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Free radicals (FRs) are formed in the high amounts result of the metabolic imbalance in cells and tissue. These radicals-induced oxidative damages constitute the basis of many diseases. Organisms have antioxidant defence systems (ADS) to eliminate the destructive effects of the oxidative damage. In addition to these antioxidant systems, dietary flavonoids have the antioxidant effect and the protective role against oxidative damage. In the present study, it was investigated whether a flavonoid derived diosmetin (10, 50, and 100 µM) have the elimination potential on hydrogen peroxide (H2O2)- induced oxidative damage in erythrocyte culture by using biomarkers such as lipid peroxidation (LP) level, catalase (CAT), total superoxide dismutase (SOD) activity and changes of SOD isozymes containing the manganese SOD (Mn SOD) and the cupper-zinc SOD (CuZn SOD). CAT, total SOD, Mn SOD and CuZn SOD activities showed a serious decline with H2O2 treatment, but diosmetin addition significantly increased their activities. While the H2O2 application critically increased LP products in erythrocytes, diosmetin considerably reduced these oxidative damage products. In conclusion, it has been determined that diosmetin can moderate oxidative damage in human erythrocytes by activating or protecting the ADS. [ABSTRACT FROM AUTHOR]

All cells produce carbon dioxide (CO2), which is released as a result of metabolism and must be removed from the body. A large part of this CO2 is converted to bicarbonate by the carbonic anhydrase (CA) enzyme in erythrocytes and is discarded from the body. So, CA has a vital role in red blood cells. In addition to, CA involved in many other pathological and physiological processes and it was determined that the inhibitors of CA were effective in the treatment and diagnosis of many diseases particularly glaucoma. Considering the importance of the CA's inhibitors, in this study it was intended to research the inhibition effects of Eosin Y on CA I and CA II isoenzymes activity purified from human erythrocytes. Eosin Y is a dye molecule commonly used in histological and medical applications. For this purpose, firstly CA I and CA II isoenzymes were purified from human erythrocytes by using Sepharose-4B-L-tyrosine-sulfanilamide affinity chromatography. Then the inhibitory effect of Eosin Y on the activity of these human erythrocyte CA I (hCA I) and CA II (hCA II) isoenzymes was investigated. It was determined that hCA I and hCA II were inhibited by Eosin Y in the millimolar range. IC50 values were found to be 3.78 mM for hCA I and 2.04 mM for hCA II and Ki values were determined as 9.65±0.968 mM and 7.52±2.88 mM for hCA I and hCA II, respectively. In conclusion, it is hoped that the results obtained in this study may be beneficial in the development of new CA inhibitors which may be drug candidates. [ABSTRACT FROM AUTHOR]

Aβ(1-42) peptide is a neurotoxic agent strongly associated with the etiology of Alzheimer’s disease (AD). Current treatments are still of very low effectiveness, and deaths from AD are increasing worldwide. Huprine-derived molecules have a high affinity towards the enzyme acetylcholinesterase (AChE), act as potent Aβ(1-42) peptide aggregation inhibitors, and improve the behavior of experimental animals. AVCRI104P4 is a multitarget donepezil-huprine hybrid that improves short-term memory in a mouse model of AD and exerts protective effects in transgenic Caenorhabditis elegans that express Aβ(1-42) peptide. At present, there is no information about the effects of this compound on human erythrocytes. Thus, we considered it important to study its effects on the cell membrane and erythrocyte models, and to examine its protective effect against the toxic insult induced by Aβ(1-42) peptide in this cell and models. This research was developed using X-ray diffraction and differential scanning calorimetry (DSC) on molecular models of the human erythrocyte membrane constituted by lipid bilayers built of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE). They correspond to phospholipids representative of those present in the external and internal monolayers, respectively, of most plasma and neuronal membranes. The effect of AVCRI104P4 on human erythrocyte morphology was studied by scanning electron microscopy (SEM). The experimental results showed a protective effect of AVCRI104P4 against the toxicity induced by Aβ(1-42) peptide in human erythrocytes and molecular models.

The intracellular antioxidant activities of millet, rice, wheat, brown rice, oat, buckwheat, corn and adlay phenolic extracts were evaluated in an ex vivo erythrocytes system. Buckwheat, corn, and brown adlay free phenolic extracts could effectively attenuate H2O2-induced erythrocyte hemolysis. The protective effects and involved mechanisms of polished adlay, brown adlay and adlay bran phenolic extracts were studied further. The intracellular antioxidant activities of adlay phenolic extracts decreased in the following order: adlay bran > brown adlay > polished adlay. Adlay bran free phenolic extract possessed stronger intracellular antioxidant activity than chlorogenic acid, p-coumaric acid and ferulic acid. The free phenolic extracts were more effective than bound phenolic extracts to suppress hemolysis and plasma oxidation through inhibition of MDA equivalents, MetHb and ROS generation and to restore the activities of Na+K+-ATPase, Ca2+Mg2+-ATPase and intracellular antioxidant enzyme to normal levels. Adlay-rich diets could reduce the incidence of cancer, cardiovascular and neurodegenerative diseases.

Solute translocation by membrane transport proteins is a vital biological process that can be tracked, on the sub-second timescale, using nuclear magnetic resonance (NMR). Fluorinated substrate analogues facilitate such studies because of high sensitivity of 19 F NMR and absence of background signals. Accurate extraction of translocation rate constants requires precise quantification of NMR signal intensities. This becomes complicated in the presence of J-couplings, cross-correlations, and nuclear Overhauser effects (NOE) that alter signal integrals through mechanisms unrelated to translocation. Geminal difluorinated motifs introduce strong and hard-to-quantify contributions from non-exchange effects, the nuanced nature of which makes them hard to integrate into data analysis methodologies. With analytical expressions not being available, numerical least squares fitting of theoretical models to 2D spectra emerges as the preferred quantification approach. For large spin systems with simultaneous coherent evolution, cross-relaxation, cross-correlation, conformational exchange, and membrane translocation between compartments with different viscosities, the only available simulation framework is Spinach. In this study, we demonstrate GLUT-1 dependent membrane transport of two model sugars featuring CF 2 and CF 2 CF 2 fluorination motifs, with precise determination of translocation rate constants enabled by numerical fitting of 2D EXSY spectra. For spin systems and kinetic networks of this complexity, this was not previously tractable., (© 2023 The Authors. ChemBioChem published by Wiley-VCH GmbH.)

Donepezil is used to treat symptomatically the Alzheimer's disease (AD). This drug is a specific inhibitor of the enzyme acetylcholinesterase (AChE), whose main physiological function is to hydrolyze the neurotransmitter acetylcholine. The main objective of this work was to study the effect of donepezil on human erythrocytes as AChE is present in its membrane. For this purpose, human erythrocytes and molecular model of its membrane built-up of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE) were used. The latter correspond to classes of phospholipids present in the outer and inner monolayers of the human erythrocyte membrane, respectively. Our experimental evidences obtained from X-ray diffraction and differential scanning calorimetry (DSC) analysis indicated that donepezil was capable of interacting with both phospholipids. Fluorescence spectroscopy results showed a moderate increase in the fluidity of the hydrophobic tails of DMPC and isolated unsealed human erythrocyte membranes (IUM). On the other hand, results by scanning electron microscopy (SEM) and optical defocusing microscopy (DM) showed that the drug changed the normal biconcave shape of the erythrocytes inducing the formation of stomatocytes (cup-shaped cells). This effect was explained by the incorporation of donepezil molecules into the erythrocyte membrane and interactions with AChE. Unlabelled Image • Donepezil interacted with lipid bilayers present in the human erythrocyte membrane. • Morphological changes in human erythrocytes were induced by donepezil. • Donepezil might interact with the AChE enzyme present in the erythrocyte membrane. [ABSTRACT FROM AUTHOR]

Alzheimer's disease remains largely unknown, and currently there is no complete cure for the disease. New synthetic approaches have been developed to create multi-target agents, such as RHE-HUP, a rhein-huprine hybrid which can modulate several biological targets that are relevant to the development of the disease. While RHE-HUP has shown in vitro and in vivo beneficial effects, the molecular mechanisms by which it exerts its protective effect on cell membranes have not been fully clarified. To better understand RHE-HUP interactions with cell membranes, we used synthetic membrane models and natural models of human membranes. For this purpose, human erythrocytes and molecular model of its membrane built-up of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE) were used. The latter correspond to classes of phospholipids present in the outer and inner monolayers of the human erythrocyte membrane, respectively. X-ray diffraction and differential scanning calorimetry (DSC) results indicated that RHE-HUP was able to interact mainly with DMPC. In addition, scanning electron microscopy (SEM) analysis showed that RHE-HUP modified the normal biconcave shape of erythrocytes inducing the formation of echinocytes. Moreover, the protective effect of RHE-HUP against the disruptive effect of Aβ(1–42) on the studied membrane models was tested. X-ray diffraction experiments showed that RHE-HUP induced a recovery in the ordering of DMPC multilayers after the disruptive effect of Aβ(1–42), confirming the protective role of the hybrid. [Display omitted] • Aβ(1–42) induces neurotoxicity in neurons, playing a crucial role in Alzheimer's disease. • RHE-HUP attenuates the effects of Aβ(1–42) in lipid bilayer models. • RHE-HUP normalizes X-ray diffraction of Aβ(1–42) disrupted phospholipids. • RHE-HUP protects human erythrocyte cell membranes from Aβ(1–42)-induced damage. [ABSTRACT FROM AUTHOR]

Labetalol is one of the most used drugs for the treatment of hypertension. This molecule is able to bind to both alpha-1 (α1) and beta (β) adrenergic receptors present in vascular smooth muscle among other tissues. It has been determined that human erythrocytes possess both alpha receptors and beta-adrenergic receptors expressed on their surface. The objective of this work was to study the effect of labetalol on the morphology of human erythrocytes. To accomplish this goal, human erythrocytes and model membranes built of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE) were used. These lipid species are present in the outer and inner monolayers of the red blood cell membrane, respectively. Our findings obtained by X-ray diffraction and differential scanning calorimetry (DSC) indicate that labetalol interacted with both lipids in a process dependent on concentration. In fact, at low concentrations labetalol preferentially interacted with DMPE. On the other hand, results obtained by scanning electron microscopy (SEM) showed that labetalol alters the normal biconcave form of erythrocytes to stomatocytes and knizocytes (cells with one or more cavities, respectively). According to the bilayers couple hypothesis, this result implied that the drug inserted in the inner monolayer of the human erythrocyte membrane. [ABSTRACT FROM AUTHOR]

Diabetes mellitus is a metabolic affliction saunter that is characterized by a nobler than normal blood glucose poise. Glucose-6-phosphate dehydrogenase (G6PD) enzyme code (E.C.1.1.1.49) is an underlying enzyme in the phosphogluconate pathway. In this study, G6PD vitality in the mortal erythrocyte of male and female patients with type 2 diabetes mellitus was assessed utilizing a spectrophotometer at 340 nm. The activity of the enzyme increased with elevated glycated haemoglobin (HbA1C) levels. G6PD activity was found to be significantly associated with type 2 diabetes mellitus. The association between G6PD and diabetes mellitus was significant (P < 0.001). Moreover, G6PD was positively correlated with HbA1C levels (r = 0.572). The following mean ± standard deviation values were obtained: G6PD activity (IU/g Hb), 3.1103 ± 0.79349; HbA1C (%), 8.6600 ± 1.63120; Hb (g/dL), 13.4933 ± 1.38836; platelet count (103/μl), 283.4667 ± 58.59312; WBC (103/μl), 7.4890 ± 1.49842; HCT (%), 45.0100 ± 2.63430; and BS (mg/dL), 230.2667 ± 75.67760. The results showed that an elevated HbA1C up leads to increased G6PD performance in the human erythrocyte, which is concerning to glucose levels in the special (blood). [ABSTRACT FROM AUTHOR]

Summary: Maca is known to be rich in macamides. The objective of this study was to analyse macamide profiles of macas with different colours cultivated in Yunnan, China, and commercial Peru maca by ultra‐high‐performance liquid chromatography–tandem mass spectrometry. The antioxidant activities of water, citric acid and alkali‐extracted polysaccharides from Yunnan and Peru macas were evaluated via DPPH radical scavenging activity, oxygen radical absorbance capacity and erythrocyte haemolysis inhibition effect assays. Six major macamides present in Peru maca were also found dominant in Yunnan maca at comparable concentrations. Maca polysaccharide effectively scavenging DPPH and peroxyl radicals protected the erythrocytes against H2O2‐induced haemolysis by inhibition of malondialdehyde (MDA) generation. Citric acid extraction was found the most suitable preparation method for generating heteropolysaccharide with strong antioxidant activity. Black maca cultivated in Yunnan possessing the most abundant macamides and polysaccharide with strong antioxidant activity could be developed as functional foods. [ABSTRACT FROM AUTHOR]

Two cytotoxic copper(II) complexes with N-H and N -methylated benzimidazole-derived ligands (Cu- L 1 and Cu- L 1Me ; L 1 = bis(2-methylbenzimidazolyl)(2-methylthioethyl)amine, L 1Me = bis(1-methyl-2-methylbenzimidazolyl)(2-methylthioethyl)amine) were synthesized and exposed to human erythrocytes and molecular models of its membrane. The latter were bilayers built-up of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE), classes of lipids present in the external and internal moieties of the human red cell membrane, respectively. Scanning electron microscopy (SEM) of erythrocytes incubated with solutions of both Cu(II) complexes showed that they induced morphological changes to the normal cells to echinocytes, and hemolysis at higher concentrations. Real-time observation of the dose-dependent effects of the complexes on live erythrocytes by defocusing microscopy (DM) confirmed SEM results. The formation of echinocytes implied that complex molecules inserted into the outer moiety of the red cell membrane. X-ray diffraction studies on DMPC and DMPE showed that none of these complexes interacted with DMPE and only Cu- L 1 interacted with DMPC. This difference was explained by the fact that Cu- L 1Me complex is more voluminous than Cu- L 1 because it has two additional methyl groups; on the other hand, DMPC molecule has three methyl groups in its bulky terminal amino end. Thus, by steric hindrance Cu- L 1Me molecules cannot intercalate into DMPC bilayer, which besides is present in the gel phase. These results, together with the increased antiproliferative capacity of the N -methylated complex Cu- L 1Me over that of Cu- L 1 are rationalized mainly based on its higher lipophilicity. [ABSTRACT FROM AUTHOR]

The antioxidant and antihemolytic properties contained in the leaves of Buddleja globosa (B. globosa), also known as "Matico," were determined. Aqueous extracts of leaves were assayed in human erythrocytes and molecular models of its membrane. The latter were bilayers built-up of lipids located in the outer and inner leaflets of the erythrocyte membrane. Observations by scanning electron microscopy showed that the extract altered the morphology of erythrocytes inducing the formation of crenated echinocytes. This result implied that the extract components were inserted into the outer leaflet of the cell membrane. This conclusion was confirmed by experiments carried out by fluorescence spectroscopy of red cell membranes and vesicles (LUV) of dimyristoylphosphatidylcholine (DMPC) and by X-ray diffraction of DMPC and dimyristoylphosphatidylethanolamine bilayers. Human erythrocytes were in vitro exposed to HClO, which is a natural powerful oxidant. Results demonstrated that low concentrations of B. globosa aqueous extract neutralized the harmful capacity of HClO. Hemolysis experiments also showed that the extract in very low concentrations reduced hemolysis induced by HClO. [ABSTRACT FROM AUTHOR]

The intracellular antioxidant activities of millet, rice, wheat, brown rice, oat, buckwheat, corn and adlay phenolic extracts were evaluated in an ex vivo erythrocytes system. Buckwheat, corn, and brown adlay free phenolic extracts could effectively attenuate H 2 O 2 -induced erythrocyte hemolysis. The protective effects and involved mechanisms of polished adlay, brown adlay and adlay bran phenolic extracts were studied further. The intracellular antioxidant activities of adlay phenolic extracts decreased in the following order: adlay bran > brown adlay > polished adlay. Adlay bran free phenolic extract possessed stronger intracellular antioxidant activity than chlorogenic acid, p -coumaric acid and ferulic acid. The free phenolic extracts were more effective than bound phenolic extracts to suppress hemolysis and plasma oxidation through inhibition of MDA equivalents, MetHb and ROS generation and to restore the activities of Na + K + -ATPase, Ca 2+ Mg 2+ -ATPase and intracellular antioxidant enzyme to normal levels. Adlay-rich diets could reduce the incidence of cancer, cardiovascular and neurodegenerative diseases. [ABSTRACT FROM AUTHOR]

Memantine is a NMDA antagonist receptor clinically used for treating Alzheimer's disease. NMDA receptors are present in the human neurons and erythrocyte membranes. The aim of the present study was to investigate the effects of memantine on human erythrocytes. With this purpose, the drug was developed to in vitro interact with human red cells and bilayers built-up of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE). The latter represent lipids respectively present in both outer and inner monolayers of the red cell membrane. Results obtained by scanning electron microscopy (SEM) showed that memantine changed the normal biconcave shape of red cells to cup-shaped stomatocytes. According to the bilayer-couple hypothesis the drug intercalated into the inner monolayer of the erythrocyte membrane. Experimental results obtained by X-ray diffraction on multibilayers of DMPC and DMPE, and by differential scanning calorimetry on multilamellar vesicles indicated that memantine preferentially interacted with DMPC in a concentration-dependent manner. Thus, it can be concluded that in the low therapeutic plasma concentration of circa 1 μM memantine is located in NMDA receptor channel without affecting the erythrocyte shape. However, at higher concentrations, once the receptors became saturated excess of memantine molecules (20 μM) would interact with phosphoinositide lipids present in the inner monolayer of the erythrocyte membrane inducing the formation of stomatocytes. However, 40–50 μM memantine was required to interact with isolated phosphatidylcholine bilayers. [ABSTRACT FROM AUTHOR]

After red blood cells lysis, hemoglobin is released to blood circulation. Hemoglobin is carried in blood by binding to haptoglobin. In normal individuals, no free hemoglobin is observed in the blood, because most of the hemoglobin is in the form of haptoglobin complex. In some diseases that are accompanied by hemolysis, the amount of released hemoglobin is higher than its complementary haptoglobin. As a result, free hemoglobin appears in the blood, which is a toxic compound for these patients and may cause renal failure, hypertensive response and risk of atherogenesis. Free hemoglobin has been determined to have peroxidase activity and considered a pseudoenzyme. In this study, the effect of methocarbamol on the peroxidase activity of human hemoglobin was investigated. Our results showed that the drug inhibited the pseudoenzyme by un-competitive inhibition. Both K m and V max decreased by increasing the drug concentration. K i and IC 50 values were determined as 6 and 10 mM, respectively. Docking results demonstrated that methocarbamol did not attach to heme group directly. A hydrogen bond linked NH 2 of carbamate group of methocarbamol to the carboxyl group of Asp126 side chain. Two other hydrogen bonds could be also observed between hydroxyl group of the drug and Ser102 and Ser133 residues of the pseudoenzyme. [ABSTRACT FROM AUTHOR]

Nuclear magnetic resonance (NMR) magnetization-transfer (MT) experiments provide a convenient tool for studying rapid sub-second membrane-transport processes in situ in metabolically active cells. These experiments are used with membrane-permeable substances when separate (resolved) NMR signals are observed from their populations inside and outside the cells. Here, we provide a description of the theory and practice of the most common NMR MT experiments that have been used to study membrane-transport processes in human erythrocytes (red blood cells; RBCs). The procedures, involved in the analysis of the experimental data for defining mechanisms of transport, and for estimating values of kinetic parameters in the corresponding mathematical models, are given special attention. [ABSTRACT FROM AUTHOR]