The splicing event generates a new core-polymerase fusion protein and removes the terminal protein domain and the spacer region of the polymerase gene. A nucleotide probe specific for the splice junction was used following RT-PCR, to further confirm the existence of this spliced RNA in the liver of seven WHV-infected woodchucks. Deleted viral DNA molecules corresponding to the 2.4 kb spliced RNA were also detected in the liver and, to a lesser extent, in the serum of infected woodchucks, suggesting that this spliced RNA can be encapsidated and reverse-transcribed during the course of natural WHV infection.


Prevention of the spread of HIV-1 infection with nonnucleoside reverse transcriptase inhibitors. Certain bisheteroarylpiperazines (BHAPs) directly inhibit the replication of human immunodeficiency virus type 1 (HIV-1) and block the spread of infection to susceptible populations of cells. At a 1 microM concentration three analogs, U-87201, U-88204, and U-89674, inhibited the replication of HIV-1 in MT-2 cells by 83, 100, and 93%, respectively. At the same concentration, U-88204 completely inhibited replication of primary HIV-1 isolates in peripheral blood mononuclear cells. Replication of 3'-azido-2',3'-dideoxythymidine (AZT)-resistant strains of HIV-1 was also inhibited by U-88204. When MT-2 cells that were lytically infected with HIV-1 were mixed with uninfected MT-2 cells, U-88204 provided complete protection to the uninfected cells. Integrated proviral DNA sequences were not detected by the polymerase chain reaction technique in this culture after 15 days in the presence of drug. The resultant healthy cell culture was subsequently maintained without drug with no evidence of latent proviral DNA. Serial passage of a laboratory strain and a primary isolate of HIV-1 in cell culture in the presence of increasing concentrations of U-88204 yielded virus populations which were at least 100-fold resistant to the drug. These resistant viruses also showed cross-resistance to the pyridinone class of nonnucleoside inhibitors but were sensitive to AZT. Analysis of the nucleotide sequence of resistant viruses revealed mutations at conserved regions of the reverse transcriptase (RT) gene. The results presented here suggest the therapeutic potential of U-88204 in the combination therapy for HIV-1 infection.


[Immunologic disorders in patients with idiopathic subretinal neovascular membrane]. The authors analyze the results of comprehensive clinical and immunologic examinations of 52 patients with this condition (SNVM) in its active and inactive phases, angiographically confirmed, aged 19 to 46; 32 of these were women (61.5%), 20 (38.4%) men. A complex of immunologic methods was employed to reveal the etiology of the condition and the immune status of SNMV patients. The origin of the condition was identified in 44.2% of patients; it was toxoplasmic in 19.2, autoimmune in 15.4, tuberculous in 7.7, and mixed toxoplasmic and tuberculous in 1.9% of cases. T lymphocyte functional activity was inhibited in 37.5% of patients with the exudative hemorrhagic phase of SNMV and in 16.7% of those in the cicatrization phase. Changes in the humoral immunity presenting as gammapathies were detected in 34% of patients, those manifesting by hyperconcentrations of circulating immune complexes in 66.7%, and of cryoglobulins in 37.8% of patients. Krypton laser coagulation was employed in multiple-modality treatment of patients with SNMV. The remissions persisted for 3-5 years.


Antibody response to human papovavirus JC (JCV) and simian virus 40 (SV40) T antigens in SV40 T antigen-transgenic mice. Human papovavirus JC (JCV) and simian virus 40 (SV40) genomes share approximately 69% homology; and there is antigenic cross-reactivity between JCV and SV40 tumor or T antigens. In order to determine whether a selective immune response to JCV T antigen could be demonstrated, transgenic mice (SV11+) that express SV40 T antigen in the choroid plexus and are partially tolerant to antigenic determinants on SV40 T antigen were immunized with SV40 or JCV T antigens and their antibody responses were analyzed. The results show that SV11+ mice responded as well as their nontransgenic litter mates to JCV T antigen. Monoclonal antibodies were derived from hybridomas generated from immunized mice which reacted specifically with epitopes in the amino and carboxy terminal halves on JCV T antigen. These studies show that transgenic mice expressing SV40 T antigen are capable of responding to determinants not shared between JCV and SV40 T antigen.


The transmembrane domain of influenza A M2 protein forms amantadine-sensitive proton channels in planar lipid bilayers. In a direct test of the hypothesis that the M2 coat protein of influenza A can function as a proton translocator, we incorporated a synthetic peptide containing its putative transmembrane domain into voltage-clamped planar lipid bilayers. We observed single proton-selective ion channels with a conductance of approximately 10 pS at a pH of 2.3, consistent with the association of several monomers around a central water-filled pore. The channels were reversibly blocked by the anti-influenza drug amantadine. These experiments imply a central role for M2 protein in virus replication and assembly and may explain the mechanism of action of amantadine. Analogous proteins may have a similar function in other viruses, and these may be susceptible to similar antiviral agents.


Herpesvirus saimiri has a gene specifying a homologue of the cellular membrane glycoprotein CD59. Herpesvirus saimiri (HSV) is a T-lymphotropic tumor virus that causes fulminant lymphomas and leukemias in various New World primates other than its natural host, the squirrel monkey (Saimiri sciureus). In the course of completing the nucleotide sequence of its genome, we identified an open reading frame of 363 nucleotides, designated HVS-15, that has no detectable homology to any other viral sequences to date. HVS-15 encodes a 121-amino-acid protein which shows significant similarities to human CD59, a phosphatidyl-inositol-glycan-anchored glycoprotein involved in T-cell activation and restriction of complement-mediated lysis. The predicted HVS-15 gene product is more similar to human CD59 than to the related murine Ly-6 antigens. A nucleotide sequence identity of 64% was found between HVS-15 and the CD59 reading frame, and a 48% identity exists between the corresponding protein sequences. The comparison of the amino acid sequences revealed a number of conserved structural features such as a similar pattern of hydrophobic termini and an identical cysteine skeleton.


HPV-1 L1 protein expressed in cos cells displays conformational epitopes found on intact virions. Seven polyclonal and monoclonal antibodies were characterized for their ability to react specifically with either conformational or nonconformational epitopes of the HPV-1 virion. Using these antibodies, it was shown that the HPV-1 L1 protein (when expressed by an SV40 vector in cos cells) displayed conformational epitopes characteristic of intact viral particles. In addition, the L1 capsid protein was translocated normally into cell nuclei, was of appropriate size (57 kDa), and could be isolated in native form by immunoprecipitation techniques. Most importantly, the screening of expressed papillomavirus capsid proteins for reactivity with conformation-dependent antibodies represents a new, general methodology for ensuring that such proteins will be suitable for use in vaccine development or in the serologic detection/typing of human papillomavirus infections.


The innervation of the great saphenous vein: an immunohistochemical study with special regard to regulatory peptides. The human great saphenous vein is often used in by-pass surgery. In spite of its importance little is known about the neuronal regulation and innervation of this vessel. On the other hand, therapeutic phlebectomy of the saphenous vein provides a good basis for investigations on the nervous supply of human veins. In former studies the human great saphenous vein has been proved to be a richly innervated portion of the low pressure vascular system. We studied the innervation of the proximal femoral saphenous vein by immunohistochemical methods and with special regard to regulatory neuropeptides. A peptidergic innervation mainly localized along the vasa vasorum and associated with immunoreactivity of substance P and CGRP was found which yields evidence for a mechanosensory innervation of this vascular segment.


The expression of insulin-like growth factor binding proteins is tissue specific during human fetal life and early infancy. The insulin-like growth factors (IGFs) are bound to multiple IGF binding proteins (IGFBPs) that are present both in the circulation and in extracellular fluids. There are at least six different IGFBP species that have been fully characterized in terms of molecular structure and amino acid sequence. The tissue distribution and local production of these proteins as well as the regulation of IGFBP production in different tissues have not been elucidated. We have studied the distribution of multiple IGFBP species in protein extracts from human kidney, skeletal muscle, lung, liver and brain by ligand blotting employing [125I]IGF-2 as the radiolabeled hormone. Five distinct IGFBP species with a respective molecular weight of 43, 38, 34, 30 and 20 kDa were detected on the ligand blots in tissues from human fetuses and infants (23 weeks of gestation till 24 months of postnatal age). The 34 kDa species and a 30-32 kDa IGFBP species were predominant in brain, whereas a 30 kDa IGFBP species was mainly detected in skeletal muscle. Immunoblotting experiments using an anti IGFBP-2 antiserum showed that the 34 kDa IGFBP species from human brain was presumably related to IGFBP-2. We conclude that IGFBPs are differentially expressed in different tissues throughout human fetal life and early infancy. Local production or accumulation of the different IGFBPs could modulate IGF action at a local level or alternatively have differential functions during development.


Histochemistry of the porcine pilosebaceous unit. The present study describes lectin and immunoreactivity in the pilosebaceous unit of porcine skin. Complex carbohydrates of mucin and biantennary Man/Gluc types were distributed among hair follicle epithelia (hair root sheaths, cuticula, shaft, and shaft matrix). Sebaceous glands expressed biantennary Man/Gluc carbohydrates and GalNAc residues. The expression of simple-type and epidermis-like keratins was confirmed by immunohistochemistry with monoclonal antibodies. Filaggrin-positive cells were found in the keratinizing zone of Henle's layer in anagen follicles. The innermost layer of the outer hair root sheath was stained with antibodies against the epidermal growth factor-receptor, keratin 10 and Ki67 antigen. The differences to humans were remarkably small.


DNA cytochemistry in polytene chromosomes: electron contrasting agents for the ultrastructural detection of chromatin DNA after alkaline hydrolysis/methylation-acetylation. Salivary glands from Chironomus tentans larvae were fixed in glutaraldehyde and either subjected to alkaline hydrolysis followed by methylation-acetylation, or dehydrated without these treatments as controls. Ultrathin sections from Durcupan-embedded samples were contrasted by means of uranyl acetate, ruthenium red, indium trichloride, or the complex indium (III)-hematoxylin. Electron microscopic observations revealed a general contrasting pattern in control sections, while after the hydrolytic and blocking procedure only chromatin from polytene chromosomes appeared selectively contrasted. The nucleolus, Balbiani ring granules and puff materials showed weak or no electron opacity. After toluidine blue staining of semithin sections, an orthochromatic blue colour was found in chromatin bands from treated samples. These results indicate that alkaline hydrolysis/methylation-acetylation followed by contrasting with cationic heavy compounds is a valuable procedure to visualize chromatin DNA in polytene chromosomes.


Influence of inorganic salts on the staining reaction of eosinophil leukocyte granules by anionic dyes. The use of salts as competing agents in staining solutions allows to evaluate to what extent ionic interactions take place between microscopical substrates and cationic or anionic dyes. We have employed this method to study the staining reaction of horse eosinophil leucocyte granules by the anionic dyes nuclear fast red, naphthol yellow S, eosin Y, indigocarmin, acid fuchsin, alizarin red S, orange G, and Evans blue in the presence of NaCl (from 0.015 to 2 mol/l). Different values of "minimal electrolyte concentration" (the least amount of salt which reduces the staining intensity) were found for these dyes. Comparative observations using ammonium sulphate as competing salt showed that it is less effective than NaCl. Staining of eosinophil granules by anionic dyes could be not suppressed at 2 mol/l NaCl, which indicates that in addition to electrostatic forces, other non-ionic interactions are also responsible for some staining reaction of these acidophilic structures.


Patterns of cytokeratins and vimentin in guinea pig and mouse eye tissue: evidence for regional variations in intermediate filament expression in limbal epithelium. The anatomical distribution of different individual cytokeratin polypeptides and of vimentin was investigated by means of immunofluorescence with 41 monoclonal antibodies in guinea pig and mouse eyes. Simple epithelial type cytokeratins 7, 8, 18, and 19 selectively decorated conjunctival goblet cell clusters in mouse specimens and a continuous superficial cell layer of the corresponding part of guinea pig conjunctiva. A changed pattern of squamous epithelial type cytokeratins was found in the limbal region of the guinea pig eye as compared to the corneal epithelium. Cytokertains 3 and 17, which stained the entire corneal epithelium, were not detected, whereas cytokeratin 4, 5 and 13 were expressed. A focal vimentin and cytokeratin coexpression in the limbus of guinea pig is interpreted as indicating corneal stem cells. Similar patterns of expressions were found in the mouse ocular surface. In both species, a cytokeratin 4 staining of basal conjunctival epithelial cells could be detected. The neuroectodermally derived epithelia of the eye such as the retinal pigment epithelium and the ciliary body epithelia expressed solely the cytokeratin pair 8/18.


Immunoelectron microscopic localization of microsomal alanine aminopeptidase. The localization of microsomal alanine aminopeptidase was investigated in the rat kidney. Resin embedding failed to demonstrate the localization of the enzyme by immunogold labelling. Using a cryo-ultramicrotomy method the enzyme could be detected on the luminal side of the brush border membrane of proximal tubular cells and to a lesser degree in their mitochondria. Furthermore, vesicular structures labelled with gold were found in the cytoplasma in the apical region of these cells.


Characterization of secretory cell glycoconjugates in the alimentary tract of the ruin lizard (Podarcis sicula campestris De Betta) by means of lectin histochemistry. Secretory cell glycoconjugates of the alimentary canal of the ruin lizard (Podarcis sicula campestris De Betta) were characterized by traditional staining methods and by lectin histochemistry. The goblet cells of the upper esophagus produced sialo- and sulfomucins, while those of the lower esophagus mainly contained sulfomucins. Lectin histochemistry demonstrated the presence of N-acetyl-D-glucosamine and terminal sialic acid. The epithelial mucous cells lining the surface of the stomach and the gastric pits contained neutral glycoproteins with glycosidic residues of N-acetyl-D-galactosamine, D-glucose, D-mannose, D-galactose, and N-acetyl-D-glucosamine. The mucous cells of the gastric glands produced neutral glycoproteins that contained stable class-III mucosubstances, as revealed by Paradoxical Con A staining, with terminal residues of L-fucose and D-galactose. They can be similar to the true neck cells of the gastric pits of other vertebrates. The goblet cells of the small intestine produced acidic glycoproteins with glycosidic residues of N-acetyl-D-glucosamine, sulfated esters on internal residues and terminal sialic acid. In the large intestine, there is a predominance of sulfated mucosubstances with D-galactose, N-acetyl-D-glucosamine, and N-acetyl-D-galactosamine. The microheterogeneity of mucins of the digestive tract, as proved by lectin histochemistry, is probably connected to their different functions.


The rib perichondrium. An anatomical study in sheep of a tissue used as transplant in the treatment of hyaline-cartilage defects. Perichondrial tissue is of great importance as a transplant for the repair of hyaline-cartilage defects of synovial joints because of its potential to produce hyaline-like cartilage. The histological and histochemical character and fiber orientation of rib perichondrial grafts prior to a transplantation procedure for the repair of full-thickness hyaline-cartilage defects is analyzed in a sheep model. Compared to other descriptions of the rib perichondrium of other different species and humans, the perichondrial architecture of different origins demonstrates the same histological, histochemical character and fiber orientation. The three different layers of this tissue are: the outer fibrous layer, the central part called proliferation zone and the inner part towards the underlying rib called transition zone. The fibrous layer demonstrates a wave-like extracellular configuration and contains a few fibrocytes. The proliferation zone is only a few cell rows thick and contains single cells with an oval shape and longitudinal fibrocyte-like nucleus. Because of the potential of these cells to produce cartilage this layer is called proliferation or cambium layer. The transition zone contains more oval-shaped cells with rounded nuclei and demonstrates a more chondrocytic character. Some cells lie in groups of 2 and more cells like in chondrons. The extracellular matrix shows an increasing stain towards the rib cartilage indicating new cartilage production. The fiber orientation is longitudinal in the fibrous part and vertical in the transition zone. Between these layers, fibers deviate from the longitudinal to the vertical orientation corresponding to the light-microscopically visible proliferation zone.


Fine structure of the dorsal lingual epithelium of the Japanese monkey Macaca fuscata fuscata. Filiform papillae, which were densely distributed all over the dorsal surface of the lingual body, were crown-shaped, with a central, circular area that sloped in the anterior direction and several branches that surrounded it in a semicircle from the back of the central area. Dome-shaped, fungiform papillae were scattered among these filiform papillae. At the posterior end of the lingual body, there were four circumvallate papillae. Prominent microridges and elevated intercellular borders were widely distributed in the central area of the filiform papillae and the interpapillar region. On the surface of the branches of the filiform papillae, microridges were rarely seen. On the surface of the fungiform papillae, indistinct microridges were observed. Histologically, the dorsal lingual epithelium revealed three different regions: the epithelium on the anterior side of the filiform papillae, the epithelium on the posterior side of the filiform papillae and the interpapillar epithelium. Whereas the basal and suprabasal cells are similar throughout, differences characterize the intermediate and surface layers. Keratohyalin granules appear predominantly in the intermediate layer in the epithelium on the anterior side of filiform papillae. In the epithelium on the posterior side of the filiform papillae, no keratohyalin granules occur and, instead, tonofibrils are prominent. The cells become significantly flattened. In the interpapillar epithelium, no keratohyalin granules are visible, and the tonofilaments occupy almost the entire cytoplasm of most cells in the intermediate and surface layers. The cells are larger in volume in these layers.


[Development of resistance of herpes simplex virus to antivirus drugs in vitro]. Resistant variants of HSV-1 wt strain to ACV, CC, DHPG and PFA can be selected after several passages in drug-containing cultures. The degree of resistance of different variants varied and emerged at different times. The combination of antiviral agents can delay and even prevent the emergence of drug-resistant variants. At the same time, the concentration, dose and, therefore, toxicity of the antiviral agents can also be reduced. So, it is a feasible method for treating resistant HSV infections. Cross-sensitivity tests showed that ACVr variants are resistant to ACV, DHPG and PFA, but sensitive to CC; PFAr variants are sensitive to ACV, CC and DHPG; CCr variants are sensitive to ACV and DHPG, but resistant to PFA; DHPGr variants are sensitive to ACV, CC and PFA. These results suggest that the study of cross-sensitivity of HSV strains in vitro provide information which will aid the design of suitable therapy for drug-resistant HSV infections.


Myocardial force interval relationships: influence of external sodium and calcium, muscle length, muscle diameter and stimulation frequency. Several inotropic interventions were studied in thin papillary muscles under dynamic conditions. The effects on mechanical restitution and postextrasystolic potentiation were analysed. The decay of postextrasystolic potentiation was taken as a measure of recirculation fraction of activator calcium. The mechanical restitution curve had a plateau phase on its rising phase which was abolished in low extracellular sodium but pronounced in increased extracellular calcium. The recirculation fraction (RF) in control was 0.35 +/- 0.03; lowering the extracellular sodium by 20% increased the RF to 0.46 +/- 0.04 (n = 10). A reduction of sodium by 40% increased the RF to 0.57 +/- 0.04, whereas increasing extracellular calcium to 4 mM gave an RF of 0.48 +/- 0.05 (n = 10 in all cases). There was no significant effect on RF of changing basic stimulation frequency or muscle preparation length. These findings support RF as a good index of myocardial contractility. Furthermore, at muscle diameters above 0.65 mm the RF was found to be reduced, suggesting this diameter as critical for muscle function. Also, postextrasystolic potentiation in relation to preceding steady state contraction was markedly increased at these diameters. In conclusion, this study shows that RF is independent of stimulation frequency and muscle length, and that it is increased when calcium extrusion by the sodium/calcium exchange is reduced. Furthermore, RF is critically dependent upon the diameter of the preparation and mechanical restitution is changed by altered extracellular sodium concentration.


Effect of free radicals on excitation-contraction coupling in isolated rat and guinea pig ventricular myocytes. Using the model of isolated myocytes, we have investigated under current- and/or voltage-clamp conditions: (i) the time-course of H2O2-induced changes in the action potentials and the contractility. (ii) the effect of pharmacological tools such as ryanodine, TTX and nifedipine on H2O2-induced changes; (iii) the effect H2O2 on ionic currents underlying the changes in APD; and (iv) the effect of deferoxamine on H2O2-induced changes.


Potassium channel activation in vascular smooth muscle. 1. Numerous compounds and changes in physical state functions shift the membrane potential of vascular smooth muscle to more negative values. The consequence is a vasodilatation because Ca2+ channels are closed. K+ channel opening frequently causes the hyperpolarization. 2. Acidification of the blood substitute solution and a fall in O2 partial pressure dilate arterial vessels. Acidosis is associated with a rise in K+ permeability and a simultaneous fall in Na+ permeability. Prostacyclin has a 20-30% share, and EDHF a 70-80% share, in hypoxic vasodilatation. Experiments with iloprost (PGI2 analogue) confirmed the K+ channel opening properties of this drug. A voltage-dependent K+ channel and a Ca(2+)-activated K+ channel, via the influence of cA-PK or cG-PK, are responsible for the hyperpolarization with iloprost and with oxygen deficiency. 3. Cicletanine and ajoene cause a concentration-dependent membrane hyperpolarization and are potent vasodilators. A cicletanine concentration, which is attained by the dosage given to patients, is sufficient to produce these effects. Ajoene exerts a hyperpolarizing and vasodilating influence even in a concentration which may occur in the extracellular space by the administration of a single garlic clove. 4. The stationary activation curve 'developed force vs. membrane potential' satisfactorily explains the effects of K+ channel openers. The tight electromechanical coupling expressed by this curve comprises a 50% vasorelaxation for a 2.5 mV hyperpolarization. In the linear part of the curve, the coupling ratio is 5.1 mV/g. 5. In the vascular smooth muscle, vasorelaxation can be evoked by membrane hyperpolarization which is linked to a simultaneous increase in K+ outward current and 42K+ efflux. In the case of substances whose influence is solely or partially receptor-mediated, cyclic nucleotides may be involved in vasorelaxation. Since cyclic nucleotides also hyperpolarize through an increase in K+ conductance, the resulting dilatation often cannot be divided into its single components. Therefore, it is sensible not to give the term "K+ channel opener" too fine a definition. The term should be applied to all substances and changes in physical states which predominantly increase the open probability of K+ channels finally via a conformational change in the cell membrane. For example, giving an acidic blood substitute solution (acidosis) is an intervention opening K+ channels. Which K+ channel and which single channel conductance is concerned in a particular case, and which 'mediator' may participate, become secondary questions.


The effect of prednisolone on substance P-induced vascular permeability in mice. The effect of prednisolone on the substance P (SP)-induced vascular permeability increase in male ddY, WBB6 F1(-)+/+ (control) and WBB6 F1-W/WV (no mast cell in skin or internal organs) mice was investigated. 1) SP (1-10,000 pg/site) increased vascular permeability in ddY, WBB6 F1(-)+/+ and WBB6 F1-W/WV mice ears. 2) SP (100 pg/site)-induced vascular permeability was inhibited by prednisolone (10 mg/kg) administered intraperitoneally 3 to 12 hours prior to the elicitation of the reaction in ddY mice. When dexamethasone at a dose of 1 mg/kg was administered intraperitoneally 2 to 24 hours prior to the elicitation of the reaction, significant inhibition was observed. When prednisolone was administered intraperitoneally 8 hours prior to the elicitation of the reaction, the SP-induced capillary permeability increase in both ddY and WBB6 F1-W/WV mice was clearly inhibited by the drug at doses of 5 and 10 mg/kg. 3) Diphenhydramine (1 and 10 mg/kg) inhibited SP-induced vascular reaction in ddY mice but not in WBB6 F1-W/WV mice. 4) Atropine (10 mg/kg) inhibited SP-induced vascular reaction in both ddY and WBB6 F1-W/WV mice. But acetylcholine did not cause an increase of vascular permeability in ddY and WBB6 F1-W/WV mice ears. 5) Prednisolone (5 mg/kg) inhibited histamine- and serotonin-induced vascular permeability in ddY and WBB6 F1-W/WV mice ears. 6) Prednisolone (5 and 10 mg/kg) inhibited the SP-induced histamine release from ddY mice peritoneal mast cells. These results suggest that the vascular effect of SP is mediated by both mast cell dependent (release of histamine from mast cells) and mast cell independent mechanisms. Prednisolone inhibits the SP-induced vascular permeability mediated by both mechanisms in mice.


Hyperosmolarity selectively enhances IgE-receptor-mediated histamine release from human basophils. Increased osmotic pressure has been reported to cause non-cytotoxic histamine release (HR) from human basophils, as well as a potentiation of HR induced by anti-IgE. In this study, the effects of hyperosmolar Na-K-acetate (300-600 mOsm/kg H2O) on HR was studied in washed human blood cells from newborns, adult volunteers and patients with severe atopic dermatitis. These three patient groups represented 3 very distinct populations with respect to total plasma IgE content, medians were less than 0.2 IU/ml, 20.5 IU/ml and 2508 IU/ml; respectively. Increasing osmolarity to 500 mOsm/kg H2O caused little HR in the absence of other stimuli, whereas at 600 mOsm/kg H2O a significant increase in spontaneous HR was seen. The HR induced by anti-IgE and Concanavalin A, acting through the IgE-receptor, was increased approximately twofold at 500 mOsm/kg H2O. Responses were highly correlated to results at 300 mOsm/kg H2O. The use of 600 mOsm/kg H2O buffers caused a further increase in most, but not all blood samples. The potentiation of IgE-receptor-mediated HR when using hyperosmolar media was clearly independent of plasma IgE contents, and did not change the concentration-response to anti-IgE. In contrast, HR induced by the IgE-receptor-independent stimuli, Formyl-met-leu-phe and calcium ionophore A 23187, were not enhanced at all by increased osmotic pressure. We conclude, that hyperosomolar media selectively enhance IgE-receptor-mediated HR. The use of hyperosmolar media may therefore be beneficial in a diagnostic application of washed blood HR assays used in allergy diagnosis.


Modulation of IgE-mediated histamine release from human leukocytes by a new class of histamine H2-agonists. A new class of phenyl (pyridylalcyl) guanidines, acting as potent histamine H2-agonists, inhibits IgE-mediated human basophil histamine release in a nanomolar range. IC30-level of three substitutes of this group (arpromidine, BUA-75, and FRA-19) were found to be 0.02, 0.015 and 0.008 microM. The inhibition appeared with a fast onset (plateau after 10 min. preincubation) and claimed its maximum (60 +/- 2.9%, 63 +/- 1.8%, and 61 +/- 3.1%, n = 7) with 10 microM of the compounds. H2-mediated inhibition was totally blocked by 10 microM famotidine, a potent histamine H2-antagonist. The amount of anti-IgE or antigen for the initiation of the immunological release influenced the strength of inhibition of H2-agonist FRA-19 (p less than 0.05). Combined preincubation of FRA-19 with zardaverine, a cAMP-specific phosphodiesterase III/IV inhibitor, produced a synergistical inhibitory effect of leukocyte histamine release, which might explained by their different sites of action on intracellular cAMP levels. The capability of histamine to inhibit its own release is mediated by H2-receptors exclusively. New, potent H2-receptor stimulating compounds with positive inotropic effects possess additional potent anti-allergic properties.


Lymphocyte migration in the mouse. I. Time course of cell accumulation and the effect of antigen sensitisation and challenge in a murine model of chronic inflammation. The effect of time and of antigen sensitisation and challenge on lymphocyte migration into a site of chronic inflammation has been examined in the mouse. Enhanced lymphocyte migration occurred at sites of chronic inflammation after sensitisation and challenge to Bordetella pertussis vaccine (BPV). Biphasic migration was observed with time (5 min to 24 h), the initial very rapid but transient localisation at the inflamed site being followed by a second slower more sustained influx of cells. Increased localisation was also obtained with time in the lymphoid tissues and was accompanied by a parallel decrease in the number of cells present in the blood. The relative importance of antigen sensitisation and challenge for lymphocyte migration to a site of chronic inflammation has also been assessed. Lymphocyte migration into the inflamed site was partially dependent on the immunological status of the injected lymphocytes, but the presence or absence of antigen at the site of inflammation was the major factor which determined the degree of migration to the site.


Effects of human recombinant IL-1 on d-tubocurarine-induced histamine release from isolated rat peritoneal mast cells. The relationship between d-tubocurarine (d-Tc) and human recombinant interleukin-1 (rIL-1) was studied on the histamine-releasing property of isolated rat peritoneal mast cells. d-Tc induced histamine release in a dose-dependent manner (1 x 10(-4) M-3 x 10(-3) M) from isolated rat mast cells. Human rIL-1 (0.3-10 ng/ml) potentiated the d-Tc-induced histamine release and shifted the dose-response curve to left without changing the maximum histamine release by d-Tc. The potentiation by human rIL-1 was completely blocked by anti-IL-1-antibody. Human rIL-1 neither released histamine nor affected IgE-related histamine release in isolated rat peritoneal mast cells. Human recombinant IL-2, IL-3, and tumour necrosis factor neither released histamine from isolated rat mast cells nor affected d-Tc-induced histamine release. These results suggested that human rIL-1 might hypersensitize d-Tc receptors distributed on plasma membrane of rat mast cells.


The roles of calmodulin and protein kinase C in histamine secretion from mast cells. The effect of a new calmodulin-antagonist, 5-iodo-1-C8, with a high selectivity for calmodulin in comparison to protein kinase C, has been investigated on histamine secretion from mast cells. It has been found to be much more sensitive for the inhibition of histamine secretion than the earlier calmodulin-antagonists, trifluoperazine and W7. The effect of four inhibitors of protein kinase C, viz. staurosporine, K252a, tamoxifen and sphingosine, has also been studied on histamine secretion from mast cells. All of them caused dose-dependent inhibition of histamine secretion induced by the three secretagogues used: antigen, compound 48/80 and the calcium ionophore A23187. K252a was tested against histamine release, induced by the stimulation of protein kinase C alone with the phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA) or the synthetic diacylglycerol, 1-oleoyl-2-acetyl-rac-glycerol (OAG). In both the cases K252a caused dose-dependent inhibition of histamine release. Staurosporine was also tested against TPA and was found to inhibit the release induced by it. Potentiation and inhibition (modulation) of secretagogue-induced histamine release by simultaneous protein kinase C stimulation with TPA or OAG have been demonstrated before. The potentiation and inhibition are shown to be antagonized by staurosporine. The observations point to the involvement of both calmodulin and protein kinase C in the histamine secretion process from mast cells.


Influence of CGS 9343B, an inhibitor of calmodulin activity, on histamine release from isolated rat mast cells. Investigations of calmodulin involvement in cell responses has been complicated by the lack of selective calmodulin antagonists. A novel inhibitor, CGS 9343B, reportedly without influence on protein kinase C, is used in the present study of mast cell responses. The histamine release induced by antigen and compound 48/80 in the presence of calcium was enhanced by 10-20 microM CGS 9343B and inhibited by higher concentrations. Only inhibitory effects on the response to compound 48/80 in the absence of calcium and to the ionophore A23187 were observed, the latter being inhibited by 20 microM CGS 9343B. The influence on responses to combinations of the phorbol ester TPA and the ionophore A23187 was more complex, giving rise to enhancement at lower and inhibition at higher concentrations of CGS 9343B in a manner which depended on the experimental conditions. Unlike previously used calmodulin antagonists, CGS 9343B is devoid of detergent effects and without serious metabolic interference. The inhibitor seems useful to reveal differences in the mechanisms involved in responses to various histamine liberators. Our results conform with an inhibition of calmodulin by CGS 9343B but are at present inconclusive.


Inhibition of IgE-mediated histamine release from human peripheral leukocytes by selective phosphodiesterase inhibitors. Several phosphodiesterase (PDE) inhibitors with the capacity to inhibit the PDE IV isoenzyme produce dose-dependent inhibition of IgE-mediated histamine release (HR) from human peripheral leukocytes in vitro. Inhibition reached a maximum after 20 min of preincubation (IC30: 6-30 microM, IC50: 30-80 microM). Motapizone--a potent inhibitor of the isoenzyme PDE III--was much less effective, thus giving indirect evidence that PDE IV plays a predominant role in the control of cAMP cleavage in human basophils. The inhibiting effect of PDE-III/IV-selective compounds on IgE-mediated HR did not exceed the action of PDE-IV-selective inhibitors. The inhibition of anti-IgE-induced HR by zardaverine (a PDE-III/IV-inhibiting compound) was synergistically enhanced in the combined presence of forskolin or the recently synthesized histamine H2-agonist FRA 19).


Inhibition of histamine release from human granulocytes by ions of the rare earth elements lanthanum and cerium. The influence of the ATPase inhibitors, lanthanum or cerium, on histamine release in basophils and mast cells was studied. Both compounds inhibited IgE- or A23187-induced histamine release. To exclude a general inhibition of calcium-dependent reactions in the cell, we tested the influence of these compounds on phagocytosis and superoxide production of neutrophil granulocytes. Phagocytosis of Candida albicans was inhibited partially, superoxide generation, measured by the INT test after stimulation with zymosan or aggregated gamma globulin, was not affected. Because of the inhibitory effect of lanthanum or cerium compounds on membrane ATPase and immunological function of epidermal Langerhans cells we propose that these compounds may be used in the treatment of atopic eczema, where both histamine-releasing mast cells and IgE-bearing Langerhans cell play a pathogenetically important role.


A 5-lipoxygenase inhibitor, FR110302, suppresses airway hyperresponsiveness and lung eosinophilia induced by Sephadex particles in rats. To study the role of chemical mediators in airway hyperresponsiveness and simultaneous eosinophilia, we examined effects of a potent 5-lipoxygenase inhibitor FR110302 and those of prednisolone, indomethacin, platelet-activating factor (PAF) antagonist (RP-59227) and leukotriene C4 (LTC4) antagonist (ONO-1078) on airway hyperresponsiveness and lung eosinophilia induced by Sephadex particles. Sephadex G200 particles (2.5 mg/kg) were injected intravenously to rats and 3 days later the airway hyperresponsiveness to acetylcholine (ACh) and the eosinophilia in the bronchoalveolar lavage (BAL) fluids were observed. FR110302 (10 mg/kg b.i.d.p.o.) significantly suppressed both of these indicators of asthma. The amounts of immunoreactive LTB4,C4 (i-LTB4, C4) in the BAL fluid were measured by radioimmunoassay. The amounts of i-LTB4,C4 in the FR110302-treated rats were significantly less compared with that in the Sephadex-injected controls. Prednisolone completely inhibited the airway hyperresponsiveness. PAF antagonist and LTC4 antagonist partially inhibited the airway hyperresponsiveness, and indomethacin had no effect. The results indicate that 5-lipoxygenase products play important roles in the Sephadex-induced airway hyperresponsiveness and lung eosinophilia in rats.


Effect of compounds from Mandevilla velutina on bradykinin-mediated contractile and relaxant responses of the isolated guinea pig trachea. This study examines the action of three putative functional bradykinin (BK) antagonists isolated from Mandevilla velutina on BK and other agonist-induced contraction or relaxation responses in intact or in epithelium-denuded strips of tracheal muscle from guinea pigs. Compound MV 8612 (8 and 16 microM) and MV 8610 (12 and 24 microM) caused a graded rightward displacement of BK dose-response curves in the isolated guinea pig trachea (dose ratio 2- to 4-fold). Compound MV 8608 (28 microM) had a small effect on BK contractile responses. At high concentrations, compound MV 8612 (24 microM) caused a slight but significant depression of the BK-induced maximal responses. These pharmacological actions appear to be quite selective towards BK, since within the same concentration range these compounds failed to interfere with the sensitivities to prostaglandin F2 alpha, carbachol and histamine. However, these compounds significantly enhanced maximal responses to the latter two agonists, an action that was not influenced by indomethacin. In addition, MV 8612 and MV 8608 (16 and 56 microM) significantly potentiated BK-mediated epithelium-dependent relaxation responses in preparations precontracted with carbachol. These compounds also significantly potentiated isoprenaline but not theophylline-relaxant responses, an action that seems to occur independently of the generation of cyclooxygenase products of arachidonic acid pathway. These results further extend our previous studies and indicate that some of the M. velutina compounds exert a weak though selective antagonistic action against BK-induced contractile responses of the isolated epithelium-denuded guinea pig tracheal smooth muscle and potentiate BK-induced relaxations in tissues with intact epithelium precontracted with carbachol.(ABSTRACT TRUNCATED AT 250 WORDS)


Inhibition by calcium antagonists of shape changes induced in vitro by pro-inflammatory mediators in venous endothelial cells. The changes in shape of endothelial cells (ECs) lining rat inferior venae cavae in response to exposure in vitro to platelet activating factor (PAF, 5 x 10(-7) M), or 5-hydroxytryptamine creatinine sulphate (5HT, 3 x 10(-5) M), or calcium ionophore A23187 (1 x 10(-5) M), or cytochalasin B (CB, 1 x 10(-5) M) were assessed using an immunofluorescence technique and semi-automated image analysis. Most of these shape changes were prevented by lowering intracellular Ca2+ levels in several ways. Thus, pre-treatment with ryanodine (1 x 10(-6) M), verapamil HCl (2 x 10(-5) M), indomethacin (1 x 10(-4) M), isoprenaline sulphate (2 x 10(-6) M) or dibutyryladenosine 3'5' cyclic monophosphate (DbcAMP, 1 x 10(-5) M) significantly reduced the effects of PAF, 5HT and A23187. Pre-treatment of ECs with 8(N,N-diethylamino)octyl 3,4,5-trimethoxybenzoate HCl (TMB-8, 1 x 10(-5) M) or the nominal absence of Ca2+ in the extracellular medium prevented the effects of PAF and 5HT, but not those of A23187. The responses to 5HT were also reduced by cyproheptadine HCl (3 x 10(-6) M). However, only the absence of extracellular Ca2+ reduced the effects of CB. The calmodulin inhibitor N-(6-aminohexyl)-5-chloro-1-naphthalene sulphonamide HCl (W7, 1 x 10(-5) M) and the protein kinase C inhibitor 1-(5-isoquinolinyl sulphanyl)-2-methylpiperazine 2HCl (H7, 1 x 10(-5) M) were without effect against all the above agonists, but H7 and verapamil prevented EC shape changes induced by phorbol myristate acetate (PMA, 1 x 10(-6) M).(ABSTRACT TRUNCATED AT 250 WORDS)


Serine-protease inhibitors modulate nitric oxide-synthase activity of alveolar macrophages. Immunostimulated peritoneal macrophages of mice and rat have been demonstrated to produce L-arginine-derived nitrogen oxides. This metabolic pathway has also recently been found in rat alveolar macrophages and is suggested to play a certain role in lung injury. In vitro nitrite production from alveolar macrophages stimulated in vitro with lipopolysaccharide and recombinant interferon-gamma was inhibited by the addition of the irreversible serine-protease inhibitors, N-tosyl-L-phenylalanine chloromethyl-ketone (3 x 10(-7)-3 x 10(-4) M) and N-tosyl-L-lysine chloromethyl-ketone (3 x 10(-7)-3 x 10(-4) M) in a concentration-dependent manner. Two reversible inhibitors, N-alpha-p-tosyl-L-arginine methyl ester hydrochloride and benzoyltyrosine ethyl ester, were also effective but to a lesser extent. These antiproteases provide an opportunity to study the modulating influence on this recently discovered inflammatory pathway in alveolar phagocytic cells.


PLA2-induced oedema in rat skin and histamine release in rat mast cells. Evidence for involvement of lysophospholipids in the mechanism of action. Injection of phospholipase A2 (PLA2) in the rat skin produced a significant rise in oedema, which was inhibited by the simultaneous coinjection of aristolochic acid (100 micrograms), mepacrine (100 micrograms) and p-bromophenacyl bromide (10 micrograms). Indomethacin, nordihydroguaiaretic acid and WEB 2086 were without inhibitory effect on this model, whereas dexamethasone (5 mg/kg, p.o.) and coinjection of chlorpheniramine (20 micrograms) inhibited the oedema formation by more than 60%. Dose-dependent histamine release by rat peritoneal cells was induced by PLA2 and by lysophosphatidylserine and this effect could be antagonized by aristolochic acid and mepacrine. Apomorphine, previously reported to be an antagonist at lysophospholipid receptors, was able to inhibit the histamine release by mast cells as well as the oedema formation in rat skin. Taken all together, these results suggest that lysophospholipids, produced by the action of PLA2, are the mediators for the histamine release in rat peritoneal cells and could play an important role in the early oedema development in rat skin after PLA2 administration.


Participation of alpha 2-macroglobulin in counter-irritation. The participation of corticosteroids and alpha 2-macroglobulin (alpha 2-M) in counter-irritation (CI) induced by carrageenin was studied. We observed that CI could be produced when the second stimulus was done after 24 h, at which time glucocorticoids had already returned to their basal levels but alpha 2-M levels were maximum. When the level of alpha 2-M decreased (48 h) the counter-irritatory effect was not observed. As the synthesis of this protein is corticosteroid-dependent, we did not observe CI in adrenalectomized rats. However, when adrenalectomy was done 20 h after the first stimulus, allowing the synthesis of large amounts of alpha 2-M, the carrageenin stimulus again produced CI. Furthermore, adrenalectomized rats treated with aprotinin, a kallikrein inhibitor, after the first carrageenin stimulus, showed a reduction of the response to the second carrageenin stimulus. These observations demonstrated that corticosterone was not directly responsible for this inhibitory effect. Nevertheless, the anti-inflammatory factors involved in CI were, in fact, corticosteroid-dependent and alpha 2-M was probably one of the main factors.


Biochemical changes in arthritic rats under the influence of vitamin E. Two phases of adjuvant arthritis, acute phase (4 days after adjuvant inoculation) and chronic phase (21 and 29 days after adjuvant inoculation) were studied in male rats. The effect of administration of vitamin E in a daily oral dose of 147 mg/kg body wt. for one week against these phases of arthritis were demonstrated before and after adjuvant inoculation. Results showed that administration of the vitamin before and after adjuvant inoculation increased the lowered serum-SH group in arthritic rats so that their level was restored to pre-arthritic values especially in chronic treated group. Meanwhile, these treatments produced no change in the increased level of blood GSH or erythrocyte SOD activity of arthritic rats. The results showed also that administration of vitamin E before adjuvant inoculation increased significantly the level of alpha 2-M while it did not alter the increased serum Cp in acute phase. However, administration of the vitamin after adjuvant inoculation failed to exert any change in these parameters. In the meantime, these treatments tended to increase the lowered A/G of arthritic rats in different phases especially in acute one. These observations suggest that antioxidants such as vitamin E may be beneficial for arthritis.


Lobenzarit disodium (CCA) inhibits the proliferation of human endothelial cells and the activity of DNA polymerase alpha. N-(2)-Carboxyphenyl-4-chloroanthranilic acid disodium [Lobenzarit disodium (CCA)] is widely used for the treatment of patients with RA in Japan; however, the pharmacological mechanism of the compound is still unclear. In this report, the effect of CCA on the proliferation and DNA synthesis of endothelial cells was examined. CCA inhibited DNA synthesis in endothelial cells at a rather lower concentration than that in fibroblasts and HeLa cells. The DNA polymerase alpha activity was inhibited by CCA at a lower concentration than E. coli DNA polymerase I and avian myeloblastosis virus reverse transcriptase. Thus, CCA is a potent inhibitor of DNA polymerase alpha and this inhibitory effect could cause the inhibition of endothelial cell proliferation, which may be related to the therapeutic and pharmacological mechanisms of CCA.


[Measurement of serum PSA values by DELFIA PSA and its clinical significance on diagnosis and follow-up of prostate cancer patients]. Serum prostate specific antigen (PSA) values detected by DELFIA PSA were evaluated for usefulness in the diagnosis and follow-up of patients with prostate cancer. The system is time-resolved fluoroimmunoassay using europium as a tracer, which has a detectable range of 0.10-500 ng/ml with a small sample volume (25 microliters) and reliable quality control data. Furthermore, serum PSA values detected by the assay were equivocal to those detected by Tandem-R PSA. From the mean +3 S.D. of serum PSA values obtained on 227 normal males, 1.98 ng/ml was decided as an upper normal level. Serum PSA values in benign prostatic hyperplasia (BPH) (n = 69) and prostate cancer (n = 86) patients were statistically higher than those in normal males. However, when 1.98 ng/ml was used as a cut-off value, the false positive rate in BPH cases elevated up to 80%. Therefore, in the differential diagnosis of prostate cancer and BPH, we recommend 11.7 ng/ml (mean + S.D. in BPH cases) as a cut-off value, in which sensitivity is 72.1%, 88.4% are true negative in BPH, and efficacy is 79.4%. Serially determined serum PSA values in following up the patients with prostate cancer were confirmed to be highly effective for diagnosing recurrence and evaluating treatment responses. These findings suggest that DELFIA PSA is a useful tool for determination of serum PSA values.


Prevention of life-threatening arrhythmias by pharmacologic stimulation of the muscarinic receptors with oxotremorine. The potential antiarrhythmic efficacy of pharmacologic parasympathetic activation is still controversial. This study assessed the antiarrhythmic effect of saline solution (n = 9) and of the muscarinic agonist oxotremorine (1.5 micrograms/kg administered intravenously) (n = 17) in a feline animal model in which malignant arrhythmias were reproducibly elicited by the combination of acute myocardial ischemia and left stellate ganglion stimulation. Although saline solution had no effect, oxotremorine significantly decreased heart rate, blood pressure, the incidence of ventricular fibrillation from 47% to 0% (p = 0.004), and the incidence of malignant arrhythmias (either ventricular tachycardia or ventricular fibrillation) from 88% to 12% (p less than 0.001). When reduction in heart rate was prevented by means of atrial pacing (n = 15), the incidence of malignant arrhythmias was still significantly reduced from 87% to 27% (p = 0.001). Arrhythmias were also graded as follows: 0 = no premature ventricular contractions; 1 = 1 to 10 premature ventricular contractions; 2 = 11 to 50 premature ventricular contractions; 3 = ventricular tachycardia; 4 = ventricular fibrillation. Arrhythmia severity was 3.29 +/- 0.16 (SEM) in the control trials and was reduced to 0.76 +/- 0.26 (p less than 0.001) by oxotremorine and to 1.53 +/- 0.34 by oxotremorine and pacing (p = 0.002). Therefore a muscarinic agonist can significantly reduce malignant arrhythmias during acute myocardial ischemia and may represent a novel approach to the prevention of sudden cardiac death.


Influence of lidocaine on human muscle sympathetic nerve activity during programmed electrical stimulation and ventricular tachycardia. Lidocaine directly affects conduction and refractoriness of ventricular myocardium, and may also indirectly affect these electrophysiologic properties by inhibition of cardiac sympathetic nerve traffic. Both effects may play important roles in preventing ventricular arrhythmias in humans. To determine if lidocaine has a direct effect on sympathetic nerve activity, the effects of a 100 mg lidocaine bolus followed by a 2 mg/min infusion of lidocaine on muscle sympathetic nerve activity was assessed in seven patients during programmed ventricular stimulation with single extrastimuli (premature ventricular contractions [PVCs]) in sinus rhythm, and in seven patients during induced hemodynamically stable monomorphic ventricular tachycardia. During single extrastimuli, the mean (+/- SEM) area of PVC-associated bursts of sympathetic nerve activity was unaffected by lidocaine (1101 +/- 16 units pre-lidocaine versus 1075 +/- 19 units following lidocaine; p = 0.30). Likewise, the transient decrease in blood pressure with induced PVCs was similar before and after lidocaine infusion (p = 0.46). In seven patients with induced monomorphic ventricular tachycardia, tachycardia cycle length did not change after the lidocaine bolus (393 +/- 18 versus 399 +/- 17 msec; p = 0.34) but increased during lidocaine maintenance infusion (428 +/- 17 msec; p = 0.01). After induction of ventricular tachycardia, systolic pressure decreased from 150 +/- 6 to 117 +/- 9 mm Hg at 1 minute of tachycardia, to 109 +/- 6 mm Hg during the lidocaine bolus, and rebounded to 126 +/- 8 mm Hg during the lidocaine maintenance infusion (p = 0.04, bolus versus infusion).(ABSTRACT TRUNCATED AT 250 WORDS)


The importance of derived 12-lead electrocardiography in the interpretation of arrhythmias detected by Holter recording. Holter monitoring has been used extensively for the detection, diagnosis, and evaluation of therapy for cardiac arrhythmias. The availability of three-channel monitors allows for the recording of vectorcardiographic leads X, Y, and Z. One method, which was recently described by Dower et al., (J Electrocardiol 1988;21:5182-7), uses modified vectorcardiographic leads and allows for the acquisition of a derived 12-lead ECG of selected rhythm strips during the recording. In the present study, we evaluated the usefulness of the derived 12-lead ECG in the detection of P-wave and ST-segment shifts, assessment of QRST changes, and distinction between ventricular ectopic and aberrant supraventricular complexes. Our preliminary findings indicate that careful analysis of the derived 12-lead ECG provides additional information for a more accurate diagnosis of arrhythmias that are detected by the Holter monitor. The clinical importance and cost-effectiveness of the derived 12-lead ECG needs further evaluation.


Elevated levels of midtrimester maternal serum alpha-fetoprotein are associated with preterm delivery but not with fetal growth retardation. Our objective was to determine if the low birth weight associated with unexplained elevations of midtrimester maternal serum alpha-fetoprotein levels is due to prematurity or to fetal growth retardation. Rates of preterm delivery and fetal growth retardation were analyzed according to incremental maternal serum alpha-fetoprotein levels in 5555 women, predominantly white, who were screened for neural tube defects (group 1) and 843 women, predominantly black, with risk factors for low birth weight (group 2). Statistical methods included chi 2, t tests, analysis of variance, and regression analysis. In both groups increasing levels of maternal serum alpha-fetoprotein are significantly associated with preterm delivery but not with fetal growth retardation. The preterm delivery rate increased in each group from 8% at levels less than 0.5 multiples of the median to 18.1% (p less than 0.001) at levels greater than or equal to 2.5 multiples of the median in group 1 and 28.1% (p = 0.01) in group 2. Women with unexplained elevations of maternal serum alpha-fetoprotein are at increased risk for preterm delivery but not fetal growth retardation. Because of the wide availability of maternal serum alpha-fetoprotein screening, women at increased risk for preterm delivery can be identified in the midtrimester of pregnancy.


Transient hyperthyroidism and hyperemesis gravidarum: clinical aspects. Our objectives were to describe the presentation and course of hyperemesis gravidarum with respect to thyroid function and to test the hypothesis that patients with biochemical hyperthyroidism differ in clinical presentation from euthyroid hyperemesis patients. Sixty-seven patients seen at Los Angeles County Women's Hospital over a 10-month period with hyperemesis gravidarum were studied prospectively with respect to thyroid function. Forty-four patients (66%) had biochemical hyperthyroidism (increased free thyroxine index [n = 39] or suppressed thyroid-stimulating hormone [n = 40]) that was self-limited, resolving by 18 weeks' gestation. Hyperthyroid patients were more likely than euthyroid patients to have abnormal electrolyte levels (23/39 [59%] vs 6/28 [21%] and increased liver enzyme levels (23/59 [59%] vs 5/28 [18%], p less than 0.01). The severity of hyperemesis was found to vary directly with the degree of hyperthyroidism. Hyperthyroidism is a common, self-limited finding in hyperemesis. The cause of the hyperthyroidism may be linked to the cause of hyperemesis itself.


Overview of current treatment of neuroblastoma. Neuroblastoma is the second most common solid tumor in infants and children. Improvement of therapy for stage IV patients remains the major goal of research in treatment of neuroblastoma. New approaches under study are focused in four main areas: (a) phase II studies; (b) mega-therapy procedures; (c) targeted therapy; and (d) immunotherapy. Future approaches will be closely linked to progress in laboratory investigation and more efficient use of currently available drugs. Of all the childhood malignancies, this is the one tumor where such an approach is most likely to be effective.


Ethanol enhances the release of dopamine and serotonin in the nucleus accumbens of HAD and LAD lines of rats. The effects of intraperitoneal administration of ethanol, 0.5, 1.0, and 2.0 g/kg body weight, on the extracellular concentrations of dopamine (DA), serotonin (5-HT), and their major metabolites were studied in the nucleus accumbens (ACC) of alcohol-naive, selectively bred high-alcohol-drinking (HAD) and low-alcohol-drinking (LAD) lines of rats using the technique of microdialysis. In both lines, the extracellular levels of DA were increased following the injection of 1.0 and 2.0 g of ethanol/kg body weight while only the 2.0-g/kg dose elevated the concentration of 5-HT. None of the ethanol doses altered the extracellular levels of the major metabolites of DA and 5-HT. Dose-response curves for DA and 5-HT release indicated no marked difference in the sensitivity to ethanol between the lines. Local perfusion with 60 mM K+ through the microdialysis probe markedly enhanced the release of DA and 5-HT in the ACC of both lines; there was a small difference between the lines in the amounts of DA, but not 5-HT, released by K(+)-stimulation. Overall, the results indicate that (1) the ACC DA system is more sensitive than the ACC 5-HT system to intraperitoneal ethanol administration in both lines and (2) there is no evidence for a relationship between alcohol preference and sensitivity of the ACC DA and 5-HT systems to acute intraperitoneal ethanol administration.


Effects of prenatal ethanol exposure on hippocampal ionotropic-quisqualate and kainate receptors. Previous studies from our laboratories have shown that the consumption of moderate quantities of ethanol by rat dams during pregnancy reduces N-methyl-D-aspartate (NMDA) agonist receptor binding and NMDA-mediated electrophysiological responses in the hippocampal formation of adult offspring. We hypothesized that prenatal ethanol exposure would produce similar effects on receptor number and agonist-mediated responses of two so-called "non-NMDA" subtypes of glutamate receptors, the ionotropic-quisqualate (RS)-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA)-sensitive and the kainate-sensitive receptors. Sprague-Dawley rats were fed either a liquid diet containing 3.35% ethanol, an isocalorically matched liquid diet, or lab chow ad libitum throughout gestation. No significant differences between offspring from these three groups in the agonist concentration-response curves for either AMPA-induced or kainate-induced depolarization of hippocampal CA1 pyramidal neurons were observed. Furthermore, no significant differences in the density of [3H]-AMPA or [3H]-vinylidene kainic acid binding sites in any of the apical dendritic field regions of dorsal or ventral hippocampal formation were observed between the groups. These results indicate that the ionotropic quisqualate and kainate receptors, located in the apical dendritic field regions of the principal hippocampal neurons, are not affected by the same degree of prenatal ethanol exposure, which is known to reduce NMDA receptor binding and function in these same regions.


[The efficacy of transcatheter arterial infusion chemotherapy in patients with hepatocellular carcinoma]. The efficacy of transcatheter arterial infusion chemotherapy was examined for patients with hepatocellular carcinoma (HCC) between April 1986 and May 1992. We investigated the serial serum AFP level in patients with HCC. In 9 of 18 patients, the AFP level decreased promptly to 70% of the pretreatment level after this treatment. In 29 HCC patients with intrahepatic metastasis or portal thrombosis, 10 cases survived more than one year. However, only 3 cases survived more than two years after treatment. Our results suggest that we should carefully select appropriate drugs sensitive to HCC and also give careful consideration to the resistance to anti-cancer drugs.


[Intra-arterial chemotherapy with granulocyte colony-stimulating factor for breast cancer before surgical treatment]. Intra-arterial chemotherapy (IAC) for far advanced breast cancer is now performed as a routine adjuvant chemotherapy before surgical treatment. However, the following problems remain unsolved; (1) serious adverse reactions such as myelosuppression, (2) unsuccessful insertion of catheter into the internal thoracic artery, and (3) long waiting-period from IAC to surgical treatment. The present studies were conducted to evaluate the utility of a one-route IAC, in which the subclavian artery alone is used, and the efficacy of granulocyte colony-stimulating factor (G-CSF). There was no significant difference in the antitumor effect between the patients with two-route IAC, in which both internal thoracic artery and subclavian artery are used, and the patients with one-route IAC. Administration of G-CSF in combination with IAC reduced both the frequency and the severity of IAC-induced side effects. G-CSF administration during IAC was more effective than after IAC. G-CSF prevented IAC-induced myelo-suppression and/or accelerated recovery from this complication and thus reduced significantly the waiting-period before surgical treatment.


[A case of complete response of locally advanced breast cancer to arterial infusion chemotherapy: cannulation through the right femoral artery and combined use of G-CSF]. We reported a case of locally advanced breast cancer who was cannulated into the right internal thoracic artery through the right femoral artery by Seldinger's method for intra-arterial chemotherapy and obtained good results. In general, aggressive chemotherapy is often accompanied by bone marrow suppression necessitating discontinuation of chemotherapy. However, this patient recovered from this complication by G-CSF. It was suggested that Seldinger's method is suitable for locally advanced breast cancer and that G-CSF has an immediate effect of bone marrow suppression caused by aggressive chemotherapy.


Y-chromosome-specific fluorescence (f-body) of poorly decondensed bovine spermatozoa. An account is given in this paper of a method of identifying the fluorescence body (f-body) as a marker of the Y-chromosome. Also covered by this method is poor decondensation of spermatozoal nuclei when exposed to action of 1.25% of papain, 0.155% of DTE, and 0.025% of DMSO. Quinacrine mustard was used as fluorescent stain, its final concentration being 0.0025%. For staining, spermatozoa were suspended for 1-5 h. Average f-body frequency accounted for 41.0 +/- 5.1% in 35 ejaculates from 22 bulls. The overall variation coefficient amounted to 12.4% and thus was higher than each of the single values individually recorded from six bulls which were involved with three or four ejaculates (3.6-7.0%). F-bodies could not be detected by the method generally used on human spermatozoa. The applicability of the f-body test to quantification of Y spermatozoa in experimental separation of androspermatozoa and gynospermatozoa is discussed and is demonstrated by an example.


[Hyperthermia for cancer with dextran magnetite using tubular implants]. Dextran magnetite particles (Meito Sangyo Corp.) are an aqueous magnetite zol and a nanometer complex consisting of dextran chains surrounding a core of ultrafine iron oxide. The tubular implants were made with polyester tube (3 mm diameter, 20 mm length) filled with DM aqueous zol (29%w/v). The temperature of an agar phantom with implants was measured in the inductive field. Heating was effected by creating an electromagnetic field with a 7 kW generator operating at 500kHz (Yamamoto Vinyter). The temperature was elevated 3.4 degrees at a distance of 5 mm from the implant at an inductive power of 2.6 kW. An area of 20 x 20 mm was heated while changing the power, the number of implants, and their arrangement. Selective heating of cancer was considered possible by inductive heating at 500 kHz and DM implants. Since the DM aqueous zol configuration can be readily changed, treatment of various cancers is possible.


[Role of polymorphonuclear leukocytes (PMN) and active oxygen species in hyperthermia--antitumor effect of hyperthermia combined with rhG-CSF]. We have reported that polymorphonuclear leukocytes (PMN) and active oxygen species from PMN may play an important role in the mechanism of the antitumor effect of hyperthermia. At this time, we focused our experimental studies on rat AH109A carcinoma treated with hyperthermia combined with arterial injection of rhG-CSF. Rats with transplantable AH109A carcinoma at the hind leg received hyperthermia. These tumors showed mild suppression of further development only by hyperthermia. However, when arterial injection of rhG-CSF was applied together with hyperthermia, marked suppression of tumor development was observed. Our data suggest that hyperthermia combined with rhG-CSF is closely related to the generation of free radical-mediated tumor cell killing, and it can be an effective treatment for cancer.