Proc Natl Acad Sci U S A, 1997 Jan 21, 94(2), 663 - 8 Constitutive production and thrombin-induced release of vascular endothelial growth factor by human megakaryocytes and platelets; Mohle R et al.; We have shown that coculture of bone marrow microvascular endothelial cells with hematopoietic progenitor cells results in proliferation and differentiation of megakaryocytes . In these long-term cultures, bone marrow microvascular endothelial cell monolayers maintain their cellular integrity in the absence of exogenous endothelial growth factors . Because this interaction may involve paracrine secretion of cytokines, we evaluated megakaryocytic cells for secretion of cytokines, we evaluated megakaryocytic cells for secretion of vascular endothelial growth factor (VEGF) . Megakaryocytes (CD41a+) were generated by ex vivo expansion of hematopoietic progenitor cells with kit-ligand and thrombopoietin for 10 days and further purified with immunomagnetic microbeads . Using reverse transcription-PCR, we showed that megakaryocytic cell lines (Dami, HEL) and purified megakaryocytes expressed mRNA of the three VEGF isoforms (121, 165, and 189 amino acids) . Large quantities of VEGF (> 1 ng/10(6) cells/3 days) were detected in the supernatant of Dami cells, ex vivo-generated megakaryocytes, and CD41a+ cells isolated from bone marrow . The constitutive secretion of VEGF by CD41a+ cells was stimulated by growth factors of the megakaryocytic lineage (interleukin 3, thrombopoietin) . Western blotting of heparin-Sepharose-enriched supernatant mainly detected the isoform VEGF165 . In addition, immunohistochemistry showed intracytoplasmic VEGF in polyploid megakaryocytes . Thrombin stimulation of megakaryocytes and platelets resulted in rapid release of VEGF within 30 min . We conclude that human megakaryocytes produce and secrete VEGF in an inducible manner . Within the bone marrow microenvironment, VEGF secreted by megakaryocytes may contribute to the proliferation of endothelial cells . VEGF delivered to sites of vascular injury by activated platelets may initiate angiogenesis. Vestn Ross Akad Med Nauk, 1997, (6), 24 - 30 {Morphology, ultrastructure and populational features of bacteria francisella}; Gerasimov VN et al.; The morphology, ultrastructure of cells and the structure of microbial populations of various bacteria of the Francisella genus were estimated by electron microscopy . The strain 503 has been found to produce a bacterial population that is most homogeneous in shape and size . It contains microbes of only round and avoid forms, 0.5-0.6 micron in size . In addition of oval and round microbes there are ellipsoid and rod-shaped ones in the strains 15/3M, A . Cole 120, 117, etc . The largest tularemia microbes are typical of the strain Schu . The bacteria of all strains are covered by a capsule-like coat with well-defined borders . A thick capsule (0.12-0.35 micron) is specific for virulent strains whereas a thin capsular coat (0.06-0.12 micron) is encountered in vaccinal and avirulent microbes . The cells of the strain 503 were also shown to have the thickest envelope . All tularemia microbes have an asymmetric structure in the outer and cytoplasmic membranes due to the location of the bulk of intramembrane particles on their inner hydrophobic surfaces . Some F . tularensis microbes are able to produce keel-like protrusions on the outer membrane . The microbial nucleotide occupies 55-65% of the cytoplasmic volume and forms about 20-30 DNA-membrane contacts . Under unfavourable conditions, the microbes are capable of producing cell envelop protrusions and involutional cells, 0.1-0.3 micron in size. Lasers Surg Med, 1997, 21(2), 159 - 65 Giant cell formation in cells exposed to 740 nm and 760 nm optical traps; Liang H et al.; BACKGROUND AND OBJECTIVE: Optical trapping is becoming a useful and widespread technique for the micromanipulation of cells and organelles . Giant cell formation following optical trapping was studied to detect the potential adverse effects . STUDY DESIGN/MATERIALS AND METHODS: The nuclei of preselected single CHO cells were exposed to 740 nm and 760 nm laser microbeam generated by a titanium-sapphire tunable laser at 88 and 176 mW and different time exposures . The irradiated single cells were recorded and observed morphologically following exposure . Giant cells were tabulated and photographed . RESULTS: The irradiated cells either failed to divide, or they underwent nuclear proliferation to form giant cells through endoreduplication . CONCLUSION: Giant cells were induced by both 740 nm and 760 nm . The frequency of giant cell formation was higher for the longer time exposures and at the higher power densities . The use of an optical etalon to remove intracavity mode beating and high peak powers of the titanium-sapphire laser caused a significant reduction in the formation of giant cells. Bioelectromagnetics, 1997, 18(6), 410 - 7 Increased resorptions in CBA mice exposed to low-frequency magnetic fields: an attempt to replicate earlier observations; Juutilainen J et al.; This paper has two aims . First, it reports the findings of a study on the effects of low-frequency magnetic fields on reproduction . Second, it serves as an example of an attempt to replicate the results of an experimental study in an independent laboratory and discusses some of the problems of replication studies . To try to replicate the findings of a study reporting increased resorptions (fetal loss) in mice exposed to 20 kHz magnetic fields with sawtooth waveform and to study the possible effects of 50 Hz sinusoidal fields, pregnant mice were exposed to magnetic fields from day 0 to 18 of pregnancy, 24 h per day . The flux densities of the vertical magnetic fields were 15 microT (peak-to-peak) at 20 kHz and 13 or 130 microT (root mean square) at 50 Hz . Two strains of animals were used: CBA/S mice imported from the laboratory reporting the original observations, and a closely related strain CBA/Ca . The CBA/S mice were cleaned of pathogenic microbes and parasites before they were imported into our laboratory . The magnetic field exposures did not affect resorption rate in CBA/Ca mice . In CBA/S, the frequency of resorptions was higher in the exposed mice than in the control group . However, the increase was not significantly different from either the no-effect hypothesis or the results of the original study we were attempting to replicate . Differences between the two studies and difficulties in interpreting the results are discussed . It is concluded that the results tend more to support than argue against increased resorptions in CBA/S mice exposed to the 20 kHz magnetic field . The results demonstrate that animal strain is an important variable in bioelectromagnetics research: even closely related strains may show different responses to magnetic field exposure. Biofactors, 1997, 6(2), 145 - 55 Pathways for oxidation of low density lipoprotein by myeloperoxidase: tyrosyl radical, reactive aldehydes, hypochlorous acid and molecular chlorine; Heinecke JW; Many lines of evidence implicate oxidation of low density lipoprotein (LDL) in the pathogenesis of atherosclerosis, a chronic inflammatory disease . The physiologically relevant mechanisms have not been identified, but phagocytic white cells may play an important role because macrophage-rich lesions characterize the disorder . Recent studies have shown that myeloperoxidase, a heme enzyme secreted only by phagocytes, is present in human atherosclerotic tissue . The enzyme is a potent catalyst of LDL oxidation in vitro, it co-localizes with macrophages in lesions, and it generates products that are detectable in atherosclerotic plaque . These findings suggest that myeloperoxidase may promote LDL oxidation in the artery wall . This article reviews the enzyme's ability to generate a range of oxidants, including tyrosyl radical, reactive aldehydes, hypochlorous acid and molecular chlorine . These products have the potential to damage host molecules as well as microbes, suggesting a mechanism that may contribute to atherosclerotic vascular disease. J Environ Pathol Toxicol Oncol, 1997, 16(1), 21 - 6 Bioactivation of promutagens by the unicellular green alga Chlamydomonas reinhardtii; Podstavkova S et al.; The aromatic amine 2-aminofluorene (2-AF) was activated by the intact Chlamydomonas reinhardtii cells to a mutagen that exhibited toxic and mutagenic effects comparable to those of the direct-acting mutagen N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) . There were different responses of the wildtype and repair-deficient strains to the toxic and mutagenic effect of 2-AF . The recombination repair plays a major role in repair of damages induced in the C . reinhardtii DNA by the aromatic amine promutagen 2-AF and the direct-acting mutagen MNNG . The 2-AF activation has also been analyzed by algal cells/microbe coincubation assay . This new assay is used in addition to animal microsome-metabolizing system (S9 fraction) and plant cell/microbe coincubation assay . This additional system is suitable for detection of environmental promutagens and their conversion to mutagens, mainly in aquatic environments. Annu Rev Nutr, 1997, 17, 1 - 18 Memories of microbes and metabolism; Broquist HP; As Jackie Gleason was wont to say: "How sweet it really is!" And--reflecting on the 1940s-1980s, when studies of microbial nutrition revealed exciting structure-function relationships of the B-complex vitamins with relevance to metabolism in humans--it really is . A chemistry degree from Beloit College and a doctorate in biochemistry from the University of Wisconsin set the stage for my life's work at Lederle Laboratories, the University of Illinois, and Vanderbilt University . At Lederle my research contributed to folic acid chemistry: coenzyme forms and function; antimetabolites and cancer chemotherapy . My subsequent university studies centered on lysine biosynthesis and metabolism, e.g . its precursor role in carnitine and in indolizidine alkaloids of physiological interest . There were also many opportunities to reach out and give something back to the system via teaching and diverse service activities, all of which has led to a happy, fulfilling career, one for which I am ever thankful. Khirurgiia (Mosk), 1997, (3), 36 - 41 {Ozone therapy of diffuse peritonitis in the early postoperative period}; Kudriavtsev EP et al.; The study of the use of ozone-containing solutions in 40 experimental animals and in 58 patients with diffuse purulent peritonitis revealed their marked detoxicant effect that manifested with early decrease of ESR, leucocytosis, plasma concentration of bilirubin and medium-size molecules, and microbes . Lethality in the experimental and control group were 5.2 and 16.6% respectively. Crit Rev Microbiol, 1997, 23(2), 143 - 78 Natural protection of spring and well drinking water against surface microbial contamination . I . Hydrogeological parameters; Robertson JB et al.; The fate and transport of microbes in groundwater are controlled by physicochemical characteristics of the microbe and of the groundwater/aquifer media . Key characteristics of the microbe include size, inactivation (die-off) rate, and surface electrostatic properties . Key properties of the groundwater/aquifer system include flow velocity, aquifer grain (or pore) size, porosity, solid organic carbon content, temperature, pH, and other chemical characteristics of water and mineral composition . Because of size and surface electrical properties, viruses are much more mobile in groundwater than Cryptosporidium and Giardia (which are about 100 times or more larger than viruses) . The inactivation or die-off rate is usually the most important factor governing how far microbes can migrate in significant numbers in groundwater . Typical half-lives of microbes in groundwater range from a few hours to a few weeks . Examples of maximum reported migration distances of microbes in groundwater include: bacteria, 600 m in a sandy aquifer: viruses, 1000 to 1600 m in channeled limestones and 250 to 408 m in glacial silt-sand aquifers; Cryptosporidium and Giardia, no confirmed reports found of significant migration distances . Investigations by the EPA have indicated that distances of 210 to 325 m away from septic tanks are necessary to achieve with high confidence an 11 order of magnitude reduction in virus concentrations. Rev Environ Contam Toxicol, 1997, 151, 67 - 115 Rubber tire leachates in the aquatic environment; Evans JJ; Tires have a deleterious effect on the environment . This review discusses the background of scrap tires discarded in the environment, including tire composition, adverse environmental effects, threats to public health and safety, and solid waste management . Despite the widespread use of scrap tires in environmental applications, both land-based and aquatic, data on the indicators of environmental degradation are extremely scarce . Indicators of environmental degradation include analysis of chemicals within the water and sediment, analysis of contaminants within organisms, and analysis of the biological effects of these compounds on plants, animals, microbes, and organelles . Although these indicators are most useful when used in parallel, a review of the available information on chemical characterization of tire leachate from tire storage facilities, manufacturing, usage in recycling applications, and toxicity exposure studies, of vegetation surveys from waste tire areas and reviews of mammalian tire product toxicity, and of toxicity, mutagenicity, and carcinogenicity of tire exposure in experimental aquatic animals, microbes, and organelles is presented . The major characteristics of these studies are discussed in specific sections . The "Discussion and Conclusions" section discusses and summarizes the biological effects and chemical characterization of tire leachates . A global environmental perspective is included to improve our understanding of the deficiency of the current knowledge of tire leachate toxicity from various sources and to encourage interdisciplinary studies to establish the pattern of pollution associated with waste tire management. Med Parazitol (Mosk), 1997 Jan-Mar, (1), 31 - 3 {An immunoenzyme test system for the identification of typical and atypical strains of the plague microbe}; Devdariani ZL et al.; The authors have developed the optimal variant of the enzyme immunoassay system that contains plague murine monoclonal and equine polyclonal immunoglobulins and identified all plague microbe strains at a high sensitivity (3.84 m.cl/ml) . The cross reactivity of monoclonal antibodies with some pseudotuberculosis agent strains does not prevent from using the developed test system for laboratory diagnosis of plague, particularly while isolating atypical (capsule- or plasmid-free) variants of Y . pestis, taking into account parallel special diagnostic tests for pseudotuberculosis. Planta, 1997, 202(1), 28 - 35 A novel proline-rich glycoprotein associated with the extracellular matrix of vascular bundles of Brassica petioles; Davies HA et al.; A panel of monoclonal antibodies (MAC204, MAC236, MAC265) which recognise extracellular matrix glycoproteins implicated in plant-microbe interactions has been used to study glycoprotein antigens in petioles of turnip (Brassica campestris L.) . While MAC204 recognised two glycoproteins (gp120 and gp45) with apparent M(r) 120,000 and 45,000 in petiole extracts made with 2-amino-2-(hydroxymethyl)-1,3-propanediol (Tris) buffer containing sodium dodecyl sulfate, MAC236 recognised gp120 but not gp45, and MAC265 gave no or only weak reactivity . Tissue dissection studies established that gp120 was predominantly associated with the vascular bundle whereas gp45 was largely associated with the pith . This was consistent with results from tissue prints probed with MAC204 and MAC236 which also suggested a vascular localisation for gp120 . Immunoelectronmicroscopy showed that MAC204 and MAC236 both labelled three-way junctions between cells of the phloem and sclerid fibres . Both gp120 and gp45 were shown to carry epitopes in common with known hydroxyproline-rich glycoproteins . Unlike gp45, gp120 could be extracted from petioles with Tris buffer alone and then isolated from this extract by trichloroacetic acid treatment (which left gp120 soluble), followed by size-exclusion and ion-exchange chromatography . Amino acid analysis revealed gp120 to be a novel glycoprotein, particularly rich in proline, lysine, valine and threonine but relatively poor in hydroxyproline . The most abundant sugars were arabinose and galactose . The potential role of this very basic cell surface glycoprotein in plant defence against microbes is discussed. Semin Thromb Hemost, 1997, 23(1), 23 - 30 Measurement of fluorescent-labeled LMM-heparin in biological fluids using protamine-linked microbeads; Malsch R et al.; A quantitative assay for fluorescent heparin in a purified system and in plasma was developed (Piazolo et al: Semin Thromb Hemostas 20:227-235, 1994) . The protamine microbeads (1.6 microns) showed a broad size distribution and a large standard variation in low concentrations . Our aim was to optimize these protamine microbeads for the measurement of fluorescent heparin . The following results were obtained: Paramagnetic protamine microbeads of different average diameters (0.8, 1.6, 2.8, and 4.5 microns) were synthesized by cyclocarbodiimide and tosyl activation . These microbeads bind heparin and are assayed using flow cytometry . The protamine concentration on the surface of the beads ranged between 2.0 and 61 mg/mL . The protamine microbeads bound fluorescent heparin and were analyzed by flow cytometry . The protamine microbeads bound LMM-heparin-tyramine-FITC dose dependently in saline solution, plasma, and blood . There are substantial differences between the microbeads of different origins with regard to the amount of protamine bound, the sensitivity of the detection, and the reliability for the determination of heparins in plasma and blood . The minimal sensitivity of the final method was 0.001 U/mL LMMH-tyramine-FITC in saline solution and in plasma . Human blood cells were not bound to protamine microbeads . The half-maximal binding of LMMH-tyramine-FITC of the different protamine-coated microbeads ranged from 1.7 to 8.0 micrograms/mL in saline solution, 2.3 to 8.7 micrograms/mL in plasma, and 3.1 to 6.4 micrograms/mL in blood . We conclude that all protamine microbeads can be used to quantify the concentration of LMMH-tyramine . Protamine Dynabeads M-450 (diameter 4.5 microns) have advantages over other microbeads because of their more homogeneous size distribution, a higher selectivity, and they can be measured together with leukocytes . They are currently used to develop a competitive binding assay for heparin in plasma. Gut, 1997 Jan, 40(1), 20 - 4 Role of Helicobacter pylori surface structures in bacterial interaction with macrophages; Chmiela M et al.; BACKGROUND: Helicobacter pylori infection is associated with a marked infiltration of the gastric epithelium by neutrophils, macrophages, lymphocytes, and plasma cells . Despite the presence of phagocytes in close vicinty to H pylori microbes a great number of people are unable to eradicate bacteria . AIMS: To investigate the involvement of multiple bacterial 'adhesins' and some phagocytic receptors in the process of the ingestion of H pylori by macrophages . BACTERIA: H pylori strains differing in the expression of sialic acid dependent (sHA) or sialic acid independent (HA) haemagglutinin and heparan sulphate binding were selected for the study . METHODS: The uptake of fluorescein labelled H pylori bacteria by a homogenous macrophage cell line J 774A.1 was estimated in a quantitative fluorometric assay . RESULTS: The ingestion of H pylori 17874 and 25 strains expressing sHA was inhibited by the pretreatment of the bacteria with anti-sHA antibodies or fetuin as well as by treatment of the macrophages with neuraminidase . In contrast the uptake of H pylori 17875 strain expressing HA remained unchanged . The phagocytosis of all investigated bacteria was inhibited after the treatment with heparin, hyaluronic acid or vitronectin with fresh but not heat inactivated serum . CONCLUSIONS: The results suggest that H pylori surface compounds binding host proteins such as fetuin, heparin/haparan sulphate, hyaluronic acid, and vitronectin in the presence of complement, could allow the bacteria to avoid phagocytosis. J Biomater Sci Polym Ed, 1997, 8(6), 411 - 20 Comparison of albumin binding capacities of three different reactive dye-derivatized poly(ethylene glycol dimethacrylate-hydroxyethyl methacrylate) microbeads; Denizli A et al.; Bovine serum albumin (BSA) adsorption onto dye-derivatized poly(ethylene glycol dimethacrylate-hydroxyethyl methacrylate) {poly(EGDMA-HEMA)} microbeads carrying three different reactive dye ligands (i.e . Congo Red . Cibacron Blue F3GA, and Alkali Blue 6B) was investigated . Swellable poly(EGDMA-HEMA) microbeads, in the size range of 150-200 microns, were produced by a modified suspension copolymerization of EGDMA and HEMA . The dyes were covalently attached to the microbeads . The maximum amounts of dye loadings were 14.5, 16.5, and 23.7 mumol g-1 for Congo Red, Cibacron Blue F3GA, and Alkali Blue 6B, respectively . The maximum BSA adsorption on the dye-derivatized microbeads from aqueous solutions containing different amounts of BSA were 90, 60.5, and 40 mg g-1 for the Congo Red, Cibacron Blue F3GA, and Alkali Blue 6B carrying microbeads, respectively . The maximum BSA adsorptions were observed at pH 6.0 in all cases . Desorption of albumin molecules were achieved by using 1.0 M NaSCN (pH 8.0) . High desorption ratios (more than 85% of the adsorbed BSA) were observed in all cases . It was possible to reuse these novel sorbents without significant losses in the adsorption capacities. Cell Motil Cytoskeleton, 1997, 37(1), 1 - 6 Traction fibre: toward a "tensegral" model of the spindle; Pickett-Heaps JD et al.; Most current hypotheses of mitotic mechanisms are based on the "PAC-MAN" paradigm in which chromosome movement is generated and powered by disassembly of kinetochore microtubules (k-MTs) by the kinetochore . Recent experiments demonstrate that this model cannot explain force generation for anaphase chromosome movement {Pickett-Heaps et al., 1996: Protoplasma 192:1-10} . Another such experiment is described here: a UV-microbeam cut several kinetochore fibres (k-fibres) in newt epithelial cells at metaphase and the half-spindle immediately shortened: in several cells, the remaining intact spindle fibres bowed outwards as they came under increased compression . Thus, severing of k-MTs can lead to increased tension between chromosomes and poles . This observation cannot be explained by models in which force is produced by motor molecules at the kinetochore actively disassembling k-MTs . Rather, we argue that tensile forces act along the whole k-fibre, which, therefore, can be considered as a classic "traction fibre." We suggest that anaphase polewards force is generated by MTs interacting with the spindle matrix and when k-MTs are severed, polewards force continues to act on the remaining kMT-stub; spindle MTs act as rigid struts concurrently resisting and being controlled by these forces . We suggest that the principles of "cellular tensegrity" {Ingber, 1993: J . Cell Sci . 104:613-627} derived from the behaviour and organization of the interphase cell apply to the spindle . In an evolutionary context, this argument further suggests that the spindle might originally have evolved as the mechanism by which a single tensegral unit (cytoplast) is divided into two cytoplasts; use of the spindle for segregating chromosomes might represent a secondary, more recent development of this primary function . If valid, this concept has implications for the way the spindle functions and for the spindle's relationship to cytokinesis. Am J Reprod Immunol, 1997 Jan, 37(1), 125 - 36 TGF beta 2 in rabbit blastocoelic fluid regulates CD4 membrane expression: possible role in the success of gestation; Ouellette MJ et al.; PROBLEM: During pregnancy, major changes occur in the decidual cell population . One of these changes involves some phenotypical transformations of lymphocyte sub-populations . Since these variations might be due to the presence of the embryo, the current study was designed to investigate the implication of blastocoelic fluid (BF) in these changes and to determine the mechanism by which this phenomenon occurs . METHOD: Lymphocytes isolated from human peripheral blood (PBL) were cultured for 72 h in RPMI-FCS 10% and with or without BF day 12 (BF d-12) or Concanavalin A (ConA) . After 72 h, T cells were labelled with anti-CD4 antibodies and Quantum Simply Cellular microbeads were used as a standard to evaluate the antibody binding capacity (ABC) . RESULTS: Treatment of human PBL with BF d-12 decreases the percentage of CD4 and TCR positive cells, as compared to non-stimulated cells, but has no significant effect on CD2, CD3, and CD8 positive cells . It was also demonstrated, for the first time, that transforming growth factor beta-2 (TGF beta 2) in BF day 12 diminishes the percentage of CD4 positive cells by downregulating CD4 membrane expression on leucocytes . CONCLUSION: These findings suggest that the embryo plays a role in its own protection . Furthermore, it is predicted that any tissue producing TGF beta 2, such as certain types of tumor, downregulates the immune response, thus allowing tumor growth. J Neural Transm Suppl, 1997, 50, 173 - 81 Pathogenesis of immune-mediated demyelination in the CNS; Hartung HP et al.; Collective evidence from studies in the animal model experimental autoimmune encephalomyelitis and pathological and immunological studies on MS patients suggest that this most common inflammatory demyelinating disorder of the central nervous system results from primarily T-lymphocyte driven aberrant immune responses to a number of myelin and possibly non-myelin antigens . These include MBP, PLP, MOG, MAG, CNP and S 100 . Autoreactive T-cells reactive with these antigens circulate in blood and upon activation can travel across the blood-brain-barrier to initiate a local immunoflammatory response provided they encounter a microglial cell that displays antigenic epitopes in the context of MHC class II gene products and accessory molecules . Demyelination probably results from antibody-induced complement activation . Repeated inflammatory episodes eventually exhaust the reparative capacities of oligodendrocytes and damage axons . As the disease evolves, an initialy focussed immune response may diversify due to a process termed epitope spreading . The initial event of T lymphocyte activation remains elusive, but molecular mimicry, cross-recognition of structures shared between microbes and myelin, appears to be crucial. Cell Motil Cytoskeleton, 1997, 36(4), 339 - 54 Amoeboid movement anchored by eupodia, new actin-rich knobby feet in Dictyostelium; Fukui Y et al.; To date, protrusion of pseudopodia has been considered to be primarily responsible for translocation of free-living amoebae and leukocytes of higher organisms . Although there is little question that the pseudopodium plays an important role, little attention has been given to the cortical structures that are responsible for cell-substratum anchorage in amoeboid movement . Here, we report on a new knobby foot-like structure in amoebae of a cellullar slime mold, Dictyostelium discoideum . These feet, each about 1 micron in diameter, appear transiently in multiple units at the base of certain pseudopodia where the amoeba contacts a partially deformable substrate . The feet were discovered, and their spatial and temporal behavior relative to pseudopodial anchorage and invasive locomotion were observed, by examining Dictyostelium amoebae using a DIC video microscope providing an 0.3 micron depth of field . Key evidence for the anchoring role of the knobby feet was obtained by investigating amoebae, flattened in a specially devised observation chamber, and attracted by chemotaxis towards 3',5' cyclic-adenosine monophosphate (cAMP) . The cAMP was released by highly localized, pulsed UV-microbeam irradiation of caged cAMP . We show by indirect immunofluorescence that the knobby feet contain a high concentration of filamentous (F-) actin, myoB (a member of Dictyostelium myosin-I family), and alpha-actinin (an actin-binding protein) . Interestingly, myoB exhibits a circular disposition around each foot . Neither myosin-II (conventional myosin) nor the 269 kD protein, which has been recently identified as a talin homologue of Dictyostelium {Kreitmeier et al., 1995: J . Cell Biol . 129:179-188}, are concentrated at the feet . We propose that the knobby feet provide anchorage to the substratum needed by lamellipodia to exert projectile forces for invading narrow spaces or otherwise for a flattened amoeba to secure itself to the deformable substratum . Some forms of adhesion plaques in higher organisms such as "podosomes" or "invadopodia" may perform functions similar to the knobby feet, but appear to differ in life time, cytoskeletal organization and composition . We have named the knobby foot "eupodium." J Assoc Nurses AIDS Care, 1997 Jan-Feb, 8(1), 23 - 31 Complementary and alternative medicine and HIV/AIDS . Part I: Issues and context; MacIntyre RC et al.; This paper reviews the definitions of complementary and alternative medicine (CAM) and adopts the definition, "Complementary and alternative medicine is defined through a social process as those practices that do not form part of the dominant system for managing health and disease" (Jonas, 1996, p.1) . The authors review the literature on CAM usage in the general and HIV/AIDS populations, and the impact of CAM on modern Western medicine and the allied health professions . Host-virus factors in HIV/AIDS are examined from the perspectives of both Western medicine and complementary and alternative medicine . Methodological issues and the historical argument over the relative importance of host and microbe are explored in relation to HIV/AIDS and the CAM and Western approaches to HIV/AIDS care. J Periodontal Res, 1997 Jan, 32(1 Pt 2), 126 - 32 Are bacterial proteases important virulence factors? Lantz MS. The contribution of bacterial proteases to virulence has been relatively understudied . It is a simple matter to argue that bacterial proteases have the potential to destroy the structural and functional proteins that constitute host tissues as well as to destroy proteins important in host defense . Systematically demonstrating that such interactions occur during disease pathogenesis is more difficult, although a few studies have suggested that the ability of a pathogen to use proteases to cross proteinaceous barriers within the host contributes to bacterial virulence . This manuscript reviews concepts of bacterial virulence . Next, it describes how the host regulates the activities of its own proteases to maintain a state of health, and examines evidence suggesting that dysregulation of host proteases results in disease . Finally, evidence supporting a role for endogenous microbial proteases or acquisition of host proteases by microbes as virulence determinants is discussed as are suggestions for future directions for research in this area. Cell Motil Cytoskeleton, 1997, 36(3), 266 - 75 Effects of ultraviolet-microbeam irradiation of kinetochores in crane-fly spermatocytes; Ilagan AB et al.; Ultraviolet (UV)-microbeam irradiation of a single kinetochore during anaphase generally causes all 6 of the half-bivalents in the cell to stop poleward motion within 1 min after the irradiation . The half-bivalents regain movement after remaining stopped for an average of 8.7 min, through different pairs in the same cell can resume at different times . Once movement resumes they usually continue movement until they reach the poles . As controls, to see if the effect is due to alteration of the kinetochore, we irradiated spindle fibers and chromosome arms using the same doses and wavelengths as for kinetochore irradiation . After spindle fiber irradiation, only the half-bivalent associated with the irradiated spindle fiber and its partner stop moving poleward while the other half-bivalents in the same cell are not affected . After irradiation of a chromosome arm, the movement of the two partner half-bivalents associated with irradiated arm either slowed or moved with unchanged velocity; no other half-bivalents in the cell were affected . Therefore, only irradiation of a kinetochore stops the movement of all the half-bivalents in the same cell . We suggest that the irradiated kinetochore sends a "stop" signal to the other kinetochores in the same cell. J Clin Periodontol, 1997 Jan, 24(1), 1 - 7 Determination of energy density threshold for laser ablation of bacteria . An in vitro study; Coffelt DW et al.; The Nd:YAG and CO2 lasers have been shown to be bactericidal at relative low energy densities . However, at energy densities exceeding 120 J/cm2 (CO2) and 200 J/cm2 (Nd:YAG), laser irradiation also causes irreparable root surface damage . The purpose of this study was to determine, in vitro, the energy density threshold at which microbial ablation could be achieved while inflicting the least amount of damage to the root surfaces of human teeth . Pairs of Escherichia coli colonies cultured on broth agar were treated with a CO2 laser using a pulsed waveform at approximate energy densities ranging from 3 to 110 J/cm2 . One of each colony-pair was then examined by scanning electron microscopy (SEM) and the other subcultured for viable microbes . Roots of extracted teeth were lightly scaled and treated by CO2 laser, again with pulsed beam using approximate energy densities of 3 to 110 J/cm2: and examined by SEM . Regardless of the level of energy density, residual bacteria could be subcultured from all laser treated microbial colonies . The inability of the laser to completely obliterate microbial colonies was likely due to: depth of energy penetration, difficulty in precisely overlapping beam focal spots, irregular beam profile, and presence of microbes at the periphery of the beam focal spot . The threshold energy density for bacterial obliteration was determined to be 11 J/cm2 and that for root damage was 41 J/cm2 . Root damage was evident by charring, crater formation, melt-down and resolidification surface mineral, and increasing surface porosity . The results of this in vitro study indicate that when used at an energy density between 11 and 41 J/cm2 the CO2 laser may destroy microbial colonies without inflicting undue damage to the tooth root surface. Int J Oral Maxillofac Implants, 1997 Jan-Feb, 12(1), 32 - 42 The clinical, microbial, and host response characteristics of the failing implant; Salcetti JM et al.; The goal of this study was to provide new data regarding levels of inflammatory and growth factor mediators and bacterial pathogens associated with failing implants, as compared to healthy implants . Twenty-one patients with failing implant sites (group 1) and 8 patients with only healthy implants (group 2) were included . Fifteen of the 21 failing implant patients (group 1) also presented with at least one stable nondiseased implant . Plaque samples were examined, using DNA oligonucleotide probes for 40 different microbes . Gingival crevicular fluid samples were collected for the analyses of catabolic bone resorbing agonists prostaglandin E2 (PGE2), interleukin-1 beta (IL-1 beta) and IL-6 and anabolic bone-forming growth factors transforming growth factor beta (TGF-beta) and platelet-derived growth factor (PDGF) . Although positive trends were noted, there were no significant differences in any of the microbial, inflammatory, or growth factors mediators comparing failing to stable implants in group 1 . This study found greater detection frequencies of P . nigrescens, P . micros, F . nucleatum ss vincentii, and F . nucleatum ss nucleatum, as well as significant elevations in GCF levels of PGE2, IL-1 beta, and PDGF in mouths with failing implant sites as compared to mouths with healthy control implants . Risk appears to be primarily at a patient level and secondarily at a site or implant level from a clinical, microbial (P . micros and P . nigrescens), and biochemical (PGE2 and IL-1 beta) perspective . Furthermore, the counts of P . nigrescens and P . micros were found to correlate with concentrations of PGE2 at a site level. Diabet Med, 1997 Jan, 14(1), 29 - 34 Impaired leucocyte functions in diabetic patients; Delamaire M et al.; This study evaluates polymorphonuclear neutrophil (PMN) cell performance in 61 diabetic patients free of infection (40 Type 1, 21 Type 2), using tests that explore all the functional steps of PMN: (1) adherence: expression of adhesion molecules, CD 11a, CD 11b, CD 11c; nylon fiber adherence test; (2) chemotaxis under agarose towards the bacterial oligopeptide FMLP and complement fractions, used as attracting agents; (3) phagocytosis of opsonized latex microbeads; (4) bactericidal activity: chemiluminescence assessment of the oxidative killing potential before and after stimulation by opsonized zymosan and PMA; nitroblue tetrazolium reduction test . Results were analysed according to potentially influential factors: metabolic control (HbA1C, glycaemia), age of patient, type of diabetes, disease duration, and existence of vascular complications . PMN chemotaxis was significantly lower in patients than in healthy controls (p < 0.001) and associated with spontaneous adherence and increased expression of adhesion molecules (CD 11b, CD 11c) . The increased response to chemiluminescence reflects spontaneous activation of PMN cells and increased free radical production; after stimulation, response was lower than in controls . The type of diabetes, the age of patients, HbA1C level and disease duration did not affect the responses . Chemotaxis and chemiluminescence were further reduced in patients with vascular complications and hyperglycaemia . We conclude that all steps of PMN functioning are altered in diabetic patients, which may increase the risk of vascular complications and infectious episodes. Cell Motil Cytoskeleton, 1997, 36(2), 136 - 48 Ultraviolet microbeam irradiations of epithelial and spermatocyte spindles suggest that forces act on the kinetochore fibre and are not generated by its disassembly; Spurck T et al.; Ultraviolet (UV) microbeam irradiations of crane-fly spermatocyte and newt epithelial spindles severed kinetochore fibres (KT-fibres), creating areas of reduced birefringence (ARBs): the remnant KT-fibre consists of two "stubs," a pole-stub attached to the pole and a KT-stub attached to the kinetochore . KT-stubs remained visible but pole-stubs soon became undetectable {Forer et al., 1996} . At metaphase, in both cell types the KT-stub often changed orientation immediately after irradiation and its tip steadily moved poleward . In spermatocytes, the chromosome attached to the KT-stub remained at the equator as the KT-stub elongated . In epithelial cells, the KT-stub sometimes elongated as the associated chromosome remained at the equator; other times the associated chromosome moved poleward together with the KT-stub, albeit only a short distance toward the pole . When an ARB was generated at anaphase, chromosome(s) with a KT-stub often continued to move poleward . In spermatocytes, this movement was accompanied by steady elongation of the KT-stub . In epithelial cells, chromosomes accelerated polewards after irradiation until the KT-stubs reached the pole, after which chromosome movement returned to normal speeds . In some epithelial cells fine birefringent fibres by chance were present along one edge of ARBs; these remnant fibres buckled and broke as the KT-stub and chromosome moved polewards . Similarly, KT-stubs that moved into pole stubs (or astral fibres) caused the pole stubs (or astral fibres) to bend sharply from the point of impact . Our results contradict models of chromosome movement that postulate that force is generated by the kinetochore disassembling the KT-fibre . Instead, these results suggest that poleward directed forces act on the KT-fibre and the KT-stub and suggest that continuity of microtubules between kinetochore and pole is not obligatory for achieving anaphase motion to the pole. Fertil Steril, 1997 Jan, 67(1), 98 - 103 Processing of semen in an antibiotic-rich culture medium to minimize microbial presence during in vitro fertilization; Cottell E et al.; OBJECTIVE: To identify and determine the prevalence of microorganisms in preprocessed and postprocessed semen in an IVF-ET program . DESIGN: Prospective study . SETTING: University Teaching Hospital . PATIENT(S): Seventy-four men undergoing preprogram evaluation, each producing two semen samples . INTERVENTION(S): Semen processing with a wash and swim-up technique in a penicillin- and streptomycin-rich medium . MAIN OUTCOME MEASURE(S): The identity and prevalence of seminal microorganisms before and after processing . RESULT(S): Sixty-three percent of individual unprocessed semen samples grew microorganisms, the majority of which were nonpathogenic . Thirty-three men (44.6%) had microbes identified in repeat samples, four had identical organisms each time . Twenty (27%) had positive cultures in one sample, negative in the other . Twenty-one (28.4%) had consistently sterile semen . After seminal processing, the recovery rate for microbes was 5% . Microbial presence after processing did not correlate with either the sperm swim-up concentration or the initial microbial concentration . CONCLUSION(S): Bacteriospermia is common . The microorganisms found rarely are replicated and most likely represent contamination . Wash and swim-up semen preparation in an antibiotic rich culture medium effectively eliminates 95% of organisms. Structure, 1996 Dec 15, 4(12), 1465 - 74 Structure of UDP-N-acetylglucosamine enolpyruvyl transferase, an enzyme essential for the synthesis of bacterial peptidoglycan, complexed with substrate UDP-N-acetylglucosamine and the drug fosfomycin; Skarzynski T et al.; BACKGROUND: UDP-N-acetylglucosamine enolpyruvyl transferase (MurA), catalyses the first committed step of bacterial cell wall biosynthesis and is a target for the antibiotic fosfomycin . The only other known enolpyruvyl transferase is 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase, an enzyme involved in the shikimic acid pathway and the target for the herbicide glyphosate . Inhibitors of enolpyruvyl transferases are of biotechnological interest as MurA and EPSP synthase are found exclusively in plants and microbes . RESULTS: The crystal structure of Escherichia coli MurA complexed with UDP-N-acetylglucosamine (UDP-GlcNAc) and fosfomycin has been determined at 1.8 A resolution . The structure consists of two domains with the active site located between them . The domains have a very similar secondary structure, and the overall protein architecture is similar to that of EPSP synthase . The fosfomycin molecule is covalently bound to the cysteine residue Cys115, whereas UDP-GlcNAc makes several hydrogen-bonding interactions with residues from both domains . CONCLUSIONS: The present structure reveals the mode of binding of the natural substrate UDP-GlcNAc and of the drug fosfomycin, and provides information on the residues involved in catalysis . These results should aid the design of inhibitors which would interfere with enzyme-catalyzed reactions in the early stage of the bacterial cell wall biosynthesis . Furthermore, the crystal structure of MurA provides a model for predicting active-site residues in EPSP synthase that may be involved in catalysis and substrate binding. Cytometry, 1996 Dec 15, 26(4), 265 - 74 Quantitative determination of surface antibody binding capacities of immune subsets present in peripheral blood of healthy adult donors; Denny TN et al.; The objective of this study was to quantitate the antibody binding capacity (ABC) of CD3, CD4, CD8, CD16, and CD19 on lymphocytes and CD4 on monocytes from healthy adult donors . Peripheral blood was collected over three consecutive days and repeated in the same format two weeks later for comparison to initial measurements . Immune subsets were labeled by direct single or two-color staining in whole blood followed by lysis of erythrocytes . Fluorescence intensity measurements were made by carefully calibrating the flow cytometer and then measuring the intensity of monoclonal antibody staining on labeled cells and on Quantum Simply Cellular Microbeads . The effect of paraformaldehyde fixation on intensity measurements and coefficient of variation of thirty replicates for each phenotype were also studied . We found a small change in calculated ABC following overnight fixation with a greater change following 48 h of fixation prior to flow cytometric analysis . We found excellent precision could be achieved for measuring the ABC of most markers with some improvement desirable for expression of CD4 on monocytes and CD16+ lymphocytes . Between donors we found a high-low range of CD3+ = 134,349-45,905; CD4+ (lymphocytes) = 54,174-36,106; CD4+ (monocytes) = 9,246-3094; CD8+ = 268,868-190,622; CD3+CD8+ = 269,858-212,024; CD16+ = 38,307-336; and CD19+ = 25,252-11,689 . For the total donor group, the observations at week 1 and week 2 were not significantly different (alpha = .05) for any of the immunophenotypes we studied . The data presented here continue to show that it is possible to perform quantitative intensity measurements of immune subsets when performing immunophenotyping studies. Forensic Sci Int, 1996 Dec 2, 83(2), 95 - 103 Bone-dust in autopsies: reduction of spreading; Kernbach-Wighton G et al.; During autopsies, an open oscillating saw produces large quantities of respirable bone-dust, which is able to carry microbes over several metres . Experiments were done using a modified (open) undulation saw (spray tube to moisten the saw-blade with water) . Saw-dust was asservated with culture media . Colonies were identified macroscopically . Microbes in the air were quantified (per unit of time) . A remarkable reduction of saw-dust is done by an integrated spray tube using water . There remains a contamination at the head of the autopsy table in the level of the table top . We found a complete decontamination 150 cm above the floor . No spreading of particles carrying microbes was seen over distances of more than 1.5 m . The risk of an airborne infection is minimal when using a manual saw (absence of grinding-dust) . The modified type of an 'oscillating saw with a spray-tube' may be considered a practicable compromise between a manual saw and an unprotected undulation saw . It is necessary to complete the precautions against airborne infections by breath masks and safety-goggles. Pol Arch Med Wewn, 1996 Dec, 96(6), 577 - 80 {An unusual case of Kartagener's syndrome associated with rheumatoid arthritis}; Porawska W et al.; We observed a 56-year-old woman with Kartagener's syndrome and severe seropositive rheumatoid arthritis . This is the third case of such association in the world literature and a second one being diagnosed in our Department . The patient was also as the previous one a carrier of HLA DR1 and B27 antigens . An electromicroscopic study showed normal bronchial cilia in contrast to classical course of the disease . A number of immunological disturbances were observed, especially defective granulocyte function . We suggest that the severe course of rheumatoid arthritis may be related to the chronic stimulation of immune system by microbes continuously present in the patients airways. Microbiologia, 1996 Dec, 12(4), 537 - 46 Fungal siderophores in plant-microbe interactions; Riquelme M; Siderophores are low-molecular-weight high specificity, ferric iron chelating agents . They are produced under iron starvation by most microorganisms . Systems such as siderophores, involved in the acquisition of iron under iron limited conditions, may play a major role in microbial interactions . Some siderophores are virulence factors in animal and in plant pathogens . Moreover, siderophores have been demonstrated to play a major role in plant disease suppression by some bacterial biocontrol agents which inhibit the growth or the activity of plant pathogens by sequestering iron . This latest type of mechanism has been extensively studied in bacteria . However, the role of these iron chelating compounds in disease suppression by fungal biocontrol agents has not been clearly determined. J Cell Sci, 1996 Dec, 109 ( Pt 12), 2855 - 63 Integrin-dependent induction of early growth response genes in capillary endothelial cells; Dike LE et al.; Studies were carried out to explore how extracellular matrix molecules, such as fibronectin (FN), promote capillary endothelial (CE) cell growth . When G0-synchronized cells were plated on FN-coated dishes, expression of the immediate-early mRNAs, c-fos, c-myc and c-jun, was rapidly induced, even in the absence of serum or soluble growth factors . Moreover, plating cells on different FN densities (5-200 micrograms/150 mm dish), resulted in a dose-dependent increase in the steady state levels of these mRNAs . Addition of FGF potentiated gene activation and was required for maximal DNA synthesis, however, the overall steady-state level of gene induction was dictated primarily by the density of immobilized FN . Expression of junB also was induced when suspended cells bound RGD-peptide coated microbeads that promote integrin clustering, but not when the suspended cells bound beads coated with other receptor ligands (e.g . acetylated low density protein) or when they were stimulated by soluble FN or FGF in the absence of substrate adhesion . c-Jun exhibited a similar requirement for gene induction except that it also was partially induced by binding to soluble FN alone . In contrast, c-fos expression was induced by all stimuli tested . Interestingly, inhibition of Na+/H+ exchange using hexamethylene-amiloride prevented most of the FN-induced increase in c-jun expression whereas it was relatively ineffective when cells were simultaneously stimulated by both FN and FGF . These data demonstrate that cell adhesion to extracellular matrix and associated integrin binding can directly activate signaling cascades in quiescent CE cells that lead to induction of immediate-early genes associated with the G0/G1 transition and thereby, stimulate these cells to reenter the growth cycle. Chem Biol, 1996 Dec, 3(12), 1011 - 9 Iron transport-mediated drug delivery using mixed-ligand siderophore-beta-lactam conjugates; Ghosh A et al.; BACKGROUND: Assimilation of iron is essential for microbial growth . Most microbes synthesize and excrete low molecular weight iron chelators called siderophores to sequester and deliver iron by active transport processes . Specific outer membrane proteins recognize, bind and initiate transport of species-selective ferric siderophore complexes . Organisms most often have specific receptors for multiple types of siderophores, presumably to ensure adequate acquisition of the iron that is essential for their growth . Conjugation of drugs to synthetic hydroxamate or catechol siderophore components can facilitate active iron-transport-mediated drug delivery . While resistance to the siderophore-drug conjugates frequently occurs by selection of mutants deficient in the corresponding siderophore-selective outer membrane receptor, the mutants are less able to survive under iron-deficient conditions and in vivo . We anticipated that synthesis of mixed ligand siderophore-drug conjugates would allow active drug delivery by multiple iron receptor recognition and transport processes, further reducing the likelihood that resistant mutants would be viable . RESULTS: Mixed ligand siderophore-drug conjugates were synthesized by combining hydroxamate and catechol components in a single compound that could chelate iron, and that also contained a covalent linkage to carbacephalosporins, as representative drugs . The new conjugates appear to be assimilated by multiple active iron-transport processes both in wild type microbes and in selected mutants that are deficient in some outer membrane iron-transport receptors . CONCLUSIONS: The concept of active iron-transport-mediated drug delivery can now be extended to drug conjugates that can enter the cell through multiple outer membrane receptors . Mutants that are resistant to such conjugates should be severely impaired in iron uptake, and therefore particularly prone to iron starvation. FEMS Microbiol Rev, 1996 Dec, 19(2), 69 - 84 Genetic manipulations of microorganisms for the degradation of hexachlorocyclohexane; Johri AK et al.; Hexachlorocyclohexane (HCH) is an organochlorine insecticide which has been banned in technologically advanced countries . However, it is still in use in tropical countries for mosquito control and thus new areas continue to be contaminated . Anaerobic degradation of HCH isomers have been well documented but until recently there have been only a few reports on aerobic microbial degradation of HCH isomers . The isolation of these microbes made it possible to design experiments for the cloning of the catabolic genes responsible for degradation . We review the microbial degradation of HCH isomers coupled with the genetic manipulations of the catabolic genes . The first part discusses the persistence of residues in the environment and microbial degradation while the second part gives an account of the genetic manipulations of catabolic genes involved in the degradation. Lab Invest, 1996 Dec, 75(6), 801 - 7 Single-cell mutation analysis of tumors from stained histologic slides; Becker I et al.; Formalin-fixed and paraffin-embedded tissues are a valuable resource for diagnosis and research . PCR is one of the most powerful methods of retrospective analysis of the DNA present in fixed tissues . One major problem with the molecular analysis of tissue samples, however, is cellular heterogeneity, ie, the large variety of cell types usually present in these specimens can mask cell-specific genetic alterations associated with disease . Herein we describe a procedure for obtaining and analyzing single cells recovered from stained histologic tissue sections without risking contamination from neighboring cells . An ultraviolet laser microbeam was used to physically destroy the tissue surrounding the single cells of interest . These cells, now freed from adjacent cells, were then easily retrieved with a motorized, computer-controlled micromanipulator and molecularly characterized through the use of PCR-based microanalysis . This accurate microdissection technique, followed by DNA amplification and direct sequencing, revealed a novel mutation in the gene coding for the cell adhesion molecule E-cadherin in single tumor cells that was absent in the adjacent single epithelial cells of a patient with early gastric cancer of the diffuse type . In this form of malignancy, tumor cells lose homophilic cell-to-cell interactions and invade the connective tissue as single cells . E-cadherin gene mutations have previously been detected in advanced diffuse-type gastric cancer and gastric carcinoma cell lines . The present study suggests that E-cadherin gene mutations may be an early event in gastric tumorigenesis . The laser-based isolation and subsequent molecular characterization of individual cells, as described herein, allows for micrometer-sized precision and should prove useful in detecting the nucleic acid abnormalities that underlie cancer, infection, and genetic disease. Thromb Haemost, 1996 Dec, 76(6), 1072 - 9 Detection of platelet adhesion/aggregation to immobilized ligands on microbeads by an aggregometer; Minamoto Y et al.; Platelet aggregation is believed to follow platelet adhesion to vascular injury sites . We have developed a turbidimetric assay for platelet aggregation following platelet adhesion to immobilized ligands using an aggregometer . The addition of polystyrene beads coated with von Willebrand factor (vWF) or fibrinogen (Fg) to platelet suspensions caused prompt aggregation of beads and platelets, which was detected as an increase in light transmission . Electron microscopic analysis revealed that platelets adhered to the bead surfaces and that additional platelets adhered to already adhering platelets, leading to the formation of platelet aggregates . vWF-coated beads induced larger aggregates than Fg-coated beads . The interaction of vWF-coated beads with platelets was abolished by both GPIb and GPIIb-IIIa blockers, while that of Fg-coated beads was abolished by GPIIb-IIIa blockers . vWF-coated beads induced modest secretion of granules from platelets but no thromboxane B2 synthesis . Fg-coated beads induced neither reaction . However, pleckstrin phosphorylation and protein tyrosine phosphorylation was induced by both types of bead . Platelet aggregation following platelet adhesion to both types of bead was inhibited by ADP scavengers, a protein kinase C inhibitor and a tyrosine kinase inhibitor, but not by aspirin . These findings suggest that vWF- and Fg-coated beads can induce platelet aggregation following platelet adhesion through specific ligand-receptor interactions and intracellular signaling . Our simple assay using these beads may represent a useful test for immobilized ligand-induced platelet adhesion and aggregation. Schweiz Rundsch Med Prax, 1996 Nov 5, 85(45), 1444 - 7 {Pathology of Helicobacter infection}; Kirchner T et al.; The colonization of gastric mucosa by Helicobacter pylori (H.p.) is a special form of chronic bacterial infection characterized by long term persistence of microbes because cause of an inefficiency of local immune responses . The resulting chronic gastritis causes decisive transformations of gastric mucosa . They comprise the acquisition of an active mucosa-associated lymphoid tissue as well as the development of glandular atrophy and intestinal metaplasia in the stomach . Both transformations change normal gastric functions, and create abnormal microenvironments with an increased risk for gastric cancer and lymphoma . Own recent investigations indicate, that the distribution and severity of chronic gastritis might be decisively influenced by a rise of antigastric autoimmune reactivity during the H.p . infection . The histologic examination of gastric biopsy samples is essential for an exact diagnosis of the complex pathogenic processes in chronic gastritis . In the future special emphasis of histologic analyses has to be put on the subtypes of gastritis, which are prone for complications and on the evaluation of gastritis remission after H.p . eradication. Dev Comp Immunol, 1996 Nov-Dec, 20(6), 393 - 406 Phagocytes from both vertebrate and invertebrate species use "coiling" phagocytosis; Rittig MG et al.; Coiling phagocytosis has been observed previously only by chance, and there has been no systematic investigation of this uptake mechanism . Therefore, a comparative electron microscopical study was performed . Different human and murine cell populations, phagocytes from various vertebrate and invertebrate species, and predatory amoebae were incubated with Borrelia burgdorferi, one of the microbes known to induce coiling phagocytosis, to study the uptake mechanisms used . In this model, coiling phagocytosis was observed with both vertebrate and invertebrate species but not with amoebae . With cells from humans and mice, this uptake mechanism was restricted to phagocytic cells of myeloid origin . The coiled membrane gaps did not give rise to phagosomes; instead, membrane fusion was followed by membrane dissipation . Thus, coiling of B . burgdorferi apparently is an alternative uptake mechanism used by metazoan phagocytes, involving special membrane processing . However, coiling phagocytosis may show different features with different microbes. Hum Reprod, 1996 Nov, 11(11), 2488 - 92 Zona opening of human embryos using a non-contact UV laser for assisted hatching in patients with poor prognosis of pregnancy; Antinori S et al.; The aim of this study was to examine the safety and the efficiency of a 'non-contact' UV laser to assist hatching through zona opening of human embryos . Between January and November 1995 we performed zona drilling for assisted hatching using a new laser system (PALM UV Laser microbeam), operating in a 'non-contact' mode to create a hole in the zona pellucida of human embryos . In a randomized study, laser zona opening was applied on embryos from two groups of patients with repeated in-vitro fertilization (IVF) failures (two to four attempts): group A was composed of 107 patients who received mixed embryos (216 laser-treated and 223 not treated) and group B of 72 patients who received 218 laser-treated embryos only . Both groups were compared with a control group of 98 patients whose embryos were not laser treated (n = 407) (group C) . The mean ages of all groups (38.1, 38.2 and 37.8 years respectively) and the number of IVF attempts (two to four attempts) were similar . The resulting clinical pregnancies were 39 (36.4%) in group A, 32 (44.4%) in group B and 19 (19.3%) in group C . The implantation rates/embryo were 9.3% in A, 16% in B and 5.1% in the control group . In total, 17 normal babies have been delivered (10 in group A and seven in group B) . These results show that laser zona drilling increased the pregnancy and implantation rates in all the treated patients . The increase was slight but significant in patients of group A (P < 0.01 and P < 0.02), it was even higher in the patients of group B (P < 0.05). Appl Environ Microbiol, 1996 Nov, 62(11), 4129 - 35 Cloning, disruption, and expression of two endo-beta 1, 4-xylanase genes, XYL2 and XYL3, from Cochliobolus carbonum; Apel-Birkhold PC et al.; In culture, the filamentous fungus Cochliobolus carbonum, a pathogen of maize, makes three cationic xylanases, XYL1, which encodes the major endoxylanase (Xyl1), was earlier cloned and shown by gene disruption to encode the first and second peaks of xylanase activity (P . C . Apel, D . G . Panaccione, F . R . Holden, and J . D . Walton, Mol . Plant-Microbe Interact . 6:467-473, 1993) . Two additional xylanase genes, XYL2 and XYL3, have now been cloned from C . carbonum . XYL2 and XYL3 are predicted to encode 22-kDa family G xylanases similar to Xyl1 . Xyl2 and Xyl3 are 60% and 42% identical, respectively, to Xyl1, and Xyl2 and Xyl3 are 39% identical . XYL1 and XYL2 but not XYL3 mRNAs are present in C . carbonum grown in culture, and XYL1 and XYL3 but not XYL2 mRNAs are present in infected plants . Transformation-mediated gene disruption was used to construct strains mutated in XYL1, XYL2, and XYL3 . Xyl1 accounts for most of the total xylanase activity in culture, and disruption of XYL2 or XYL3 does not result in the further loss of any xylanase activity . In particular, the third peak of cationic xylanase activity is still present in a xyl1 xyl2 xyl3 triple mutant, and therefore this xylanase must be encoded by yet a fourth xylanase gene . A minor protein of 22 kDa that can be detected immunologically in the xyl1 mutant disappears in the xyl2 mutant and is therefore proposed to be the product of XYL2 . The single xylanase mutants were crossed with each other to obtain multiple xylanase disruptions within the same strain . Strains disrupted in combinations of two and in all three xylanases were obtained . The triple mutant grows at the same rate as the wild type on xylan and on maize cell walls . The triple mutant is still fully pathogenic on maize with regard to lesion size, morphology, and rate of lesion development. Fertil Steril, 1996 Nov, 66(5), 776 - 80 Microbial contamination in an in vitro fertilization-embryo transfer system; Cottell E et al.; OBJECTIVE: To determine sources and transmission of microorganisms in IVF-ET and efficacy of in-place controlling systems . DESIGN: Prospective study . SETTING: In Vitro Fertilization-Embryo Transfer Unit at a university teaching hospital . PATIENTS: Twenty-eight couples undergoing 30 completed IVF-ET cycles . INTERVENTIONS: Gamete and embryo processing in a penicillin and streptomycin-rich medium . MAIN OUTCOME MEASURES: Presence of microorganisms at various stages of IVF-ET . Fertilization, cleavage, and pregnancy rates . RESULTS: In 50% of cycles no microorganisms were isolated and in the other 50% microbes were cultured from various loci . Cultures of four preprocessed semen samples were positive and corresponding postprocessed samples negative . Microbes were detected in 27% of needle washes after oocyte collection; in 40% and 32% of follicular fluids from left and right ovaries, respectively; and in two culture media from egg-sperm incubations at 20 hours after insemination . No microorganisms were grown from media from zygote incubations . Fertilization, cleavage, and pregnancy rates were independent of microbial presence . CONCLUSION: Seminal fluid and transvaginally collected oocytes are potential sources of microbial contamination of the IVF-ET culture system . A penicillin- and streptomycin-rich culture medium is effective in removing contaminating microbes . End point measures are not affected by commensal contamination. J Immunol Methods, 1996 Oct 16, 197(1-2), 97 - 107 Isolation technique alters eosinophil migration response to IL-8; Rozell MD et al.; Disparate reports exist on the eosinophil chemotactic capacity of interleukin-8 (IL-8) . We hypothesized that the difference is due to the methods used to purify eosinophils . We therefore compared the eosinophilotactic capacity of IL-8 on human cells isolated by Percoll (positive selection) vs . magnetic cell separation system (MACS) (negative selection) . Discontinuous Percoll gradients were preceded by dextran and Ficoll-Paque steps, and followed by gelatin wash and red blood cell (RBC) lysis . MACS isolation included: Percoll 1.090 g/ml layering and RBC lysis; incubation with CD16 antibody conjugated to magnetic beads (to bind neutrophils); and isolation of eluate from column positioned in magnet . Percoll isolated eosinophils migrated to IL-8 in a dose-responsive fashion . Although MACS isolation provided a greater number and higher purity of eosinophils, these eosinophils did not migrate to IL-8 . Neither dextran sedimentation, Ficoll-Paque and Percoll prior to, nor Percoll discontinuous gradients subsequent to, MACS isolation reversed the negative chemotactic response . Moreover, Percoll-isolated eosinophils further purified with CD16 MicroBeads no longer chemotactically responded to IL-8 . This inhibition was not due to change in eosinophil purity, a loss of eosinophil adhesion molecules or activation markers, the presence of a soluble neutrophil or eosinophil inhibitor or the effect of the magnet . Thus, the technique used to isolate eosinophils clearly affects the chemotactic responsiveness of this cell to IL-8 . Since several in vivo studies suggest that IL-8 is an eosinophil chemoattractant, Percoll isolation of these cells might be more appropriate for studies involving eosinophil chemotactic responses to IL-8. J Chromatogr A, 1996 Oct 4, 746(1), 137 - 45 High-speed particle separation and steric inversion in thin flow field-flow fractionation channels; Jensen KD et al.; The performance of thin (down to 71 microns thick) flow field-flow fractionation (flow FFF) channels was examined at both high channel flow-rates (up to approximately 47 ml/min) and cross flow-rates (up to approximately 11 ml/min) . High levels of retention were observed, suggestive of good performance characteristics . Supporting this expectation, four sizes (0.04-0.30 microns) of polystyrene latex microbeads were baseline separated in under 3 min . Nonetheless, a plate height analysis showed that performance was still less than theoretically expected . Fast steric-hyperlayer separations of larger latex beads were observed in the same systems . Furthermore, it was shown that the steric inversion diameter was shifted down to approximately 0.23 micron thus expanding the size range to which this FFF mode is applicable . The steric inversion phenomenon observed using narrow latex standards was shown to be consistent with that found for a polydisperse polyvinylchloride latex. Patol Fiziol Eksp Ter, 1996 Oct-Dec, (4), 28 - 30 {Activity of glutamate dehydrogenase, gamma-glutamyltranspeptidase and creatine kinase in saliva in gingivitis}; Petrovich IuA et al.; In the human saliva (oral fluid), the activity of glutamate dehydrogenase tends to decrease in gingivitis and diminishes in periodontitis . That of gamma-glutamyl transpeptidase and creatine kinase increases in gingivitis . In periodontitis, their activity is higher than that in gingivitis . The above changes in enzymatic activities reflect metabolic changes in the gingiva and periodontium in inflammation . The saliva from the salivary ducts shows no activity of the above enzymes . They enter the mixed saliva (oral fluid) together with epithelial and other oral tissue cells, with leukocytes and microbes which manifold in gingivitis. Dtsch Tierarztl Wochenschr, 1996 Oct, 103(10), 438 - 43 {The epidemiology of helicobacteriosis in humans; studies of the survival capacity of the microbe in food}; Bohmler G et al.; In man suffering from diseases of the stomach and the duodenum (gastritis, ulcus, enteritis, neoplasms), Helicobacter pylori (H..pylori) is frequently detected in the mucous membrane of the stomach . Up to now the spread of this agent is not quite clear . Since the direct transmission in humans can be taken for granted, the following study was to find out whether and for how long the agent mentioned above is able to survive in selected food and whether an infection of the consumer by these contaminated food is possible . 376 samples of secretions from the udder of healthy cows and those with mastitis where tested for the presence of H . pylori along with 100 stomachs of chicken from different flocks . In no case H . pylori could be detected . H . pylori was inoculated in high concentrations into milk and some milk-products . From cooled milk samples the agent could still be reisolated after six days in a density up to 10(3) CFU/ml of milk . At room-temperature or 37 degrees C resp . the pathogen could be detected in milk for three to four days only . In yoghurt the agent kept viable for three hours only, whereas in kefir for 24 hours . Mean survival time of then hours was found in pH-neutral curd cheese . The incubation of H.pylori in sterile drip from chicken and in physiologic saline resulted in maximal survival time of at least 48 hours at room temperature . But in H.pylori-broth the number of microorganisms had dropped below the limit of detectability only after 72 hours . At refrigerator-temperature (7 degrees C) H . pylori could still be detected within these three media after 72 hours in high concentrations . In drip from chicken kept at-20 degrees C before thawing H . pylori showed a considerable survival time . After four weeks its number had only dropped by one to two log cycles, whereas in saline and in broth the agent could not be detected anymore after one week at the most . Experiments concerning tenacity showed: On culture-media with different pH-values the growth-optimum of H . pylori was between pH 6.1 and 7.3 H . pylori was suspended in melting water from chicken and brought in thin layers onto wooden board, plastic and ceramic tiles . The bacterium could be recultured from these surfaces only as long as these were moist . At room-temperature the bacterium could not be detected anymore on wood after 30 minutes, on plastic or ceramic tiles after 90 minutes . At refrigerator-temperature the administered suspensions dried more slowly, so that H . pylori survived longer, but it still could not be isolated anymore on wood after 240 minutes, on plastic or ceramic tiles after 300 minutes . The decimal reduction-time for H . pylori suspensions in broth were . 72 sec . at +50 degrees C 43 sec . at +52 degrees C 20 sec . at +55 degrees C 10 sec . at +57 degrees C 4 sec . at +60 degrees C from which data z = 7.9 +/- 0.01 degrees C can be calculated . From these experiments on can conclude, that in all probability fresh milk and chicken do not contain H . pylori and thus do not represent a source of infection for man . After contamination of slaughtered chicken within the abattoir or from milk and milk-products within dairy industry by insufficient hygiene-management of infected personnel it can not be excluded, that H . pylori gets into households by these foods . An infection of the consumer by this route is not very likely, but can not be excluded with complete certainly. Med Parazitol (Mosk), 1996 Oct-Dec, (4), 3 - 8 {The interrelations of mucosal epithelial cells with microbes--intracellular parasites}; Domaradskii IV et al.; On the basis of information at hand, it can be concluded that there are principal limits on the capture of bacteria by epithelial cells in the gastrointestinal tract and other cavities . Although entry is, however, a result both of bacteria and epithelial cells, it is frequently induced by parasite-directed endocytosis and may also accompanied by passive entry of nonpathogenic bacteria . The induction of parasite-induced endocytosis should be regarded as a consequence of a fundamental concept in biology, namely: molecular and cellular recognition wherein bacterial adhesion to the host's cell receptors is of great importance . In this connection, discussion covers the origin of corresponding receptors . The authors share the opinion in that epithelial cells, those of the gastrointestinal tract in particular, are part of the human common mucosal immune system. Biosci Biotechnol Biochem, 1996 Oct, 60(10), 1604 - 7 Efficient L-serine production from methanol and glycine by resting cells of Methylobacterium sp . strain MN43; Hagishita T et al.; Resting cells of methanol-utilizing microorganisms isolated from soils were examined for L-serine production under conditions in which L-serine-degradation was suppressed . Strain MN43, a facultative methylotrophic bacterium identified as a Methylobacterium sp., was selected for further studies . Under the optimal conditions, 65 mg/ml L-serine was produced by this bacterium from 50 mg/ml glycine and 104 mg/ml methanol in 5 days, with a molar conversion ratio from glycine to L-serine of 93% . This production is the highest so far reported for microbes producing L-serine. Int J Parasitol, 1996 Oct, 26(10), 1063 - 74 Infection of sheep with adult and larval Ostertagia circumcincta: effects on abomasal pH and serum gastrin and pepsinogen; Lawton DE et al.; Infection of sheep with adult or larval O . circumcincta increased serum pepsinogen and gastrin and abomasal pH . The upper limits of the normal range, calculated from over 1000 samples collected from parasite-naive sheep, were set at 2 standard deviations above the mean; these were for serum pepsinogen, 454 mU tyrosine l-1; serum gastrin, 64 pM and abomasal pH, 3.26 . Five infection regimes were used: sheep previously exposed to field parasitism were infected with 30,000 larvae intraruminally (Group A), while parasite-naive sheep were administered either 50,000 larvae intraruminally (Group B), 150,000 larvae intraruminally followed by a trickle infection of 10,000 larvae thrice weekly from days 21 to 45 (Group C), 150,000 exsheathed larvae via an abomasal cannula (Group D) or 15,000 adult worms via an abomasal cannula (Group E) . Whereas the presence of adult worms rapidly increased serum pepsinogen (after 8 h) and abomasal pH and serum gastrin (after about 19 h), the early infective larval stages, regardless of the infection regime, had minimal effects until the abrupt rise in all parameters 5-6 days after infection . Abomasal pH returned to near normal levels when the infections became patent and was not re-elevated by a subsequent trickle infection, whereas serum gastrin and pepsinogen remained high . The initial hypergastrinaemia was coincident with the increased abomasal pH, but was preceded by the increase in serum pepsinogen . In several sheep, serum pepsinogen increased very little during the parasitism, although there were typical effects on abomasal pH and serum gastrin . Serum gastrin was depressed when the abomasal pH exceeded about 5.5 . It is suggested that an inhibitor of gastrin release is generated by proliferating abomasal microbes under these conditions and that this is a limitation to the use of elevated serum gastrin in the diagnosis of parasitism in individual sheep. APMIS, 1996 Oct, 104(10), 689 - 97 Virus-induced autoimmune disease: transgenic approach to mimic insulin-dependent diabetes mellitus and other autoimmune diseases; Oldstone MB et al.; The technology of cloning viral genes and expressing them in vivo under cell-specific promoters allows to dissect the role of viruses, host self proteins, host genetics and immune responses in the complex etiology of autoimmune disease . Expression of a viral transgene, that is really a marker for a host "self" protein per se in beta cells of the islets of Langerhans, need not cause disease . In our model, expression of a viral gene was not associated with disease over the lifetime of the animal . However, when the host becomes infected with a virus encoding the same gene as the transgene or one closely related to it, a resultant immune response directed against the virus also recognizes the transgene leading to progressive T-cell-mediated response and destruction of the tissue expressing the viral ("self") gene, leading to autoimmune disease . This multifactorial process is influenced by whether the viral transgene is expressed in the thymus as well as in the disease-related cell or target tissue . Thymic expression influences negative selection of responder lymphocytes and thus delays the onset of the autoimmune disorder . Further, the MHC haplotype or other background genes of an individual undergoing autoimmune dysfunction play a role in the affinity of binding of the transgene products to the MHC molecule and influence the degree of negative selection that occurs, thereby influencing the vigor of the resulting immune response . The current ability to express host or viral genes in unique cell populations, and to make double- or triple-tg mice in which various cytokine genes or lymphocyte activation genes can be expressed along with the viral gene, offers a unique possibility for molecular dissection of autoimmunity . With the information on hand, approaches to the prevention and treatment of human autoimmune disease are likely to be uncovered . Finally, animal models are available in which the onset, progression and control of molecular mimicry can be evaluated . Future studies should define roles played by cytokines, bystander and immune-specific cross-reactivity to viruses and other microbes in several autoimmune diseases. Hum Reprod, 1996 Oct, 11(10), 2162 - 4 Effects of ultraviolet exposure and near infrared laser tweezers on human spermatozoa; Konig K et al.; Photostress has to be considered during optical micromanipulation of gametes . Ultraviolet light, including low-energy UVA (32-400 nm) radiation, as well as high-intensity near infrared (NIR) laser radiation may induce cell damage . A total number of 580 light-exposed sperm cells were studied in single-cell photostress experiments . Low-power (1.5 mW, 5.3 W/cm2) UVA exposure with 365 nm radiation of a standard mercury microscopy lamp to human spermatozoa resulted within 109 +/- 30 s in paralysis and within 310 +/- 110 s in cell death . Cytotoxic effects during cell manipulation with laser microbeams were found to be partly based on non-linear excitation phenomena, in particular two-photon absorption by endogenous cell chromophores . Two-photon absorption will be more intense in the case of pulsed laser microradiation, but occur also during micromanipulation with highly focused continuous wave (cw) microbeams used as laser tweezers ('optical traps') . In particular, short-wavelength NIR traps < 800 nm induce UVA-like biological effects (oxidative stress) . For example, sperm trapping with 760 nm microbeams resulted in UVA-like autofluorescence modifications, paralysis within 35 +/- 20 s and cell death within 65 +/- 20 s . In contrast, laser microbeams at 800-1064 nm may act as relatively safe micromanipulation tools . In most optical traps multifrequency cw lasers are employed . Radiation of these lasers can magnify cytotoxic effects . Therefore, single-frequency laser operation should be preferred . In general, laser assisted cell micromanipulation requires a new understanding of microbeam-cell interaction, including aspects of non-linear optics. Infect Immun, 1996 Oct, 64(10), 4123 - 8 Cellular immune responses to recombinant heat shock protein 70 from Histoplasma capsulatum; Allendoerfer R et al.; Heat shock protein (hsp) 70 from several microbes is antigenic in mammals . In this study we sequenced and expressed the gene encoding this protein from Histoplasma capsulatum to study its immunological activity . The deduced amino acid sequence of the gene demonstrated 71 and 76% identity to hsp7O from humans and Saccharomyces cerevisiae, respectively . A cDNA was synthesized by reverse transcription-PCR and was expressed in Escherichia coli . Recombinant protein reacted with a mouse monoclonal antibody raised against human hsp7O . Splenocytes from C57BL/6 mice immunized with recombinant hsp7O emulsified in adjuvant, but not yeast cells, reacted in vitro to the antigen . Recombinant hsp7O elicited a cutaneous delayed-type hypersensitivity response in mice immunized with protein or with viable yeast cells . Mice were injected with recombinant hsp7O and challenged intranasally with a sublethal inoculum of yeast cells . Vaccination did not confer protection in this model . Thus, recombinant hsp7O can induce a cell-mediated immune response but does not induce a protective response. Clin Microbiol Rev, 1996 Oct, 9(4), 489 - 98 Pneumocystis carinii: what is it, exactly? Stringer JR. The identity of Pneumocystis carinii has been uncertain for many years . Until recently, it was widely regarded to be a protozoan because it does not grow in culture and is not susceptible to antifungal drugs . Although these and a number of other phenotypic characteristics of P . carinii differ from those of typical fungi, analysis of DNA sequences has shown that P . carinii is a member of the fungal lineage of eukaryotes . However, a close phylogenetic relative of P . carinii has not yet been found . Analysis of gene sequences has also revealed that P . carinii is not a single entity but that the genus Pneumocystis contains a complex group of organisms . P . carinii organisms from one host species do not grow when introduced into another host species, and P . carinii isolates from different host species are more genetically divergent from one another than might be expected for members of the same species . Genetic variation of a lesser degree also occurs among P . carinii organisms from the same host species, suggesting that it may be possible to identify strains and to conduct transmission and epidemiological studies . Results of early studies exploiting genetic variation among P . carinii isolates from patients have suggested that recurrent P . carinii pneumonia may not always be caused by reactivation of latent organisms, as is commonly believed . However, other features of P . carinii suggest that this microbe has established a long-term relationship with its host . A striking new development in this regard is the discovery of a genetic system that is designed to allow variation in the structure of a major antigen on the surface of P . carinii. Biophys J, 1996 Oct, 71(4), 2158 - 67 Physiological monitoring of optically trapped cells: assessing the effects of confinement by 1064-nm laser tweezers using microfluorometry; Liu Y et al.; We report the results of microfluorometric measurements of physiological changes in optically trapped immotile Chinese hamster ovary cells (CHOs) and motile human sperm cells under continuous-wave (CW) and pulsed-mode trapping conditions at 1064 nm . The fluorescence spectra derived from the exogenous fluorescent probes laurdan, acridine orange, propidium iodide, and Snarf are used to assess the effects of optical confinement with respect to temperature, DNA structure, cell viability, and intracellular pH, respectively . In the latter three cases, fluorescence is excited via a two-photon process, using a CW laser trap as the fluorescence excitation source . An average temperature increase of < 0.1 +/- 0.30 degrees C/100 mW is measured for cells when held stationary with CW optical tweezers at powers of up to 400 mW . The same trapping conditions do not appear to alter DNA structure or cellular pH . In contrast, a pulsed 1064-nm laser trap (100-ns pulses at 40 microJ/pulse and average power of 40 mW) produced significant fluorescence spectral alterations in acridine orange, perhaps because of thermally induced DNA structural changes or laser-induced multiphoton processes . The techniques and results presented herein demonstrate the ability to perform in situ monitoring of cellular physiology during CW and pulsed laser trapping, and should prove useful in studying mechanisms by which optical tweezers and microbeams perturb metabolic function and cellular viability. Am J Respir Crit Care Med, 1996 Oct, 154(4 Pt 2), S133 - 6 Carbohydrate recognition by Mycoplasma pneumoniae and pathologic consequences; Feizi T et al.; Mycoplasma pneumoniae infection in the human is often followed by a transient autoimmune hemolytic disorder characterized by high titer autoantibodies to a carbohydrate antigen, the I antigen . Because the major host cell receptor for the Mycoplasma is the sialylated form of this antigen, it is likely that the immunologic disorder is initiated by the microbe-saccharide interaction . Here we review briefly knowledge on the autoantibodies and the structures and distribution of the saccharide antigens and receptors . We discuss possible mechanisms for the triggering of autoantibody production and consider ways in which perturbation of various glycoprotein carriers of the carbohydrate ligands may elicit a variety of pathobiologic responses . We conclude by highlighting ideas on further molecular dissections of the elements of the microbe-host interaction. J Chromatogr B Biomed Appl, 1996 Sep 20, 684(1-2), 25 - 49 Methods for the separation of lactate dehydrogenases and clinical significance of the enzyme; Kopperschlager G et al.; Lactate dehydrogenase (LDH), an ubiquitous enzyme among vertebrates, invertebrates, plants and microbes was discovered in the early period of enzymology . The enzyme has been dissolved in several distinguishable molecular forms . In mammals, three types of subunits encoded by the genes Ldh-A, Ldh-B and Ldh-C give rise to a selected number of tetrameric isoenzymes . LDH-A4, LDH-B4 and the mixed hybrid forms of the A- and B-subunits are present in many tissues but with certain distribution patterns . LDH-C4 is confined in mammals to testes and sperm . Numerous techniques have been employed to purify, characterize and separate the different forms of the enzyme . This report deals with the main protocols and procedures of purification of LDH and its isoenzymes including chromatographic and electrophoretic methods, partitioning in aqueous two-phase systems and precipitation approaches . In particular, affinity separation techniques based on natural and pseudo-biospecific ligands are described in detail . In addition, basic physico-chemical and kinetic properties of the enzyme from different sources are summarized in a second part, the clinical significance of the determination of LDH in diverse body fluids in respect to the total activity and the isoenzyme distribution in different organs is discussed. Ukr Biokhim Zh, 1996 Sep-Oct, 68(5), 103 - 5 {Determination of the content of common protein in biological samples using chemical stabilizers}; Iarmol'chuk HM; Proteins of biological samples in the process of storage are subject to microbe contamination as well as the destruction by their own lipoprotein lipases . It is established that N-thymol compounds of formulas 52 and 82 stabilize blood serum samples of donors and preserve the content of total protein in them unchanged for 7, 15 and 40 days, respectively . The obtained stabilizers can be used when studying protein metabolism in the organism of cosmonauts performing long-term space flight, depots of nuclear submarines, participants of long-term Alpine, desert, polar and other expeditions. J Exp Med, 1996 Sep 1, 184(3), 803 - 10 Limited role of CD28-mediated signals in T helper subset differentiation; Brown DR et al.; The role of CD28-mediated signals in T helper cell maturation is not fully understood . We tested the requirement for costimulation through CD28 in several systems of CD4+, T cell differentiation . In vivo priming of mice with genetic disruption of CD28 (CD28-/-) yielded normal levels of antigen-specific interferon gamma production but markedly diminished levels of interleukin 4 (IL-4) after in vitro restimulation . In response to the pathogenic microbe, Leishman a major, C57BL6 CD28-/- mice were fully capable of controlling infection and exhibited a normal T helper 1 response . BALB/c CD28-/- mice unexpectedly exhibited normal susceptibility to L . major . BALB/c CD28-/- mice developed high levels of IL-4 mRNA and protein induction in the draining lymph nodes . In addition, susceptibility of BALB/c CD28-/- mice was reversed by neutralization of IL-4 in vivo . We also activated transgenic CD28-bearing T cells from the BALB and C57BL background in vitro in the presence of CTLA4Ig . BALB cells had greater IL-4 producing capacity than C57BL cells in the absence of costimulation . Diverse factors including costimulatory signals, genetic polymorphism, and the nature of the immunogen all influence T helper phenotype commitment, but these results provide evidence that CD28 is not an absolute requirement for generating either Th1 or Th2 responses. Sante, 1996 Sep-Oct, 6(5), 269 - 74 {Perception of lepers by non-lepers in an urban center in Cameroon}; Touko A et al.; We investigated the knowledge and perceptions the general population have about leprosy in a sociological study between 1994 and 1995 in the city of Yaounde (Cameroon) . The sample consisted of 251 respondents aged between 15 and 73 (mean 30 +/- 18) years old . Asked about the causes of leprosy, 27.8% knew that the disease was caused by microbes, 33.4% did not know the cause, 17.4% thought the disease to be hereditary and 7.6% due to ill luck . Attitudes to lepers depended on the type of behavior considered . Thus, interactions that did not involve physical contact were generally welcomed (94%), whereas physical contact was shunned: only 45% would tolerate it . Intimacy was rejected by almost all subjects . Analysis of the opinion of leprosy victim-oriented social projects indicated that most subjects showed humanitarian concern for lepers, but preferred public to individual support . The study group did not want to be individually or personally committed to any form of special support for leprosy victims, but clearly saw the need and justification for public concern for these patients. Clin Lab Haematol, 1996 Sep, 18(3), 161 - 9 Usefulness of flow cytometric detection of cell surface interleukin-6 receptors in human myeloma cell lines; Chen YH et al.; A flow cytometric procedure was used to analyse the cell surface interleukin-6 receptor (IL-6R) based on the principle of detecting the binding of IL-6 to IL-6R . The bound IL-6 was visualized by reacting with anti-IL-6 antibody, a second biotinylated antibody to immunoglobulins, fluorescein-conjugated avidin and biotinylated, fluorescein-conjugated bovine serum albumin . Studies with a number of human myeloma cell lines showed that the fluorescence signal from cellular binding of IL-6, and thus IL-6R, could be specifically discerned from the background . The specific IL-6R signal could be semi-quantitatively expressed as the mean channel number of the fluorescence histogram or as relative IL-6R density in relation to a reference cell line, such as U266-BL cells . The relative IL-6R density of various myeloma cell lines thus determined was found to correlate with their sensitivity to the growth inhibitory effect of glucocorticoids in vitro . Quantitatively, calibration of the staining procedure with microbeads that have a defined binding capacity for anti-IL-6 antibody allowed calculation of cellular IL-6R density that yielded results close to that reported with conventional radio-ligand binding assay . Similarly, the cytometric method was applied to the studies of kinetics of IL-6/IL-6R interaction and the cellular IL-6R changes after ligand-binding to provide estimates of IL-6R binding affinity and IL-6R turnover rate . It is suggested that flow cytometric measurement of cellular IL-6R is useful in providing biologically relevant information on myeloma cells. Chromosome Res, 1996 Sep, 4(6), 411 - 26 Characterization of DNA sequences constituting the terminal heterochromatin of the chicken Z chromosome; Hori T et al.; Two clones, pCZTH5-8 and pCZTH12-8, were isolated from a female chicken genomic library by screening with sequences obtained from genomic libraries which had been constructed from a terminal region of a single Z chromosome of chicken utilizing laser microbeam irradiation and PCR amplification . Fluorescence in situ hybridization to the mitotic Z chromosome and the lampbrush ZW bivalent of chicken demonstrated that both the cloned sequences are located in the heterochromatic region of the Z chromosome at the end opposite to the pairing region with the W chromosome . The sequences pCZTH-8 and pCZTH12-8 are distributed widely on both the telomeric bow-like loops (TBL) and the region I (short loops region) of the Z lampbrush chromosome . These clones, pCZTH12-8 particularly notably, hybridized also to the TBLs of lampbrush bivalents 1-4 of chicken . Both sequences are transcribed in the lampbrush stage oocytes on the Z chromosome and on other macrobivalents . The subfragment of pCZTH5-8 which hybridizes to the TBLs and the insert of pCZTH12-8 contain regions that are closely similar in sequence . The pCZTH-8 sequence has no internal repeats and may be part of the 24-kb macrosatellite repeating unit that is evident after Nhel digestion of the genomic DNA . A cloned 24-kb unit, pFN-1, does not show significant DNA curvature, but cytosines of its CpG dinucleotides may be highly methylated in vivo . This contrasts with the repeat sequences of the W heterochromatin which not only have highly methylated CpG but are also strongly curved . The 24-kb unit is repeated about 830 times in the diploid genome of a female chicken, suggesting that nearly the entire terminal heterochromatin on the Z chromosome consists of this macrosatellite family . Sequences of the greater part of the pCZTH-8 are restricted to the genus Gallus but the sequence of one subregion which hybridizes to TBLs is present in the genomes of the order Galliformes. J Microsc, 1996 Sep, 183 ( Pt 3), 197 - 204 Two-photon excited lifetime imaging of autofluorescence in cells during UVA and NIR photostress; Konig K et al.; By monitoring coenzyme autofluorescence modifications, as an indicator of cell damage, the cellular response to femtosecond near-infrared (NIR) radiation (two-photon absorption) was compared with exposure to low-power UVA radiation (one-photon absorption) . Excitation radiation from a tunable Ti-sapphire laser, focused through high-numerical-aperture microscope optics, provided diffraction-limited microbeams of an adjustable peak power . Laser scanning NIR microscopy was used to detect spatially the intracellular distribution of fluorescent coenzymes by fluorescence intensity imaging as well as fluorescence lifetime imaging (tau-mapping) . Upon the onset of UV or NIR exposure, Chinese hamster ovary cells exhibited blue/green autofluorescence with a mean lifetime of 2.2 ns, which was attributed to NAD(P)H in mitochondria . Exposure to 365 nm radiation from a high-pressure mercury lamp (1 mW, 300 J cm-2) resulted in oxidative stress correlated with increased autofluorescence intensity, onset of nuclear fluorescence, and a fluorescence lifetime decrease . The cellular response to femtosecond NIR microbeams depended significantly on peak power . Peak powers above a threshold value of about 0.5 kW (average power: 6 mW) . 0.55 kW (7 mW) and 0.8 kW (10 mW) at 730 nm, 760 nm and 800 nm, respectively, resulted in the onset of short-lived luminescence with higher intensity (100 x) than the intracellular NAD(P)H fluorescence . This luminescence, accompanied by destruction of cellular morphology, was localized and occurred in the mitochondrial region . In contrast, beams at a power of less than 0.5 kW allowed nondestructive fluorophore detection with high spatial and temporal resolution without modification of cellular redox state or cell morphology. Microbiol Rev, 1996 Sep, 60(3), 473 - 82 Virus-encoded superantigens; Huber BT et al.; Superantigens are microbial agents that have a strong effect on the immune response of the host . Their initial target is the T lymphocyte, but a whole cascade of immunological reactions ensues . It is thought that the microbe engages the immune system of the host to its own advantage, to facilitate persistent infection and/or transmission . In this review, we discuss in detail the structure and function of the superantigen encoded by the murine mammary tumor virus, a B-type retrovirus which is the causative agent of mammary carcinoma . We will also outline what has more recently become known about superantigen activity associated with two human herpesviruses, cytomegalovirus and Epstein-Barr virus . It is likely that we have only uncovered the tip of the iceberg in our discovery of microbial superantigens, and we predict a flood of new information on this topic shortly. J Neurosci, 1996 Sep 1, 16(17), 5466 - 77 Regenerative proliferation in organ cultures of the avian cochlea: identification of the initial progenitors and determination of the latency of the proliferative response; Warchol ME et al.; Sensory hair cells in the cochleae of birds are regenerated after the death of preexisting hair cells caused by acoustic over-stimulation or administration of ototoxic drugs . Regeneration involves renewed proliferation of cells in an epithelium that is otherwise mitotically quiescent . To determine the identity of the first cells that proliferate in response to the death of hair cells and to measure the latency of this proliferative response, we have studied hair-cell regeneration in organ culture . Cochleae from hatchling chicks were placed in culture, and hair cells were killed individually by a laser microbeam . The culture medium was then replaced with a medium that contained a labeled DNA precursor . The treated cochleae were incubated in the labeling media for different time periods before being fixed and processed for the visualization of proliferating cells . The first cells to initiate DNA replication in response to the death of hair cells were supporting cells within the cochlear sensory epithelium . All of the labeled supporting cells were located within 200 microns of the hair-cell lesions . These cells first entered S-phase approximately 16 hr after the death of hair cells . The results indicate that supporting cells are the precursors of regenerated hair cells and suggest that regenerative proliferation of supporting cells is triggered by signals that act locally within the damaged epithelium. Infect Immun, 1996 Sep, 64(9), 3491 - 6 Partial characterization of a cell proliferation-inhibiting protein produced by Helicobacter pylori; Knipp U et al.; Despite the induction of an immunological reaction, Helicobacter pylori-associated gastritis is a chronic disease, suggesting that this microbe can evade the host immune defense . Previous studies by our group showed that H . pylori suppresses the in vitro proliferative response of human mononuclear cells to mitogens and antigens . Here we demonstrate that the antiproliferative activity of H . pylori also affects the proliferation of various mammalian cell lines (U937, Jurkat, AGS, Kato-3, HEP-2, and P388D1) . This effect is detectable in the first 16 h of incubation and maximal between 24 and 48 h . In addition, the presence of H . pylori significantly diminished the protein synthesis of cells in the first 6 h of incubation, comparable to the results with cycloheximide and diphtheria toxin . The urease enzyme, the cagA gene product, and the vacuolizing cytotoxin of H . pylori were excluded as causative agents of the antiproliferative effect by using isogenic knockout mutant strains . The inhibitory effect was not due to a lytic activity of this bacterium . The results reported here indicate that the responsible factor is a protein with an apparent native molecular mass of 100 +/- 10 kDa . Our work implicates the presence of a protein factor in H . pylori (termed PIP {for proliferation-inhibiting protein}) with antiproliferative activity for mammalian cells, including immunocompetent and epithelial cells . Thus, it is reasonable to presume that this property may contribute to the pathogenesis of H . pylori-induced diseases . It may be involved on the one hand in immune response evasion and on the other hand in the suppression of epithelial repair mechanisms. J Neurotrauma, 1996 Aug, 13(8), 417 - 37 Effects of methylprednisolone on lesioned and uninjured mammalian spinal neurons: viability, ultrastructure, and network electrophysiology; Rosenberg LJ et al.; An in vitro investigation was undertaken to provide information regarding the effectiveness of methylprednisolone sodium succinate (MPSS) as a treatment for the primary mechanical injury of spinal cord (SC) trauma . Exposure of uninjured mouse SC cells to MPSS for 24 h caused neuronal stress when the concentration exceeded 150 micrograms/mL; neuronal death occurred at concentrations above 600 micrograms/mL . The concentration range for MPSS protection of SC neurons subjected to a defined physical injury (laser microbeam transection of a primary dendrite 100 microns from the perikaryon) was very narrow: survival in the 30 micrograms/mL group differed significantly from the untreated control group (68.5% +/- 14.1 vs . 47.1% +/- 14.1), treatment with 20 or 60 micrograms/mL MPSS did not increase survival, and treatment with 100 micrograms/mL MPSS accelerated ultrastructural deterioration and increased the likelihood of death . Enhanced survival of lesioned neurons was observed when 30 micrograms/mL MPSS was applied within 15 min of dendrotomy but not when MPSS was administered 2 h after lesioning . Multimicroelectrode plate (MMEP) studies of SC network electrical activity indicated that MPSS associated readily with neuronal membranes . This finding was consistent with the hypothesis that MPSS may protect lesioned neurons by stabilizing damaged membranes, enhancing lesion resealing, and limiting the spread of ion-mediated damage . However, comparisons of neurite die-back 24 h after dendrotomy found no significant difference between MPSS-treated and control neurons . Application of 30 or 100 micrograms/mL MPSS increased the spontaneous burst activity of SC networks grown on MMEPs, however, there was no evidence that the increased excitability at these concentrations was the result of specific actions of MPSS on GABA or NMDA synapses. J Dairy Sci, 1996 Aug, 79(8), 1496 - 502 Why don't ruminal bacteria digest cellulose faster? Weimer PJ. The bacteria Fibrobacter succinogenes, Ruminococcus flavefaciens, and Ruminococcus albus generally are regarded as the predominant cellulolytic microbes in the rumen . Comparison of available data from the literature reveals that these bacteria are the most actively cellulolytic of all mesophilic organisms described to date from any habitat . In light of numerous proposals to improve microbial cellulose digestion in ruminants, it is instructive to examine the characteristics of these species that contribute to their superior cellulolytic capabilities and to identify the factors that prevent them from digesting cellulose even more rapidly . As a group, these species have extreme nutritional specialization . They are able to utilize cellulose (or in some cases xylan) and its hydrolytic products as their nearly sole energy sources for growth . Moreover, each species apparently has evolved to similar maximum rates of cellulose digestion (first-order rate constants of 0.05 to 0.08 h-1) . Active cellulose digestion involves adherence of cells to the fibers via a glycoprotein glycocalyx, which protects cells from protozoal grazing and cellulolytic enzymes from degradation by ruminal proteases while it retains-at least temporarily-the cellodextrin products for use by the cellulolytic bacteria . These properties result in different ecological roles for the adherent and nonadherent populations of each species, but overall provide an enormous selective advantage to these cellulolytic bacteria in the ruminal environment . However, major constraints to cellulose digestion are caused by cell-wall structure of the plant (matrix interactions among wall biopolymers and low substrate surface area) and by limited penetration of the nonmotile cellulolytic microbes into the cell lumen . Because of these constraints and the highly adapted nature of cellulose digestion by the predominant cellulolytic bacteria in the rumen, transfer of cellulolytic capabilities to noncellulolytic ruminal bacteria (e.g., by genetic engineering) that display other desirable properties offers limited opportunities to improve ruminal digestion of cellulose. J Vet Pharmacol Ther, 1996 Aug, 19(4), 295 - 9 Stability of ivermectin in rumen fluids; Andrew NW et al.; To determine whether ivermectin is metabolized in the rumen, in vitro studies were conducted with the tritium-labelled H2B1a component of ivermectin in rumen fluid from sheep and cattle . No detectable metabolism occurred over 24 h in in vitro incubations at 38 degrees C . The viability of the microbes in the rumen fluids was demonstrated by the conversion of 17% and 11% of {14C}cellulose to 14CO2 in 24 h in the incubations with sheep and steer rumen fluids respectively . The results indicate that ivermectin is not metabolized in the rumen . Based on the lack of in vitro metabolism of ivermectin in rumen fluid, the similarity of in vitro liver microsomal metabolism with in vivo metabolism of the avermectins and the physicochemical properties of the avermectins, any disappearance of ivermectin in vitro from rumen fluid is probably a result of binding to solids or surfaces . Apparent discrimination by dung beetles, where observed, between control faeces and faeces from cattle or sheep treated with ivermectin or abamectin therefore must be attributable to chance, to factors unrelated to treatment or to factors such as changes in amino acid composition rather than the production of volatile metabolites of ivermectin. Can J Microbiol, 1996 Aug, 42(8), 811 - 8 Effect of heat flushing on the concentrations of Legionella pneumophila and other heterotrophic microbes in hot water systems of apartment buildings; Zacheus OM et al.; The decontamination of Legionella pneumophila and other heterotrophic microbes by heat flushing in four legionellae-positive hot water systems was studied . Before the decontamination procedure, the concentration of legionellae varied from 3.0 x 10(3) to 3.5 x 10(5) cfu/L and the hot water temperature from 43.6 to 51.5 degrees C . During the contamination the temperature was raised to 60-70 degrees C . All taps and showers were cleaned from sediments and flushed with hot water twice a day for several minutes . The decontamination lasted for 2-4 weeks . In a few weeks the heat-flushing method reduced the concentration of legionellae below the detection limit (50 cfu/L) in the hot circulating water system just before and after the heat exchanger . The high hot water temperature also decreased the viable counts of heterotrophic bacteria, fungi, and total microbial cells determined by the epifluorescent microscopy . However, the eradication of legionellae failed in a water system where the water temperature remained below 60 degrees C in some parts of the system . After the decontamination, the temperature of hot water was lowered to 55 degrees C . Thereafter, all the studied hot water systems were recolonized by legionellae within a few months, showing that the decontamination by heat flushing was temporary . Also, the contamination of other bacteria increased in a few months to the level before decontamination. Braz J Med Biol Res, 1996 Jul, 29(7), 873 - 5 The role of trehalose in cell stress; de-Araujo PS; Water is usually thought to be required for the living state, but many organisms can withstand anhydrobiosis when essentially all of their body water has been removed . The mechanisms for survival to this kind of stress could be similar in microbes, plants and animals . One common feature is the accumulation of sugars by anhydrobiotic organisms . Trehalose, which is one of the most effective saccharides in preventing phase transition events in the lipid bilayer, is accumulated by anhydrobiotic organisms in large amounts . It lowers membrane phase transitions in dry yeast cells, thus preventing imbibitional damages when cells are rehydrated . Yeast cells have a trehalose carrier in the plasma membrane which endows them with the ability to protect both sides of the membrane . Kinetic analysis of the trehalose transport activity in Saccharomyces cerevisiae cells revealed the existence of a multicomponent system with a constitutive low-affinity uptake component and a high-affinity H(+)-trehalose symporter regulated by glucose repression. Gut, 1996 Jul, 39(1), 100 - 4 High intracolonic acetaldehyde values produced by a bacteriocolonic pathway for ethanol oxidation in piglets; Jokelainen K et al.; BACKGROUND: Human colonic contents and many colonic microbes produce considerable amounts of acetaldehyde from ethanol in vitro . AIMS: To examine in piglets if acetaldehyde is produced in the colon also in vivo, and if so, what is the fate of intracolonically formed acetaldehyde . ANIMALS: Seventeen native, non-fasted female piglets (20-25 kg) were used . METHODS: Six piglets received either 1.5 g/kg bw or 2.5 g/kg bw of ethanol intravenously . In seven piglets, 0.7 g or 1.75 g of ethanol/kg bw was administered intravenously, followed by a subsequent intragastric ethanol infusion of 1.8 g/kg bw and 4.5 g/kg bw, respectively . The samples of colonic contents for the assessment of ethanol and acetaldehyde concentrations were obtained up to seven hours . In four additional piglets, the intracolonic values of ethanol, acetaldehyde, and acetate were observed for 60 minutes after an intracolonic infusion of acetaldehyde solution . RESULTS: A raised intracolonic, endogenous acetaldehyde concentration (mean (SEM); 36 (9) microM) was found in all piglets before ethanol infusion . After the infusion of ethanol, intracolonic ethanol and acetaldehyde values increased in parallel, reaching the peak values 57 (4) mM of ethanol and 271 (20) microM of acetaldehyde in the group that received the highest dose of ethanol . A positive correlation (r = 0.45; p < 0.001) was found between intracolonic ethanol and acetaldehyde values . Acetaldehyde administered intracolonically was mainly metabolised to acetate but also to ethanol in the colon . CONCLUSIONS: Significant endogenous intracolonic acetaldehyde values can be found in the normal porcine colon . Furthermore, our results suggest the existence of a bacteriocolonic pathway for ethanol oxidation . Increased amounts of acetaldehyde are formed intracolonically from ingested ethanol by this pathway. Curr Opin Rheumatol, 1996 Jul, 8(4), 334 - 40 Reactive arthritis; Toivanen A et al.; During the past year, new information has been obtained regarding the pathogenetic process in reactive arthritis . Characterization of the triggering microbes as well as of their interaction with T cells and other host components has brought us closer to achieving a comprehensive picture of the events leading from infection to reactive arthritis . Efforts are ongoing to define clinically applicable diagnostic criteria . It is becoming apparent that chronic forms of the disease are not uncommon. Curr Opin Rheumatol, 1996 Jul, 8(4), 322 - 6 Septic bone and joint infections; Hedstrom SA; Three main trends are apparent in the study of septic bone and joint infections . First, molecular microbiology has advanced our understanding of how microorganisms find structures in the joints and connective tissue on which to adhere, reproduce, and exert an infectious process . Second, modern diagnostic methods such as DNA hybridization and polymerase chain reaction are facilitating the identification of causative microbes . Finally, unusual and previously unknown causes of infection are being identified in joints and bone, often through a time-consuming search in which modern diagnostic techniques may be of valuable help. Curr Opin Rheumatol, 1996 Jul, 8(4), 309 - 15 Etiopathogenesis and biochemical and immunologic evaluation of spondyloarthropathies; Scofield RH; Spondyloarthropathies are closely related to genetics and certain bacteria . The ability to study these relationships at the molecular level has revealed increasing complexity in the interplay of genetics, microbes, and the diseases . Advances have been made in the genetics at the major histocompatibility complex . The mediators of inflammation in spondyloarthropathies have begun to be elucidated . The role of bacterial pathogenesis in the joints is being better defined and implies that viable organisms are in the joints of persons with reactive arthritis at some point in the illness . Disease-prone transgenic animals have been developed that demonstrate the need for an environmental trigger . The nature of the interaction of HLA-B27 with the microbes associated with the disease is