Biomedical subjects

H Tapiero

Publications and source records attributed to H Tapiero.

At least 37 records · Page 2Linked to original sources

PubMed1997

Potentialization of IL-2 effects on immune cells by oyster extract (JCOE) in normal and HIV-infected individuals.

Peripheral blood mononuclear cells (PBMCs) are physiologically activated by interleukin (IL)-2. We found that oyster extract (JCOE) currently used as a functional nutrient enhanced in vitro the IL-2 dependent activation as measured by cell count. 3H-thymidine uptake and up-regulation of a IL-2 receptor. In human immunodeficiency virus (HIV) seropositive individuals, this oyster extract-induced effect was marked in asymptomatic individuals with quasi-normal CD4 cell counts, but was weakly reflected in acquired immunodeficiency syndrome (AIDS) patients.

Animals↗

PubMed1997

Accumulation of simple organic cations correlates with differential cytotoxicity in multidrug-resistant and -sensitive human and rodent cells.

Structure/functional studies previously reported showed that in a series of simple organic cations in which the charge is delocalized, an aromatic ring and a minimal degree of lipophilicity (log P > -1) were required for recognition by murine cells which express P-glycoprotein (p-gp)-mediated multidrug resistance (MDR). In the present report we find that 3H-octylpyridinium, the simple aromatic cation which has been shown to be preferentially toxic to MDR- as compared to MDR+ cells, accumulates 4.7-fold greater in the MDR- cell line. In contrast, we find that 3H-guanidinium which displays no selective toxicity between MDR+ and MDR- cells, shows no significant uptake differences between these two cell types. We also present data which demonstrate that other organic cations which contain aromatic rings, a minimal degree of lipophilicity (log P> -1) and carry a delocalized (Rho 123) or shielded (triphenylmethyl phosphonium) positive charge, also accumulate to a greater degree in MDR- vs MDR+ cells. Additionally, we find that human cells which express p-gp MDR, have similar requirements for recognition of these simple compounds. In fact, the sensitivity profiles of these compounds closely correlate between murine and human cell lines. It was also found that none of the series of simple organic compounds tested showed modulatory activity in MDR+ cells, as assayed by monitoring retention of Rho 123. Thus, the requirements for MDR recognition vs those for MDR modulation are clearly distinguished with these simple structured compounds. In comparison, the calcium channel antagonist, verapamil, and a calcium channel agonist, Bay K 8644, both showed modulatory activity by increasing Rho 123 retention in MDR+ cells, further supporting the interpretation that verapamil's modulation of MDR is unrelated to its action on calcium flux. Overall, the data presented here add further information for defining the structural requirements of compounds for their recognition by, or modulation of, human cells expressing p-gp-mediated MDR.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗

PubMed1997

Dysregulation of ornithine decarboxylase activity, apoptosis and Bcl-2 oncoprotein in Syrian hamster embryo cells stage-exposed to di(2-ethylhexyl)phthalate and tetradecanoylphorbol acetate.

Perturbations of cell proliferation and death are considered as essential events in the process of carcinogenesis. Thus, two parameters, ornithine decarboxylase (ODC), an enzyme closely related to cell proliferation and transformation, and apoptotic phenomenon are profoundly modified. Using Syrian hamster embryo (SHE) cells, we have examined in the framework of two-stage carcinogenesis (initiation-promotion) the effects of a non-genotoxic [diethylhexylphthalate (DEHP)] or genotoxic [benzo[a]pyrene (BaP)] carcinogen or a non-carcinogenic compound [phthalic anhydride (AP)] on these parameters. Immunoreactive Bcl-2 and Bcl-xL proteins were also investigated following two-stage exposures. Whereas exposures to BaP, DEHP or AP alone did not provoke any modification of ODC activity, the phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA), strongly increased it. Using two-stage exposure protocol (xenobiotics first, then replacement by TPA-promoter), the ODC activity was higher than that obtained with TPA alone. This superinduction of ODC activity was observed only with the carcinogenic compounds DEHP and BaP. Following the same exposure protocol, spontaneous cellular apoptosis was decreased. Furthermore, Bcl-2 oncoprotein was also upregulated approximately 8- and 11-fold for BaP and DEHP respectively; meanwhile Bcl-xL protein rate did not change. The non-carcinogenic compound AP slightly inhibited spontaneous SHE cell death without ODC superinduction. Exogenous polyamines, putrescine, spermidine and spermine diluted in the medium did not inhibit spontaneous apoptosis. Although inhibition of apoptosis was not specific of carcinogenic compound, both superinduction of ODC activity and inhibition of apoptosis via Bcl-2 upregulation, may cooperate during early stages of the carcinogenic process.

Animals↗

PubMed1996

Effects of fluoro-doxorubicin (ME2303) on microtubules: influence of different classes of microtubule-associated proteins.

Anthracyclines are among the most useful agents for the treatment of neoplastic disease, but their clinical use is limited by progressive cardiomyopathy. A few studies have suggested the role of microtubules for the understanding of this toxicity. By using kinetic and structural studies, we demonstrate the disorganizing action of fluoro-doxorubicin, a novel anthracycline, on the microtubule system. Microtubules have a rich and complex composition in relation to their numerous functions in cells. In the present study, we investigate the role of two major microtubule-associated protein (MAP) families, Tau and MAP2. Both MAP families are responsible for the properties of different classes of microtubules. We show the differential effect of fluoro-doxorubicin on these two classes of microtubules. Furthermore, we show that fluoro-doxorubicin is able to affect the capacity of purified tubulin to form normal microtubules. This study confirms that anthracyclines may interfer with the microtubule organization. We suggest that some classes of microtubules, with regard to their MAP composition, may be affected more specifically in cardiac myocytes.

Animals↗

PubMed1996

Increased glutathione expression in cells induced by Crassostera gigas extract (JCOE).

The amino-acid composition of Crassostera gigas extract powder (JCOE, Japan clinic oyster extract) is high in glutamic acid and taurine. In order to study whether these components present in JCOE could be contributory factors in enhancing cellular glutathione-stimulating hormone (GSH) and glutathione S-transferase (GST), HL60 cells were exposed to increasing concentrations of JCOE. Statistically significant increases in GSH occurred at extract concentrations of 0.05 and 0.1% with no significant change in GST activity. Because of the high taurine levels measured in the extract, the effects of micromolar taurine supplementation of the medium were studied. A modest, but not statistically significant increase in both GSH levels and GST activity was measured. We therefore conclude that while the taurine present in JCOE could be a contributory factor in enhancing GSH and GST, it was not the only stimulus.

Animals↗

PubMed1994

Modulatory effect of dexamethasone on ornithine decarboxylase activity and gene expression: a possible post-transcriptional regulation by a neutral metalloprotease.

The intracellular effect of dexamethasone (DXME) on the activity and gene expression of ornithine decarboxylase (ODC) was studied in Syrian hamster embryo cells (SHE). The ODC activity (expressed as nmoles decarboxylated ornithine mg-1 protein h-1) was 4.61 +/- 0.14 in untreated cells, whereas it increased to 14.38 +/- 0.26 after 5 h treatment with 1.6 x 10(-7) M TPA. In contrast, DXME (2.5 x 10(-5) M) reduced the ODC activity by 50 per cent to 2.35 +/- 0.22. In cells co-treated for 5 h with TPA and DXME, ODC activity decreased to the level of the untreated cells. However, when DXME was added 3 h after TPA treatment for 2 h, in the continuous presence of TPA, the ODC activity unexpectedly increased further to 16.44 +/- 1.05. The modulation of ODC activity correlated partly with the level of ODC mRNA. Thus when cells were treated with TPA, the ODC mRNA increased threefold, whereas it decreased by 30 per cent when the cells were exposed to DXME. In TPA-DXME co-treated cells, as in TPA pretreated cells followed by DXME for 2 h, a decrease (31.25 per cent and 12.5 per cent respectively) was observed in ODC mRNA. In turnover studies, DXME was found to increase the stability of ODC; the discrepancy between ODC activity and ODC mRNA levels could result from an inhibitory effect of the corticoid on proteolysis of ODC. Studies of lysosomal protease showed that the activities of cathepsins L, B and H decreased following TPA treatment. DXME also inhibited cathepsin L and B activities, but stimulated cathepsin H.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1994

Effect of DNA-repair-enzyme modulators on cytotoxicity of L-phenylalanine mustard and cis-diamminedichloroplatinum (II) in mammary carcinoma cells resistant to alkylating drugs.

We investigated the effect of DNA-repair-enzyme inhibitors on L-phenylalanine mustard (L-PAM) and cis-diamminedichloroplatinum (II) (CDDP) cytotoxicity in rat mammary-carcinoma MatB cells sensitive (WT) and resistant (MLNr) to bifunctional alkylating drugs. Among the modulators tested, the combination of arabinofuranosylcytosine (Ara-C) and hydroxyurea (HU) significantly increased the sensitivity of the cells to CDDP and, to a lesser extent, L-PAM as compared with cells treated with drug alone. The modulation effect of HU+Ara-C on CDDP and L-PAM cytotoxicity was more effective when intracellular glutathione (GSH) was depleted by L-buthionine-(S,R)-sulfoximine (BSO). This was also associated with a significant increase in DNA-DNA interstrand cross-links. Caffeine also sensitized both WT and MLNr cells to the cytotoxic effect of L-PAM and CDDP, and this effect was potentiated in GSH-depleted cells. No significant effect was observed with other repair modulators such as aphidicolin, 3-aminobenzamide, novobiocin, or etoposide. These results show (a) that inhibition of DNA repair by HU+Ara-C or caffeine could be a target for modulation of bifunctional alkylating-drug resistance and (b) that GSH depletion renders resistant cells more susceptible to the repair-enzyme modulators, suggesting that intracellular GSH may be involved in the regulation of some of these enzymes. Our results also indicate that a combination of a number of modulators may offer an advantage over the use of a single modulator in tumor resistance that may be associated with multifactorial mechanisms.

Adenocarcinoma↗

PubMed1994

Cross resistance relevance of the chemical structure of different anthracyclines in multidrug resistant cells.

Positively charged doxorubicin (DOX) and non-positively charged anthracyclines, aclarubicin (ACR) and morpholino-carminomycin (KRN 8602), have been investigated with respect to pharmacological parameters, cytotoxicity, DNA damage and repair in DOX-sensitive and -resistant murine and human cells. Friend leukemia cells (FLC) resistant to high concentrations of doxorubicin (DOX-RFLC3) or daunorubicin (DNR-RFLC3) (1771 and 1543 fold resistance respectively) express less than 10 fold resistance to aclarubicin (ACR). In these cells, the intracellular accumulation of ACR is similar in sensitive and resistant cells. Resistance to ACR was not observed in either DOX-RFLC1 or DNR1 with a lower level of resistance (27 fold). Increased expression of a 170,000-dalton surface antigen (gp-170) was found to be correlated with the level of resistance. However, when the selective agent in ACR, despite the low level of resistance (2.8 fold) both high expression of gp 170 and resistance to DOX (77 fold) or DNR (62 fold) are observed. It is assumed therefore that induction of multidrug resistance phenotype can be achieved by compounds which do not display cross resistance with DOX or DNR. Reduced levels or absence of cross-resistance can be related to the electrical charge of the compound. This assumption is supported by further studies on DOX-sensitive or -resistant human K562 cells exposed to another non-positively charged anthracycline, KRN 8602. In the continuous presence of drug, K562/DOX were less resistant to KRN 8602 (2.9 fold) than to DOX (31 fold). After short time exposure followed by growth in drug-free medium, absence of cross-resistance to KRN 8602 has been observed in K562/DOX. Furthermore, accumulation experiments showed that high intracellular drug concentrations were rapidly achieved (within 15 min) in both DOX-sensitive and -resistant cells. In cells exposed to DOX, DNA single-strand break (DNA-SSBs) frequencies were related to time and drug concentration while those produced by KRN 8602 or ACR were maximal after short time incubation. DNA-SSBs produced by these anthracyclines are not repaired when cells are incubated in drug free medium. In DOX resistant cells, DNA-SSBs produced by DOX were repaired whereas those produced by ACR or KRN 8602 were not. It is suggested, therefore, that absence of cross resistance to various anthracyclines is related to differences in the chemical electrical charge, which may influence drug accumulation and DNA repair in resistant cells.

Aclarubicin↗

PubMed1993

The (8-thiotheophyllinate) (triphenylphosphine) gold (I), (tTAuP): a new gold complex as an anticancer agent.

In continuous Friend leukemia cell exposure to tTAuP, the IC50 was 0.2 microM whereas in cells exposed 15 or 60 min to tTAuP followed by 72 h in drug-free medium the IC50 was 2.2 and 1.3 microM respectively. A combination of tTAuP and cisplatin (CDDP) is shown to be more active than either agent alone. Intracellular accumulation studies analysed by SXRF have shown that to achieve a cytotoxic effect, large concentrations of gold are necessary to accumulate in both the nuclear and cytoplasmic fraction. However, at cytostatic doses (1 microM), tTAuP has no effect on the cell cycle but does affect DNA synthesis. At a higher dose, greater than that necessary to induce cytotoxicity, it produces DNA damage, as observed by alkaline elution method. When cells were exposed to toxic doses of tTAuP (10 microM) in the presence of albumin, cytotoxicity was significantly reduced. Similar results were obtained when cells were co-treated with L-cysteine, dithiothreitol or reduced gluthatione. Reduced cytotoxic effect can be related to the interaction with free thiol groups. According to these data it is concluded that tTAuP is highly effective in vitro; whether it is active in vivo remains to be determined.

Antineoplastic Agents↗

PubMed1992

Structural requirements of simple organic cations for recognition by multidrug-resistant cells.

We previously noted that a wide variety of drugs which are recognized by multidrug-resistant cells (MDR+) are positively charged. However, it remains unclear why and how such a large number of structurally different compounds can be distinguished by MDR+ cells. The majority of the diverse compounds subject to MDR are complex and thereby complicate definitive structure/function characterization of the P-glycoprotein-mediated MDR mechanism. Using a series of simple aromatic (alkypyridiniums) and nonaromatic (alkylguanidiniums) organic cations differing in their lipophilicity by stepwise additions of single alkyl carbons, we demonstrate by growth inhibition studies that a single aromatic moiety and a critical degree of lipophilicity (log P > -1) are required for recognition of these simple organic cations by MDR+ cells. Thus, MDR+ cells are not cross-resistant to the nonaromatic guanidiniums but do show cross-resistance to those aromatic pyridiniums with chain lengths > four. Resistance ratios, as determined by comparison of 50% inhibitory doses in MDR- versus MDR+ cells, increase as a function of increasing chain lengths of these latter simple aromatic compounds. Resistance to pyridinium analogues in MDR+ cells is reversible by co-treatment with nontoxic doses of verapamil. Preliminary uptake data with radioactive analogues further implicate the MDR mechanism of lowered drug accumulation in accounting for resistance to the pyridinium homologues. Utilization of these simple organic cations provides a rational basis for better defining the physical chemical properties of more complex compounds processed by the MDR mechanism and suggests a strategy for designing chemotherapeutic agents with reduced susceptibility to MDR.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1992

Variation of choline-substituted lipid metabolism in doxorubicin-resistant leukemia cells.

In adriamycin-resistant murine (FLC) and human (K562) leukemia cells, phosphatidylcholine increases and phosphatidylethanolamine decreases compared to adriamycin-sensitive lines. This change is due to an increase in phosphatidylethanolamine methylation. The choline pathway of phosphatidylcholine biosynthesis is also disturbed in resistant cells with a blocking step of CDP-phosphocholine transferase and a decrease in sphingomyelinase activity. These changes in phospholipid metabolism are suggested to be responsible for the changes in membrane fluidity reported previously (Tapiero et al, 1986) for resistant cells.

Animals↗

PubMed1992

Cytotoxic and photodynamic effects of Photofrin on sensitive and multi-drug-resistant Friend leukaemia cells.

To study cross-resistance to Photofrin (PF) photosensitization, a Friend leukaemia cell line (ADM-RFLC) with a high level of multi-drug resistance (MDR) and the parental sensitive cell line (FLC) have been used. PF uptake measured by HPLC shows a similar intracellular drug accumulation in both cell lines. The ID50s for cell growth inhibition by PF are also similar after exposure in the dark in the two cell lines, while after illumination they are slightly lower in ADM-RFLC than in FLC cells. Moreover, verapamil, known to reverse the MDR phenotype induced by P-glycoprotein over-expression (the drug efflux mechanism), affects equally ADM-RFLC and FLC cells sensitivity to PF. In addition, photodynamic treatment with PF did not reverse the resistance to rhodamine 123 and aclarubicin, but partly reverses resistance of ADM-RFLC cells to antitubulin drugs such as vinblastine or vincristine. These latter results could have clinical application in the treatment of tumours expressing the MDR phenotype.

Animals↗

PubMed1990

3'-Deamino-3'-morpholino-13-deoxo-10-hydroxycarminomycin conquers multidrug resistance by rapid influx following higher frequency of formation of DNA single- and double-strand breaks.

The mechanism of action of 3'-deamino-3'-morpholino-13-deoxo-10-hydroxycarminomycin (MX2) was examined in a human leukemia cell line (K562) and its Adriamycin (ADM)-resistant subline (K562/ADM). ADM and MX2 showed an equivalent antitumor effect against K562. K562/ADM was highly resistant to ADM. In cellular pharmacokinetic studies, MX2 showed faster and greater influx than did ADM in both K562 and K562/ADM. The efflux of ADM was rapid in K562/ADM but not in K562. On the other hand, the efflux of MX2 was rapid in both cell lines. The formation of DNA single-strand breaks and double-strand breaks by ADM was significantly lower in K562/ADM than K562. On the other hand, formation of those breaks by MX2 was not decreased. Although some of the DNA breaks induced by MX2 were resealed, there was no difference in the degree of resealing in K562 and K562/ADM cells. On the other hand, most of the small number of DNA breaks in K562/ADM induced by ADM were resealed. The topoisomerase II activity in K562 and K562/ADM was not significantly different. It is concluded that MX2 conquers multidrug resistance by rapid influx following a higher frequency of formation of DNA single- and double-strand breaks in K562/ADM cells.

Antibiotics, Antineoplastic↗

PubMed1990

Can cytotoxic activity of anthracyclines be related to DNA damage?

Accumulation, cytotoxicity, and DNA damages produced by doxorubicin (DOX), pirarubicin (THP-DOX), fluoro-doxorubicin (ME2303) or its isolated metabolite M1 have been investigated in human myelogenous leukemia cells, sensitive (K562) and resistant to DOX (K562/DOX). These compounds differed by lipophilicity and/or sugar moiety either with (DOX, THP-DOX) or without (ME2303, M1) amino group. In K562 cells, the cytotoxicity was correlated to DNA single-stranded breaks and the intracellular drug amount of DOX or M1. This was not true when the cells were treated with THP-DOX or ME2303. In addition, THP-DOX produced total DNA protein cross-linking. In K562 cells DNA damage was not repaired, while in K562/DOX repair of DNA damage produced by all drugs could be observed. Although in K562/DOX cells drug accumulation was much reduced, higher intracellular drug concentration was required to induce similar level of cytotoxicity and DNA damage. Thus, cytotoxicity produced by anthracycline is not always associated with DNA damage. Different level of resistance to DOX, THP-DOX, ME2303 or M1 is associated with reduced drug accumulation which varies with the structure.

Antibiotics, Antineoplastic↗

PubMed1989

Relevance of the chemical charge of rhodamine dyes to multiple drug resistance.

Previously, we have shown that multiple drug resistant (MDR) Friend leukemia cells (FLC) are cross-resistant to the positively-charged dye, Rhodamine 123 (Rho 123), and that this resistance can be reversed by verapamil (VER). In the present study we used two zwitterionic rhodamine analogs, Rhodamine 116 and Rhodamine 110, and another positively-charged analog, Rhodamine 6G, to determine whether drug accumulation, resistance and modulation were affected by changes in the charge of these compounds. While there was no differential sensitivity between sensitive and resistant FLC to zwitterionic rhodamines, there was marked differential toxicity between these cell types for the positively-charged analogs. The IC50 values were 1000- and 100-fold greater in resistant than in sensitive cells for Rho 123 and Rho 6G respectively. Intracellular drug accumulation was significantly higher in sensitive as compared to resistant cells for both Rho 123 and Rho 6G, but little difference in drug uptake between these two cell types was observed for Rho 110 and Rho 116. It was also found that the intracellular to extracellular ratio of the positively-charged compounds was greater than unity in both sensitive and resistant cells whereas for the zwitterionic analogs this ratio was less than 1. Furthermore, this ratio of drug uptake was found to be significantly higher for Rho 6G than for Rho 123, which correlated with the high oil:water partition coefficient of Rho 6G (115.6). In MDR cells, verapamil increased Rho 123 and Rho 6G accumulation by 9.4- and 8.6-fold respectively. In addition, IC50 values in resistant cells were reduced greater than 100-fold for Rho 6G and greater than 1000-fold for Rho 123 in the presence of 10 micrograms/ml of verapamil. In contrast, less than 2-fold reduction of IC50 values for both of the zwitterionic analogs could be obtained under the same conditions. These results indicate that the chemical charge of rhodamines plays an important role in their differential accumulation, cytotoxicity and sensitivity to modulators such as verapamil, in sensitive and multi-drug resistant cells. The data also suggest that increased lipophilicity of the positively-charged rhodamines may increase their ability to accumulate in, and subsequently kill, MDR cells.

Animals↗

PubMed1989

Effect of doxorubicin on mouse hybridoma B cells: stimulation of immunoglobulin synthesis and secretion.

The purpose of these studies was to investigate whether the cell-differentiating effect of anthracyclines can trigger an over-production and secretion of molecules that may interfere with the tumor-host relationship. We exposed mouse hybridoma B-cells, which are devoted to immunoglobulin production, to doxorubicin (10-40 ng/ml). We found that most doxorubicin-treated cells secreted 3- to 5-fold higher amounts of immunoglobulin than untreated cells, along with an accumulation of 50% of them in the G2+M phase of the cell cycle. The antigenic specificity of the immunoglobulin and its size pattern as determined by polyacrylamide gel electrophoresis were similar whether or not cells were treated with doxorubicin. The enhancement of immunoglobulin secretion by doxorubicin was associated with an increase of the intracellular pool of heavy and light chains of the immunoglobulin. Furthermore, an elevated synthesis of immunoglobulin was observed. The synthesis of other proteins also appeared to be modified in these circumstances. These data suggest that doxorubicin can potentiate the biological functions of target cells when used at low concentrations, elevating the production and secretion of effector molecules that interfere with the tumor-host relationship.

Animals↗

PubMed1989

Membrane potential differences between adriamycin-sensitive and -resistant cells as measured by flow cytometry.

Using the fluorescent membrane potential probe, 3,3'-dihexyl-oxacarbocyanine (DiOC6(3], we found a 4-fold higher uptake in Adriamycin (ADM)-sensitive versus -resistant Friend leukemia cells (FLC). When sensitive cells were treated in the presence of high potassium (120 mM K+), there was a greater than 80% reduction of DiOC6(3) uptake. Using carbonylcyanide 4-trifluoromethoxy-phenylhydrazone (FCCP), a specific inhibitor of mitochondrial membrane potential, DiOC6(3) accumulation was reduced by less than 30% in these cells. Both results support the conclusion that a greater uptake of DiOC6(3) in ADM-sensitive than in -resistant cells indicates an increased plasma transmembrane potential. Since electronegative plasma membrane potentials are a driving force for the transport of lipophilic positively-charged compounds, differences in membrane potentials between sensitive and multiple drug resistant (MDR) tumor cells could have an important influence on drug accumulation and cytotoxicity. The drugs which our ADM-resistant FLC display multiple drug resistance to are positively charged. In MDR FLC, the calcium channel antagonist, verapamil, has been shown to block the efflux of Rhodamine 123 (Rho 123) and other positively-charged compounds. Since DiOC6(3) is also positively-charged, we used verapamil to investigate its effects on drug uptake. In MDR FLC, verapamil increased DiOC6(3) accumulation by 1.9-fold, whereas in sensitive cells it was increased 1.5-fold. In contrast, verapamil increased the levels of Rho 123 in resistant cells 7.8-fold but lowered them in sensitive cells 1.5-fold. The minimal loss of DiOC6(3) from both sensitive and MDR cells and the above results can best be interpreted as indicating that DiOC6(3) is not transported by the efflux "pump" system but that verapamil induces a plasma membrane potential increase in sensitive and resistant cells that DiOC6(3) is sensitive to. On the other hand, since Rho 123 did appear to be actively effluxed from these resistant cells, the enhancement of this compound by verapamil was more likely due to inhibition of the MDR "pump." How, or whether, plasma membrane potentials and the MDR efflux "pump" are related remains to be investigated. In the resistant cells, verapamil also induced an increase (13-fold) in the accumulation of the electrically neutral fluorescent probe for calcium, INDO-1/AM. However, verapamil had no effect on the efflux of this compound, which was equivalent in both resistant and sensitive cells. Thus, a new effect of verapamil on drug accumulation in MDR cells is identified here.

Animals↗

PubMed1989

Glutathione, glutathione S-transferases, and related redox enzymes in Adriamycin-resistant cell lines with a multidrug resistant phenotype.

Friend erythroleukemia cells (FLC) selected by exposure to Adriamycin (doxorubicin) express an approximate 2.5-fold (ARN1) or 13-fold (ARN2) resistance to the drug with various degrees of cross-resistance to other anthracyclines, vinca alkaloids, and epipodophyllotoxins. Because the redox cycling of the quinone moiety of Adriamycin is known to produce oxidative stress, however, an analysis of glutathione (GSH) and related enzyme systems was undertaken in the wild-type and selected resistant cells. In ARN1 and ARN2, superoxide dismutase (SOD) and catalase activities were slightly decreased, intracellular GSH and GSH reductase were essentially unchanged, and total GSH peroxidase, glutathione S-transferase (GST), and DT-diaphorase activities were slightly elevated. In each case there was no stoichiometric relationship between degree of resistance and level of activity. GST isozymes were purified from each cell line by HPLC GSH affinity column chromatography. Two-dimensional gel electrophoresis and western blot immunoreactivity against a battery of GST isozyme polyclonal antibodies determined that both the resistant and sensitive cells expressed isozymes of the alpha, pi, and mu classes (alternative murine nomenclature: M1, M2, M3). Of significance, both ARN1 and ARN2 cell lines expressed a unique alpha subunit which was absent from the parent FLC cell line. This isozyme presumably accounted for the increased GSH peroxidase activity (cumene hydroperoxide as substrate) found in ARN1 and ARN2 and may play a role in the small incremental resistance to melphalan found for both resistant lines. Expression of the isozyme was not stoichiometric with respect to degree of resistance. The presence of this isozyme may contribute to the resistant phenotype or may be the consequence of a more general cellular response to oxidative stress.

Animals↗