PubMed Health⌕ Search

Biomedical subjects

A Aszalos

Publications and source records attributed to A Aszalos.

At least 37 records · Page 2Linked to original sources
PubMed1993

The effect of ion channel blockers, immunosuppressive agents, and other drugs on the activity of the multi-drug transporter.

The MDRI protein is an energy-dependent transport protein responsible for the multi-drug resistance seen in many tumors. A variety of drugs have been shown to inhibit the function of this pump, including compounds known to block various ion channels. The mouse lymphoma cell line L5178Y has been transduced with the human mdrI gene. Using this cell line, we have tested a number of compounds to determine whether there is a correlation between the ability to block a specific type of ion channel, or shift membrane potential, and the ability to act as an MDR-reversing agent using the fluorescent substrates Rhodamine 123 and daunorubicin as test compounds. Our results show no apparent correlation between the ability to block a specific ion channel and reversal of MDR transport ability. We have found active MDR inhibitors in compounds that affect K+, Na+, Ca++, H+, but not Cl- channels. Our data suggest that Cl- channel activity may be distinct from MDR activity. Several immunosuppressive compounds and analogs were also tested and found to be active reversing agents. Measurements suggest a significant difference in resting membrane potential between the L5178YvMDR line and the L5178Y parental cell line used in these experiments. No correlation was found between the ability of drugs to alter membrane potential and to inhibit MDR transport activity. Our results suggest that MDR transport function may be independent of the physiological movement of ions and show that a wide variety of compounds can inhibit MDR transport.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1993

Specific disengagement of cell-bound anti-LAM-1 (anti-L-selectin) antibodies by aurintricarboxylic acid.

Brief treatment of human peripheral blood lymphocytes with the potential anti-HIV compound aurintricarboxylic acid (ATA) prompts the selective release of already bound L-selectin-specific anti-Leu8 and anti-LAM1-1 antibodies from the cells. Two other anti-LAM1 antibodies, anti-LAM1-3 and anti-LAM1-5 stay antigen-bound at the same time. Interestingly, the ATA-sensitive anti-Leu8 strongly competes with the ATA-resistant anti-LAM1-3 for binding. Photobleaching fluorescence resonance energy transfer (pFRET) measurements on flow-sorted cells suggests that these two antibodies compete for the same epitope, while anti-LAM1-5-FITC and anti-Leu8-PE bind to distinct sites, although they also compete for binding. Combining the data on competition, pFRET and ATA effect, we suggest that the ATA sensitive anti-Leu8 and resistant anti-LAM1-3 bind to overlapping but non-identical epitopes. This remarkably specific effect may be exploited for designing anti-inflammatory drugs that modulate leukocyte adhesion.

Antigen-Antibody Reactions↗
PubMed1992

CD4 changes conformation upon ligand binding.

Aurintricarboxylic acid (ATA) has been shown to block the binding site for both HIV gp120 and mAb anti-Leu 3a on CD4. We have unexpectedly found that brief treatment with > or = 1 micrograms/ml ATA rapidly disengages another mAb, OKT4E, after it has been bound to CD4 on human PBL. OKT4E is specific for a discontinuous epitope overlapping the MHC class II-binding region in the N-terminal CD4 domain. Interestingly, among 10 other mAb tested, only anti-Leu 8, specific for a leukocyte homing receptor is also quickly released from the cells by ATA treatment. Disengagement of the OKT4E mAb is also seen on a CD4-positive cell line (HPB-ALL) and with recombinant soluble CD4 (sCD4) bound to immobilized OKT4E. In all of these cases, disengagement is prevented if OKT4E is cross-linked, or the Leu 3a site is blocked by the mAb, but not by gp120. Photobleaching fluorescence resonance energy transfer (pFRET) measurements suggest that OKT4E is released as an indirect consequence of ATA-evoked conformational changes of CD4. Similar changes were detected as a result of gp120 binding to PBL. These data raise the possibility of a novel type of immunomodulation: induced disengagement of a bound ligand from its Ag.

Antibodies, Monoclonal↗
PubMed1992

Modulation of the anti-proliferative signal of interferon-alpha by tamoxifen in U937 cells.

Several clinical protocols are attempting to utilize the combined anti-proliferative signal of interferon-alpha (INF-alpha) and tamoxifen on cancer cells. We demonstrated here that the effect of these two agents on the growth of the premacrophage U937 cells is antagonistic. This antagonistic effect is paralleled by the ability of tamoxifen to modulate the INF-alpha-induced hyperpolarization in these cells. INF-alpha-induced hyperpolarization was shown before to be an integral part of the anti-proliferative signal of this agent. Tamoxifen liberates Ca2+ from intracellular stores of U937 cells but this effect of the drug is not the cause of its antagonistic effect with the anti-proliferative signal of IFN-alpha. We suggest therefore, that the combined effect of these two anti-cancer drugs could also be advantageous for macrophage proliferation.

Calcium↗
PubMed1992

Prevention of binding of rgp120 by anti-HIV active tannins.

Several tannins with anti-HIV activity have been described previously (Nonaka et al., J Nat Prod 53: 587-595, 1990). We have shown that the tannins chebulinic acid and punicalin were able to block the binding of HIV rgp120 to CD4. These compounds were not toxic to stimulated human peripheral blood lymphocytes at concentrations ten times above their maximal effective concentration.

Antiviral Agents↗
PubMed1992

Epitope mapping by photobleaching fluorescence resonance energy transfer measurements using a laser scanning microscope system.

The donor photobleaching method (T. M. Jovin and D. J. Arndt-Jovin. 1989. Annu. Rev. Biophys. Biophys. Chem. 18:271-308.) has been adapted to an ACAS 570 (laser scanning microscope) system to measure fluorescence resonance energy transfer (FRET) on individual human peripheral blood T cells. Photobleaching was completed in approximately 100 ms in our case and it followed double-exponential kinetics. The energy transfer efficiency (E) was approximately 20% between the CD4 epitopes OKT4-FITC and Leu-3a-PE as well as between OKT4E-FITC and OKT4-PE. E was approximately 8% between OKT4-FITC and Leu-4-PE (alpha CD3) and barely detectable (approximately 4%) from OKT4-FITC to Leu-5b-PE (alpha CD2). The E values obtained by the photobleaching method were highly reproducible both in repeated measurement of identical samples and in experiments with different batches of cells and were in agreement with the flow cytometric donor quenching measurements. As expected, E measured between primary and secondary layers of antibodies increased (from approximately 14% to approximately 28%) when F(ab')2 fragments were substituted for whole antibody molecules as the donor. On a T cell line we mapped the distance between the idiotypic determinant of the T cell receptor (TcR) and the Leu-4 epitope of CD3 as proximal as E = 28%, as compared to E = 4% between a framework TcR epitope and Leu-4. In the latter case, however, approximately 40% less Leu-4 was bound suggesting that the antigen binding site of TcR is in close proximity with one of the two CD3 epsilon chains, which hence are not equivalent.

Antibodies, Monoclonal↗
PubMed1992

Cytoskeletal modulation of plasma membrane events induced by interferon-alpha.

Cytochalasin B, a drug that alters microfilament structure, was found to modulate interferon-alpha (IFN-alpha)-induced changes in ion fluxes, in motional freedom of spin probes, and lateral diffusion of surface antigens. These changes occur in Daudi cells inherently sensitive to the antiproliferative signal of IFN-alpha, but not in insensitive cells, and were associated with the antiproliferative signal previously. The biophysical effects of cytochalasin B were detected by flow cytometric quantitation of membrane potential using an oxonol dye, by electron spin resonance (ESR) spectrometry, and by measurements of fluorescence recovery after photobleaching (FRAP) of surface antigens using a laser-interactive cell imaging system. Cytochalasin B treatment increased an IFN-alpha-induced membrane potential shift by -5 mV. The motional freedom of 5-doxyl-stearic acid changed from 0.67 to 0.63, as expressed by the order parameter, S, with IFN-alpha treatment and was prevented by cytochalasin B. Changes in the lateral diffusion of surface antigens induced by IFN-alpha treatment, D = 5.3 x 10(-10) without treatment and D = 7.8 x 10(-10) cm2/s with treatment, was blocked by cytochalasin B. In contrast, the microtubule stabilizers taxol and D2O and the microtubule depolymerizing colcemid were ineffective at dose levels sufficient to cause the characteristic cell physiological alterations of these agents. These results implicate microfilaments but not the microtubule system in transduction of the antiproliferative signal by IFN-alpha in Daudi cells.

Actin Cytoskeleton↗
PubMed1991

Cyclosporin elicits a non-responsive state and a shift in K+ fluxes in the early phase of activation of human lymphocytes with anti-CD3.

The effect of cyclosporin A on the initial phase of activation of human peripheral blood lymphocytes (PBL) by anti-CD3 was studied in a two-step incubation process. Cyclosporin A treatment in the initial phase of activation blocked the second phase activation of a cell population by anti-CD3, IL-2, or concanavalin A (ConA). In contrast, similar treatment with the calcium ionophore, ionomycin, enhanced the activation of anti-CD3. Only a marginal synergistic increase of intracellular [Ca2+] was elicited by cyclosporin A during anti-CD3 stimulation and this drug prevented activation-induced depolarization of lymphocytes by its ability to hyperpolarize cells. The hyperpolarization effect of cyclosporin A is related to the K+ flux but not the Na+ or Ca2+ flux and is unlikely to be mediated through calmodulin and protein kinase C. We postulate that the K+ flux-modulating ability of cyclosporin A renders T cells non-responsive in the initial phase of activation.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1991

Laser scanning and confocal microscopy of daunorubicin, doxorubicin, and rhodamine 123 in multidrug-resistant cells.

The multidrug-resistant gene (MDR1) encodes an energy-dependent drug efflux pump (P-glycoprotein) for many anti-cancer drugs. We have studied the intracellular distribution of rhodamine 123 (R123), daunorubicin (DN), and doxorubicin (DOX) in cells expressing a human MDR1 gene. The distribution of these fluorescent drugs was measured by laser scanning microscopy and confocal microscopy. We devised a new method for analysis of fluorescence line scan data to determine the intracellular distribution of fluorescent probes. This method and confocal microscopy showed that R123, DN, and DOX are localized to both plasma membrane and intracellular compartments in multidrug-resistant cells. When the cells are treated with verapamil, an inhibitor of the multidrug transporter, the amount of DOX, DN, and R123 associated with the cell rises. After inhibition, the relative distribution of DOX and DN between the cell surface and intracellular structures does not change dramatically. However, R123 tends to relocalize to intracellular sites from predominantly plasma membrane sites, indicating that this dye behaves differently than the anti-cancer drugs. These results show the subcellular distributions of R123, DN, and DOX in plasma membrane, cytoplasm, and intracellular membrane systems, but do not allow definitive distinctions among existing models of how P-glycoprotein affects the distribution of drugs.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1991

Effect of an interferon inhibitor on the antiproliferative signal of interferon-alpha.

We have described a nonantibody type inhibitor of interferons (IFN) in the blood of patients with AIDS, advanced neoplastic disorders, and lupus erythematosus in earlier reports (1,2). In the present study we show that the semipurified inhibitor blocks the antiproliferative signal of IFN-alpha in Daudi cells and the membrane potential shifting ability of IFN-alpha is modulated by the interferon inhibitor preparation (IFI).

Cell Division↗
PubMed1991

Comparison of the in vitro and biophysical effects of cyclosporine A, FK-506, and mycophenolic acid on human peripheral blood lymphocytes.

The immunosuppressive drugs FK-506 and mycophenolic acid (MPA) have recently been described, but their mode(s) of action are not well understood. We have compared them to cyclosporine A (CsA) in several assays. We have shown that CsA (1 microgram/ml), MPA (0.1 microgram/ml), and FK-506 (0.5 microgram/ml) all induce a state of unresponsiveness to anti-CD3 stimulation as measured by [3H]-thymidine uptake. This suggests that the target of these drugs may be present only after mitogenic stimulation. These drugs also cause a hyperpolarization of the plasma membrane of lymphocytes. This effect is blocked by quinine or verapamil. All three immunosuppressors only slightly modulate the increase in intracellular Ca++ caused by Con-A or by anti-CD3 stimulation but do not affect Ca++ levels alone. They also decrease expression of IL-2 receptors on alpha CD3-stimulated lymphocytes. Similarities in their modes of action, as measured by these biophysical and cell biological tests, indicate the possibility that these three drugs will show similarities in their clinical performance.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1991

On the mechanism of action of interferons: interaction with nonsteroidal anti-inflammatory agents, pentoxifylline (Trental) and cGMP inducers.

In earlier studies, we showed that when interferon alpha (IFN alpha) binds to cell receptors, it alters membrane potentials. In IFN-sensitive Daudi cells derived from Burkitt's lymphoma patients, IFNs produce dose-dependent hyperpolarization, decreased plasma membrane viscosity, modulation of the microfilament system, and altered synthesis of cAMP, cGMP and prostaglandins. No such changes were seen in IFN alpha-resistant Daudi cells. In this study, we found that indomethacin, pentoxifylline, or the cGMP inducer sodium azide (NaN3) had no significant effect on the IFN alpha induced membrane potential changes, or on membrane viscosity changes as measured by flow cytometry and electron spin resonance spectrometry. In tissue culture, indomethacin did not alter the anti-proliferative effect of IFN alpha on Daudi cells. Indomethacin may influence IFN synthesis, but not it's antiproliferative actions. Relatively high doses of pentoxifylline slightly inhibited proliferation of Daudi cells and synergized with IFN alpha. Measurement of early biophysical changes induced by IFNs may represent a new screening method to rapidly explore certain types of IFN alpha potentiating agents.

Azides↗
PubMed1990

Aurin tricarboxylic acid, the anti-AIDS compound, prevents the binding of interferon-alpha to its receptor.

Binding of HIV to its receptor, the CD4 molecule of lymphocytes, can be prevented by chemical agents. These agents could be considered as potential anti-AIDS drugs. We have shown that aurin tricarboxylic acid (ATA, 3 microM) specifically blocks the binding of gp120, the HIV coat protein, to the CD4 molecule. We have also found that ATA prevents the binding of interferon-alpha to its receptor in a dose-dependent manner (12-50 microM range). Membrane potential shift, associated with binding of interferon-alpha to its receptor, was also blocked by ATA in a dose-dependent fashion. Our results indicate that potential anti-AIDS drugs should be screened for such undesired side effects.

Antiviral Agents↗
PubMed1990

Polyionic compounds selectively alter availability of CD4 receptors for HIV coat protein rgp120.

We studied the ability of several polyionic compounds, previously shown to have activity in vitro against human immunodeficiency virus (anti-HIV) to block binding of anti-CD4 and recombinant HIV gp120 to the CD4 receptor on human lymphocytes. We found that Evans blue and aurin tricarboxylic acid could completely inhibit binding of anti-CD4 (Leu3a) and rgp120 and have selectivity for the CD4 receptor. A number of other compounds, including dextran sulfate and heparin had no effect on binding of rgp120 and were shown to be nonspecific for inhibition of binding of monoclonal antibodies to different T-cell receptors. Studies using a number of membrane-active drugs showed that changes in membrane potential or ion fluxes were not involved in the inhibition of binding of rgp120 by Evans blue or aurin tricarboxylic acid.

Antibodies, Monoclonal↗
PubMed1990

Protein kinase C is involved in the early signals of interferon-alpha but not of interferon-gamma in U937 cells.

Increasing interest is being focused on the role of protein kinase C (PKC) in the mode of action of interferons (IFNs). Here we report that IFN-alpha induced a transient translocation of PKC from the cytosol to the particulate fraction of U937 cells. In contrast, after IFN-gamma treatment, no significant change in PKC activity could be observed. IFN-induced changes in membrane potential were also examined by means of a potential sensitive oxonal dye and flow cytometry. Hyperpolarization was induced by both IFN-alpha and IFN-gamma. The protein kinase inhibitor H-7 blocked the hyperpolarization induced by IFN-alpha but not by IFN-gamma. These concordant results suggest that PKC is involved in the early signals of IFN-alpha but not of IFN-gamma in U937 cells.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1989

Lymphocyte subpopulation with low membrane potential in the blood of cyclosporin- and prednisone-treated patients: in vivo selectivity for T4 subset.

Previous work on the mode of action of CsA revealed that this drug shifts the membrane potential of human periferal blood lymphocytes in vitro. Recently we have analyzed lymphocytes of cyclosporin-treated transplant patients. Membrane potential analysis with the DIOC6(3) fluorescence dye indicates that all the studied patients have a subpopulation of lymphocytes with low membrane potential and that this population is made up predominently of OKT4+ cells. However there is no correlation between the clinical events and the percentage distribution of lymphocyte subpopulations as defined by the membrane potential and the T4/T8 ratio of the total lymphocyte population.

CD4-Positive T-Lymphocytes↗
PubMed1989

Effect of cyclosporin A and ionophores on the intracellular pH of lymphocytes as measured by flow cytometry.

Because of the known importance of pHi changes on the biology of cells, we have investigated the effect of CsA, the clinically important immunomodulator, on the pHi of resting mouse spleen and human peripheral blood lymphocytes. We have found that pharmacological doses (0.8 microM) of CsA caused no change in the pHi in these cells. Similarly, the Ca2+ ionophore, A23187, up to 5 microns concentration, had no effect on the pHi of these cells, in contrast to previous reports. However, higher doses of CsA (2.4-8.0 microM) did change the pHi transiently or permanently. These results were obtained with flow cytometric measurements of fluorescence intensity of the pH-sensitive dye, BCECF.

Animals↗