Coupling of lateral and vertical electron motion in GaAs-AlxGa1-xAs quantum wires and dots.
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Journal Article↗
Biomedical subjects
K Keller
Publications and source records attributed to K Keller.
Interaction of LS174T human colon cancer cell mucins with Entamoeba histolytica: an in vitro model for colonic disease.
BACKGROUND & AIMS: Colonic mucins secreted by goblet cells protect the colon by preventing the attachment of enteric pathogens to the epithelium. Entamoeba histolytica overcomes this protective barrier and causes ulcerations, allowing the parasite to disseminate to the liver and form abscesses. An in vitro model is used to study the interaction between E. histolytica and colonic mucins. METHODS: Secretory mucins from the colonic adenocarcinoma cell line LS174T were collected and their functions assessed by their ability to inhibit amebic adherence to target cells and killing. The cytoprotective effect of mucus against E. histolytica cytolysis of LS174T monolayers was studied at 37 degrees C. RESULTS: Sepharose 4B column chromatography, metabolic labeling with [3H]glucosamine, cesium chloride density gradient centrifugation, and amino acid and carbohydrate compositional analysis revealed that LS174T cell mucins were typical of native colonic mucins. Mucin O-linked oligosaccharides bound to and inhibited the adherence of amebae to Chinese hamster ovary cells. E. histolytica killing of Chinese hamster ovary cell monolayers occurred rapidly, whereas killing of LS174T monolayers with an intact mucus layer was significantly retarded. CONCLUSIONS: Our results show that colonic mucins serve as the first line of host defense against amebic invasion and provide a useful model to study pathogen-mucin interactions.
Animals↗
Hyperthermia occurring with paradoxical development of cerebral tuberculomas.
A 59-year-old male Moroccan who had been mechanically ventilated for extensive pulmonary tuberculosis, suddenly developed a generalized seizure with hyperthermia. He had been receiving adequate antituberculosis treatment for 2 weeks. Brain CT scan demonstrated the development of multiple cerebral tuberculomas. The brain CT scan that had been performed at the initiation of antituberculosis therapy was normal. Corticosteroids resulted in rapid improvement of the patient's clinical condition. The pathogenic mechanisms of the development of such lesions as well as their therapy and CT appearance are discussed.
Adrenal Cortex Hormones↗
The tar fraction of cigarette smoke does not promote arteriosclerotic plaque development.
In addition to being the single greatest known environmental cause of cancer, cigarette smoke (CS) is also a major contributor to heart disease. We reported previously that 1) inhalation of either mainstream or sidestream CS promotes aortic arteriosclerotic plaque development; 2) 1,3 butadiene, a vapor-phase component of CS, promotes plaque development at 20 ppm, which at the time was only 2 times higher than the threshold limit value; and 3) individual tar fraction carcinogens in CS, including polynuclear aromatic hydrocarbons (PAHs) and nitrosamines, either do not promote plaque development or do so only at high concentrations. These results suggested that the tar fraction is not the primary source of plaque-promoting agents in CS. We asked whether repeated exposure to the tar fraction of CS, collected in a cold trap (TAR), promotes plaque development in an avian model of arteriosclerosis. Acetone extracts of mainstream CS tar from burning, unfiltered reference cigarettes were solubilized in dimethyl sulfoxide (DMSO) and injected weekly into cockerels for 16 weeks (25 mg/kg/week). Positive controls were injected weekly with the synthetic PAH carcinogen, 7,12 dimethylbenz(a)anthracene (DMBA) dissolved in DMSO and negative controls were injected with DMSO. Plaque location and prevalence did not differ from group to group. Morphometric analysis of plaque cross-sectional areas showed that plaque sizes, which are log-normally distributed, were significantly larger in the DMBA cockerels compared to both the TAR and DMSO groups. There were no significant differences in plaque size between DMSO and TAR cockerels. The results reported here, combined with other recent findings, support the conclusion that the primary arteriosclerotic plaque-promoting components of CS are in the vapor phase.
9,10-Dimethyl-1,2-benzanthracene↗
Women with "squamous atypia".
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Cervix Uteri↗
Identification of the galactose-adherence lectin epitopes of Entamoeba histolytica that stimulate tumor necrosis factor-alpha production by macrophages.
The 170-kDa subunit of the galactose-adherence lectin (Gal-lectin) of Entamoeba histolytica mediates adherence to human colonic mucins and intestinal epithelium as a prerequisite to amebic invasion. The Gal-lectin is an immunodominant molecule and a protective antigen in the gerbil model of amebiasis. Tumor necrosis factor alpha (TNF-alpha) produced by activated macrophages enhances nitric oxide-dependent cytotoxicity in host defense against E. histolytica. The purpose of this study was to identify the Gal-lectin epitopes which stimulate TNF-alpha production by macrophages. Murine bone marrow-derived macrophages (BMMs) exposed to Gal-lectin (100-500 ng/ml) stimulated stable expression of TNF-alpha mRNA (8-fold increase) and TNF-alpha production similar to that of lipopolysaccharide-stimulated cells (100 ng/ml). Polyclonal anti-lectin serum specifically inhibited TNF-alpha mRNA induction in response to the Gal-lectin but not to lipopolysaccharide. Anti-lectin monoclonal antibodies 8C12, H85 and 1G7, which recognize nonoverlapping epitopes of the cysteine-rich region of the 170-kDa heavy subunit, inhibited both amebic adherence to mammalian cells and Gal-lectin-stimulated TNF-alpha mRNA expression by BMMs,but monoclonal antibody 7F4 did neither. As these inhibitory antibodies map to amino acids 596-1082 of the 170-kDa Gal-lectin, our results have identified the functional region that mediates amebic adherence and TNF-alpha mRNA induction in BMMMs; thus, this region of the Gal-lectin is a subunit vaccine candidate.
Animals↗
From triple cysteine mutants to the cysteine-less glucose transporter GLUT1: a functional analysis.
Two triple cysteine mutants containing Cys-less N- or C-terminal halves and the Cys-less GLUT1 were generated by site-directed mutagenesis. Following expression in Xenopus oocytes, the intrinsic transport activities of the multiple cysteine mutants were slightly decreased when either the cysteine residues of the C-terminal half or all six residues were changed; substitution of serine for cysteine residues located at the N-terminal half was without consequence for the catalytic activity. The exofacial ligand ethylidene glucose inhibited 2-deoxy-D-glucose uptake of wild-type and Cys-less GLUT1-expressing Xenopus oocytes with comparable half-saturation constants (11.5 and 13.2 mM). However, each of the multiple cysteine mutants exhibited an increase in affinity for the endofacial inhibitor cytochalasin B, with the greatest effect being observed for the Cys-less construct (decrease in Ki by the factor 5-6).
Animals↗
Proinflammatory cytokines regulate cyclooxygenase-2 mRNA expression in human macrophages.
Prostaglandins play a major role in the inflammatory and immune response. Macrophages constitutively produce prostaglandins by the enzyme cyclooxygenase-1 (cox) whereas inflammatory mediators and cytokines stimulate the inducible enzyme cox-2 for high output prostaglandin production. In this study, we investigated the effect of LPS, IFN-gamma and TNF-alpha in the regulation of cox-1 and cox-2 mRNA expression in PMA-differentiated U937 human macrophages. LPS, but not IFN-gamma or TNF-alpha, caused the induction of cox-2 mRNA in a dose- and time-dependent fashion. In IFN-gamma-primed macrophages, LPS stimulated a marked increase in the accumulation of cox-2 mRNA, whereas TNF-alpha and IFN-gamma induced a moderate level. However, IFN-gamma in combination with either LPS or TNF-alpha induced a synergistic increase in the accumulation of cox-2 mRNA after 24 hours. Regardless of the conditions, cox-1 mRNA remained unchanged. These results indicate that IFN-gamma priming is required for full expression of cox-2 mRNA in response to LPS or TNF-alpha stimulation and suggest that cox-2 mRNA is highly regulated by cytokines during macrophage activation.
Cells, Cultured↗
Functional consequences of proline mutations in the putative transmembrane segments 6 and 10 of the glucose transporter GLUT1.
Proline residues are thought to play a characteristic structural and/or dynamic role in various membrane proteins [Williams, K.A. & Deber, C.M. (1991) Biochemistry 30, 8919-8923]. By use of site-directed mutagenesis and functional expression of mutant glucose transporters in Xenopus oocytes, we investigated the effects of single proline substitutions in the putative helices 6 and 10 on GLUT1-mediated glucose transport. Proline residues of helix 6, that are conserved in all human glucose-transporter isoforms except for the human GLUT2, were mutated either to alanine or to the corresponding residues of GLUT2, i.e. to histidine (P187H), arginine (P196R) or phenylalanine (P205F). In addition, the three proline amino acids within the domain G382-P-G-P-I-P of helix 10 were individually replaced with either alanine or glutamine residues. In all cases, transport function was retained when each individual proline residue was replaced with alanine. Substitution of proline 196 by arginine (P196R), however, resulted in reduction of 2-deoxy-D-glucose uptake rates by approximately 70%. Since the amount of this mutant transporter protein in plasma membrane and total membrane preparations was found to be decreased, as detected by immunoblotting, no single proline residue seemed to play a critical role in maintaining the catalytic activity of GLUT1. However, structural changes introduced by incorporation of the neutral polar amino acid glutamine at each single proline position of helix 10 almost completely abolished 2-deoxy-D-glucose uptake. Thus, the specific chemical structure of the side chain of the substituted amino acid rather than the unique property of proline residues for cis-trans isomerization seemed to determine the consequences on glucose transport.
Animals↗
Estimation of water content and water mobility in the nucleus and cytoplasm of Xenopus laevis oocytes by NMR microscopy.
NMR microscopy is a noninvasive approach for studying cell structure and properties. Spatially resolved measurements of the relaxation times T1 and T2 provided information on the water proton spin density and water mobility in different parts of Xenopus laevis oocytes. The spin-lattice relaxation time T1 was determined using a saturation-recovery sequence and the common spin-echo sequence with increasing repetition times, while the transverse relaxation time T2 was measured by means of the spin-echo sequence with varying echo times. From the relaxation times, the mole fractions of possible reorientational correlation times tau c for different types of intracellular water were calculated according to a simple two-phase model. The values for T1, T2, and proton spin density (i.e., water content) are: nucleus >> animal cytoplasm > vegetal cytoplasm. Based on the estimation of tau c, nearly 90% of the nuclear water and 74.4% of the water of the animal pole was considered as free mobile water, whereas 55.5% of the water of the vegetal pole appeared as bound water.
Animals↗
The relationships between skin hardness, pressure perception and two-point discrimination in the fingertip.
With the recent introduction of computer-assisted sensibility testing devices, it is possible to investigate the hypothesis that the neurological mechanisms responsible for perception of pressure and two-point discrimination are influenced by skin hardness. This study examined the relationships between the skin hardness of the human index and little finger pulp and the perception of pressure in 25 adults. Skin hardness was measured using the Derma-Durometer. Pressure perception was measured with the Pressure-Specified Sensory Device for both static and moving touch stimuli and for two-point discrimination. The mean hardness of the fingertip pulp was 12.5 +/- 0.6 gm/mm2. There was no statistically significant difference in hardness between the dominant and non-dominant, right and left, index and little, or male and female fingertips. Skin hardness was independent of age for this population. The highest correlation, which was r = 0.46, was found between the cutaneous pressure threshold for one-point static touch and skin hardness. While this degree of correlation is significant at the P < 0.02 level, the r2 = 21%, demonstrating that this degree of correlation leaves 79% of the variability unexplained. These results suggest a physical interaction between mechanoreceptors and dermis that is only partially explained by the hardness of the skin.
Adolescent↗
Transforming growth factor-beta 1 primes macrophages for enhanced expression of the nitric oxide synthase gene for nitric oxide-dependent cytotoxicity against Entamoeba histolytica.
Nitric oxide (NO) produced by activated macrophages is the major cytotoxic molecule for in vitro cytotoxicity against Entamoeba histolytica trophozoites. Transforming growth factor-beta 1 (TGF-beta 1) is a potent negative regulator of several macrophage functions, including NO production. In this study, we investigated the effect of TGF-beta 1 on macrophage nitric oxide synthase (mac-NOS) mRNA expression and NO production for macrophage cytotoxicity against E. histolytica trophozoites. TGF-beta 1 by itself was incapable of inducing mouse bone marrow-derived macrophage (BMM) amoebicidal activity and NO production (as measured by nitrite). In contrast, TGF-beta 1 pretreatment (4 hr) primed BMM for an enhanced amoebicidal activity of 15% and 23% in response to (interferon-gamma) IFN-gamma+tumour necrosis factor-alpha (TNF-alpha) or IFN-gamma+lipopolysaccharide LPS, concomitant with increased NO production of 85% and 27%, respectively. TGF-beta 1 pretreatment increased NO production in response to IFN-gamma+TNF-alpha/LPS stimulation in a time- and dose-dependent manner. By Northern blot analysis, the increased NO production of TGF-beta 1-pretreated BMM was preceded by markedly enhanced expression of mac-NOS mRNA. The priming effect of TGF-beta 1 on NO production was critically dependent on both a TNF-alpha (> or = 100 U) and a LPS (> or = 100 ng) triggering dose in the presence of IFN-gamma. TGF-beta 1 pretreatment enhanced TNF-alpha mRNA expression, but had no effect on TNF-alpha production in culture supernatants after 4 hr of stimulation with IFN-gamma+TNF-alpha/LPS; however, at a later time-point (16-48 hr), even though the levels of TNF-alpha mRNA expression were unaffected, TNF-alpha production was reduced. These data demonstrate that TGF-beta 1 priming for increased mac-NOS mRNA expression for NO-dependent cytotoxicity against E. histolytica in response to IFN-gamma+TNF-alpha/LPS stimulation may be involved in the modulation of a TNF-alpha triggering signal by TGF-beta 1.
Animals↗
Entamoeba histolytica stimulates the unstable transcription of c-fos and tumour necrosis factor-alpha mRNA by protein kinase C signal transduction in macrophages.
Macrophages play an important role in the control of and resistance to Entamoeba histolytica (E. histolytica). However, E. histolytica infections are characterized by suppression of cell-mediated immunity. To elucidate the molecular mechanisms whereby amoebae modulate host defences, we investigated whether the parasite elicits the 'immediate early' gene c-fos and cytokine tumour necrosis factor-alpha (TNF)-alpha mRNA and determined the signal transduction pathways involved in naive bone marrow-derived macrophages (BMM). E. histolytica stimulated a rapid and transient expression of c-fos and low levels of TNF-alpha mRNA, whereas the non-pathogenic Entamoeba moshkovshii (E. moshkovskii) did not. Inhibition of the protein kinase C (PKC) pathway with the pharmacological inhibitor H7 and by PKC depletion with phorbol myristate acetate showed that E. histolytica modulates TNF-alpha and c-fos gene expression through a PKC-dependent stimulus-response coupling event. E. histolytica activated and translocated PKC to the membrane fraction in BMM demonstrating a rapid and direct effect on PKC enzyme activity. Unlike lipopolysaccharide (LPS), BMM stimulated with E. histolytica had reduced stability of both c-fos and TNF-alpha mRNA transcripts (> 50%) and failed to secrete TNF-alpha protein. BMM treated with amoebic proteins and stimulated with LPS, or interferon-gamma (IFN-gamma)+LPS, resulted in a 33% and 50% reduction in TNF-alpha mRNA levels, respectively. These data argue that although E. histolytica stimulates c-fos and TNF-alpha gene expression through PKC signal transduction, the rapid degradation of the mRNAs, the lack of secreted TNF-alpha protein and the observed decreased responsiveness to a stimulatory signal may be a novel mechanism whereby the parasite modulates host defence mechanisms.
Animals↗
The role of cysteine residues in glucose-transporter-GLUT1-mediated transport and transport inhibition.
The role of cysteine residues in transport function of the glucose transporter GLUT1 was investigated by a mutagenesis-expression strategy. Each of the six cysteine residues was individually replaced by site-directed mutagenesis. Expression of the heterologous wild-type or mutant glucose transporters and transport measurements at two hexose concentrations (50 microM and 5 mM) were undertaken in Xenopus oocytes. The catalytic activity of GLUT1 was retained, despite substitution of each single cysteine residue, which indicated that no individual residue is essential for hexose transport. This finding questions the involvement of oligomerization or intramolecular stabilization by a single disulphide bond as a prerequisite for transporter activation under basal conditions. Application of the impermeant mercurial thiol-group-reactive reagent p-chloromercuribenzenesulphonate (pCMBS) to the external or internal surface of plasma membrane demonstrated that cysteine-429, within the sixth external loop, and cysteine-207, at the beginning of the large intracellular loop which connects transmembrane segments 6 and 7, are the residues which are involved in transport inhibition by impermeant thiol-group-reactive reagents from either side of the cell. These data support the predicted membrane topology of the transport protein by transport measurements. If residues other than the cysteines at positions 429 or 207 are exposed to either side of the plasma membrane by conformational changes, they do not contribute to the transport inhibition by pCMBS. Application of pCMBS to one side of the plasma membrane also inhibited transport from the opposite direction, most likely due to the hindrance of sugar-induced interconversion of transporter conformation.
4-Chloromercuribenzenesulfonate↗
Type I hypersensitivity to beer.
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Adult↗
Tumor necrosis factor alpha augments nitric oxide-dependent macrophage cytotoxicity against Entamoeba histolytica by enhanced expression of the nitric oxide synthase gene.
Nitric oxide (NO measured as nitrite, NO2-) is the major effector molecule produced by activated macrophages for in vitro cytotoxicity against Entamoeba histolytica trophozoites. In this study, we determine whether tumor necrosis factor alpha (TNF-alpha) produced by activated bone marrow-derived macrophages (BMM) is involved in the induction of the inducible NO synthase gene (mac-NOS) for NO-dependent amebicidal activity. TNF-alpha alone did not directly induce macrophage NO2- production to kill amebae; however, in combination with increasing concentrations of TNF-alpha and gamma interferon (IFN-gamma), BMM amebicidal activity and NO2- production progressively increased and showed a significant linear correlation. Antiserum to TNF-alpha and the NO synthase inhibitor NG-monomethyl L-arginine (L-NMMA) inhibited the synergistic effects of TNF-alpha and IFN-gamma. BMM activated with increasing concentrations of lipopolysaccharide (LPS) and IFN-gamma showed a significant linear correlation between TNF-alpha release and NO2- production. Antiserum to TNF-alpha suppressed TNF-alpha release, NO2- production, and amebicidal activity by 93, 53, and 86%, respectively. L-NMMA diminished NO2- production by 74% and macrophage amebicidal activity by 83% but had no effect on TNF-alpha release. Quantification by Northern (RNA) blot analyses demonstrated that IFN-gamma in combination with TNF-alpha or LPS increased markedly the accumulation of mac-NOS and TNF-alpha mRNAs in a time-dependent manner with a concomitant increase in NO and TNF-alpha production. Peak induction of mac-NOS occurred after 24 h, whereas TNF-alpha mRNA was rapidly expressed after 4 h and remained stable for 48 h. Taken together, these data argue that TNF-alpha augments NO-dependent macrophage cytotoxicity against E. histolytica via elevated levels of mac-NOS mRNA expression which may be associated with the accumulation of TNF-alpha mRNA.
Amino Acid Oxidoreductases↗
Acute asthma management in children: factors identifying patients at risk for intensive care unit treatment.
The hospitalization and mortality rates incurred from childhood asthma continue to rise despite recent progress into the pathophysiology and treatment of reactive airway disease. We believe that there are specific factors that identify children at risk for death from asthma. The objective of the study was to determine those risk factors that identify children at increased risk for nonfatal, but life-threatening asthma exacerbations resulting in pediatric intensive care unit (PICU) admission. Patients aged 10 months-16 years admitted to Brenner Children's Hospital for status asthmaticus between April 1991 and December 1992 were evaluated with regard to the prevalence of eight different factors using two categories for asthma: (1) life-threatening asthma requiring PICU admission and (2) non-life-threatening asthma requiring routine hospitalization. Risk factors associated with an increased prevalence of non-fatal, but life-threatening asthma requiring PICU sensitivity state. The classification employed here seemed to provide some merits in delineating the features of adulthood asthma.
Acute Disease↗
Entamoeba histolytica modulates the nitric oxide synthase gene and nitric oxide production by macrophages for cytotoxicity against amoebae and tumour cells.
Nitric oxide (NO) is the major cytotoxic molecule produced by activated macrophages for cytotoxicity against Entamoeba histolytica trophozoites. In the present study, we determined whether E. histolytica infection and soluble amoebic proteins affected macrophage cytotoxicity against amoebae and tumour cells by modulating the inducible NO synthase gene (iNOS) and NO (measured as nitrite, NO2-) and tumour necrosis factor-alpha (TNF-alpha) production. Amoebic liver abscess-derived macrophages [days 10, 20, 30 post-infection (p.i.)] stimulated with interferon-gamma (IFN-gamma) and lipopolysaccharide (LPS) showed increased cytotoxicity against L929 cells (TNF-alpha-sensitive), but were refractory for killing amoebae and P815 cells (both NO-sensitive), concomitant with low NO2- production (< 4 microM/10(6) cells). In contrast, peritoneal and spleen macrophages at 10 and 20 days p.i. activated with IFN-gamma and LPS demonstrated increased killing of amoebae, and L929 and P815 cells concomitant with high NO2- production (> 12 microM/10(6) cells). Pretreatment of mouse bone marrow-derived macrophages with amoebic proteins suppressed IFN-gamma and LPS-induced amoebicidal (33%) and tumoricidal (44-49%) activities, with a corresponding decrease in TNF-alpha (56%) and NO (41%) production as well as TNF-alpha (41%) and iNOS (27%) mRNA by Northern blot analyses as compared to untreated activated controls. Inhibition of prostaglandin E2 (PGE2) biosynthesis in abscess and naive macrophages pretreated with amoebic proteins augmented IFN-gamma- and LPS-induced killing of L929 cells and TNF-alpha production, but failed to increase killing of P815 cells and amoebae as well as iNOS mRNA levels or NO production. These results suggest that E. histolytica selectively induces dysfunction of macrophage cytotoxicity by modulating iNOS mRNA expression and NO production independent from TNF-alpha and PGE2 allowing the parasites to survive within the host by impairing host immune responses.
Amino Acid Oxidoreductases↗