Biomedical subjects

S Lei

Publications and source records attributed to S Lei.

At least 19 recordsLinked to original sources

PubMed2002

Induction of apoptosis by norcantharidin in human colorectal carcinoma cell lines: involvement of the CD95 receptor/ligand.

PURPOSE: Cantharidin, a natural toxin, is the active substance of mylabris and has antitumor effects in man. Norcantharidin, the demethylated analogue of cantharidin, has been used in the treatment of patients with primary hepatoma and those with leukopenia in China. The present study was designed to investigate whether norcantharidin exerts cytotoxic activity against colorectal cancer cells by inducing apoptosis and to examine the possible mechanism in the phenomenon. METHODS: Inhibition of proliferation of norcantharidin on Colo205, HT-29, and SW480 colorectal cancer cells was determined by the trypan blue dye exclusion test. Apoptosis of norcantharidin-treated cells was determined by morphological analysis, agarose gel DNA electrophoresis, and quantitated by flow cytometry after staining with propidium iodide. Cell cycle and the cell surface expression of the CD95/CD95 ligand were evaluated by flow cytometry. Caspase 8-like protease and protein phosphatase 1 and 2A activities were also analyzed. RESULTS: Treatment with norcantharidin of colorectal cancer cells not only inhibited cell proliferation, but also induced apoptosis. Norcantharidin induced apoptosis mainly in two phases: rapid apoptosis in S-phase cells and delayed apoptosis in G2/M arrested cells. Treatment with norcantharidin resulted in an upregulation of the CD95 receptor and CD95 ligand on the cell surface. Furthermore, stimulation with anti-CD95 monoclonal antibody (mAb) resulted in further induction of apoptosis after treatment with norcantharidin. In addition, the apoptosis-inducing effect of norcantharidin was almost completely inhibited by anti-CD95 ligand mAb. Norcantharidin-treated cells showed the activation of caspase 8. Both zVAD-FMK (a broad range caspase inhibitor) and IETD-FMK (a caspase-8 inhibitor) showed apparent inhibition of the apoptosis-inducing effect. Norcantharidin did not show an inhibitory effect on protein phosphatase. CONCLUSIONS: These results suggest that norcantharidin triggers apoptosis in colorectal cancer cell lines via the activation of the CD95 receptor/ligand system, and that this agent may be useful for developing new therapeutic regimens for the treatment of colorectal carcinoma.

Antibodies, Monoclonal↗

PubMed2002

A gene therapy for cancer based on the angiogenesis inhibitor, vasostatin.

The growth and persistence of solid tumors and their metastasis are angiogenesis-dependent. Vasostatin, the N-terminal domain of calreticulin inclusive of amino acids 1-180, is a potent angiogenesis inhibitor. To investigate whether intramuscular administration of vasostatin gene has the antitumor activity in mouse tumor models, we constructed a plasmid DNA encoding vasostatin and a control vector. Production and secretion of vasostatin protein by COS cells transfected with the plasmid DNA encoding vasostatin (pSecTag2B-vaso) were confirmed by Western blot analysis and ELISA. Conditioned medium from vasostatin-transfected COS cells apparently inhibited human umbilical vein endothelial cell (HUVEC) and mouse endothelial cell (SVEC4-10) proliferation, compared with conditioned medium from the COS cells transfected with control vector or non-transfected cells. Treatment with pSecTag2B-vaso twice weekly for 4 weeks resulted in the inhibition of tumor growth and the prolongation of the survival of tumor-bearing mice. The sustained high level of vasostatin protein in serum could be identified in ELISA. Angiogenesis was apparently inhibited in tumor by immunohistochemical analysis. Angiogenesis was also inhibited in the chicken embryo CAM assay and mouse corneal micropocket assay. The increased apoptotic cells were found within the tumor tissues from the mice treated with plasmid DNA encoding vasostatin. Taken together, the data in the present study indicate that the cancer gene therapy by the intramuscular delivery of plasmid DNA encoding vasostatin, is effective in the inhibition of the systemic angiogenesis and tumor growth in murine models. The present findings also provide further evidence of the anti-tumor effects of the vasostatin, and may be of importance for the further exploration of the application of this molecule in the treatment of cancer.

Angiogenesis Inhibitors↗

PubMed2001

Regulation of NMDA receptor activity by F-actin and myosin light chain kinase.

The postsynaptic density (PSD) at excitatory dendritic synapses comprises a protein complex of glutamate receptors, scaffolding elements, and signaling enzymes. For example, NMDA receptors (NMDARs) are linked to several proteins in the PSD, such as PSD-95, and are also tethered via binding proteins such as alpha-actinin directly to filamentous actin of the cytoskeleton. Depolymerization of the cytoskeleton modulates the activity of NMDARs, and, in turn, strong activation of NMDARs can trigger depolymerization of actin. Myosin, the motor protein of muscular contraction and nonmuscle motility, is also associated with NMDARs and the PSD. We show here that constitutively active myosin light chain kinase (MLCK) enhances NMDAR-mediated whole-cell and synaptic currents in acutely isolated CA1 pyramidal and cultured hippocampal neurons, whereas inhibitors of MLCK depress these currents. This MLCK-dependent regulation was observed in cell-attached patches but was lost after excision to inside-out patches. Furthermore, the enhancement induced by constitutively active MLCK and the depression of MLCK inhibitors were eliminated after depolymerization of the cytoskeleton. NMDARs and MLCK did not colocalize in clusters on the dendrites of cultured hippocampal neurons, further indicating that the effects of MLCK are mediated indirectly via actomyosin. Our results suggest that MLCK enhances actomyosin contractility to either increase the membrane tension on NMDARs or to alter physical relationships between the actin cytoskeleton and the linker proteins of NMDARs.

Actins↗

PubMed2001

Immunogene therapy of tumors with vaccine based on Xenopus homologous vascular endothelial growth factor as a model antigen.

Overcoming immune tolerance of the growth factors associated with tumor growth should be a useful approach to cancer therapy by active immunity. We used vascular endothelial growth factor (VEGF) as a model antigen to explore the feasibility of the immunogene tumor therapy with a vaccine based on a single xenogeneic homologous gene, targeting the growth factors associated with angiogenesis. To test this concept, we constructed a plasmid DNA encoding Xenopus homologous VEGF (XVEGF-p) and control vectors. We found that immunogene tumor therapy with a vaccine based on XVEGF was effective at both protective and therapeutic antitumor immunity in several tumor models in mice. VEGF-specific autoantibodies in sera of mice immunized with XVEGF-p could be found in Western blotting analysis and ELISA assay. The purified immunoglobulins were effective at the inhibition of VEGF-mediated endothelial cell proliferation in vitro, and at antitumor activity and the inhibition of angiogenesis by adoptive transfer in vivo. The elevation of VEGF in the sera of the tumor-bearing mice could be abrogated with XVEGF-p immunization. The antitumor activity and production of VEGF-specific autoantibodies, significantly elevated IgG1 and IgG2b, could be abrogated by the depletion of CD4(+) T lymphocytes. The observations may provide a vaccine strategy for cancer therapy through the induction of autoimmunity against the growth factors associated with tumor growth in a cross reaction with single xenogeneic homologous gene and may be of importance in the further exploration of the applications of other xenogeneic homologous genes identified in human and other animal genome sequence projects in cancer therapy.

Animals↗

PubMed2001

Differential synaptic processing separates stationary from transient inputs to the auditory cortex.

Sound features are blended together en route to the central nervous system before being discriminated for further processing by the cortical synaptic network. The mechanisms underlying this synaptic processing, however, are largely unexplored. Intracortical processing of the auditory signal was investigated by simultaneously recording from pairs of connected principal neurons in layer II/III in slices from A1 auditory cortex. Physiological patterns of stimulation in the presynaptic cell revealed two populations of postsynaptic events that differed in mean amplitude, failure rate, kinetics and short-term plasticity. In contrast, transmission between layer II/III pyramidal neurons in barrel cortex were uniformly of large amplitude and high success (release) probability (Pr). These unique features of auditory cortical transmission may provide two distinct mechanisms for discerning and separating transient from stationary features of the auditory signal at an early stage of cortical processing.

Action Potentials↗

PubMed2001

Preparation of a ribonucleic acid-(polyamidoamine)-(zirconia-urea-formaldehyde resin) high-performance liquid affinity chromatographic stationary phase.

A preparative method for a high-performance liquid affinity chromatographic (HPLAC) stationary phase is described. The 3- to 5-microm nonporous composite spherical microparticles of zirconia and urea-formaldehyde (UF) resin are synthesized through the reaction of zirconyl chloride with hexamethylene tetra-amine and urea, and then it is used as the matrix of the HPLAC stationary phase of which the diameter and structure are determined by scanning electron microscopy. In a methanol medium, the polyamidoamine (PAMAM) starburst dentritic spacer arms are linked with the imido-groups on the surface of the matrix by the Michael addition reaction with methyl acrylate and the amination reaction with ethylene diamine. After repeating these steps in triplets, amine-terminated dentritic spacer arms with a generation of 3 are obtained. The topological structure of the spacer arms is examined by solid-state 13C NMR. The Br-substituted ribonucleic acid (RNA) ligand is obtained by the reaction of liquid bromine with RNA and bonded to the dendritic spacer arms of the matrix in a solution of NaOH (pH 9-11). The binding capacity of RNA is measured by UV spectrophotometry. A new type of stationary phase--RNA-(PAMAM)-(zirconia-UF resin--for HPLAC, which possesses starburst dendritic spacer arms, is synthesized and used for the separation of biological macromolecules.

Adenine Nucleotides↗

PubMed2001

Previous pulmonary diseases and risk of lung cancer in Gansu Province, China.

BACKGROUND: Although active smoking is well established as the main cause of lung cancer, there is accumulating evidence that history of prior lung diseases may be an independent risk factor for lung cancer. METHODS: A population-based case-control study in Gansu Province, China identified 886 lung cancer cases (656 male, 230 female) diagnosed between January 1994 and April 1998. A standardized interview collected information on a variety of potential risk factors including a history of physician-diagnosed non-malignant lung diseases (pulmonary tuberculosis, chronic bronchitis/emphysema, asthma, pneumonia), age and year in which each condition was first diagnosed, and any therapy or hospitalization received. RESULTS: Pulmonary tuberculosis (odds ratio [OR] = 2.1, 95% CI : 1.4-3.1) and chronic bronchitis/emphysema (OR = 1.4, 95% CI : 1.1-1.8) were associated with increased risk of lung cancer, after adjustment for active smoking and socioeconomic status. The OR for asthma (OR = 1.4, 95% CI : 0.9-2.1) and pneumonia (OR = 1.5, 95% CI : 1.0-2.3) were also elevated. The risk of lung cancer remained significant for pulmonary tuberculosis and chronic bronchitis/emphysema when analysis was limited to the pathologically confirmed cases and self-responders. CONCLUSIONS: This study provides additional evidence that previous pulmonary tuberculosis and chronic bronchitis/emphysema are causally related to lung cancer, although the precise mechanism is still unclear. The results for asthma and pneumonia, while suggestive of a positive association, did not reach the traditional level of statistical significance and should be interpreted with caution.

Adult↗

PubMed2001

Human bronchial epithelial and endothelial cells express alpha7 nicotinic acetylcholine receptors.

The epithelial or endothelial cells that line the human bronchi and the aorta express nicotinic acetylcholine receptors (nAChRs) of alpha3 subtypes. We report here that human bronchial epithelial cells (BEC) and aortic endothelial cells (AEC) express also the nAChR alpha7 subunit, which forms functional nAChRs. Polymerase chain reaction and in situ hybridization experiments detected alpha7 subunit mRNA in cultured human BEC and AEC and in sections of rat trachea. The binding of radiolabeled alpha-bungarotoxin revealed a few thousand binding sites per cell in cultured human BEC and human and bovine AEC. Western blot and immunohistochemistry experiments demonstrated that cultured BEC and AEC express a protein(s) recognized by anti-alpha7 antibodies. Whole-cell patch-clamp studies of cultured human BEC demonstrated the presence of fast-desensitizing currents activated by choline and nicotine that were blocked reversibly by methyllycaconitine (1 nM) and irreversibly by alpha-bungarotoxin (100 nM), consistent with the expression of functional alpha7 nAChRs. In some cells, choline activated also slowly decaying currents, confirming previous reports that BEC express functional alpha3beta4 nAChRs. Exposure of cultured BEC to nicotine (1 microM) for 3 days up-regulated functional alpha7 and alpha3 nAChRs, as indicated by the increased number of cells responding to acetylcholine and choline, with both fast-desensitizing currents, which were blocked irreversibly by alpha-bungarotoxin, and with slowly desensitizing currents, which are alpha-bungarotoxin-insensitive currents. The presence of alpha7 nAChRs in BEC and AEC suggests that some toxic effects of tobacco smoke could be mediated through these nicotine-sensitive receptors.

Animals↗

PubMed2001

Positive allosteric modulators of AMPA receptors reduce proton-induced receptor desensitization in rat hippocampal neurons.

Whole-cell or outside-out patch recordings were used to investigate the effects of protons and positive modulators of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors on the desensitization of glutamate-evoked AMPA receptor currents in isolated hippocampal CA1 neurons. Protons inhibited glutamate-evoked currents (IC(50) of 6.2 pH units) but also enhanced the apparent rate and extent of AMPA receptor desensitization. The proton-induced enhancement of desensitization could not be attributed to a reduction in the rate of recovery from desensitization or to a change in the kinetics of deactivation. Non-stationary variance analysis indicated that protons reduced maximum open probability without changing the conductance of AMPA channels. The positive modulators of AMPA receptor desensitization, cyclothiazide and GT-21-005 (an organic nitrate), reduced the proton sensitivity of AMPA receptor desensitization, which suggests that they interact with protons to diminish desensitization. In contrast, the effects of wheat germ agglutinin and aniracetam on AMPA receptor desensitization were independent of pH. These results demonstrate that a reduction in the proton sensitivity of receptor desensitization contributes to the mechanism of action of some positive modulators of AMPA receptors.

Allosteric Regulation↗

PubMed2001

Gadolinium reduces AMPA receptor desensitization and deactivation in hippocampal neurons.

The actions of the trivalent cation Gd(3+) on whole cell AMPA receptor-mediated currents were studied in isolated hippocampal neurons, in nucleated or outside-out patches taken from cultured hippocampal neurons, and on miniature excitatory postsynaptic currents (mEPSCs) recorded in cultured hippocampal neurons. Glutamate, AMPA, or kainate was employed to activate AMPA receptors. Applications of relatively low concentrations of Gd(3+) (0.1-10 microM) substantially enhanced steady-state whole cell glutamate and kainate-evoked currents without altering peak currents, suggesting that desensitization was reduced. However, higher concentrations (>30 microM) depressed steady-state currents, indicating an underlying inhibition of channel activity. Lower concentrations of Gd(3+) also increased the potency of peak glutamate-evoked currents without altering that of steady-state currents. An ultrafast perfusion system and nucleated patches were then used to better resolve peak glutamate-evoked currents. Low concentrations of Gd(3+) reduced peak currents, enhanced steady-state currents, and slowed the onset of desensitization, providing further evidence that this cation reduces desensitization. In the presence of cyclothiazide, a compound that blocks desensitization, a low concentration Gd(3+) inhibited both peak and steady-state currents, indicating that Gd(3+) both reduces desensitization and inhibits these currents. Gd(3+) reduced the probability of channel opening at the peak of the currents but did not alter the single channel conductance calculated using nonstationary variance analysis. Recovery from desensitization was enhanced, and glutamate-evoked current activation and deactivation were slowed by Gd(3+). The Gd(3+)-induced reduction in desensitization did not require the presence of the GluR2 subunit as this effect was seen in hippocampal neurons from GluR2 null-mutant mice. Gd(3+) reduced the time course of decay of mEPSCs perhaps as a consequence of its slowing of AMPA receptor deactivation although an increase in the frequency of mEPSCs also suggested enhanced presynaptic release of transmitter. These results demonstrate that Gd(3+) potently reduces AMPA receptor desensitization and mimics a number of the properties of the positive modulators of AMPA receptor desensitization such as cyclothiazide.

Animals↗

PubMed2001

Nerve growth factor regulates sodium but not potassium channel currents in sympathetic B neurons of adult bullfrogs.

The TTX-sensitive and -resistant components of the voltage-gated Na(+) current (TTX-s I(Na) and TTX-r I(Na)) are increased within 2 wk of cutting the axons of B-cells in bullfrog paravertebral sympathetic ganglia (BFSG). Axotomy also increases the noninactivating, voltage-activated K(+) current (M current I(M)), whereas delayed rectifier K(+) current (I(K)) is reduced. We found that similar effects were produced when BFSG B cells were dissociated from adult bullfrogs and maintained in a defined-medium, neuron-enriched, low-density, serum-free culture. Thus the density of TTX-s I(Na), TTX-r I(Na), and I(M) were transiently increased, whereas I(K) density was decreased. Reduction in voltage-sensitive, Ca(2+)-dependent K(+) current (I(C)) was attributed to previously documented decreases in Ca(2+) channel current (I(Ca)). To test whether axotomy- or culture-induced changes in ion channel function reflect loss of retrograde influence of nerve growth factor (NGF), we examined the effect of murine beta-NGF on TTX-s I(Na), TTX-r I(Na), I(K), and I(M). Culture of neurons for 15 days in the presence of NGF (200 ng/ml), more than doubled total I(Na) density but did not enhance neurite outgrowth. The TTX-r I(Na) density was increased about threefold and the TTX-s I(Na) density increased 2.4-fold. NGF did not affect the activation or inactivation kinetics of the total Na(+) conductance. Effects of NGF were blocked by the transcription inhibitors, cordycepin (20 microM) and actinomycin D (0.01 microg/ml). I(K) and I(M) were unaffected by NGF, and although I(C) was enhanced, this likely reflected the known effect of NGF on I(Ca) in BFSG neurons. Na(+) channel synthesis and/or expression in adult sympathetic neurons is therefore subject to selective regulation by NGF. Despite this, the increase in I(Na) and I(M) as well as the decrease in I(K) seen in BFSG neurons in culture or after axotomy cannot readily be explained in terms of alterations in the availability of target-derived NGF.

Age Factors↗

PubMed2001

[Effect of flow shear stress on the expression of adhesion molecules of endothelial cells].

Adherence of various leukocytes, including monocyte, to vascular endothelial cells may play an important role in the development of atherosclerosis. In vivo, the hemodynamic shear forces have a critical effect on the surface expression of adhesion proteins. In order to elucidate the effect of flow shear stress on the expression of adhesion molecules of endothelial cells, we investigated the effect of flow shear stress (2.23-6.08 dyne/cm2) on the expression of adhesive molecules, intercellular adhesion molecule(ICAM-1), vascular cell adhesion molecule (VCAM-1), and E-selectin on cultured human umbilical vein endothelial cells(HUVECs). The expression of adhesion molecules on the surface of HUVECs induced by shear stress was analyzed using flow cytometry. The results showed that compared with stationary control, the surface expression of ICAM-1 was significantly increased (P < 0.05) on HUVECs after exposure to different shear stress (2.23, 4.20, 6.08 dyne/cm2), and it was found to be in close relationship with the shearing time (r = 0.992, 0.997, 0.997; P < 0.05). After exposure to shear(2.23 dyne/cm2), VCAM-1 expression was significant increased(P < 0.05), and it was positively correlated with the shearing time(r = 0.930; P < 0.05), while VCAM-1 expression dropped down to basal level(P < 0.05) after it was sheared at 4.20 or 6.08 dyne/cm2, and the magnitude of the reduction of VCAM-1 expression was negatively correlated with the shearing time (r = -0.975, -0.989; P < 0.05). E-selectin expression was less sensitive to shear stress, especially at the lower magnitudes of shear. These results indicate that the increase of ICAM-1 and VCAM-1 expression in endothelial cells induced by low shear stress may play a prominent role in the development of both inflammation and atherosclerosis.

Cells, Cultured↗

PubMed2001

[Effects of IL-8 on neutrophils under fluid shear].

OBJECTIVE: In order to elucidate the effect of IL-8 on neutrophils in a systematic and all-round way, we introduced an important physiological environment of neutrophils, fluid flow, into this study on the changes in some responses of neutrophils to IL-8 and the inter relationship of those responses under the conditions of different patterns of flow field. METHODS: After using IL-8 and steady flow or sinusoidal-oscillate fluid shear rate simultaneously acting on separated neutrophils for one minute, we measured the expression of CD18, CD62L and the F-actin content with flow cytometry and measured the free calcium concentration inside cells ([Ca2+]i) with the fluorescent stain Fura2/AM using the fluorescent spectrophotometer. RESULTS: It was found that flow shear stress could influence the expression of surface adherence molecule of neutrophils under the action of IL-8. CD18 surface expression increased, and CD62L shed under different flow shear rates, but no significant relationship of these changes with the intensity and pattern of shear stress was noted. On the other hand, the F-actin content in neutrophils under the action of shear stress and IL-8 decreased significantly at low shear rate and then gradually increased at high shear rate (600s -1) to the control value (without shear). The [Ca2+]i concentration under the action of shear stress and IL-8 decreased significantly at low shear rate followed by a quick increase at high shear rate. CONCLUSION: The flow shear stress can influence the activation effects of IL-8 on neutrophils and regulate the magnitude of neutrophil reactions to a great extent, and among these reactions, the change of [Ca2+]i concentration is the most sensitive one and the greatest one. This is in consistence with its second messenger role in the signal transduction pathway. Our results could serve as a basis on which to understand better the mechanism of inflammatory reaction wherein the neutrophils under flow condition of blood are involved.

Actins↗

PubMed2001

[Effects of estradiol and isoflavoid on the expression of adhesion molecules on neutrophils].

OBJECTIVE: To elucidate the effects of estradiol and isoflavoid on the expression of adhesion molecules on neutrophils. METHODS: Neutrophils of healthy subjects and ischemic stroke patients were treated with tumor necrosis factor alpha (TNF alpha), in the presence or absence of different concentrations of isoflavoid (WZ1, WZ2) and estradiol (WZ3, WZ4). Flow cytometry was used to detect the expression of CD18 and CD62L on neutrophil surface. RESULTS: 1. 10 ng/ml TNF alpha could activate neutrophils of healthy subjects; it increased the expression of CD18 by 10% on neutrophil surface and shed CD62L from the surface as shown by a 15% decrease of the fluorescence intensity and a 30% decrease of the percentage of positive cell. 2. Isoflavoid (WZ1, WZ2) had no significant effect on the expression of CD18 and CD62L on neutrophils. 3. Pretreatment of neutrophils with estradiol (WZ3, WZ4) could inhibit the activation of neutrophils by TNF alpha, which decreased the fluorescence intensity of CD18 by 8%, increased the fluorescence intensity of CD62L by 15% and increased the percentage of CD62L positive cell by 20%. 4. TNF alpha could activate the neutrophils of ischemic stroke patients strikingly; it increased the fluorescence intensity of CD18 by 20% and decreased the fluorescence intensity and percentage of positive cell of CD62L by 30%, and there was a significant difference when the patients were compared with the healthy subjects. Estradiol had the same effect on the expression of CD18 and CD62L as on those of healthy subjects. CONCLUSION: 1. TNF alpha is a strong activator of neutrophils; it plays an important role in the occurrence and development of ischemic stroke. 2. Isoflavoid has no obvious effect on the expression of adhesion molecules on neutrophils, so its role in protecting the cardiovascular system may come into play not by the way of affecting adhesion molecules expression. 3. Pretreatment of neutrophils with estradiol could protect them from activation by TNF alpha, thus decreasing the expression of adhesion molecules, the adhesion of neutrophils -endothelial cells, and hence the risk of ischemic stroke. However, in case that the neutrophils have been activated by TNF alpha, estradiol has no effect on the expression of adhesion molecules; this implies that estradiol possibly has no valid anti-adhesion therapeutic effect on ischemic stroke.

Adult↗

PubMed2001

[A kinetic study on the relationship between of IL-5, IL-10 and eosinophil apoptosis in asthmatic airway inflammation].

OBJECTIVE: To observe time-course profile of eosinophil apoptosis, in murine asthma models in airway explicate its implication for asthmatic inflammation remission, and investigate the role of IL-5 and IL-10 in modulating cell apoptosis. METHODS: An animal model of asthma was established by OVA sensitizing-challenging BALB/c mice. Histologic study of lung tissue was made with the use of electronmicroscope. At seven points (0 h, 8 h, 24 h, 48 h, 96 h, Day 7 and Day 14) in the time course after challenge, bronchoalveolar lavage (BAL) was performed obtain BAL cells and recover fluid. Levels of IL-5 and IL-10 were detected by ELISA. Cell apoptosis was assayed by PI stain on a flow cytometer. RESULTS: After challenge, apoptosis of eosinophils, lymphocytes, endothelial cells could be seen under the electronmicroscope. IL-5 in BALF elevated significantly whereas no significant difference was found between the levels of IL-10 in BALF before and after challenge. Eosinophil apoptosis was identified and a two-phase elevation of apoptosis percentages was observed, although no significant difference was found between average apoptosis in asthmatic group and that in control. Data analyses revealed a negative correlation between apoptosis percentages and the levels of IL-5, and a closer correlation between the percentages and IL-5/IL-10 ratio. CONCLUSION: IL-5 and IL-10 could up- and down-regulate asthmatic airway inflammation, respectively. EOS underwent apoptosis in situ and this was modulated by IL-5 and IL-10 interaction.

Animals↗

PubMed2001

[Effects of calcium channel antagonist on gastrin-induced proliferation of HT29 colon carcinoma cells].

OBJECTIVE: This study aimed to assess the effects of nifedipine on gastrin-induced proliferation of HT29 colon carcinoma cells and inquire into the possible mechanisms. METHODS: Flow cytometry was used to monitor the cytoplasmic free calcium; MTT colorimetry was used to determine the proliferation of HT29 cells. RESULTS: The results showed that 2.5 x 10(-6) mol/L pentagastrin (PG) induced a quick rise of intracellular free calcium ([Ca2+]i) (P < 0.01). 10(-5) mol/L nifedipine can significantly inhibited the rise of [Ca2+]i induced by pentagastrin (P < 0.01), in parallel, at growth assay we demonstrated that 10(-5)-10(-6) mol/L nifedipine could obviously block the increase in cell number elicited by 2.5 x 10(-6) mol/L PG. CONCLUSION: These data indicate that nifedipine can stop the influx of Ca2+ and hence inhibit the pentagastrin-induced proliferation.

Biological Transport, Active↗

PubMed2000

Activation of vanadium nitrogenase expression in Azotobacter vinelandii DJ54 revertant in the presence of molybdenum.

Azotobacter vinelandii carries three different and genetically distinct nitrogenase systems on its chromosome. Expression of all three nitrogenases is repressed by high concentrations of fixed nitrogen. Expression of individual nitrogenase systems is under the control of specific metal availability. We have isolated a novel type of A. vinelandii DJ54 revertant, designated A. vinelandii BG54, which carries a defined deletion in the nifH gene and is capable of diazotrophic growth in the presence of molybdenum. Inactivation of nifDK has no effect on growth of this mutant strain in nitrogen-free medium suggesting that products of the nif system are not involved in supporting diazotrophic growth of A. vinelandii BG54. Similar to the wild type, A. vinelandii BG54 is also sensitive to 1 mM tungsten. Tn5-B21 mutagenesis to inactivate the genes specific to individual systems revealed that the structural genes for vnf nitrogenase are required for diazotrophic growth of A. vinelandii BG54. Analysis of promoter activity of different nif systems revealed that the vnf promoter is activated in A. vinelandii BG54 in the presence of molybdenum. Based on these data we conclude that A. vinelandii BG54 strain utilizes vnf nitrogenase proteins to support its diazotrophic growth.

Azotobacter vinelandii↗

PubMed2000

Identification of a second site compensatory mutation in the Fe-protein that allows diazotrophic growth of Azotobacter vinelandii UW97.

Azotobacter vinelandii UW97 is defective in nitrogen fixation due to a replacement of serine at position 44 by phenylalanine in the Fe-protein [Pulakat, L., Hausman, B.S., Lei, S. and Gavini, N. (1996) J. Biol. Chem. 271, 1884-1889]. Serine residue 44 is located in a conserved domain that links the nucleotide binding site and the MoFe-protein docking surface of the Fe-protein. Therefore, it is possible that the loss of function by A. vinelandii UW97-Fe-protein may be caused by global conformational disruption or disruption of the conformational change upon MgATP binding. To determine whether it is possible to generate a functional nitrogenase complex via a compensating second site mutation(s) in the Fe-protein, we have attempted to isolate genetic revertants of A. vinelandii UW97 that can grow on nitrogen-free medium. One such revertant, designated A vinelandii BG9, encoded a Fe-protein that retained the Ser44Phe mutation and also had a second mutation that caused the replacement of a lysine at position 170 by glutamic acid. Lysine 170 is highly conserved and is located in a conserved region of the Fe-protein. This region is implicated in stabilizing the MgATP-induced conformation of the Fe-protein and in docking to the MoFe-protein. Further complementation analysis showed that the Fe-protein mutant that retained serine 44 but contained the substitution of lysine at position 170 by glutamic acid was also non-functional. Thus, neither Ser44Phe nor Lys170Glu mutants of Fe-protein were functional; however, the Fe-protein in A. vinelandii BG9 that contained both substitutions could support diazotrophic growth on the strain.

Adenosine Triphosphate↗