Red Cell Protein Research Articles

Opportunistic infections changed before and after SARS-CoV-2 infection in inflammatory bowel disease patients: a retrospective single-center study in China.

SARS-CoV-2 transmission has become a serious worldwide public health concern. However, there is currently insufficient data to determine whether SARS-CoV-2 infection would affect opportunistic infections in inflammatory bowel disease (IBD) patients. A retrospective study included 451 IBD patients (294 UC and 157 CD). The IBD patients were divided into two groups: before SARS-CoV-2 infection and after SARS-CoV-2 infection, and outcomes were measured for these groups. The primary outcome was the presence and distribution of opportunistic infections. The secondary outcomes included factors associated with opportunistic infections, based on which a nomogram prediction model was developed and validated. After SARS-CoV-2 infection, the proportion of IBD patients with opportunistic infections by Clostridium difficile (21.31% vs. 14.01%, p = 0.044) and Epstein-Barr virus (13.93% vs. 4.35%, p = 0.001) was significantly higher compared to that before. Conversely, the proportion of patients with hepatitis B virus (3.69% vs. 10.14%, p = 0.006) and herpes simplex virus type I (1.23% vs. 4.35%, p = 0.04) infections was significantly lower after the infection. Additionally, pre-SARS-CoV-2 infection factors associated with opportunistic infections in IBD include duration of illness, red blood cell count, the presence of comorbid chronic illnesses, and alcohol consumption, while post-SARS-CoV-2 infection, the primary risk factors involve corticosteroid use, red blood cell count, protein level, and high-sensitivity C-reactive protein. After the SARS-CoV-2 infection, there has been a shift in the occurrence of opportunistic infections among IBD patients. It might be attributed to the use of corticosteroids and also the strengthening of containment measures, heightened public health awareness, and widespread vaccination.

WITHDRAWN: Recent Advances in the Microbial Synthesis of Human Hemoglobin

Hemoglobin is a major binding protein in human red blood cells and plays an important role in the storage and transportation of oxygen. In recent years, the use of recombinant hemoglobin as a new type of oxygen carrier has attracted widespread attention. So far, the research on recombinant hemoglobin technology is in its early stages and there are still many problems, such as low yield and easy degradation. In this review, we summarize the molecular strategies for recombinant synthesis of human hemoglobin in microorganisms through metabolic engineering and synthetic biology techniques in recent years, and explore the possibility of preparing hemoglobin-based oxygen carriers based on fetal hemoglobin, in order to improve the yield of recombinant human hemoglobin and the sustained stability during protein production through protein engineering strategies, and to possibly reduce the toxic side effects of recombinant hemoglobin products.

Role of Quercetin Supplementation on Iron Parameters in Blood Transfusion-Dependent Thalassemia Patients

Background: Thalassemia is a group of inherited blood disorders that affect the production of hemoglobin, a protein in red blood cells that carries oxygen throughout the body. Iron overload is a condition in which the body absorbs and stores too much iron. In addition to repeated blood transfusions, increased gastrointestinal tract (GIT) iron absorption plays an important role in iron overload with thalassemia. Quercetin, a common flavonoid present in fruits and vegetables, exhibits diverse biological effects. Objective: To assess the effect of quercetin on iron overload parameters in blood transfusion-dependent thalassemia patients (TDT). Methods: A randomized, double-blind, placebo-placebo group-led study was conducted on 110 TDT patients, more than 12 years of age, who were supplemented with either quercetin or a placebo capsule daily (500 mg) for 3 months. A blood sample was obtained for laboratory parameters at baseline and at the end of 3 months. Results: At the baseline time of the study, the demographic features and iron overload parameters of patients and the placebo group were not statistically different, while after three months of supplementation, there was a significant decrease in levels of serum iron, UIBC, serum ferritin and ferritin saturation rate, and a significant increase in TIBC in the patients compared with the placebo group. Conclusions: The study shows the significant role of quercetin on iron overload parameters in blood transfusion-dependent thalassemia patients.

Effect of Degree of Smoking Based on Brinkman Index on Hemoglobin Levels in Adults

Cigarette consumption is the main cause of death in the world because it affects various physiological conditions in the body. Various ingredients in cigarette smoke such as tar, nicotine, carbon monoxide, and others can cause health problems, especially in the O2 transportation system. Hemoglobin is a protein in red blood cells that carries oxygen from the lungs to all tissues. The carbon monoxide contained in cigarette smoke is competitive with O2 in binding to hemoglobin so that the oxygen distributed to the tissues is reduced. The body will compensate for peripheral hypoxia that occurs by increasing hemoglobin levels in the body so that oxygen transport to the tissues can be maintained. Research regarding the relationship between the degree of smoking and hemoglobin levels is still controversial. Analytical research with cross-sectional design. A total of 184 respondents were taken using purposive sampling and it was found that the majority (94%) were men who worked as security officers, cleaners, parking attendants, or technicians. Based on the Brinkman Index, 73.9% of respondents were classified as light smokers, only 1.6% were classified as heavy smokers and 50.5% of respondents had low hemoglobin levels. Analysis showed that the majority of respondents in the light smoker group had normal hemoglobin levels (58.1%), while moderate and heavy smokers had lower hemoglobin levels. In statistical analysis, a significant p-value (0.001) was found between the degree of smoking and hemoglobin levels.

Microstrip isoelectric focusing with deep learning for simultaneous screening of diabetes, anemia, and thalassemia

BackgroundHemoglobin (Hb) is an important protein in red blood cells and a crucial diagnostic indicator of diseases, e.g., diabetes, thalassemia, and anemia. However, there is a rare report on methods for the simultaneous screening of diabetes, anemia, and thalassemia. Isoelectric focusing (IEF) is a common separative tool for the separation and analysis of Hb. However, the current analysis of IEF images is time-consuming and cannot be used for simultaneous screening. Therefore, an artificial intelligence (AI) of IEF image recognition is desirable for accurate, sensitive, and low-cost screening. ResultsHerein, we proposed a novel comprehensive method based on microstrip isoelectric focusing (mIEF) for detecting the relative content of Hb species. There was a good coincidence between the quantitation of Hb via a conventional automated hematology analyzer and the one via mIEF with R2 = 0.9898. Nevertheless, our results showed that the accuracy of disease diagnosis based on the quantification of Hb species alone is as low as 69.33 %, especially for the simultaneous screening of multiple diseases of diabetes, anemia, alpha-thalassemia, and beta-thalassemia. Therefore, we introduced a ResNet1D-based diagnosis model for the improvement of screening accuracy of multiple diseases. The results showed that the proposed model could achieve a high accuracy of more than 90 % and a good sensitivity of more than 96 % for each disease, indicating the overwhelming advantage of the mIEF method combined with deep learning in contrast to the pure mIEF method. SignificanceOverall, the presented method of mIEF with deep learning enabled, for the first time, the absolute quantitative detection of Hb, relative quantitation of Hb species, and simultaneous screening of diabetes, anemia, alpha-thalassemia, and beta-thalassemia. The AI-based diagnosis assistant system combined with mIEF, we believe, will help doctors and specialists perform fast and precise disease screening in the future.

Protein fibril aggregation on red blood cells: a potential biomarker to distinguish neurodegenerative diseases from healthy aging.

Neurodegenerative diseases like Alzheimer's disease are characterized by the accumulation of misfolded proteins into fibrils in the brain. Atomic force microscopy is a nanoscale imaging technique that can be used to resolve and quantify protein aggregates from oligomers to fibrils. Recently, we characterized protein fibrillar aggregates adsorbed on the surface of red blood cells with atomic force microscopy from patients with neurocognitive disorders, suggesting a novel Alzheimer's disease biomarker. However, the age association of fibril deposits on red blood cells has not yet been studied in detail in healthy adults. Here, we used atomic force microscopy to visualize and quantify fibril coverage on red blood cells in 50 healthy adults and 37 memory clinic patients. Fibrillar protein deposits sporadically appeared in healthy individuals but were much more prevalent in patients with neurodegenerative disease, especially those with Alzheimer's disease as confirmed by positive CSF amyloid beta 1-42/1-40 ratios. The prevalence of fibrils on the red blood cell surface did not significantly correlate with age in either healthy individuals or Alzheimer's disease patients. The overlap in fibril prevalence on red blood cells between Alzheimer's disease and amyloid-negative patients suggests that fibril deposition on red blood cells could occur in various neurodegenerative diseases. Quantifying red blood cell protein fibril morphology and prevalence on red blood cells could serve as a sensitive biomarker for neurodegeneration, distinguishing between healthy individuals and those with neurodegenerative diseases. Future studies that combine atomic force microscopy with immunofluorescence techniques in larger-scale studies could further identify the chemical nature of these fibrils, paving the way for a comprehensive, non-invasive biomarker platform for neurodegenerative diseases.

Building Synthetic Biosensors Using Red Blood Cell Proteins.

As the use of engineered cell therapies expands from pioneering efforts in cancer immunotherapy to other applications, an attractive but less explored approach is the use of engineered red blood cells (RBCs). Compared to other cells, RBCs have a very long circulation time and reside in the blood compartment, so they could be ideally suited for applications as sentinel cells that enable in situ sensing and diagnostics. However, we largely lack tools for converting RBCs into biosensors. A unique challenge is that RBCs remodel their membranes during maturation, shedding many membrane components, suggesting that an RBC-specific approach may be needed. Toward addressing this need, here we develop a biosensing architecture built on RBC membrane proteins that are retained through erythropoiesis. This biosensor employs a mechanism in which extracellular ligand binding is transduced into intracellular reconstitution of a split output protein (including either a fluorophore or an enzyme). By comparatively evaluating a range of biosensor architectures, linker types, scaffold choices, and output signals, we identify biosensor designs and design features that confer substantial ligand-induced signal in vitro. Finally, we demonstrate that erythroid precursor cells engineered with our RBC-protein biosensors function in vivo. This study establishes a foundation for developing RBC-based biosensors that could ultimately address unmet needs including noninvasive monitoring of physiological signals for a range of diagnostic applications.

Pyruvate kinase deficiency and PKLR gene mutations: Insights from molecular dynamics simulation analysis

Pyruvate kinase deficiency is a rare hereditary erythrocyte enzyme disease caused by mutations in the pyruvate kinase liver and red blood cell gene. The clinical presentations of pyruvate kinase deficiency are significantly heterogeneous, ranging from just mild anemia to hemolytic crisis or even death. The proband in our study was a 2-year-old girl for severe skin and scleral icterus with progressive aggravation. We collected the family's data for further analysis. Whole exome genome sequencing of the pedigree revealed a novel compound heterozygous mutation, c.1097del (p.P366Lfs*12) and c.1493G > A (p.R498H), in the pyruvate kinase liver and red blood cell gene. Furthermore, molecular dynamics simulations were employed to uncover differences between the wild type and mutant pyruvate kinase liver and red blood cell proteins, focusing on structural stability, protein flexibility, secondary structure, and overall conformation. The combined bioinformatic tools were also utilised to assess the effects of the missense mutation on protein function. Thereafter, wild type and mutant plasmids were constructed and transfected into 293T cells, and Western blot assay was conducted to validate the impact of the mutations on the expression of pyruvate kinase liver and red blood cell protein. The data presented in our study enriches the genotype database and provides evidence for genetic counseling and molecular diagnosis of pyruvate kinase deficiency.

Pharmacologic Therapies in Beta Hemoglobinopathies: Fetal Globin Gene Induction in the First Molecular Diseases

Beta hemoglobinopathies and thalassemias are caused by diverse molecular mutations of the βA globin chains and modulated by polymorphisms. These are important disorders in medical history, as they became prevalent globally in regions where P. falciparum malaria was endemic. Heterozygous or carrier states conferred a survival advantage; however, doubly heterozygous or homozygous states cause severe hemolytic anemia and multi-organ complications. These disorders are somewhat unique in that all humans have alternate genes for fetal globin chains, which are expressed in fetal life but are silenced on a developmental clock in infancy. An established approach to ameliorating conditions of abnormal adult globin genes is to reactivate, or increase expression, of the endogenous fetal globin genes to replace the missing protein chains in beta thalassemias or inhibit polymerization of hemoglobin S and reduce the red cell abnormalities. This review provides a high-level overview of cell and molecular mechanisms that mediate fetal to adult globin gene switching and pharmacologic therapies that can reactivate fetal globin through different actions. Intermittent or metronomic dosing regimens have overcome challenges of undesirable off-target effects by agents that cause erythroid cell growth arrest. Multiple pharmacologic candidates reactivate or increase the expression of fetal globin protein and proportions of red blood cells containing HbF (F-cells). Hydroxyurea maintains high levels of HbF if begun in infancy, with some decline in mid-childhood. It elicits lower responses in HbF in adult patients, but still has broad clinical benefit in reducing many complications. However, many adult patients do not tolerate optimal hydroxyurea dosing due to cytopenias. Parenterally administered therapies with differing molecular actions, such as demethylating agents and histone deacetylase inhibitors, have shown proof-of-principle in reactivating HbF. Orally bioavailable candidates with complementary molecular mechanisms are in trials. Combining fetal globin-inducing agents with other therapies with complementary mechanisms, such as recombinant erythropoietin that promotes the survival of red blood cells and therapeutics that promote erythroid cell metabolism, should have additive effects. These pharmaceutical candidates offer great clinical potential and global feasibility for ameliorating these serious diseases.

Peran Tenaga Kesehatan Dalam Menurunkan Prevalensi Anemia Pada Remaja, dan Faktor Yang Mempengaruhi Terjadinya Anemia

Anemia is a disease characterized by lower-than-normal hemoglobin (Hb) levels in the blood. Hemoglobin is a protein in red blood cells that functions to transport oxygen from the lungs to the rest of the body. Based on the research findings, it is known that the role of Posyandu adolescents is to provide health services by health workers. The services provided are in the form of facilitators who will conduct health checks and refer adolescent girls diagnosed with anemia to the Puskesmas for hemoglobin testing. Since adolescent girls are at risk of anemia during menstruation, Posyandu Remaja needs to motivate them to take blood product tablets and undergo regular health checks every month. This study uses a literature review methodology that focuses on themes or variables obtained from previous research, namely written research (Wahyudi, 2022). Data regarding the role of health workers in reducing the prevalence of anemia in adolescents and factors influencing the occurrence of anemia were identified from research articles and literature reviews. Similar to Bangladesh and India, the prevalence of anemia in India is very high at 45.9%. One of the main factors causing anemia in adolescent girls is improper nutritional habits.

Abstract 12733: The Effect of Mavacamten Treatment on Hemolysis Biomarkers in Patients With Obstructive Hypertrophic Cardiomyopathy in the EXPLORER-HCM Study

Introduction: In patients with obstructive hypertrophic cardiomyopathy (HCM), blood flow changes from left ventricular outflow tract (LVOT) obstruction can lead to hemolysis and release of red blood cell proteins into circulation. Increased cell-free plasma hemoglobin (Hb) and reduced haptoglobin (Hp) are biomarkers of intravascular hemolysis. Mavacamten, a cardiac myosin inhibitor, reduced LVOT obstruction in the phase 3 EXPLORER-HCM study; however, its effect on hemolysis biomarkers is unknown. Aim: To assess the effect of 30 weeks of mavacamten treatment versus placebo on levels of intravascular hemolysis biomarkers in patients with obstructive HCM. Methods: Hemolysis biomarkers were measured as part of exploratory proteomic analyses from randomized EXPLORER-HCM (NCT03470545) patients at baseline and Week 30 (mavacamten, n = 115; placebo, n = 120) using a proteomic platform (SomaScan v4.1). Results: At Week 30, mavacamten treatment showed decreases in cell-free plasma Hb (Log 2 fold change = -1.57, FDR = 1.7х10 -4 ) and increases in cell-free plasma Hp (Log 2 fold change = 1.83, FDR = 1.2х10 -4 ) vs placebo (Figure). Hb and Hp levels were negatively correlated at baseline and at Week 30. In general, patients can be grouped into high- or low- hemolysis biomarker profile (high Hb with low Hp or low Hb with high Hp, respectively) at baseline and Week 30. Patients who received mavacamten and changed from high- to low hemolysis biomarker profile at Week 30 trended towards greater LVOT reductions under resting and Valsalva conditions. No changes were detected for either group in whole blood Hb or hematocrit. Conclusions: Reduced cell-free plasma Hb and increased cell-free plasma Hp levels, suggest a reduction in intravascular hemolysis by mavacamten in obstructive HCM. These results are consistent with improved hemodynamic effects (less outflow obstruction) and indicate that mavacamten may reduce shear forces on red blood cells in the heart.

Changes in Hemoglobin Properties in Complex with Glutathione and after Glutathionylation.

Hemoglobin is the main protein of red blood cells that provides oxygen transport to all cells of the human body. The ability of hemoglobin to bind the main low-molecular-weight thiol of the cell glutathione, both covalently and noncovalently, is not only an important part of the antioxidant protection of red blood cells, but also affects its affinity for oxygen in both cases. In this study, the properties of oxyhemoglobin in complex with reduced glutathione (GSH) and properties of glutathionylated hemoglobin bound to glutathione via an SS bond were characterized. For this purpose, the methods of circular dichroism, Raman spectroscopy, infrared spectroscopy, tryptophan fluorescence, differential scanning fluorimetry, and molecular modeling were used. It was found that the glutathionylation of oxyhemoglobin caused changes in the secondary structure of the protein, reducing the alpha helicity, but did not affect the heme environment, tryptophan fluorescence, and the thermostability of the protein. In the noncovalent complex of oxyhemoglobin with reduced glutathione, the secondary structure of hemoglobin remained almost unchanged; however, changes in the heme environment and the microenvironment of tryptophans, as well as a decrease in the protein's thermal stability, were observed. Thus, the formation of a noncovalent complex of hemoglobin with glutathione makes a more significant effect on the tertiary and quaternary structure of hemoglobin than glutathionylation, which mainly affects the secondary structure of the protein. The obtained data are important for understanding the functioning of glutathionylated hemoglobin, which is a marker of oxidative stress, and hemoglobin in complex with GSH, which appears to deposit GSH and release it during deoxygenation to increase the antioxidant protection of cells.

Physicochemical evaluation of interaction behavior of a series of biocompatible gemini surfactants with hemoglobin: Insights from spectroscopic and computational studies

In this study, we investigated the interaction behavior of biodegradable, cationic, oxy-diester functionalized gemini surfactants N1,N1,N14,N14-tetramethyl-2,13-dioxo-N1,N14-dialkyl-3,6,12-tetraoxateradecane-1,14-diaminium dichloride (in short Cm-E2O2-Cm, m = 12, 14 or 16 is the number of carbon atoms in the alkyl chain) with one of the most important proteins in red blood cells viz. hemoglobin. We employed a variety of in-vitro and in-silico methodologies, such as fluorescence, UV–vis, FT-IR and circular dichroism spectroscopies, DFT, molecular docking, and molecular dynamics simulations, to study these interactions. Our results show that the chain length of the surfactants have distinct implications for the interaction behavior, as evidenced by the estimated physical parameters, such as Kb and ∆G. The order of Kb values (0.24 × 103 < 3.11 ×103 < 5.85 × 103 M-1) is in accordance with the increasing alkyl chain length of the surfactants. The negative Gibbs free energy change values and their order, − 13.58 < −19.93 < −21.49, further support the spontaneity of interactions and the effect of alkyl chain length. Theoretical insight into the environment at the quantum mechanical level, binding site, and energy stability of the protein-surfactant interactions were produced by computational methods. This research contributes significantly to the field of knowledge concerning surfactants' effects on protein structure and behavior.

Multispectroscopic studies of binding interaction of phosmet with bovine hemoglobin

Phosmet is a phthalimide derived broad spectrum organophosphate pesticide which is vastly used across the globe to protect several ornamental or horticulture crops. The toxicity of phosmet is of utmost concern because of its direct effect on the nervous system of the victim after exposure. The mechanism of phosmet toxicity was explored by the interaction with the model blood protein which is hemoglobin. Bovine Hemoglobin (BHb) is a major protein of red blood cells (RBCs) that plays an important role in the exchange of gases for respiration and ensures adequate oxygen supply to tissues for oxygenation. In the current study, the interaction of BHb with phosmet was revealed using various spectroscopic techniques. Circular dichroism (CD) and Fourier transform infrared spectroscopy (FTIR) studies of BHb in the presence of phosmet showed secondary structural changes in the protein post binding, Fluorescence study shows the involvement of the dynamic quenching predominantly, Van't Hoffs thermodynamic study showed negative enthalpy value and free energy change and negative entropy change that revealed the involvement of hydrogen bonding and van der Waal forces predominantly further revealing spontaneous nature of binding interaction. The shift in Ultraviolet–visible spectra also revealed the nature of the interaction. In-silico study finally deduced the involvement of hydrogen bonding and polar interaction. The study inferred the moderate interaction of BHb with phosmet.

Immunostaining-Based Detection of Dynamic Alterations in Red Blood Cell Proteins.

Antibody labeling of red blood cell (RBC) proteins is a commonly used, semi-quantitative method to detect changes in overall protein content or acute alterations in protein activation states. It facilitates the assessment of RBC treatments, characterization of differences in certain disease states, and description of cellular coherencies. The detection of acutely altered protein activation (e.g., through mechanotransduction) requires adequate sample preparation to preserve otherwise temporary protein modifications. The basic principle includes immobilizing the target binding sites of the desired RBC proteins to enable the initial binding of specific primary antibodies. The sample is further processed to guarantee optimal conditions for the binding of the secondary antibody to the corresponding primary antibody. The selection of non-fluorescent secondary antibodies requires additional treatment, including biotin-avidin coupling and the application of 3,3-diaminobenzidine-tetrahydrochloride (DAB) to develop the staining, which needs to be controlled in real-time under a microscope in order to stop the oxidation, and thus staining intensity, on time. For staining intensity detection, images are taken using a standard light microscope. In a modification of this protocol, a fluorescein-conjugated secondary antibody can be applied instead, which has the advantage that no further development step is necessary. This procedure, however, requires a fluorescence objective attached to a microscope for staining detection. Given the semi-quantitative nature of these methods, it is imperative to provide several control stains to account for non-specific antibody reactions and background signals. Here, we present both staining protocols and the corresponding analytical processes to compare and discuss the respective results and advantages of the different staining techniques.

In Vitro and In Vivo Anti-Inflammatory Properties of the Hydroethanolic Extract of the Roots of Vernonia guineensis (Asteraceae).

In traditional Cameroonian medicine, to relieve many inflammations, parts of Vernonia guineensis, are very widely used. This study considered the evaluation of acute toxicity and anti-inflammatory properties of the hydroethanolic extract of the roots of Vernonia guineensis. In an acute toxicity study, 250, 2500, and 5000 mg/kg were administered orally to mice in a single dose, and general behavior, adverse effects, and mortality were monitored. In vitro and in vivo anti-inflammatory tests were performed, and then histological, serum, hematological, and oxidative stress parameters have been evaluated. In an acute toxicity, all groups revealed neither mortality nor any significant alteration in behavior; only drowsiness, sedation, and lethargy were observed at 5000 mg/kg. For in vitro tests, the extract inhibited anti-inflammatory activity. In the formalin test, at 250 mg/kg, the extract inhibited edema with a percentage of 56.41% (4th hour) in an acute treatment and 74.44% (10th day). Joint edema was reduced by 67.24% (24th hour) in a single treatment and by 74.25% (7th day) in repeated treatment. The extract caused an increase in red blood cell, hemoglobin, and serum protein levels and reduced the white blood cells as well as the activities of alkaline phosphatase and alanine aminotransferase. The extract modulated oxidative stress parameters in the brain, spinal cord, liver, and kidneys. The extract protected the joint by reducing the bone and cartilage erosion. The present work highlights the anti-inflammatory, antioxidant, and antianemic properties of the hydroethanolic extract of the roots of Vernonia guineensis, which supports its empirical use in traditional medicine for the treatment of inflammatory pathologies.

Proteome Analysis of Whole Blood Collected by Volumetric Absorptive Microsampling.

Proteomic biomarker discovery and analysis from human biofluids using liquid chromatography-mass spectrometry (LC-MS) is an area of intense biomedical research. There is a growing interest to analyze microsampled patient blood specimens as this is potentially more patient-friendly enabling at-home and bedside self-collection of small blood volumes. However, there are limited studies applying LC-MS proteomic analysis of whole blood as it is dominated by red blood cell proteins such as hemoglobin which suppresses the detection of other less abundant proteins. Volumetric absorptive microsampling (VAMS) devices overcome this issue in part by providing a trapping matrix which allows depletion of abundant blood cell proteins through washing, prior to proteolysis and LC-MS. This approach allows the analysis of proteins from erythrocytes, leukocytes, and plasma and leads to deeper proteomic coverage compared to conventional plasma proteomics, increasing the prospects to discover novel biomarker proteins.

The in vitro red blood cell microvesiculation exerts procoagulant activity of blood cell storage in Southeast Asian ovalocytosis

Southeast Asian ovalocytosis (SAO) is characterized by the misfolding of band 3 protein in red blood cells (RBC). The abnormal structure of the band 3 protein results in dysmorphic RBC and related functions. Previous data showed that in vitro storage under hypothermic conditions alters band 3 protein structure and function. Microvesiculation includes shedding of RBC membranes, called RBC-derived microparticles/extracellular vesicles (RMP/EVs), and storage lesions. Unfortunately, there is no evidence of RBC microvesiculation under in vitro storage conditions in heterozygous SAO individuals. This study determined the generation of REVs and procoagulant activity during the storage of SAO blood samples in southern Thailand. Venous blood was collected from eight SAO and seven healthy individuals, preserved in citrate phosphate dextrose-adenine 1 (CPDA-1) at 4 °C for 35 days. The absolute numbers of REVs and PS-expressing RBCs were analyzed using flow cytometry. The procoagulant activity of the produced extracellular vesicles was determined by a clotting time assay. The results showed a significant increase in the number of REVs and PS-expressing RBCs in the SAO blood samples. Significantly correlated PS externalization and procoagulant activity were observed in the SAO blood samples. These lines of evidence indicate that the abnormality of the Band 3 protein is possibly involved in aberrant microvesiculation, exerting procoagulant activity in vitro. Increased pools of REV production and abnormal storage lesions in SAO blood samples should be a concern. Notably, the mechanisms underlying membrane vesiculation depend on the extent of blood cell storage under hypothermic conditions.

CryoEM structures of anion exchanger 1 capture multiple states of inward- and outward-facing conformations

Anion exchanger 1 (AE1, band 3) is a major membrane protein of red blood cells and plays a key role in acid-base homeostasis, urine acidification, red blood cell shape regulation, and removal of carbon dioxide during respiration. Though structures of the transmembrane domain (TMD) of three SLC4 transporters, including AE1, have been resolved previously in their outward-facing (OF) state, no mammalian SLC4 structure has been reported in the inward-facing (IF) conformation. Here we present the cryoEM structures of full-length bovine AE1 with its TMD captured in both IF and OF conformations. Remarkably, both IF-IF homodimers and IF-OF heterodimers were detected. The IF structures feature downward movement in the core domain with significant unexpected elongation of TM11. Molecular modeling and structure guided mutagenesis confirmed the functional significance of residues involved in TM11 elongation. Our data provide direct evidence for an elevator-like mechanism of ion transport by an SLC4 family member.

Effects of low-dose radiation on human blood components after in vitro exposure to gamma radiation from 137Cs radioactivity

This current study was designed to determine the effects of in vitro exposure to radioactive cesium-137 on human blood components. Whole blood samples were given a radiation dose of 0.02, 0.05, 0.1, 0.2, and 0.3 mGy of gamma radiation using a 137Cs radioactive standard source. The whole blood samples that were exposed to 0 mGy served as sham-controls. The spectrofluoroscopic technique was used to determine the autofluorescence spectrum of protein in plasma or red blood cells by using excitation wavelength and range of emission wavelengths at 280 nm and 300–550 nm, respectively. The hemolysis of red blood cells was evaluated by determination of the release of hemoglobin from the red blood cells to the supernatant. Complete blood counts were also determined in whole blood. The results showed that there was no change in the ratio of fluorescence emission intensity at 340 nm of wavelength of protein extract from irradiated whole blood or red blood cells compared to the corresponding non-irradiated control. The hemolysis value did not change in irradiated whole blood when compared to the corresponding non-irradiated group. In addition, complete blood count values in irradiated groups did not differ from non-irradiated group. These current results suggested that there were no harmful effects of the low-dose gamma radiation from radioactive 137Cs on blood components when human whole blood was exposed to gamma radiation in an in vitro condition.