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Biomedical subjects

H Chan

Publications and source records attributed to H Chan.

At least 73 records · Page 4Linked to original sources
PubMed1996

Effect of application of benzyl benzoate on house dust mite allergen levels.

BACKGROUND: Several acaricides have become available for reducing house dust mite allergen levels. OBJECTIVE: The purpose of this study was to assess whether the use of benzyl benzoate (Acarosan) provides additional benefit to the usual mite control measures including encasement of mattress and pillows with vinyl covers. METHODS: A randomized controlled trial was carried out in 26 homes (14 control versus 12 treatment) of asthmatic patients in two cities (Vancouver and Winnipeg). The control group had the usual house dust mite control measures including the use of vinyl covers for mattresses and pillows while the treatment group had application of benzyl benzoate to mattresses and carpets in the bedroom and the most commonly used room, in addition to the above control measures. Mite allergen levels were measured 3 months and immediately before, 1 week, and 1 and 3 months after the application of house dust mite control measures. Patients kept diary cards on asthma symptoms and peak expiratory flow rates morning and evening one month before and three months after the onset of mite allergen control measures. RESULTS: A reduction of mite allergen level was found in mattress samples in both groups, statistically significant at all times in the treatment group and at one and three months in the control group. Mite allergen levels on floor carpets also showed progressive reduction in both groups, but were significantly different in the treatment group (compared with controls) at 1 week, and were lower compared with baseline in the treatment group up to 3 months. No significant changes in asthma symptoms, peak expiratory flow rates, spirometric measurements, or bronchial hyperresponsiveness were observed among treatment or control group subjects. CONCLUSION: The addition of benzyl benzoate to conventional house dust mite control measures resulted in a significant reduction in floor carpet dust mite levels that persisted for 3 months. The results of this study should be confirmed in a larger and longer study.

Adult↗
PubMed1996

Occupational asthma due to latex in health care workers.

Immediate hypersensitivity reactions ranging from mild urticaria to life threatening anaphylaxis after exposure to natural rubber latex have been reported frequently in health care workers while occupational asthma due to latex exposure is less well studied. The results of specific challenge tests and immunological tests in four health care workers with work related respiratory and skin disorders induced by the use of latex gloves are described. Occupational asthma was confirmed in three subjects by specific challenge tests. All had a positive skin test reaction to the latex extract; specific IgE antibodies were detected in only one subject. The fourth subject had a negative specific inhalation and skin test reaction to the latex extract. Peak expiratory flow monitoring at work and away from work showed a pattern consistent with work related asthma. These findings confirm that latex is a cause of occupational asthma in health care workers.

Adult↗
PubMed1996

Autoimmune polyglandular syndrome and primary biliary cirrhosis.

Liver involvement in autoimmune polyglandular syndrome (APS) in the form of chronic active hepatitis has been well described. However, to our knowledge, primary biliary cirrhosis in APS has not been reported. Here we report the case of a 27-year-old man who presented with classical insulin-dependent diabetes mellitus and subsequently developed Hashimoto's thyroiditis, hypogonadism, and primary biliary cirrhosis. The latter diagnosis was confirmed by a cholestatic pattern of liver enzymes, positive anti-mitochondrial antibody, normal cholangiogram, and characteristic liver biopsy findings. Primary biliary cirrhosis should probably be regarded as a possible, though uncommon, component of APS.

Adult↗
PubMed1995

Protease inhibitors differentially regulate tumor necrosis factor-induced apoptosis, nuclear factor-kappa B activation, cytotoxicity, and differentiation.

We investigated the effect of various protease inhibitors on several tumor necrosis factor (TNF)-mediated cellular responses. Treatment of a human myelogenous leukemia cell line, ML-1a, with TNF in the presence of cycloheximide triggers endonucleolytic activity and apoptotic cell death within 90 minutes. The general serine protease inhibitor diisopropyl fluorophosphate (DFP) and the chymotrypsin-like protease inhibitor N-tosyl-L-lysyl chloromethyl ketone (TPCK) completely abrogated TNF-induced DNA fragmentation and the formation of apoptotic bodies. However, 13 other protease inhibitors, including serine protease inhibitors, did not. The addition of TPCK to cells 30 minutes after TNF treatment completely inhibited the cytokine action, indicating that TPCK-sensitive proteases are not involved in the early stages of signal transduction. TNF is cytotoxic and induces differentiation in ML-1a cells after a 3-day incubation. TPCK had no effect on the TNF-induced cytotoxicity and differentiation, indicating that TPCK-sensitive proteases are specific for DNA fragmentation. TPCK also blocked TNF-induced activation of nuclear factor (NF)-kappa B. The dose-response and the time-course of the inhibitor, however, indicated that the site of action of TPCK for NF-kappa B activation and for DNA fragmentation are quite distinct. Therefore, we conclude that TNF activates two distinct TPCK-sensitive pathways, one leading to apoptosis and the other to NF-kappa B activation.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1995

Desmosome assembly and disassembly are regulated by reversible protein phosphorylation in cultured epithelial cells.

Desmosomes are one component of the intercellular junctional complex in epithelia. In cultures of epithelial cells, desmosome assembly can be regulated by modulating the calcium concentrations of the growth media. At present, very little is known about the intracellular signal transduction mechanisms that regulate desmosome assembly and disassembly in response to changing extracellular calcium concentrations. We have used inhibitors of protein kinases and phosphatases in a combined biochemical and morphological approach to analyze the role of protein phosphorylation in the assembly and disassembly of desmosomes in Madin-Darby canine kidney epithelial cells. Our results suggest that desmosomal proteins (desmoplakins I/II and desmoglein 1) are primarily phosphorylated on serine residues. Electron microscopic analyses of desmosome assembly upon induction of cell-cell contact, in the presence of protein kinase inhibitor, H-7, revealed an apparently normal assembly of desmosomes. However, complete disassembly of desmosomes was inhibited by H-7 upon removal of extracellular calcium. Under these conditions, although desmosomes split, desmosomal plaques and their associated cytokeratin filaments can not be internalized. In contrast, treatment of the cultures with okadaic acid (OA), an inhibitor of protein phosphatases, inhibited desmosome assembly but had no effect on disassembly. In addition, the inhibitory effect of okadaic acid on desmosome assembly was specific to this junction since we observed apparently normal tight junction and adherens junction in okadaic acid-treated cultures. These results suggest that assembly and disassembly of desmosomes may be regulated by extracellular Ca2+ via reversible protein phosphorylation involving both protein kinase and protein phosphatases.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1995

High-level production from a baculovirus expression system and biochemical characterization of human GMP synthetase.

GMP synthetase, a key enzyme in the de novo synthesis of guanine nucleotides, is a potential target for immunosuppression and anticancer chemotherapy. In order to closely examine the catalytic mechanism and active-site topography of this enzyme, large amounts of pure protein are needed. Catalytically active human GMP synthetase was expressed in a baculovirus system. A high-level production system has been established from which the yield of pure protein is routinely more than 50 mg/10 liters of cell culture. The recombinant enzyme was purified to homogeneity and characterized. Like native GMP synthetase, the recombinant enzyme was resolved into two forms by ion-exchange chromatography. The two forms are both monomers and they differ in their isoelectric points. There is no evidence that these forms are in equilibrium or interconvertible. Protein sequence analysis reveals that both forms are blocked at the amino-terminus and they are essentially identical in sequence. Since they can be produced by a cDNA with a single open reading frame, we believe that they represent post-translational modification variants. The recombinant GMP synthetase is not distinguishable from the native enzyme in terms of chromatographic profiles, subunit composition, molecular weight, and kinetic properties. The inhibition constants and the modes of inhibition toward decoyinine, a selective inhibitor of GMP synthetase, are also the same as the native enzyme. The high-level production of active enzyme is invaluable to the determination of the three-dimensional structure and the discovery of potent and selective drug candidates.

Animals↗
PubMed1995

Changes in fast axonal transport in sensory neurons during tadpole metamorphosis.

Fast axonal transport of radiolabeled protein was examined in lumbar and tail dorsal root ganglion (DRG) neurons at progressive stages of bullfrog tadpole metamorphosis. Accumulation of [35S]methionine-labeled protein proximal to a lumbar peripheral nerve ligature (at a fixed distance from the DRG) increased as tadpoles advanced from premetamorphosis through prometamorphosis to metamorphic climax. The rate of increase was steeper when expressed as a percentage of protein synthesized in the neurons of origin than when expressed as a percentage of total DRG protein synthesis. Further, the increase was not secondary to a rise in protein synthesis. In contrast, fast axonal transport decreased in DRG neurons of the tail at the onset of metamorphic climax, when tail resorption is initiated. The stage-related increase in protein transport in lumbar nerves is due, at least in part, to an increased rate of transport. As determined from optically detected anterograde organelles in individual lumbar nerve axons, an approximate doubling of the fast transport rate occurred between the premetamorphic stage and metamorphic climax. In addition, the rates of organelle transport in lumbar axons of adult bullfrogs were significantly greater than in corresponding axons of tadpoles at metamorphic climax, further suggesting that organelle velocity is a developmentally regulated parameter of fast axonal transport.

Animals↗
PubMed1995

Bronchial inflammation in occupational asthma due to western red cedar.

Bronchoalveolar lavage cells and bronchial biopsies were obtained from nine patients with red cedar asthma, six atopic asthmatics and six non-atopic, non-asthmatic control subjects. There were similar proportions of neutrophils, mast cells, lymphocytes, and macrophages in BAL samples from all three groups, but eosinophil numbers were elevated in patients with cedar asthma and atopic asthma (3.0 and 2.5% respectively versus 0.5% in control subjects; p < 0.05 for each group). In bronchial mucosal biopsies, mean numbers of T cells were elevated in both asthmatic groups (cedar asthma 9.8 times, and atopic asthma 2.6 times, control values). CD4+ cells accounted for most of the increase in T-cell numbers, while CD8+ cell numbers were elevated in biopsies from a minority of cedar asthma patients. Absolute numbers of CD25+ (IL-2 receptor-bearing) cells were increased in cedar asthma but the proportion of T cells expressing CD25, was similar in all three groups. Activated eosinophils (EG2+) were increased in both asthmatic groups, with mean numbers 2.5 times greater in the cedar asthma biopsies than in atopic asthmatics. Thus both cedar asthma and atopic asthma are associated with increased numbers of T-cells and activated eosinophils in the bronchial mucosa. There was no major histologic difference between atopic asthma and red cedar asthma.

Adult↗
PubMed1995

Major cat allergen (Fel d I) levels in the homes of patients with asthma and their relationship to sensitization to cat dander.

BACKGROUND: Exposure and sensitization to the major cat allergen, Felis domesticus allergen I (Fel d I), significant causes of allergic respiratory disease. Many patients who are allergic to cats, however, do not own a cat and there is not an obvious source of allergen in their home environment. OBJECTIVE: We investigated the levels of Fel d I in dust from homes of 120 subjects with asthma in two climatologically diverse Canadian cities (Vancouver and Winnipeg). Fel d I levels were related to exposure to cats as well as to skin reactivity to cat dander. METHODS: Dust samples from bedroom floors and mattresses were collected in four different seasons and Fel d I content was determined by 2-site monoclonal antibody based-ELISA. RESULTS: Although only 18 patients (15%) were cat owners, detectable levels of Fel d I were found on at least one occasion in all homes. The geometric mean concentration of Fel d I on floors was 1.15 micrograms/g of dust (range 0.07 to 26.3 micrograms/g) and on mattresses 0.89 micrograms/g (range 0.01 to 17.4 micrograms/g). Seasonal variation of Fel d I levels was only observed in Winnipeg, where Fel d I concentrations were highest in the winter and spring compared with either summer (P < .05) or autumn (P < .005). The highest Fel d I levels were found in homes with a cat (P < .05), however, rather high levels were also found in homes of patients who did not have a cat but visited others with cats. Cat dander was the most frequent sensitizer (60%) in these patients but no correlation was found between the size of the wheal induced by cat dander extract and Fel d I levels in dust samples. CONCLUSION: Cat allergen was universally found in homes of asthmatic patients and this may explain the high frequency of cat sensitization among patients with asthma in these two cities. A seasonal variation in cat allergen was observed in Winnipeg with no variation in Vancouver.

Adolescent↗
PubMed1994

Role of tumor necrosis factor receptors in the activation of nuclear factor kappa B in human histiocytic lymphoma U-937 cells.

Tumor necrosis factor (TNF) has been shown to mediate numerous cellular responses through its interaction with two distinct types of receptor. However, the relationship between TNF receptor and the biological response is not well understood. Modulation of the number of cell surface receptors by various agents has shown a lack of direct correlation with biological responses to the cytokine. In this report, we used several approaches to investigate the relationship between TNF receptor number and an early response in human histiocytic lymphoma U-937 cells. When we examined the activation of the nuclear transcription factor kappa B (NF-kappa B), an event mediated by TNF within 10-15 min, we discovered a correlation between TNF receptor occupancy up to a certain threshold and the extent of activation of the transcription factor. In addition, by kinetically down-regulating TNF receptor expression with phorbol esters, cycloheximide, or trypsin, we determined that receptors were necessary for transduction of the TNF signal. However, 10-25% of total receptors were sufficient for optimum induction of the NF-kappa B signal. When examined in different cell lines, the activation of an early biological response was found to be related not only to the TNF receptor number but also to the type of TNF receptor. These results, overall, suggest that although TNF receptors are essential for induction of NF-kappa B, a small percentage is sufficient to fully transduce this signal.

Cycloheximide↗
PubMed1994

Reconstitution of nuclear factor kappa B activation induced by tumor necrosis factor requires membrane-associated components. Comparison with pathway activated by ceramide.

Tumor necrosis factor (TNF) is known to induce the activation of a nuclear transcription factor, nuclear factor kappa B (NF-kappa B), in a wide variety of cell types. The post-receptor binding events that culminate in TNF-dependent NF-kappa B activation are not understood. To dissect this pathway, we developed a reconstitution system consisting of membrane, cytosolic, and post-nuclear fractions. Our results indicate that when incubated with the post-nuclear fraction derived from TNF-untreated cells, the membrane fraction from TNF-treated cells causes the activation of NF-kappa B with kinetics similar to that observed in intact cells. Under these conditions, the cytosolic fraction has no effect. This activation is tyrosine kinase-dependent since erbstatin completely abolished the effect. Furthermore, as revealed by immunoblotting, no degradation of the inhibitory subunit of NF-kappa B was observed. In this reconstitution system, we can also demonstrate the activation of NF-kappa B by ceramide, but this activation is not tyrosine kinase-dependent. Overall, our results indicate that intermediates required for NF-kappa B activation by TNF or ceramide are membrane-bound, but the mechanism of activation by TNF is most likely different from that of ceramide.

Cell Line↗
PubMed1994

Inhibition of endothelial cell adhesion molecule expression with antisense oligonucleotides.

In response to inflammatory stimuli, expression of a group of proteins that bind circulating leukocytes (endothelial-leukocyte adhesion molecules) are induced on the luminal surface of vascular endothelium. A series of phosphorothioate oligonucleotides 18 to 21 bases in length were designed and synthesized to hybridize selectively to the mRNA, which encodes three such endothelial-leukocyte adhesion molecules; human intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and E-selectin. Antisense oligonucleotides were identified that selectively inhibited ICAM-1, VCAM-1, and E-selectin expression in HUVEC. Oligonucleotides that hybridized to the 3'-untranslated region of either ICAM-1, VCAM-1, or E-selectin mRNAs promoted a selective reduction in the respective mRNA levels. In contrast, oligonucleotides that hybridized to 5'-untranslated sequences did not significantly reduce target mRNA levels, although they did promote a reduction in protein expression. With the use of flow cytometry to measure cell surface expression, ICAM-1 and E-selectin were selectively inhibited by their respective antisense oligonucleotide. At low concentrations of oligonucleotides, only VCAM-1 antisense oligonucleotides inhibited VCAM-1 expression. However, at an oligonucleotide concentration of 50 nM or greater, phosphorothioate oligonucleotides not predicted to hybridize to VCAM-1 mRNA also reduced VCAM-1 expression. The sequence-independent inhibition of VCAM-1 expression by phosphorothioate oligonucleotides could be the result of a perturbation in the transcriptional regulation of the VCAM-1 gene. ICAM-1, VCAM-1, and E-selectin antisense oligonucleotides reduced adhesion of HL-60 cells to TNF-activated HUVEC. These data demonstrate that phosphorothioate oligonucleotides are capable of selectively inhibiting the expression of ICAM-1, VCAM-1, and E-selectin in HUVEC.

Base Sequence↗
PubMed1994

Concurrent RhGM-CSF does not offset myelosuppression from intensive chemotherapy: randomized placebo-controlled study in childhood acute lymphoblastic leukemia.

To determine whether recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) can offset the myelosuppressive effects of intensive chemotherapy, we carried out a double-blind placebo-controlled trial in which 40 patients with acute lymphoblastic leukemia (ALL) were randomized into two groups of 20 each. One group received rhGM-CSF (5.5 micrograms/kg SC) coadministered with chemotherapy and the other, placebo coadministered with chemotherapy from day 5 to day 11 and from day 19 to day 25 of the 28-day intensification phase of our institutional high-risk protocol for childhood ALL. The results indicate that, at the dose and schedule used, rhGM-CSF did not prevent neutropenia or shorten the number of days required to complete this phase of therapy. In addition, the treated and placebo groups showed no significant difference in absolute neutrophil counts, number of days with neutropenia, number of days with fever, number of days spent in hospital, or number of days on antibiotics during the 28-day study period. There was also no difference between the two groups in the number, type, or severity of infectious episodes. Two of 20 patients in the treatment group have relapsed, whereas none of the patients in the placebo group has yet relapsed (follow-up: 3-37 months), but these events were not statistically significant. We conclude that treatment with rhGM-CSF at the dose and schedule employed is not clinically beneficial.

Adolescent↗
PubMed1994

Disruption of fast axonal transport in vivo leads to alterations in Schwann cell gene expression.

Following nerve injury, Schwann cells distal to the site of injury down-regulate genes associated with myelination. We hypothesized that at least some of these alterations were due to the loss of ongoing axon:Schwann cell homeostatic signals, as opposed to loss of physical contact and/or inflammatory responses. To directly test this hypothesis, we perturbed axonal physiology by selectively blocking fast axonal transport via locally cooling the sciatic nerve to 5-8 degrees C (a cold block). Immunostaining with the monoclonal antibody ED1, which recognizes mononuclear phagocytic cells, demonstrated that macrophages did not invade the cold-blocked nerve, indicating the lack of an inflammatory response. Morphological studies demonstrated that the nerve distal to the cold block showed no signs of Wallerian degeneration, with maintenance of normal axon and myelin profiles, and confirmed the absence of invading macrophages. Thus, any effects of a cold-block treatment were not likely due to inflammatory responses or to loss of physical contact between axons and Schwann cells. To determine whether this treatment affected Schwann cell phenotype, we examined expression of the major myelin protein P0, and p75 NGF receptor, both of which are regulated as a function of axon:Schwann cell interactions. Levels of p75 NGF receptor mRNA were unaffected by the cold block, while p75 NGF receptor protein levels were increased in the region of the nerve immediately adjacent to the cold block, presumably reflecting protein accumulation as a consequence of the block to fast axonal transport. In contrast, levels of P0 mRNA were decreased in the distal nerve in a fashion that indicated modulation of Schwann cell phenotype as a function of local axonal microenvironment. These data therefore suggest that P0 and p75 NGF receptor are regulated as a function of two different aspects of Schwann cell:axon communication. Furthermore, these data demonstrate that the presence of axon:Schwann cell contact alone is insufficient to maintain Po gene expression and indicate that at least some myelin-specific Schwann cell responses are dependent upon ongoing biochemical signals generated by the axon and maintained by fast axonal transport.

Animals↗
PubMed1994

Genetic analyses of temperature-sensitive mutations in baculovirus late expression factors.

Two temperature-sensitive mutants of Autographa californica nuclear polyhedrosis virus were characterized with respect to late and very late gene expression. Cells infected with ts317 or ts538 at the nonpermissive temperature revealed slightly reduced intracellular levels of late structural gene products including gp64 envelope, vp39 capsid, and p6.9 DNA binding proteins. However, with the exception of gp64, there was a dramatic reduction in extracellular levels of the structural proteins, suggesting that assembly or transport of extracellular virions was impaired. Electron microscopy confirmed the dramatic effect the mutations exhibited on viral morphogenesis. The ts538 mutation was physically mapped to the 60-65 mu region and DNA sequence analysis identified a single nucleotide alteration converting Leu to Phe in amino acid 105 of the late expression factor called lef-4. The mutation was positively correlated with the temperature-sensitive phenotype by analysis of DNA from ts+ revertants generated by marker rescue. Transient expression assays confirmed that lef-4 was required for activation of the late vp39 gene in virus infected cells. The defect in activation of the ts317 and ts538 polyhedrin genes could be overcome by coinfection with helper virus indicating that the mutations affected functions involved in activating the very late class of baculovirus genes as well. These results demonstrated a clear functional differentiation between genes regulating viral DNA replication which was normal in the mutant infected cells and those regulating late gene expression and extracellular virus production which were defective in these same infected cells.

Animals↗
PubMed1994

Involvement of immunologic mechanisms in a guinea pig model of western red cedar asthma.

Western red cedar asthma is the most common form of occupational asthma in the Pacific Northwest. Plicatic acid (PA) is the chemical component of Western red cedar that causes asthma. The role of immunologic processes involved in the PA-induced asthmatic reaction has not been established. To characterize the mechanisms of PA-induced asthmatic reaction, guinea pigs were sensitized to PA through biweekly injection of PA-ovalbumin conjugate with aluminum hydroxide as an adjuvant for a period of 6 months. Specific IgG1 antibodies to PA were detected in the blood 3 months after sensitization of animals. The level of specific IgG1 antibodies to ovalbumin after 6 months was about two times the level of specific IgG1 to PA. At 6 months, tracheal tissue from PA-ovalbumin-sensitized guinea pigs contracted after exposure to either PA or ovalbumin in vitro. The degree of contraction induced by PA was two to three times less than the contraction induced by ovalbumin. PA caused histamine, prostaglandin D2, and leukotriene D4 release from both lung mast cells and blood basophils. The amount of histamine and eicosanoids released by PA was also two to three times less than the amount of mediators released by ovalbumin. When the trachea of normal guinea pigs was passively sensitized with serum from PA-ovalbumin-sensitized guinea pigs, it contracted in response to PA or ovalbumin in an organ bath. When the serum of PA-ovalbumin-sensitized guinea pigs was depleted of immunoglobulins and then used for passive sensitization of normal trachea, no contraction was observed when challenged with PA, suggesting that IgG1 antibodies mediate the tracheal reaction to PA.(ABSTRACT TRUNCATED AT 250 WORDS)

Allergens↗
PubMed1994

Action of brefeldin A on amphibian neurons: passage of newly synthesized proteins through the Golgi complex is not required for continued fast organelle transport in axons.

The relation between the availability of newly synthesized protein and lipid and the axonal transport of optically detectable organelles was examined in peripheral nerve preparations of amphibia (Rana catesbeiana and Xenopus laevis) in which intracellular traffic from the endoplasmic reticulum to the Golgi complex was inhibited with brefeldin A (BFA). Accumulation of fast-transported radio-labeled protein or phospholipid proximal to a sciatic nerve ligature was monitored in vitro in preparations of dorsal root ganglia and sciatic nerve. Organelle transport was examined by computer-enhanced video microscopy of single myelinated axons. BFA reduced the amount of radiolabeled protein and lipid entering the fast-transport system of the axon without affecting either the synthesis or the transport rate of these molecules. The time course of the effect of BFA on axonal transport is consistent with an action at an early step in the intrasomal pathway, and with its action being related to the observed rapid (< 1 h) disassembly of the Golgi complex. At a concentration of BFA that reduced fast-transported protein by > 95%, no effect was observed on the flux or velocity of anterograde or retrograde organelle transport in axons for at least 20 h. Bidirectional axonal transport of organelles was similarly unaffected following suppression of protein synthesis by > 99%. The findings suggest that the anterograde flux of transport organelles is not critically dependent on a supply of newly synthesized membrane precursors. The possibilities are considered that anterograde organelles normally arise from membrane components supplied from a post-Golgi storage pool, as well as from recycled retrograde organelles.

Animals↗