The GWAS SNP rs80207740 modulates erythrocyte traits via allele-specific binding of IKZF1 and targeting XPO7 gene.

Genome-wide association studies have identified many single nucleotide polymorphisms (SNPs) associated with erythrocyte traits. However, the functional variants and their working mechanisms remain largely unknown. Here, we reported that the SNP of rs80207740, which was associated with red blood cell (RBC) volume and hemoglobin content across populations, conferred enhancer activity to XPO7 gene via allele-differentially binding to Ikaros family zinc finger 1 (IKZF1). We showed that the region around rs80207740 was an erythroid-specific enhancer using reporter assays, and that the G-allele further enhanced activity. 3D genome evidence showed that the enhancer interacted with the XPO7 promoter, and eQTL analysis suggested that the G-allele upregulated expression of XPO7. We further showed that the rs80207740-G allele facilitated the binding of transcription factor IKZF1 in EMSA and ChIP analyses. Knockdown of IKZF1 and GATA1 resulted in decreased expression of Xpo7 in both human and mouse erythroid cells. Finally, we constructed Xpo7 knockout mouse by CRISPR/Cas9 and observed anemic phenotype with reduced volume and hemoglobin content of RBC, consistent to the effect of rs80207740 on erythrocyte traits. Overall, our study demonstrated that rs80207740 modulated erythroid indices by regulating IKZF1 binding and Xpo7 expression.

Hypertension Increases Susceptibility to Experimental Malaria in Mice.

Global prevalence of hypertension is on the rise, burdening healthcare, especially in developing countries where infectious diseases, such as malaria, are also rampant. Whether hypertension could predispose or increase susceptibility to malaria, however, has not been extensively explored. Previously, we reported that hypertension is associated with abnormal red blood cell (RBC) physiology and anemia. Since RBC are target host cells for malarial parasite, Plasmodium, we hypothesized that hypertensive patients with abnormal RBC physiology are at greater risk or susceptibility to Plasmodium infection. To test this hypothesis, normotensive (BPN/3J) and hypertensive (BPH/2J) mice were characterized for their RBC physiology and subsequently infected with Plasmodium yoelii (P. yoelii), a murine-specific non-lethal strain. When compared to BPN mice, BPH mice displayed microcytic anemia with RBC highly resistant to osmotic hemolysis. Further, BPH RBC exhibited greater membrane rigidity and an altered lipid composition, as evidenced by higher levels of phospholipids and saturated fatty acid, such as stearate (C18:0), along with lower levels of polyunsaturated fatty acid like arachidonate (C20:4). Moreover, BPH mice had significantly greater circulating Ter119+ CD71+ reticulocytes, or immature RBC, prone to P. yoelii infection. Upon infection with P. yoelii, BPH mice experienced significant body weight loss accompanied by sustained parasitemia, indices of anemia, and substantial increase in systemic pro-inflammatory mediators, compared to BPN mice, indicating that BPH mice were incompetent to clear P. yoelii infection. Collectively, these data demonstrate that aberrant RBC physiology observed in hypertensive BPH mice contributes to an increased susceptibility to P. yoelii infection and malaria-associated pathology.

Evaluation of CSF 8-iso-prostaglandin F2α and erythrocyte anisocytosis as prognostic biomarkers for delayed cerebral ischemia after aneurysmal subarachnoid hemorrhage.

Delayed cerebral ischemia (DCI) is a serious, life-threatening, complication affecting patients who have survived the initial bleeding from a ruptured intracranial aneurysm. Due to the challenging diagnosis, potential DCI prognostic markers should be of value in clinical practice. According to recent reports isoprostanes and red blood cell distribution (RDW) showed to be promising in this respect. We conducted a prospective study of 27 aSAH patients and control group (n = 8). All patients from the study group were treated within the first day of the initial bleeding. We collected data regarding clinical status and results of biochemical, and radiological examinations. We measured cerebrospinal fluid (CSF) concentration of 8-iso-prostaglandin F2α (F2-IsoP) and RDW on days 1, 3, and 5. Both CSF F2-IsoP level and RDW-SD measured on day 1 were significant predictors of DCI. The receiver operating characteristics curve for DCI prediction based on the multivariate model yielded an area under the curve of 0.924 (95% CI 0.824-1.000, p < 0.001). In our study, the model based on the combination of RDW and the level of isoprostanes in CSF on the first day after the initial bleeding showed a prognostic value for DCI prediction. Further studies are required to validate this observation.

Effect of Garlic Extract on the Erythrocyte as a Simple Model Cell.

Garlic is known to have diverse effects on mammalian cells, being cytotoxic, especially to cancer cells, but also protect against oxidative stress. Mammalian erythrocyte is a simple cell devoid of intracellular organelles, protein synthesis ability, and most signaling pathways. Therefore, examination of the effects of garlic on erythrocytes allows for revealing primary events in the cellular action of garlic extract. In this study, human erythrocytes or erythrocyte membranes were exposed to garlic extract at various dilutions. Hemoglobin oxidation to methemoglobin, increased binding of hemoglobin to the membrane, and formation of Heinz bodies were observed. Garlic extract depleted acid-soluble thiols, especially glutathione, and induced a prooxidative shift in the cellular glutathione redox potential. The extract increased the osmotic fragility of erythrocytes, induced hemolysis, and inhibited hemolysis in isotonic ammonium chloride, indicative of decreased membrane permeability for Cl- and increased the membrane fluidity. Fluorescent probes indicated an increased level of reactive oxygen species and induction of lipid peroxidation, but these results should be interpreted with care since the extract alone induced oxidation of the probes (dichlorodihydrofluorescein diacetate and BODIPY C11). These results demonstrate that garlic extract induces oxidative changes in the erythrocyte, first of all, thiol and hemoglobin oxidation.

Preanalytical considerations for clinical assays of circulating human miRNA-451a, miRNA-423-5p and miRNA-199a-3p for diagnostic purposes.

Circulating miRNA has recently emerged as important biomolecules with potential clinical values as diagnostic markers for several diseases. However, to be used as such, it is critical to accurately quantify miRNAs in the clinic. Yet, preanalytical factors that can affect an error-free quantification of these miRNAs have not been explored. This study aimed at investigating several of these preanalytical factors that may affect the accurate quantification of miRNA-451a, miRNA-423-5p and miRNA-199a-3p in human blood samples. We initially evaluated levels of these three miRNAs in red blood cells (RBCs), white blood cells (WBCs), platelets, and plasma by droplet digital PCR (ddPCR). Next, we monitored miRNA levels in whole blood or platelet rich plasma (PRP) stored at different temperatures for different time periods by ddPCR. We also investigated the effects of hemolysis on miRNA concentrations in platelet-free plasma (PFP). Our results demonstrate that more than 97% of miRNA-451a and miRNA-423-5p in the blood are localized in RBCs, with only trace amounts present in WBCs, platelets, and plasma. Highest amount of the miRNA-199a-3p is present in platelets. Hemolysis had a significant impact on both miRNA-451a and miRNA-423-5p concentrations in plasma, however miRNA-199a levels remain unaffected. Importantly, PRP stored at room temperature (RT) or 4°C showed a statistically significant decrease in miRNA-451a levels, while the other two miRNAs were increased, at days 1, 2, 3 and 7. PFP at RT caused statistically significant steady decline in miRNA-451a and miRNA-423-5p, observed at 12, 24, 36, 48 and 72 hours. Levels of the miRNA-199a-3p in PFP was stable during first 72 hours at RT. PFP stored at -20°C for 7 days showed declining stability of miRNA-451a over time. However, at -80°C miRNA-451a levels were stable up to 7 days. Together, our data indicate that hemolysis and blood storage at RT, 4°C and -20°C may have significant negative effects on the accuracy of circulating miRNA-451a and miRNA-423-5p quantification.

Evaluation of anti-sickling effects of two varieties of Cajanus cajan (L.) Huth on sickle cell beta thalassemia.

ETHNO-PHARMACOLOGICAL RELEVANCE: Globally, the prevalence of sickle cell disease is on the rise, with developing countries experiencing particularly alarming mortality rate compared to developed nations. The World Health Organization (WHO) and United Nations (UN) have acknowledged sickle cell disease as a significant global public health concern. Unfortunately, a cure for this condition is yet to be discovered, and existing allopathic treatments, while offering relief, come with serious side effects. In recent times, there has been a growing interest in exploring the potential of medicinal plants for treating sickle cell disease due to their content of secondary metabolites that may impact the disease's mechanisms. Cajanus cajan, a crucial grain legume in rain-fed agriculture in semi-arid tropics, has been traditionally used in folk medicine to manage various illnesses and is suggested to possess anti-sickling properties. AIM OF THE STUDY: The present study investigated two varieties of C. cajan for their effectiveness in treating sickle cell beta thalassemia, a variant of sickle cell disease. MATERIALS AND METHODS: The study was divided into four groups consisting of the untreated group (group 1), group treated with standard drug (group 2), group treated with white C. cajan (group 3) and group treated with brown C. cajan (group 4). The effects of the two variety of C. cajan were measured by polymerization test, reversibility test, osmotic fragility test, deoxygenation and beta globin synthesis test. RESULT: The results revealed that both varieties of C. cajan demonstrated a reduction in polymerization rates, reversed sickled red blood cells, increased the oxygen affinity of Hb-S/ß, elevated the Fe2+/Fe3+ ratio, and maintained the membrane stability of red blood cells. Notably, the white variety exhibited superior anti-sickling properties compared to the brown variety. CONCLUSION: This suggests that this significant leguminous crop could be utilized for the treatment and management of sickling disorders, particularly in low-income countries where conventional treatments may be financially inaccessible to patients.

Exogenous polyserine fibrils change membrane properties of phosphatidylcholine-liposome and red blood cells.

The causative genes for neurodegenerative polyglutamine (polyQ) diseases produce homopolymeric polyglutamine (polyQ), polyserine (polyS), polyalanine (polyA), polycysteine (polyC), and polyleucine (polyL) sequences by repeat-associated non-AUG (RAN) translation. The cytotoxicity of the intracellular polyQ and RAN products has been extensively investigated. However, little is known about the toxicity of the extracellular polyQ and RAN products on the membranes of viable cells. Because polyQ aggregates induce a deflated morphology of a model membrane, we hypothesized that extracellular polyQ and RAN products might affect the membrane properties of viable cells. In this study, we demonstrated that exogenous polyS fibrils but not polyS or polyQ non-fibril aggregates altered the thermal phase transition behavior of a model membrane composed of a phosphatidylcholine bilayer using differential scanning calorimetry. PolyS fibrils induced morphological changes in viable red blood cells (RBCs). However, both polyS and polyQ non-fibril aggregates had no effects on RBCs. These results highlight the possibility that extracellular fibrils generated from RAN products may alter the properties of neuronal cell membranes, which may contribute to changes in the brain pathology.

Progresses in overcoming the limitations of in vitro erythropoiesis using human induced pluripotent stem cells.

Researchers have attempted to generate transfusable oxygen carriers to mitigate RBC supply shortages. In vitro generation of RBCs using stem cells such as hematopoietic stem and progenitor cells (HSPCs), embryonic stem cells (ESCs), and induced pluripotent stem cells (iPSCs) has shown promise. Specifically, the limited supplies of HSPCs and ethical issues with ESCs make iPSCs the most promising candidate for in vitro RBC generation. However, researchers have encountered some major challenges when using iPSCs to produce transfusable RBC products, such as enucleation and RBC maturation. In addition, it has proven difficult to manufacture these products on a large scale. In this review, we provide a brief overview of erythropoiesis and examine endeavors to recapitulate erythropoiesis in vitro using various cell sources. Furthermore, we explore the current obstacles and potential solutions aimed at enabling the large-scale production of transfusable RBCs in vitro.

Impact of transcription factors KLF1 and GATA1 on red blood cell antigen expression: a review.

KLF transcription factor 1 (KLF1) and GATA binding protein 1 (GATA1) are transcription factors (TFs) that initiate and regulate transcription of the genes involved in erythropoiesis. These TFs possess DNA-binding domains that recognize specific nucleotide sequences in genes, to which they bind and regulate transcription. Variants in the genes that encode either KLF1 or GATA1 can result in a range of hematologic phenotypes-from benign to severe forms of thrombocytopenia and anemia; they can also weaken the expression of blood group antigens. The Lutheran (LU) blood group system is susceptible to TF gene variations, particularly KLF1 variants. Individuals heterozygous for KLF1 gene variants show reduced Lutheran antigens on red blood cells that are not usually detected by routine hemagglutination methods. This reduced antigen expression is referred to as the In(Lu) phenotype. For accurate blood typing, it is important to distinguish between the In(Lu) phenotype, which has very weak antigen expression, and the true Lunull phenotype, which has no antigen expression. The International Society of Blood Transfusion blood group allele database registers KLF1 and GATA1 variants associated with modified Lutheran expression. Here, we review KLF1 and recent novel gene variants defined through investigating blood group phenotype and genotype discrepancies or, for one report, investigating cases with unexplained chronic anemia. In addition, we include a review of the GATA1 TF, including a case report describing the second GATA1 variant associated with a serologic Lu(a-b-) phenotype. Finally, we review both past and recent reports on variations in the DNA sequence motifs on the blood group genes that disrupt the binding of the GATA1 TF and either remove or reduce erythroid antigen expression. This review highlights the diversity and complexity of the transcription process itself and the need to consider these factors as an added component for accurate blood group phenotyping.

Membrane-Bound Ferric Hemoglobin in Nucleated Erythrocytes of the Black Scorpionfish Scorpaena porcus, Linnaeus 1758.

The content of membrane-bound methemoglobin (MtHb) in nucleated erythrocytes was studied in the black scorpionfish Scorpaena porcus (Linnaeus, 1758) in vitro. Spectral characteristics were determined for a whole hemolysate, a hemolysate obtained by stroma precipitation (a clarified hemolysate), and a resuspended stroma. The MtHb proportion in the erythrocyte stroma was found to exceed 80% (6.20 ± 0.59 µM). Clarified hemolysates were nearly free of MtHb (0.5 ± 0.2 µM). Membrane-bound ferric hemoglobin did not affect the erythrocyte resistance to osmotic shock. The osmotic fragility range was determined using a LaSca-TM laser microparticle analyzer (BioMedSystems, Russia) to be 102-136 mOsm/kg, much the same as in other bony fish species. A nitrite load (10 mg/L) significantly increased the MtHb content in the blood. However, the membrane-bound ferric hemoglobin content did not change significantly, amounting to 6.34 ± 1.09 µM (approximately 95%). The finding suggested a functional importance for MtHb present in the plasma membrane of nucleated erythrocytes. Membrane-bound MtHb was assumed to neutralize the external oxidative load and the toxic effect of hydrogen sulfide in bottom water layers, where the species lives.

APEX2-based proximity proteomic analysis identifies candidate interactors for Plasmodium falciparum knob-associated histidine-rich protein in infected erythrocytes.

The interaction of Plasmodium falciparum-infected red blood cells (iRBCs) with the vascular endothelium plays a crucial role in malaria pathology and disease. KAHRP is an exported P. falciparum protein involved in iRBC remodelling, which is essential for the formation of protrusions or "knobs" on the iRBC surface. These knobs and the proteins that are concentrated within them allow the parasites to escape the immune response and host spleen clearance by mediating cytoadherence of the iRBC to the endothelial wall, but this also slows down blood circulation, leading in some cases to severe cerebral and placental complications. In this work, we have applied genetic and biochemical tools to identify proteins that interact with P. falciparum KAHRP using enhanced ascorbate peroxidase 2 (APEX2) proximity-dependent biotinylation and label-free shotgun proteomics. A total of 30 potential KAHRP-interacting candidates were identified, based on the assigned fragmented biotinylated ions. Several identified proteins have been previously reported to be part of the Maurer's clefts and knobs, where KAHRP resides. This study may contribute to a broader understanding of P. falciparum protein trafficking and knob architecture and shows for the first time the feasibility of using APEX2-proximity labelling in iRBCs.

What is the role of complement in bystander hemolysis? Old concept, new insights.

INTRODUCTION: Bystander hemolysis occurs when antigen-negative red blood cells (RBCs) are lysed by the complement system. Many clinical entities including passenger lymphocyte syndrome, hyperhemolysis following blood transfusion, and paroxysmal nocturnal hemoglobinuria are complicated by bystander hemolysis. AREAS COVERED: The review provides data about the role of the complement system in the pathogenesis of bystander hemolysis. Moreover, future perspectives on the understanding and management of this syndrome are described. EXPERT OPINION: Complement system can be activated via classical, alternative, and lectin pathways. Classical pathway activation is mediated by antigen-antibody (autoantibodies and alloantibodies against autologous RBCs, infectious agents) complexes. Alternative pathway initiation is triggered by heme, RBC microvesicles, and endothelial injury that is a result of intravascular hemolysis. Thus, C5b is formed, binds with C6-C9 compomers, and MAC (C5b-9) is formulated in bystander RBCs membranes, leading to cell lysis. Intravascular hemolysis, results in activation of the alternative pathway, establishing a vicious cycle between complement activation and bystander hemolysis. C5 inhibitors have been used effectively in patients with hyperhemolysis syndrome and other entities characterized by bystander hemolysis.

Reduction of hemagglutination induced by a SARS-CoV-2 spike protein fragment using an amyloid-binding benzothiazole amphiphile.

COVID-19 infection is associated with a variety of vascular occlusive morbidities. However, a comprehensive understanding of how this virus can induce vascular complications remains lacking. Here, we show that a peptide fragment of SARS-CoV-2 spike protein, S192 (sequence 192-211), is capable of forming amyloid-like aggregates that can induce agglutination of red blood cells, which was not observed with low- and non-aggregated S192 peptide. We subsequently screened eight amyloid-binding molecules and identified BAM1-EG6, a benzothiazole amphiphile, as a promising candidate capable of binding to aggregated S192 and partially inhibiting its agglutination activity. These results provide new insight into a potential molecular mechanism for the capability of spike protein metabolites to contribute to COVID-19-related blood complications and suggest a new therapeutic approach for combating microvascular morbidities in COVID-19 patients.

Survival of transfused red blood cells from a donor with alpha-thalassemia trait in a recipient with sickle cell disease.

BACKGROUND: Post-transfusion survival of donor red blood cells (RBCs) is important for effective chronic transfusion therapy in conditions including sickle cell disease (SCD). Biotin labeling RBCs allows direct in vivo measurement of multiple donor RBC units simultaneously post-transfusion. STUDY DESIGN AND METHODS: In an observational trial of patients with SCD receiving monthly chronic transfusion therapy, aliquots of RBCs from one transfusion episode were biotin-labeled and infused along with the unlabeled RBC units. Serial blood samples were obtained to measure RBC survival. Donor units were tested for RBC indices, hemoglobin fractionation, and glucose-6-phosphate dehydrogenase (G6PD) enzyme activity. For microcytic donor RBCs (MCV < 70 fL), HBA1 and HBA2 genetic testing was performed on whole blood. RESULTS: We present one recipient, a pediatric patient with SCD and splenectomy who received two RBC units with aliquots from each unit labeled at distinct biotin densities (2 and 18 µg/mL biotin). One donor unit was identified to have microcytosis (MCV 68.5 fL after biotinylation); whole blood sample obtained at a subsequent donation showed 2-gene deletion alpha-thalassemia trait (ɑ-3.7kb/ɑ-3.7kb) and normal serum ferritin. G6PD activity was >60% of normal mean for both. The RBCs with alpha-thalassemia RBC had accelerated clearance and increased surface phosphatidylserine post-transfusion, as compared with the normocytic RBC (half life 65 vs. 86 days, respectively). DISCUSSION: Post-transfusion RBC survival may be lower for units from donors with alpha-thalassemia trait, although the impact of thalassemia trait donors on transfusion efficacy requires further study.

In vivo genotoxicity assessment of N-(-9 acridinyl)-b-alanine hydrochloride (S-300) using a validated Pig-a mutagenesis assay.

BACKGROUND: N-(-9 acridinyl)-b-alanine hydrochloride (S-300) is the main byproduct of red blood cell (RBC) amustaline/glutathione(GSH) pathogen reduction, currently undergoing phase III US clinical trials following successful European studies(1-3). Phosphatidylinositol glycan, class A (Pig-a) X-linked gene mutagenesis is a validated mammalian in vivo mutation assay for genotoxicity, assessed as clonal loss of glycosylphosphatidylinositol-linked CD59 cell-surface molecules on reticulocytes (RETs) and RBCs. METHODS: Male Sprague-Dawley rats received continuous infusion of S-300 up to the maximum feasible dose (240 mg/kg/day-limited by solubility and volume) for 28 days. Positive controls received a known mutagen by oral gavage on Days 1-3. Plasma levels of S-300 were assessed by HPLC before, during and after infusion. CD59-negative RBCs and RETs were enumerated in pre-dose and Day 28 samples, using a flow cytometric method. Outcome was evaluated by predetermined criteria using concurrent and historical controls. Toxicity was assessed by laboratory measures and necropsy. RESULTS: S-300 reached maximum, dose-dependent levels (3-15 µmol/L) within 2-8 h that were sustained for 672 h and undetectable 2 h after infusion. Circulating RET levels indicated a lack of hematopoietic toxicity. Necropsy revealed minimal-mild observations related to poor S-300 solubility at high concentrations. Pig-a assessment met the preset acceptability criteria and revealed no increase in mutant RBCs or RETs. CONCLUSIONS: Maximum feasible S-300 exposure of rats by continuous infusion for 28 days was not genotoxic as assessed by an Organization for Economic Cooperation and Development-compliant, mammalian, in vivo Pig-a gene mutation assay that meets the requirements of International Conference on Harmonization (ICH) S2(R1) and FDA guidances on genotoxicity testing.

Structural investigation of interactions between halogenated flavonoids and the lipid membrane along with their role as cytotoxic agents.

This study focuses on understanding the structural and molecular changes in lipid membranes under the influence of six halogenated flavonoid derivatives differing in the number and position of substitution of chlorine and bromine atoms (D1-D6). Utilizing various analytical techniques, including fluorometric methods, dynamic light scattering (DLS), attenuated Fourier transform infrared spectroscopy (ATR- FTIR), and FT-Raman spectroscopy, the research aims to elucidate the mechanisms underlying the interaction of flavonoids with cell membranes. Additionally, the study includes in silico analyses to explore the physicochemical properties of these compounds and their potential pharmaceutical applications, along with toxicity studies to assess their effects on cancer, normal, and red blood cells. Our study showed the ability of halogenated derivatives to interact mostly with the outer part of the membrane, especially in the lipid heads region however, some of them were able to penetrate deeper into the membrane and affect the fluidity of hydrocarbon chains. The potential to reduce cancer cell viability, the lack of toxicity towards erythrocytes, and the favourable physicochemical and pharmacokinetic properties suggest these halogenated flavonoids potential candidates for exploring their potential for medical use.

Cognitive impairment and hippocampal neuronal damage in ß-thalassaemia mice.

ß-Thalassaemia is one of the most common genetic diseases worldwide. During the past few decades, life expectancy of patients has increased significantly owing to advance in medical treatments. Cognitive impairment, once has been neglected, has gradually become more documented. Cognitive impairment in ß-thalassaemia patients is associated with natural history of the disease and socioeconomic factors. Herein, to determined effect of ß-thalassaemia intrinsic factors, 22-month-old ß-thalassaemia mouse was used as a model to assess cognitive impairment and to investigate any aberrant brain pathology in ß-thalassaemia. Open field test showed that ß-thalassaemia mice had decreased motor function. However, no difference of neuronal degeneration in primary motor cortex, layer 2/3 area was found. Interestingly, impaired learning and memory function accessed by a Morris water maze test was observed and correlated with a reduced number of living pyramidal neurons in hippocampus at the CA3 region in ß-thalassaemia mice. Cognitive impairment in ß-thalassaemia mice was significantly correlated with several intrinsic ß-thalassaemic factors including iron overload, anaemia, damaged red blood cells (RBCs), phosphatidylserine (PS)-exposed RBC large extracellular vesicles (EVs) and PS-exposed medium EVs. This highlights the importance of blood transfusion and iron chelation in ß-thalassaemia patients. In addition, to improve patients' quality of life, assessment of cognitive functions should become part of routine follow-up.

Continuous blood exchange in rats as a novel approach for experimental investigation.

Blood exchange therapy, specifically Whole blood exchange (WBE), is increasingly being utilized in clinical settings to effectively treat a range of diseases. Consequently, there is an urgent requirement to establish convenient and clinically applicable animal models that can facilitate the exploration of blood exchange therapy mechanisms. Our study conducted continuous WBE in rats through femoral and tail vein catheterization using dual-directional syringe pumps. To demonstrate the applicability of continuous WBE, drug-induced hemolytic anemia (DIHA) was induced through phenylhydrazine hydrochloride (PHZ) injection. Notability, the rats of DIHA + WBE group all survived and recovered within the subsequent period. After the implementation of continuous WBE therapy day (Day 1), the DIHA + WBE group exhibited a statistically significant increase in red blood cells (RBC) (P = 0.0343) and hemoglobin (HGB) levels (P = 0.0090) compared to DIHA group. The rats in the DIHA + WBE group exhibited a faster recovery rate compared to the DIHA group, indicating the successful establishment of a continuous blood exchange protocol. This experimental approach demonstrates not just promising efficacy in the treatment of DIHA and offers a valuable tool for investigating the underlying mechanisms of blood exchange. Furthermore, it has a great potential to the advancement of biomedical research such as drug delivery exploration.

Glycated LDL generates reactive species that damage cell components, oxidize hemoglobin and alter surface morphology in human erythrocytes.

Low-density lipoprotein (LDL) transports cholesterol to various tissues via the blood. Glycation of LDL occurs during hyperglycemic condition which is characterised by persistently high blood glucose level. Circulating erythrocytes can come in direct contact with glycated LDL (G-LDL). The objective of this study was to investigate the effect of G-LDL on human erythrocytes, specifically on hemoglobin, intracellular generation of reactive species and the antioxidant defence system. Isolated erythrocytes were incubated with G-LDL (3 and 6 mg/ml) and native LDL (6 mg/ml) at 37 °C for 24 h. Native LDL and G-LDL untreated erythrocytes were similarly incubated at 37 °C and served as control. G-LDL treatment increased hemolysis compared to control and native LDL-treated erythrocytes. Incubation of erythrocytes with G-LDL led to an increase in protein oxidation and lipid peroxidation while greatly decreasing the total sulfhydryl content. It also significantly enhanced hemoglobin oxidation, heme degradation, and the release of free iron moiety. Treatment with G-LDL led to an appreciable increase in the production of reactive oxygen and nitrogen species. The antioxidant power and activities of major antioxidant enzymes were drastically reduced, while critical membrane-bound enzymes were inhibited. The surface morphology of G-LDL-treated erythrocytes was altered leading to the formation of echinocytes. Importantly, treatment of erythrocytes with native LDL did not significantly affect the above-mentioned parameters and values were similar to the corresponding controls. Thus, G-LDL is cytotoxic to human erythrocytes and causes oxidative damage to cell components. This can reduce the oxygen-transporting ability of blood and also result in red cell senescence and anemia.

The miR-144/Hmgn2 regulatory axis orchestrates chromatin organization during erythropoiesis.

Differentiation of stem and progenitor cells is a highly regulated process that involves the coordinated action of multiple layers of regulation. Here we show how the post-transcriptional regulatory layer instructs the level of chromatin regulation via miR-144 and its targets to orchestrate chromatin condensation during erythropoiesis. The loss of miR-144 leads to impaired chromatin condensation during erythrocyte maturation. Among the several targets of miR-144 that influence chromatin organization, the miR-144-dependent regulation of Hmgn2 is conserved from fish to humans. Our genetic probing of the miR-144/Hmgn2 regulatory axis establish that intact miR-144 target sites in the Hmgn2 3'UTR are necessary for the proper maturation of erythrocytes in both zebrafish and human iPSC-derived erythroid cells while loss of Hmgn2 rescues in part the miR-144 null phenotype. Altogether, our results uncover miR-144 and its target Hmgn2 as the backbone of the genetic regulatory circuit that controls the terminal differentiation of erythrocytes in vertebrates.