Asya Makhro - Academia.edu (original) (raw)

Papers by Asya Makhro

Frontiers in Physiology, 2016

Transportation of blood samples is unavoidable for assessment of specific parameters in blood of ... more Transportation of blood samples is unavoidable for assessment of specific parameters in blood of patients with rare anemias, blood doping testing, or for research purposes. Despite the awareness that shipment may substantially alter multiple parameters, no study of that extent has been performed to assess these changes and optimize shipment conditions to reduce transportation-related artifacts. Here we investigate the changes in multiple parameters in blood of healthy donors over 72 h of simulated shipment conditions. Three different anticoagulants (K3EDTA, Sodium Heparin, and citrate-based CPDA) for two temperatures (4°C and room temperature) were tested to define the optimal transportation conditions. Parameters measured cover common cytology and biochemistry parameters (complete blood count, hematocrit, morphological examination), red blood cell (RBC) volume, ion content and density, membrane properties and stability (hemolysis, osmotic fragility, membrane heat stability, patch-clamp investigations, and formation of micro vesicles), Ca(2+) handling, RBC metabolism, activity of numerous enzymes, and O2 transport capacity. Our findings indicate that individual sets of parameters may require different shipment settings (anticoagulants, temperature). Most of the parameters except for ion (Na(+), K(+), Ca(2+)) handling and, possibly, reticulocytes counts, tend to favor transportation at 4°C. Whereas plasma and intraerythrocytic Ca(2+) cannot be accurately measured in the presence of chelators such as citrate and EDTA, the majority of Ca(2+)-dependent parameters are stabilized in CPDA samples. Even in blood samples from healthy donors transported using an optimized shipment protocol, the majority of parameters were stable within 24 h, a condition that may not hold for the samples of patients with rare anemias. This implies for as short as possible shipping using fast courier services to the closest expert laboratory at reach. Mobile laboratories or the travel of the patients to the specialized laboratories may be the only option for some groups of patients with highly unstable RBCs.

Please cite this article in press as: A. Makhro, et al., Comparing the impact of an acute exercis... more Please cite this article in press as: A. Makhro, et al., Comparing the impact of an acute exercise bout on plasma amino acid composition, intraerythrocytic Ca 2+ handling, and red cell function in athletes and untrained subjects, Cell Calcium (2016), a b s t r a c t The N-methyl d-aspartate receptors (NMDARs) mediating Ca 2+ uptake upon stimulation with gluta-mate and glycine were recently discovered in red blood cells (RBC) of healthy humans. Activation of these receptors with agonists triggered transient Ca 2+-dependent decrease in hemoglobin oxygen affinity in RBC suspension. The aim of this study was to assess the potential physiological relevance of this phenomenon. Two groups formed by either healthy untrained volunteers or endurance athletes were subjected to a stepwise incremental cycling test to exhaustion. Plasma glutamate levels, activity of the NMDARs, and hemoglobin O 2 affinity were measured in blood samples obtained before and after the exercise in both groups. Increase in plasma glutamate levels following exercise was observed in both groups. Transient Ca 2+ accumulation in response to the NMDAR stimulation with NMDA and glycine was followed by facilitated Ca 2+ extrusion from the RBC and compensatory decrease in cytosolic Ca 2+ levels. Short-term activation of the receptors triggered a transient decrease in O 2 affinity of hemoglobin in both groups. These exercise-induced responses were more pronounced in athletes compared to the untrained subjects. Athletes were initially presented with lower basal intracellular Ca 2+ levels and hemoglobin oxygen affinity compared to non-trained controls. High basal plasma glutamate levels were associated with induction of hemolysis and formation of echinocytes upon stimulation with the receptor agonists. These findings suggest that glutamate release occurring during exhaustive exercise bouts may acutely facilitate O 2 liberation from hemoglobin and improve oxygen delivery to the exercising muscle.

Journal of cardiovascular pharmacology, Jan 10, 2016

This study focuses on characterization of the cardiac N-methyl D-aspartate receptors (NMDARs) as ... more This study focuses on characterization of the cardiac N-methyl D-aspartate receptors (NMDARs) as a target for endogenous and synthetic agonists and antagonists. Using isolated perfused rat hearts, we have shown that intracoronary administration of the NMDAR agonists and antagonists has a pronounced effect on autonomous heart function. Perfusion of rat hearts with autologous blood supplemented with NMDAR agonists was associated with induction of tachycardia, sinus arrhythmia and ischemia occurring within physiological plasma concentration range for glutamate and glycine. Intracoronary administration of the NMDAR antagonists exerted an anti-arrhythmic effect and resulted in bradycardia and improvement of capillary perfusion. Action of antagonists eliprodil, Ro25-6981, memantine, ketamine, and MK-801 on autonomous heart function diverged strikingly from that of L-type Ca channel blockers. Cardiac NMDAR subunit composition differed from that of neuronal receptors and was age- and chambe...

Neurotoxicity Research, Feb 1, 2004

Under extreme conditions, these molecules induce oxidative stress, which may stimulate (or accomp... more Under extreme conditions, these molecules induce oxidative stress, which may stimulate (or accompany) a number of neurodegenerative processes. In the glutamatergic system, ROS levels are under control of ionotropic and metabotropic glutamate receptors, which modulate ion fluxes through the neuronal membrane. The Na + /K + -pump is also one of the important participants affecting stationary ROS levels through several distinct mechanisms. This review describes the involvement of the Na + /K + -pump in intracellular signaling mechanisms via cross-talk between the pump and glutamate receptors in cerebellum granule cells. Selective dysfunction of mGlu II receptors may also lead to abnormal protein phosphorylation (i.e., tau phosphorylation), culminating in neurodegenerative disorders (i.e., Alzheimer disease). Also, unregulated production of intracellular ROS resulting from an imbalance of ionotropic and metabotropic receptors may activate one or more protein kinases. In summary, Glu receptor dysfunction, leading to a deficit in glutamate-mediated signal transduction may represent one of the earliest stages of neurodegenerative disorders. The Na + /K + -pump is able to prevent over-production of intracellular ROS, thus increasing oxidative stability of neuronal cells.

Journal of Biological Chemistry, 2012

Background: Na,K-ATPase activity is extremely sensitive to changes in the redox state. Results: B... more Background: Na,K-ATPase activity is extremely sensitive to changes in the redox state. Results: Binding of glutathione to the regulatory cysteine residues of the catalytic subunit completely inhibits the Na,K-ATPase by blocking the ATP-binding site. Conclusion: S-Glutathionylation of the catalytic subunit is revealed as a mechanism controlling the Na,K-ATPase function. Significance: Regulatory S-glutathionylation adjusts Na,K-ATPase activity to the changes in intracellular redox state and ATP levels.

Interactive Cardiovascular and Thoracic Surgery

Human congenital erythroid disorders are caused by various abnormalities such as hemoglobin polym... more Human congenital erythroid disorders are caused by various abnormalities such as hemoglobin polymorphisms, increase in membrane permeability to ions, cytoskeletal dissociation and malfunction of enzymes involved in energy metabolism and redox state. Beta-globin gene variants giving rise to various hemoglobinopathies (sickle cell disease, beta-thalassemia) are naturally restricted to the regions currently or historically populated by Plasmodium parasite. Protection against malaria in people with sickle cell trait results from the ability of deoxygenated hemoglobin S (Hb) to polymerize as substitution of charged Glu to uncharged Val in position 6 highly reduces its hydrophilicity. Irreversible HbS polymerization is triggered by deoxygenation, and markedly facilitated by dehydration that increases mean corpuscular hemoglobin concentration. This strategy to resist infection (protozoa and viruses) is not restricted to humans. Several fish, reptile and mammalian species are capable of rev...

Precise control over the intracellular free Ca 2+ level is of key importance for any cell, red bl... more Precise control over the intracellular free Ca 2+ level is of key importance for any cell, red blood cells (RBCs) included. Cell fate of erythroid precursor cells, as well as volume, redox state, deformability, aggregability, adherence to the endothelial layer, and membrane stability of circulating RBCs, and as a result RBC longevity, are regulated by Ca 2+ . Several hereditary forms of hemolytic anemia, including sickle cell disease (SCD), are associated with abnormally high Ca 2+ uptake, and as a result, Ca 2+ overload. Multiple organ failure, stroke, and myocardial infarction caused by thrombosis as well as repeated hemolytic and pain crises are induced by irreversible polymerization of deoxygenated hemoglobin S in RBCs of these patients. Dehydration of RBCs speed up sickle cell transformation, extensive oxidation and facilitated proteolysis, as well as increased adherence of circulating RBCs to the endothelial layer, substantially contribute to the sickle cell disease manifestation. As all of these processes are driven by the high intraerythrocytic Ca 2+ levels, inhibition of pathologically high Ca 2+ uptake would be a powerful therapeutic strategy. Molecular identity of ion channels involved in pathologically high permeability of RBC membranes of patients for Ca 2+ is a subject of intensive investigation. Among the identified channels are Piezo channels and N-methyl D-aspartate (NMDA) receptors. The contribution of transient voltage receptor channels (TRPCs), voltage-gated calcium channels Cav2.1, and even voltage-dependent anion channels (VDACs) into the Ca 2+ uptake is currently being debated. This chapter is an overview of the current progress in research in this area followed by an outlook into the potential use of blockers of the cation channels for therapy of SCD patients. 

Recently we showed that N-methyl D-aspartate receptors (NMDARs) are expressed in erythroid precur... more Recently we showed that N-methyl D-aspartate receptors (NMDARs) are expressed in erythroid precursors (EPCs) and present in the circulating red blood cells (RBCs) of healthy humans, regulating intracellular Ca 2+ in these cells. This study focuses on investigating the possible role of NMDARs in abnormally high Ca 2+ permeability in the RBCs of patients with sickle cell disease (SCD). Protein levels of the NMDAR subunits in the EPCs of SCD patients did not differ from those in EPCs of healthy humans. However, the number and activity of the NMDARs in circulating SCD-RBCs was substantially up-regulated, being particularly high during haemolytic crises. The number of active NMDARs correlated negatively with haematocrit and haemoglobin levels in the blood of SCD patients. Calcium uptake via these non-selective cation channels was induced by RBC treatment with glycine, glutamate and homocysteine and was facilitated by de-oxygenation of SCD-RBCs. Oxidative stress and RBC dehydration followed receptor stimulation and Ca 2+ uptake. Inhibition of the NMDARs with an antagonist memantine caused re-hydration and largely prevented hypoxia-induced sickling. The EPCs of SCD patients showed higher tolerance to memantine than those of healthy subjects. Consequently, NMDARs in the RBCs of SCD patients appear to be an attractive target for pharmacological intervention.

Neurochemical Journal, 2008

Prenatal hyperhomocysteinemia induced in rats by overloading of dietary methionine (1 g/kg body m... more Prenatal hyperhomocysteinemia induced in rats by overloading of dietary methionine (1 g/kg body mass daily) results in systemic disordering in progeny related to an increase in the excitotoxic feature of NMDA-receptors in cerebellar neurons and memory suppression. Administration of carnosine (100 mg/kg body mass daily) in the diet of pregnant rats with hyperhomocysteinemia prevents both cognitive function in pups and protects cerebellar neurons from oxidative stress. The effect of carnosine is accompanied by with the restoration of superoxide dismutase in rat brain, which is decreased during hyperhomocysteinemia from 2.07 units (control) to 1.54 units.

Neurochemical Journal, 2010

Incubation of cerebellar neurons with homocysteine or homocysteic acid at a concentration of 500 ... more Incubation of cerebellar neurons with homocysteine or homocysteic acid at a concentration of 500 μM for 30 min increases the intracellular level of free radicals. Specific antagonists of metabotropic glutamate receptors of the groups I (AIDA) and III (MSOP) decreased the accumulation of reactive oxygen species in cells, which suggests that homocysteine and homocysteic acid can interact with these receptors.

Neurochemical Journal, 2007

Homocysteine (HC) and homocysteinic acid (HCA) were capable of elevating the intracellular level ... more Homocysteine (HC) and homocysteinic acid (HCA) were capable of elevating the intracellular level of calcium ions and active oxygen forms (AOF) in rat cerebellum neurons similarly to NMDA. The NMDAreceptor antagonist, MK-801, prevented the rise of both indexes induced by HC or HCA, whereas α -methylcarboxyphenylglycine (an antagonist of metabotropic glutamate receptors of group I) did not interfere with the effect HC or HCA. Chelation of intracellular calcium by 1,2-bis(2-aminophenoxy)ethen-N , N , N ', N '-tetraacetic acid (the BAPTA intracellular calcium buffer) decreased AOF generation during neuron activation by HC or HCA to the control level. NMDA, HC, and HCA induced phosphatidylserine externalization in neurons, which was one of the early stages of apoptosis induced by AOF. Induction of apoptosis by HCA was partially prevented by BAPTA, N-acetylcysteine or cyclosporine A. These facts could be the basis of the neurotoxic effects of HC and its derivatives.

Transfusion, 2015

BACKGROUND: Prolonged storage of red blood cells (RBCs) leads to storage lesions, which may impai... more BACKGROUND: Prolonged storage of red blood cells (RBCs) leads to storage lesions, which may impair clinical outcomes after transfusion. A hallmark of storage lesions is progressive echinocytic shape transformation, which can be partially reversed by washing in albumin solutions. Here we have investigated the impact of this shape recovery on biorheologic variables.

Doklady Biochemistry and Biophysics, 2008

In this work, we demonstrate that homocysteic acid provokes oxidative stress in erythrocytes and ... more In this work, we demonstrate that homocysteic acid provokes oxidative stress in erythrocytes and decreases their hemolytic resistance, whereas the natural antioxidant carnosine [9] protects erythrocytes from its toxic effect. The discovered property can be used for development of new methods for protecting erythrocytes in hyperhomocysteinemia. Homocysteine and homocysteic acid (HCA), the product of spontaneous homocysteine oxidation, are important risk factors for neurodegenerative and cardiovascular diseases [1]. These compounds induce oxidative stress in brain neurons [2, 3] and lymphocytes [4], resulting from their toxic effect on the nervous and immune systems. The prooxidant effect of homocysteine and homocysteic acid on cell structures can be realized via both the glutamate receptors [3, 4] and the activation of NO synthase or inhibition of Na/KATPase, as was demonstrated for the vascular endothelium [5, 6]. It is known that erythrocytes are capable of accumulating homocysteine and excreting it into the extracellular medium [7, 8]. Erythrocytes constitute the main part of blood cells; correspondingly, the toxic effect of homocysteine and HCA may cover these cells as well. However, the effect of these compounds on the erythrocyte stability has not been studied. The goal of this work was to analyze the effect of HCA on human erythrocytes and to assess the effect of carnosine on the oxidative stress in erythrocytes caused by HCA. Various factors can cause erythrocyte hemolysis, including a decreased ambient osmotic pressure, decreased pH, and oxidants [10‐12]. The resistance of erythrocytes to hemolytic action is an integral parameter characterizing their integrity and viability as well as a criterion of their physiologically native state. Two models of hemolysis were used in this work, namely, the osmotic hemolysis and the hemolysis caused by hydrochloric acid. The former takes place with a decrease in the tonicity of medium produced by diluting cell suspension with distilled water. In this process, cells swell with subsequent disruption of the cell membrane. The latter is induced by supplementing cell suspension with hydrochloric acid, which leads to a decrease in the pH in the cytoplasm, impairments in the cytoskeleton structure, and, eventually, cell swelling and destruction [13].

Neurotoxicity Research, 2004

Journal of the American Academy of Dermatology, 2014

European Journal of Cardio-Thoracic Surgery, 2014

OBJECTIVES: A severe donor organ shortage leads to the death of a substantial number of patients ... more OBJECTIVES: A severe donor organ shortage leads to the death of a substantial number of patients who are listed for transplantation. The use of hearts from donors after circulatory death could significantly expand the donor organ pool, but due to concerns about their viability, these are currently not used for transplantation. We propose short-term ex vivo normothermic machine perfusion (MP) to improve the viability of these ischaemic donor hearts.

Bulletin of Experimental Biology and Medicine, 2008

We found that methionine added to the ration of pregnant rats (1 g/kg body weight) induced sustai... more We found that methionine added to the ration of pregnant rats (1 g/kg body weight) induced sustained hyperhomocysteinemia and led to the formation of sustained oxidative stress in the brain of their progeny. Newborn animals were characterized by lower body weight, SOD deficiency in the brain, increased neuronal death, and desensitization of NMDA receptors. These factors are associated with impaired cognitive capacity in the Morris test.