Ecstasy metabolites and monoamine neurotransmitters upshift the Na+/K+ ATPase activity in mouse brain synaptosomes.

3,4-Methylenedioximethamphetamine (MDMA; "ecstasy") is a psychotropic drug with well-known neurotoxic effects mediated by hitherto not fully understood mechanisms. The Na+- and K+-activated adenosine 5'-triphosphatase (Na+/K+ ATPase), by maintaining the ion gradient across the cell membrane, regulates neuronal excitability. Thus, a perturbation of its function strongly impacts cell homeostasis, ultimately leading to neuronal dysfunction and death. Nevertheless, whether MDMA affects the Na+/K+ ATPase remains unknown. In this study, we used synaptosomes obtained from whole mouse brain to test the effects of MDMA, three of its major metabolites [α-methyldopamine, N-methyl-α-methyldopamine and 5-(glutathion-S-yl)-α-methyldopamine], serotonin (5-HT), dopamine, 3,4-dihydroxy-L-phenylalanine (L-Dopa) and 3,4-dihydroxyphenylacetic acid (DOPAC) on the Na+/K+ ATPase function. A concentration-dependent increase of Na+/K+ ATPase activity was observed in synaptosomes exposed to the tested compounds (concentrations ranging from 0.0625 to 200 µM). These effects were independent of protein kinases A and C activities. Nevertheless, a rescue of the compounds' effects was observed in synaptosomes pre-incubated with the antioxidant N-acetylcysteine (1 mM), suggesting a role for reactive species-regulated pathways on the Na+/K+ ATPase effects. In agreement with this hypothesis, a similar increase in the pump activity was found in synaptosomes exposed to the chemical generator of superoxide radicals, phenazine methosulfate (1-250 µM). This study demonstrates the ability of MDMA metabolites, monoamine neurotransmitters, L-Dopa and DOPAC to alter the Na+/K+ ATPase function. This could represent a yet unknown mechanism of action of MDMA and its metabolites in the brain.

Protective effects of small heat shock proteins in Daphnia magna against heavy metal exposure.

Daphnia magna is one of the most commonly used model organisms to assess toxicity of heavy metal and other xenobiotics. However, the lack of knowledge about important stress-resistant molecules limits our understanding of the alteration of phenotypic and physiological traits of D. magna upon stress exposures. In this study, we focused on a chaperone family of small heat shock protein (sHSP) that has been found in archaea, bacteria and eukaryotes and plays an important role in stress tolerance. A total of eleven sHSP genes (termed DmsHSP1 - DmsHSP11) were identified from the D. magna genome, whose expression profiles during exposure to heavy metal (Cd2+, Cu2+ and Zn2+) and a few other potential pollutants were evaluated via qRT-PCR and RNA-Seq analysis. The results highlighted the predominant role of DmsHSP1 with the highest basal expression level in adults and robust upregulation upon exposure to heavy metals (Cu2+ > Cd2+ > Zn2+). In vivo, recombinant protein rDmsHSP1-21 and rDmsHSP11-12.8 could not only prevent model substrates agglutination induced by heavy metals or reducer dithiotreitol (DTT), but also protect tissue proteins and enzymes from denaturation and inactivation caused by heavy metals or high temperature. Ectopically expression of DmsHSP1-21 or DmsHSP11-12.8 in E. coli conferred host enhanced resistance against various abiotic stresses including Cd2+, Cu2+ and phenazine methosulfate (PMS). Knockdown of DmsHSP1-21 by RNAi, but not for DmsHSP11-12.8, significantly increased the vulnerability of D. magna to heavy metal exposure. Our work provides systematic information on the evolution and function of sHSPs in D. magna and leads to important insights into the mechanisms by which D. magna survive in adverse environments.

Oxidative stress antagonizes fluoroquinolone drug sensitivity via the SoxR-SUF Fe-S cluster homeostatic axis.

The level of antibiotic resistance exhibited by bacteria can vary as a function of environmental conditions. Here, we report that phenazine-methosulfate (PMS), a redox-cycling compound (RCC) enhances resistance to fluoroquinolone (FQ) norfloxacin. Genetic analysis showed that E. coli adapts to PMS stress by making Fe-S clusters with the SUF machinery instead of the ISC one. Based upon phenotypic analysis of soxR, acrA, and micF mutants, we showed that PMS antagonizes fluoroquinolone toxicity by SoxR-mediated up-regulation of the AcrAB drug efflux pump. Subsequently, we showed that despite the fact that SoxR could receive its cluster from either ISC or SUF, only SUF is able to sustain efficient SoxR maturation under exposure to prolonged PMS period or high PMS concentrations. This study furthers the idea that Fe-S cluster homeostasis acts as a sensor of environmental conditions, and because its broad influence on cell metabolism, modifies the antibiotic resistance profile of E. coli.

Phenazine methosulphate-treated red blood cells activate NF-κB and upregulate endothelial ICAM-1 expression.

Although enhanced Red Blood Cell (RBC) - Endothelial Cell (EC) interaction, as well as RBC induced EC activation, have been extensively studied in several RBC-linked pathologies, the specific individual effects of oxidatively modified RBC on EC activation has not yet been documented. However, increasing evidence in both experimental and clinical studies suggests that oxidatively modified RBC could be considered potential pathogenic determinants in several acute and chronic diseases displaying systemic oxidative stress. Therefore, the present study aimed to explore the specific effects of oxidized RBC interaction with endothelial cells on intracellular signaling pathways that promote EC activation. RBC were exposed to oxidative stress induced by phenazine methosulphate (PMS). It is shown that the interaction of oxidatively modified RBC with cultured human umbilical vein endothelial cells (HUVEC) results in: a) EC activation as indicated by the increased surface expression of intercellular adhesion molecule -1 (ICAM-1); b) the activation of transcription factor NF-κB, an indicator of cellular oxidant stress. These results emphasize the specific contribution of oxidatively modified RBC interaction to EC activation and their possible pathological role in vascular diseases and oxidative stress.

Serine racemase inhibition induces nitric oxide-mediated neurovascular protection during cerebral ischemia.

There are no effective neuroprotectant drugs for acute cerebral ischemia. Serine racemase (SR) synthesizes d-serine, which is involved in N-methyl-d-aspartate (NMDA) receptor-induced neurotoxicity. Recently, SR deletion was reported to protect against focal cerebral ischemia. However, regulatory mechanisms controlling SR-activity in the neurovascular unit (NVU) during cerebral ischemia remain to be clarified. We investigated the effects of SR inhibition on neurovascular protection after ischemia. The SR inhibitor phenazine methosulfate (PMS) alleviated neuronal damage in an ex vivo ischemic model (oxygen glucose deprivation [OGD]) using primary neuronal cultures, and in an in vivo mouse model of ischemia (middle cerebral artery occlusion [MCAO]). Ischemic preconditioning (IP) and PMS-treatment inhibited SR phosphorylation after ischemia ex vivo. In addition, SR phosphorylation after MCAO was also decreased in PMS-treated mice. Reductions in regional cerebral blood flow (CBF) after MCAO were improved by administration of PMS. Treatment with PMS increased phosphorylation of endothelial nitric oxide synthase (eNOS) in the ischemic core and penumbra region. In neuron-endothelial cell co-cultures, PMS promoted nitric oxide production after OGD. These findings indicate that SR inhibition acts as a neuroprotectant in the NVU and ameliorant of CBF abnormalities post-stroke. Thus, pharmacologic SR inhibition has potential clinical applications.

Effect of α-asarone on angiogenesis and matrix metalloproteinase.

α-Asarone is a main component of Acorus gramineus widely known as an oriental traditional medicinal stuff. A. gramineus has been known to have a variety of medicinal efficacies such as anti-gastric ulcer and anti-allergic activities, inhibition of histamine release and antioxidant effect. However, its effect on angiogenesis remains unclear. The aim of this study was to investigate the effect of α-asarone on induction of angiogenesis through modulation of matrix metalloproteinase (MMP). First of all, MTT assay was performed to evaluate the effect of α-asarone on cell viability using MTT assay, and then tube formation assay with human umbilical vein endothelial cells (HUVEC) in vitro and rat aorta ring assay ex vivo were carried out to elucidate its effect on angiogenesis. Treatment with α-asarone below 6µM showed no cytotoxicity in human fibrosarcoma cells (HT1080) and HUVEC. It was observed that α-asarone not only promotes tube formation of HUVEC but also induces angiogenesis of rat aorta. In addition, the effects of α-asarone on the expressions of protein and gene were evaluated using western blot analysis and RT-PCR assay. α-Asarone increased the expression levels of MMP-2 and MMP-9 stimulated by phenazine methosulfate (PMS) and phorbol 12-myristate 13-acetate (PMA) in HT1080. Especially, the expression level of antioxidant enzyme such as glutathione reductase was increased in the presence of α-asarone. Therefore, above findings suggest that α-asarone may play an important role in pathological diseases related to MMP and angiogenesis.

Phenazine methosulfate decreases HIF-1α accumulation during the exposure of cells to hypoxia.

In HEK293 cells, exposure to various NAD(P)H oxidants, including phenazine methosulfate (PMS), that non-enzymatically oxidize intracellular NAD(P)H to NAD(P), decreased hypoxia-induced hypoxia-inducible factor 1 (HIF-1α) accumulation. RT-PCR and cycloheximide inhibition experiments indicated that PMS-induced HIF-1α decrease is involved in post-translational degradation during hypoxia. The decrease in HIF-1α caused by PMS was not eliminated by proteasome inhibitor MG132. Moreover, the increase in HIF-1α induced by exposure to MG132 alone in normoxia was diminished by PMS. In contrast, calpastatin peptide, a calpain inhibitor, fully prevented PMS-induced reduction in HIF-1α in hypoxic cells. These data suggest that the decreased stability of HIF-1α induced by PMS is due to the activation by PMS of a protein degradation system that is independent of the ubiquitin-proteasome pathway.

Calcineurin expression and activity is regulated by the intracellular redox status and under hypertension in human neutrophils.

Calcineurin (protein phosphatase 2B) (CN) comprises a family of serine/threonine phosphatases that play a pivotal role in signal transduction cascades in a variety of cells, including neutrophils. Angiotensin II (Ang II) increases both activity and de novo synthesis of CN in human neutrophils. This study focuses on the role that intracellular redox status plays in the induction of CN activity by Ang II. Both de novo synthesis of CN and activity increase promoted by Ang II were downregulated when cells were treated with L-buthionine-(S,R)-sulfoximine, an inhibitor of synthesis of the antioxidant glutathione. We have also investigated the effect of pyrrolidine dithiocarbamate and phenazine methosulfate, which are antioxidant and oxidant compounds, respectively, and concluded that the intracellular redox status of neutrophils is highly critical for Ang II-induced increase of CN expression and activity. Results obtained in neutrophils from hypertensive patients were very similar to those obtained in these cells on treatment with Ang II. We have also addressed the possible functional implication of CN activation in the development of hypertension. Present findings indicate that downregulation of hemoxygenase-1 expression in neutrophils from hypertensive subjects is likely mediated by CN, which acts by hindering translocation to the nucleus of the transcription factor NRF2. These data support and extend our previous results and those from other authors on modulation of CN expression and activity levels by the intracellular redox status.

PMS: photosystem I electron donor or fluorescence quencher.

Light energy harvested by the pigments in Photosystem I (PSI) is used for charge separation in the reaction center (RC), after which the positive charge resides on a special chlorophyll dimer called P700. In studies on the PSI trapping kinetics, P700(+) is usually chemically reduced to re-open the RCs. So far, the information available about the reduction rate and possible chlorophyll fluorescence quenching effects of these reducing agents is limited. This information is indispensible to estimate the fraction of open RCs under known experimental conditions. Moreover, it would be important to understand if these reagents have a chlorophyll fluorescence quenching effects to avoid the introduction of exogenous singlet excitation quenching in the measurements. In this study, we investigated the effect of the commonly used reducing agent phenazine methosulfate (PMS) on the RC and fluorescence emission of higher plant PSI-LHCI. We measured the P700(+) reduction rate for different PMS concentrations, and show that we can give a reliable estimation on the fraction of closed RCs based on these rates. The data show that PMS is quenching chlorophyll fluorescence emission. Finally, we determined that the fluorescence quantum yield of PSI with closed RCs is 4% higher than if the RCs are open.

Inactivation of the AMP-activated protein kinase by glucose in cardiac myocytes: a role for the pentose phosphate pathway.

Incubation of adult rat cardiac myocytes with increasing glucose concentrations decreased phosphorylation (αThr172) and activity of AMPK (AMP-activated protein kinase). The effect could be demonstrated without measurable changes in adenine nucleotide contents. The glucose effect was additive to the decrease in AMPK activity caused by insulin, was attenuated by adrenaline, was not mimicked by glucose analogues, lactate or pyruvate and was not due to changes in myocyte glycogen content. AMPK activity was decreased by xylitol and PMS (phenazine methosulfate) and was increased by the glucose-6-phosphate dehydrogenase inhibitor DHEA (dehydroepiandrosterone) and by thiamine. PMS and DHEA respectively, increased and decreased CO2 formation by the PPP (pentose phosphate pathway). AMPK activity was inversely related to the myocyte content of Xu5P (xylulose 5-phosphate), an intermediate of the non-oxidative arm of the PPP. Endothall, an inhibitor of PP2A (protein phosphatase 2A), abolished the glucose effect on AMPK activity. Further studies are needed to define the 'active component' that mediates the glucose effect and whether its site of action is PP2A.

A shared mechanism of SoxR activation by redox-cycling compounds.

In this issue of Molecular Microbiology, Gu and Imlay show that a class of compounds known as redox-cycling agents directly activate the transcription factor SoxR of Escherichia coli and cause cellular toxicity independent of the production of the reactive oxygen species superoxide. Despite the fact that redox-cycling agents increase formation of superoxide in E. coli, the results described in this new publication revise the long-held assumption that superoxide is responsible for the activation of SoxR and for all of the major toxic effects of redox-cycling drugs. This study also suggests that the critical function of the SoxRS regulon in E. coli is in protection against redox-cycling agents and not exclusively the defence against superoxide.

The SoxRS response of Escherichia coli is directly activated by redox-cycling drugs rather than by superoxide.

When Escherichia coli is exposed to redox-cycling drugs, its SoxR transcription factor is activated by oxidation of its [2Fe-2S] cluster. In aerobic cells these drugs generate superoxide, and because superoxide dismutase (SOD) is a member of the SoxRS regulon, superoxide was initially thought to be the activator of SoxR. Its many-gene regulon was therefore believed to comprise a defence against superoxide stress. However, we found that abundant superoxide did not effectively activate SoxR in an SOD⁻ mutant, that overproduced SOD could not suppress activation by redox-cycling drugs, and that redox-cycling drugs were able to activate SoxR in anaerobic cells as long as alternative respiratory acceptors were provided. Thus superoxide is not the signal that SoxR senses. Indeed, redox-cycling drugs directly oxidized the cluster of purified SoxR in vitro, while superoxide did not. Redox-cycling drugs are excreted by both bacteria and plants. Their toxicity does not require superoxide, as they poisoned E. coli under anaerobic conditions, in part by oxidizing dehydratase iron-sulfur clusters. Under these conditions SoxRS induction was protective. Thus it is physiologically appropriate that the SoxR protein directly senses redox-cycling drugs rather than superoxide.

Assessment of oxidant susceptibility of red blood cells in various species based on cell deformability.

The present study was designed to investigate the oxidant susceptibility of red blood cells (RBC) from four species (echidna, human, koala, Tasmanian devil) based on changes in cellular deformability. These species were specifically chosen based on differences in lifestyle and/or biology associated with varied levels of oxidative stress. The major focus was the influence of superoxide radicals generated within the cell (phenazine methosulfate, PMS, 50 µM) or in the extracellular medium (xanthine oxidase-hypoxanthine, XO-HX, 0.1 U/ml XO) on RBC deformability at various shear stresses (SS). RBC deformability was assessed by laser-diffraction analysis using a "slit-flow ektacytometer". Both superoxide-generating treatments resulted in significant increases of methemoglobin for all species (p < 0.01), with Tasmanian devil RBC demonstrating the most sensitivity to either treatment. PMS caused impaired RBC deformability for all species, but vast interspecies variations were observed: human and koala cells exhibited a similar sigmoid-like response to SS, short-beaked echidna values were markedly lower and only increased slightly with SS, while Tasmanian devil RBC were extremely rigid. The effect of XO-HX on RBC deformability was less when compared with PMS (i.e., smaller increase in rigidity) with the exception of Tasmanian devil RBC which exhibited essentially no deformation even at the highest SS; Tasmanian devil RBC response to XO-HX was thus comparable to that observed with PMS. Our findings indicate that ektacytometry can be used to determine the oxidant susceptibility of RBC from different species which varies significantly among mammals representing diverse lifestyles and evolutionary histories. These differences in susceptibility are consistent with species-specific discrepancies between observed and allometrically-predicted life spans and are compatible with the oxidant theory of aging.

Antifungal mechanisms by which a novel Pseudomonas aeruginosa phenazine toxin kills Candida albicans in biofilms.

Pseudomonas aeruginosa produces several phenazines including the recently described 5-methyl-phenazine-1-carboxylic acid (5MPCA), which exhibits a novel antibiotic activity towards pathogenic fungi such as Candida albicans. Here we characterize the unique antifungal mechanisms of 5MPCA using its analogue phenazine methosulphate (PMS). Like 5MPCA, PMS induced fungal red pigmentation and killing. Mass spectrometry analyses demonstrated that PMS can be covalently modified by amino acids, a process that yields red derivatives. Furthermore, soluble proteins from C. albicans grown with either PMS or P. aeruginosa were also red and demonstrated absorbance and fluorescence spectra similar to that of PMS covalently linked to either amino acids or proteins in vitro, suggesting that 5MPCA modification by protein amine groups occurs in vivo. The red-pigmented C. albicans soluble proteins were reduced by NADH and spontaneously oxidized by oxygen, a reaction that likely generates reactive oxygen species (ROS). Additional evidence indicated that ROS generation precedes 5MPCA-induced fungal death. Reducing conditions greatly enhanced PMS uptake by C. albicans and killing. Since 5MPCA was more toxic than other phenazines that are not modified, such as pyocyanin, we propose that the covalent binding of 5MPCA promotes its accumulation in target cells and contributes to its antifungal activity in mixed-species biofilms.

Liver X receptors interfere with cytokine-induced proliferation and cell survival in normal and leukemic lymphocytes.

Liver X receptors (LXRs) are nuclear receptors regulating lipid and cholesterol metabolism. Recent data indicate an additional role of LXR in immunity by controlling dendritic cell and T-cell function and in breast and prostate cancer cells. Here, we show that LXR activation interferes with IL-2 and IL-7-induced proliferation and cell cycle progression of human T-cell blasts mainly through inhibited phosphorylation of the retinoblastoma protein and decreased expression of the cell cycle protein cyclin B. Comparable results were obtained with IL-2-dependent chronic lymphoblastic leukemia (CLL) T cells. Furthermore, we show for B-CLL cells that LXR are functionally active and inhibit expression of survival genes bcl-2 and MMP-9, and significantly reduce cell viability, suggesting an interference of LXR with cytokine-dependent CLL cell survival. In conclusion, our data reveal LXR as a potent modulator of cytokine-dependent proliferation and survival of normal and malignant T and B lymphocytes. This novel LXR action could find clinical application in immunosuppressive and antileukemic therapies.

[Activation of enzymes of soxRS-regulon by hydrogen peroxide in Escherichia coli].

A mutant strain (delta soxR), a derivative of strain of Escherichia coli AB 57, was obtained by general transduction. The authors have investigated the effect of oxidative stress on activity of enzymes of regulons oxyR (catalase, peroxidase, glutathione reductase) and soxRS(superoxide dismutase (SOD) and glucose-6-phosphate dehydrogenase (G6PDH) in the donor strain DJ901, recipient strain AB1157 and transductant. Oxidative stress induced by phenasine metasulfate had no effect on activity of the studied enzymes of regulon oxyR in either used strain, and on enzymes of regulon soxRS in DJ901 transductant, but caused an 1.6- and 1.3-fold increase in activity of SOD and G6PDH in AB 1157, respectively. The stress induced by 20 microM hydrogen peroxide in the last strain increased the enzyme activity of both regulons, and SOD activity increased by 28 % and that of G6PDH--by 42%. Under the same kind of stress only SOD activity increased by 20% in the transductant. Possible ways of regulation of activity in enzymes being the components of regulon soxRS are discussed.

Modulation of RBC volume distributions by oxidants (phenazine methosulfate and tert-butyl hydroperoxide): role of Gardos channel activation.

A study was made comparing the effects of two oxidants--phenazine methosulfate (50-1500 microM)+10 mM ascorbate and t-butyl hydroperoxide (1-3 mM)--on the volume-related parameters of normal human red blood cells. Incubation with either oxidative system for 20-30 min resulted in red blood cell density and osmotic resistance distribution shifts. Treatment with the phenazine methosulfate+ascorbate system in the presence of Ca(2+) led to cell shrinking, with the maximum effect being more than 20%. In contrast, under the same conditions, t-BHP caused cell swelling by up to 15%. Modification of the suspending medium (Ca(2+) removing, clotrimazole addition, or enrichment with K(+)) modulated the redistribution effects, suggesting that they were mediated to some extent by Gardos channel activation. These findings are important for understanding how oxidants modulate RBC cation channels.

Biosensor assay of neuropathy target esterase in whole blood as a new approach to OPIDN risk assessment: review of progress.

Organophosphates (OPs) that inhibit neuropathy target esterase (NTE) with subsequent ageing can produce OP-induced delayed neuropathy (OPIDN). NTE inhibition in lymphocytes can be used as a biomarker of exposure to neuropathic OPs. An electrochemical method was developed to assay NTE in whole blood. The high sensitivity of the tyrosinase carbon-paste biosensors for the phenol produced by hydrolysis of the substrate, phenyl valerate, allowed NTE activity to be measured in diluted samples of whole blood, which cannot be done using the standard colorimetric assay. The biosensor was used to establish correlations of NTE inhibitions in blood with that in lymphocytes and brain after dosing hens with a neuropathic OP. The results of further studies demonstrated that whole blood NTE is a reliable biomarker of neuropathic OPs for up to 96 hours after exposure. These validation results suggest that the biosensor NTE assay for whole blood could be developed to measure human exposure to neuropathic OPs as a predictor of OPIDN. The small blood volume required (100 microL), simplicity of sample preparation and rapid analysis times indicate that the biosensor should be useful in biomonitoring and epidemiological studies. The present paper is an overview of our previous and ongoing work in this area.

Effects of the deficiency of the rhodanese-like protein RhdA in Azotobacter vinelandii.

In Azotobacter vinelandii the rhdA gene codes for a protein (RhdA) of the rhodanese-homology superfamily. By combining proteomics, enzymic profiles and ultrastructural observations, the phenotype of an A. vinelandii rhdA mutant was analyzed. We found that the A. vinelandii rhdA mutant, and not the wild-type strain, accumulated polyhydroxybutyrate. RhdA deficiency enhanced the expression of enzymes of the polyhydroxybutyrate biosynthetic operon, and affected the activity of specific tricarboxylic acid cycle enzymes. The effect was dramatic on aconitase, in spite of comparable expression of aconitase polypeptides in both strains. By using a model system, we found that RhdA triggered protection from oxidants.

Expression of multidrug resistance (MDR) proteins and in vitro drug resistance in acute leukemias.

The expression of the multidrug resistance (MDR) proteins may influence the outcome of treatment in patients with acute leukemia. The aim of this study was to determine the IC50 of cytotoxic drugs (cytosine arabinoside, ara-C and daunorubicin, dnr) using the in vitro 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)2H-tetrazolium, inner salt (MTS) assay method. A total of 82 newly diagnosed acute leukemia cases (43 adult myeloid leukaemia, AML cases and 39 acute lymphoblastic leukaemia, ALL cases) and 16 relapsed cases (8 AML cases and 8 ALL cases) were studied. The MTS assay was performed using two cytotoxic drugs, dnr and ara-C. Cells were incubated with different concentrations of drugs for 4 days and the IC50 was extrapolated from the viability curve. In newly diagnosed cases, we found that childhood ALL samples showed higher IC50 values of dnr (0.040 +/- 2.320) compared to adult AML samples (0.021 +/- 0.158). In contrast, newly diagnosed adult AML samples showed higher IC50 values of ara-C (0.157 +/- 0.529) compared to childhood ALL samples (0.100 +/- 2.350). In relapsed cases, two samples of childhood ALL showed IC50 values of dnr (0.910 +/- 1.760) and ara-C (1.310 +/- 2.390), which was higher compared to childhood AML samples (0.129 +/- 0.214 and 0.210 +/- 0.003, respectively). However, there was no correlation between IC50 values of these drugs tested with clinical outcome. In conclusion, we found that MTS assay is an easy, rapid and non laborious method to study in vitro drug resistance in acute leukaemia cases.