Sterile α-motif domain requirement for cellular signaling and survival.

Hundreds of sterile α-motif (SAM) domains have predicted structural similarities and are reported to bind proteins, lipids, or RNAs. However, the majority of these domains have not been analyzed functionally. Previously, we demonstrated that a SAM domain-containing protein, SAMD14, promotes SCF/proto-oncogene c-Kit (c-Kit) signaling, erythroid progenitor function, and erythrocyte regeneration. Deletion of a Samd14 enhancer (Samd14-Enh), occupied by GATA2 and SCL/TAL1 transcription factors, reduces SAMD14 expression in bone marrow and spleen and is lethal in a hemolytic anemia mouse model. To rigorously establish whether Samd14-Enh deletion reduces anemia-dependent c-Kit signaling by lowering SAMD14 levels, we developed a genetic rescue assay in murine Samd14-Enh-/- primary erythroid precursor cells. SAMD14 expression at endogenous levels rescued c-Kit signaling. The conserved SAM domain was required for SAMD14 to increase colony-forming activity, c-Kit signaling, and progenitor survival. To elucidate the molecular determinants of SAM domain function in SAMD14, we substituted its SAM domain with distinct SAM domains predicted to be structurally similar. The chimeras were less effective than SAMD14 itself in rescuing signaling, survival, and colony-forming activities. Thus, the SAMD14 SAM domain has attributes that are distinct from other SAM domains and underlie SAMD14 function as a regulator of cellular signaling and erythrocyte regeneration.

PPAR-α and glucocorticoid receptor synergize to promote erythroid progenitor self-renewal.

Many acute and chronic anaemias, including haemolysis, sepsis and genetic bone marrow failure diseases such as Diamond-Blackfan anaemia, are not treatable with erythropoietin (Epo), because the colony-forming unit erythroid progenitors (CFU-Es) that respond to Epo are either too few in number or are not sensitive enough to Epo to maintain sufficient red blood cell production. Treatment of these anaemias requires a drug that acts at an earlier stage of red cell formation and enhances the formation of Epo-sensitive CFU-E progenitors. Recently, we showed that glucocorticoids specifically stimulate self-renewal of an early erythroid progenitor, burst-forming unit erythroid (BFU-E), and increase the production of terminally differentiated erythroid cells. Here we show that activation of the peroxisome proliferator-activated receptor α (PPAR-α) by the PPAR-α agonists GW7647 and fenofibrate synergizes with the glucocorticoid receptor (GR) to promote BFU-E self-renewal. Over time these agonists greatly increase production of mature red blood cells in cultures of both mouse fetal liver BFU-Es and mobilized human adult CD34(+) peripheral blood progenitors, with a new and effective culture system being used for the human cells that generates normal enucleated reticulocytes. Although Ppara(-/-) mice show no haematological difference from wild-type mice in both normal and phenylhydrazine (PHZ)-induced stress erythropoiesis, PPAR-α agonists facilitate recovery of wild-type but not Ppara(-/-) mice from PHZ-induced acute haemolytic anaemia. We also show that PPAR-α alleviates anaemia in a mouse model of chronic anaemia. Finally, both in control and corticosteroid-treated BFU-E cells, PPAR-α co-occupies many chromatin sites with GR; when activated by PPAR-α agonists, additional PPAR-α is recruited to GR-adjacent sites and presumably facilitates GR-dependent BFU-E self-renewal. Our discovery of the role of PPAR-α agonists in stimulating self-renewal of early erythroid progenitor cells suggests that the clinically tested PPAR-α agonists we used may improve the efficacy of corticosteroids in treating Epo-resistant anaemias.

Heme Oxygenase-1 Deficiency and Oxidative Stress: A Review of 9 Independent Human Cases and Animal Models.

Since Yachie et al. reported the first description of human heme oxygenase (HO)-1 deficiency more than 20 years ago, few additional human cases have been reported in the literature. A detailed analysis of the first human case of HO-1 deficiency revealed that HO-1 is involved in the protection of multiple tissues and organs from oxidative stress and excessive inflammatory reactions, through the release of multiple molecules with anti-oxidative stress and anti-inflammatory functions. HO-1 production is induced in vivo within selected cell types, including renal tubular epithelium, hepatic Kupffer cells, vascular endothelium, and monocytes/macrophages, suggesting that HO-1 plays critical roles in these cells. In vivo and in vitro studies have indicated that impaired HO-1 production results in progressive monocyte dysfunction, unregulated macrophage activation and endothelial cell dysfunction, leading to catastrophic systemic inflammatory response syndrome. Data from reported human cases of HO-1 deficiency and numerous studies using animal models suggest that HO-1 plays critical roles in various clinical settings involving excessive oxidative stress and inflammation. In this regard, therapy to induce HO-1 production by pharmacological intervention represents a promising novel strategy to control inflammatory diseases.

Erythroid glucose transport in health and disease.

Glucose transport is intimately linked to red blood cell physiology. Glucose is the unique energy source for these cells, and defects in glucose metabolism or transport activity are associated with impaired red blood cell morphology and deformability leading to reduced lifespan. In vertebrate erythrocytes, glucose transport is mediated by GLUT1 (in humans) or GLUT4 transporters. These proteins also account for dehydroascorbic acid (DHA) transport through erythrocyte membrane. The peculiarities of glucose transporters and the red blood cell pathologies involving GLUT1 are summarized in the present review.

Risk of piperacillin-induced hemolytic anemia in patients with cystic fibrosis and antipseudomonal treatment: a prospective observational study.

BACKGROUND: Drug-induced immune hemolytic anemia (DIIHA) is a rare but severe side effect caused by numerous drugs. Case reports and case series suggest that piperacillin-related DIIHA may be more common among patients with cystic fibrosis (CF). However, the prevalence is speculative. The aim of this prospective, observational study was determine the prevalence of DIIHA in such affected patients. METHODS AND MATERIALS: Patients with CF hospitalized for parenteral antibiotic therapy at Charité Universitätsmedizin Berlin, who had previously been exposed to IV antibiotics, were enrolled. Blood samples were collected on Days 3 and 12 of antibiotic treatment courses. Serological studies were performed using standard techniques with gel cards. Screening for drug-dependent antibodies (ddab) was performed in the presence of the drugs and their urinary metabolites. RESULTS: A total of 52 parenteral antibiotic cycles in 43 patients were investigated. Ddab against piperacillin were detected in two patients (4.7%). The direct AHG was positive with anti-IgG only in both patients. However only one of these patients developed mild immune hemolytic anemia. Both patients had been repeatedly treated with piperacillin without any evident hemolysis. There was no correlation between the exposure to piperacillin and the prevalence of ddab. CONCLUSION: Our prospective study indicates that piperacillin-induced ddab occur more frequently in patients with CF than previously suggested. The question related to the significance of piperacillin-dependent antibodies may reflect new aspects in this field.

Podocytes are new cellular targets of haemoglobin-mediated renal damage.

Recurrent and massive intravascular haemolysis induces proteinuria, glomerulosclerosis, and progressive impairment of renal function, suggesting podocyte injury. However, the effects of haemoglobin (Hb) on podocytes remain unexplored. Our results show that cultured human podocytes or podocytes isolated from murine glomeruli bound and endocytosed Hb through the megalin-cubilin receptor system, thus resulting in increased intracellular Hb catabolism, oxidative stress, activation of the intrinsic apoptosis pathway, and altered podocyte morphology, with decreased expression of the slit diaphragm proteins nephrin and synaptopodin. Hb uptake activated nuclear factor erythroid-2-related factor 2 (Nrf2) and induced expression of the Nrf2-related antioxidant proteins haem oxygenase-1 (HO-1) and ferritin. Nrf2 activation and Hb staining was observed in podocytes of mice with intravascular haemolysis. These mice developed proteinuria and showed podocyte injury, characterized by foot process effacement, decreased synaptopodin and nephrin expression, and podocyte apoptosis. These pathological effects were enhanced in Nrf2-deficient mice, whereas Nrf2 activation with sulphoraphane protected podocytes against Hb toxicity both in vivo and in vitro. Supporting the translational significance of our findings, we observed podocyte damage and podocytes stained for Hb, HO-1, ferritin and phosphorylated Nrf2 in renal sections and urinary sediments of patients with massive intravascular haemolysis, such as atypical haemolytic uraemic syndrome and paroxysmal nocturnal haemoglobinuria. In conclusion, podocytes take up Hb both in vitro and during intravascular haemolysis, promoting oxidative stress, podocyte dysfunction, and apoptosis. Nrf2 may be a potential therapeutic target to prevent loss of renal function in patients with intravascular haemolysis. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Thermal sensitivity and haemolysis of erythrocytes with membranopathy.

Measuring the impedance of heated suspensions of erythrocytes and erythrocyte ghost membranes, two thermally-induced alterations are registered in the plasma membrane at TA (denaturation of spectrin with inducing temperature at 49,5 °C) and TG (hyperthermic activation of basal ion permeability with inducing temperature at 60.7 °C). In this study erythrocytes from 9 healthy patients and 15 patients with hemolytic anemia were studied and divided into four groups depending on their TA and TG top temperatures. The TA and TG of erythrocytes with hemoglobinopathy were the same as those of control erythrocytes while those of erythrocytes with membranopathy were significantly reduced. In erythrocytes with severe membranopathy, the TG was decreased by about 5 °C. In latter cells the normal value of TG was restored and the resistance to thermal haemolysis was increased by 90% after the specific stabilization of band 3 protein by 4,4'-diisothiocyanato-stilbene-2,2'-disulfonic acid (DIDS). Obtained results indicate the involvement of band 3 in the membrane alteration at TG and in the heat target responsible for thermal haemolysis.

Gene disruption of dematin causes precipitous loss of erythrocyte membrane stability and severe hemolytic anemia.

Dematin is a relatively low abundance actin binding and bundling protein associated with the spectrin-actin junctions of mature erythrocytes. Primary structure of dematin includes a loosely folded core domain and a compact headpiece domain that was originally identified in villin. Dematin's actin binding properties are regulated by phosphorylation of its headpiece domain by cyclic adenosine monophosphate-dependent protein kinase. Here, we used a novel gene disruption strategy to generate the whole body dematin gene knockout mouse model (FLKO). FLKO mice, while born at a normal Mendelian ratio, developed severe anemia and exhibited profound aberrations of erythrocyte morphology and membrane stability. Having no apparent effect on primitive erythropoiesis, FLKO mice show significant enhancement of erythroblast enucleation during definitive erythropoiesis. Using membrane protein analysis, domain mapping, electron microscopy, and dynamic deformability measurements, we investigated the mechanism of membrane instability in FLKO erythrocytes. Although many membrane and cytoskeletal proteins remained at their normal levels, the major peripheral membrane proteins spectrin, adducin, and actin were greatly reduced in FLKO erythrocytes. Our results demonstrate that dematin plays a critical role in maintaining the fundamental properties of the membrane cytoskeleton complex.

Heme oxygenase-1 deficiency promotes severity of sepsis in a non-surgical preterm mouse model.

BACKGROUND: Sepsis in preterm infants is associated with systemic inflammatory responses. The stress-response protein heme oxygenase-1 (HO-1) has protective anti-inflammatory properties. Recently, we reported a protective role of HO-1 using our non-surgical cecal slurry (CS) model in wild-type (WT) mouse pups. Here, we extend these findings to investigate the association of HO-1 deficiency with sepsis severity. METHODS: Adapting the Wynn model, we induced sepsis in 4-day-old HO-1-deficient (HO-1+/-, Het) pups to determine if HO-1 deficiency affected survival rates at the LD40 (2.0 mg/g) of WT pups. To see if HO-1 induction affected sepsis severity, we gave 30-µmol heme/kg subcutaneously to 3-day-old mice 24 h prior to sepsis induction. RESULTS: Post-sepsis induction, Het pups had a mortality of 85.0% (n = 20) and increased expression of the pro-inflammatory gene in the livers and affected hematologic profiles. Heme treatment 24 h prior to sepsis induction significantly increased liver HO activity, reduced mortality to 24.5% (n = 17), attenuated inflammatory responses, reduced spleen bacterial counts, and significantly increased peripheral neutrophils. CONCLUSIONS: A partial deficiency in HO-1 increased the progression and mortality in sepsis. Furthermore, induction of HO-1 significantly reduced the mortality even in Het pups. Thus, we conclude that HO-1 plays an important role in the protection against preterm sepsis.

Hemolytic Anemia and Gastric Carcinoid in a Russian Seafarer: Highlighting the Role of Diagnostic Technologies in Modern Clinical Practice.

Vitamin B-12 deficiency, most commonly due to pernicious anemia, can cause intramedullary hemolysis. The pathogenesis is thought to be due to increased membrane rigidity and reduced red blood cell elasticity, which predisposes the patient to hemolysis and microangiopathic hemolytic anemia. In this article, we discuss a Russian engineer who worked aboard a petroleum tanker that presented from his ship with profound B-12 deficiency, microangiopathic anemia, elevated lactate dehydrogenase levels, low haptoglobin, and reticulocyte count in the setting of normal renal and neurologic function. The patient traveled around the world seven months of the year for work and had occupational exposure to fluorinated hydrocarbons. Extensive diagnostic work-up, including endoscopic biopsy, and a radio-labeled octreotide scan was performed. The patient was found to have autoimmune gastritis and a gastric carcinoid tumor. With assistance from his global health insurance provider and a local hospital near his hometown in Russia, care was coordinated to be transitioned there with a plan for repeat endoscopy and mapping biopsies to determine the extent of his tumor burden. This study adds to the now growing base of literature describing this atypical presentation of pernicious anemia with normal neurologic function and underscores the importance of screening for B-12 deficiency in these patients. It also highlights the increased risk of gastric carcinoids in patients with autoimmune gastritis. With the collaboration of different medical specialists, the full gamut of medical technology was utilized in the care of the patient. This included in vitro diagnostics, advanced endoscopic tools, pathology, and radio-isotope based imaging studies.

Hemolytic anemia repressed hepcidin level without hepatocyte iron overload: lesson from Günther disease model.

Hemolysis occurring in hematologic diseases is often associated with an iron loading anemia. This iron overload is the result of a massive outflow of hemoglobin into the bloodstream, but the mechanism of hemoglobin handling has not been fully elucidated. Here, in a congenital erythropoietic porphyria mouse model, we evaluate the impact of hemolysis and regenerative anemia on hepcidin synthesis and iron metabolism. Hemolysis was confirmed by a complete drop in haptoglobin, hemopexin and increased plasma lactate dehydrogenase, an increased red blood cell distribution width and osmotic fragility, a reduced half-life of red blood cells, and increased expression of heme oxygenase 1. The erythropoiesis-induced Fam132b was increased, hepcidin mRNA repressed, and transepithelial iron transport in isolated duodenal loops increased. Iron was mostly accumulated in liver and spleen macrophages but transferrin saturation remained within the normal range. The expression levels of hemoglobin-haptoglobin receptor CD163 and hemopexin receptor CD91 were drastically reduced in both liver and spleen, resulting in heme- and hemoglobin-derived iron elimination in urine. In the kidney, the megalin/cubilin endocytic complex, heme oxygenase 1 and the iron exporter ferroportin were induced, which is reminiscent of significant renal handling of hemoglobin-derived iron. Our results highlight ironbound hemoglobin urinary clearance mechanism and strongly suggest that, in addition to the sequestration of iron in macrophages, kidney may play a major role in protecting hepatocytes from iron overload in chronic hemolysis.

Rapid conditional targeted ablation model for hemolytic anemia in the rat.

Effective methods for cell ablation are important tools for examining the anatomical, functional, and behavioral consequences of selective loss of specific cell types in animal models. We have developed an ablation system based on creating genetically modified animals that express human CD59 (hCD59), a membrane receptor, and administering intermedilysin (ILY), a toxin produced by Streptococcus intermedius, which binds specifically to hCD59 to induce cell lysis. As proof-of-concept in the rat, we generated an anemia model, SD-Tg(CD59-HBA1)Bryd, which expresses hCD59 on erythrocytes. Hemolysis is a common complication of inherited or acquired blood disorders, which can result in cardiovascular compromise and death. A rat model that can replicate hemolysis through specific ablation of erythrocytes would allow further study of disease and novel treatments. In vitro, complete lysis of erythrocytes expressing hCD59 was observed at and above 250 pM ILY, while no lysis was observed in wild-type erythrocytes at any ILY concentration (8-1,000 pM). In vivo, ILY intravenous injection (100 ng/g body wt) dramatically reduced the hematocrit within 10 min, with a mean hematocrit reduction of 43% compared with 1.4% in the saline control group. Rats injected with ILY at 500 ng/g intraperitoneally developed gross signs of anemia. Histopathology confirmed anemia and revealed hepatic necrosis, with microthrombi present. These studies validate the hCD59-ILY cell ablation technology in the rat and provide the scientific community with a new rapid conditional targeted ablation model for hemolytic anemia and hemolysis-associated sequelae.

A new model of an arteriovenous fistula in chronic kidney disease in the mouse: beneficial effects of upregulated heme oxygenase-1.

The arteriovenous fistula (AVF) is the preferred hemodialysis vascular access, but it is complicated by high failure rates and attendant morbidity. This study provides the first description of a murine AVF model that recapitulates two salient features of hemodialysis AVFs, namely, anastomosis of end-vein to side-artery to create the AVF and the presence of chronic kidney disease (CKD). CKD reduced AVF blood flow, observed as early as 3 days after AVF creation, and increased neointimal hyperplasia, venous wall thickness, thrombus formation, and vasculopathic gene expression in the AVF. These adverse effects of CKD could not be ascribed to preexisting alterations in blood pressure or vascular reactivity in this CKD model. In addition to vasculopathic genes, CKD induced potentially vasoprotective genes in the AVF such as heme oxygenase-1 (HO-1) and HO-2. To determine whether prior HO-1 upregulation may protect in this model, we upregulated HO-1 by adeno-associated viral gene delivery, achieving marked venous induction of the HO-1 protein and HO activity. Such HO-1 upregulation improved AVF blood flow and decreased venous wall thickness in the AVF. Finally, we demonstrate that the administration of carbon monoxide, a product of HO, acutely increased AVF blood flow. This study thus demonstrates: 1) the feasibility of a clinically relevant murine AVF model created in the presence of CKD and involving an end-vein to side-artery anastomosis; 2) the exacerbatory effect of CKD on clinically relevant features of this model; and 3) the beneficial effects in this model conferred by HO-1 upregulation by adeno-associated viral gene delivery.

EPO-dependent induction of erythroferrone drives hepcidin suppression and systematic iron absorption under phenylhydrazine-induced hemolytic anemia.

Hemolytic anemia is a common form of anemia due to hemolysis, resulting in disordered iron homeostasis. In this study, a dose of 40mg/kg phenylhydrazine (PHZ) was injected into mice to successfully establish a pronounced anemia animal model, which resulted in stress erythropoiesis and iron absorption. We found that serum erythropoietin (EPO) concentration was dramatically elevated by nearly 5000-fold for the first 2days, and then drop to the basal level on day 6 after PHZ injection. Mirrored with serum EPO concentration, the mRNA expression of erythroferrone (ERFE) was rapidly increased in the bone marrow and spleen 3days after injection of PHZ, and then gradually decreased but was still higher than baseline on day 6. In addition, we also found that the hepcidin mRNA levels were gradually reduced almost up to 8-fold on day 5, and then was ameliorated compared to the untreated control. Mechanistic investigation manifested that the increase of serum EPO essentially determined the induction of ERFE expression particular at the first 3days after PHZ treatment. Lentiviral mediated ERFE knockdown significantly restrained hepcidin suppression under PHZ treatment. Thus, our data unearthed EPO-dependent ERFE expression acts as an erythropoiesis-driven regulator of iron metabolism under PHZ-induced hemolytic anemia.

Effect of Heme Oxygenase-1 Deficiency on Glomerular Proteomics.

BACKGROUND: The cytoprotective effect of heme oxygenase (HO)-1 in various forms of renal glomerular injury is established. However, little is known on the role of HO-1 in preserving glomerular structural/functional integrity in the absence of injury. The present study addressed this question in HO-1-deficient rats. METHODS: HO-1-deficient rats were generated using zinc finger nuclease-mediated HO-1 gene (Hmox1) disruption and studied. Glomeruli were isolated from HO-1-deficient (Hmox1-/-) rats and their wild type (WT) littermates for proteomic analysis. RESULTS: Glomerular lesions were characterized and differentially expressed proteins important for preserving integrity of the glomerular filtration barrier were identified. HO-1-deficient (Hmox1-/-) rats developed albuminuria with decreased glomerular filtration rate. In albuminuric rats, there were lesions resembling focal and segmental glomerulosclerosis (FSGS). Western blot analysis of the integral slit diaphragm proteins, nephrin and podocin revealed a significant decrease in nephrin, with no change in podocin. Proteomic analysis of glomerular protein lysates from Hmox1-/- and WT rats revealed differential expression of proteins previously linked with FSGS pathogenesis. Specifically, α-actinin-4, actin related protein 3, cytokeratins and novel candidates including transgelin-2 and lamins. Bioinformatic analysis predicted the upregulation of pathways implicated in platelet aggregation and fibrin clot formation. CONCLUSION: HO-1 is a putative regulator of proteins important in preserving glomerular structural stability and integrity, and in minimizing the activity of proinflammatory pathways.

Deficiency of nicotinamide mononucleotide adenylyltransferase 3 (nmnat3) causes hemolytic anemia by altering the glycolytic flow in mature erythrocytes.

NAD biosynthesis is of substantial interest because of its important roles in regulating various biological processes. Nicotinamide mononucleotide adenylyltransferase 3 (Nmnat3) is considered a mitochondria-localized NAD synthesis enzyme involved in de novo and salvage pathways. Although the biochemical properties of Nmnat3 are well documented, its physiological function in vivo remains unclear. In this study, we demonstrated that Nmnat3 was localized in the cytoplasm of mature erythrocytes and critically regulated their NAD pool. Deficiency of Nmnat3 in mice caused splenomegaly and hemolytic anemia, which was associated with the findings that Nmnat3-deficient erythrocytes had markedly lower ATP levels and shortened lifespans. However, the NAD level in other tissues were not apparently affected by the deficiency of Nmnat3. LC-MS/MS-based metabolomics revealed that the glycolysis pathway in Nmnat3-deficient erythrocytes was blocked at a glyceraldehyde 3-phosphate dehydrogenase (GAPDH) step because of the shortage of the coenzyme NAD. Stable isotope tracer analysis further demonstrated that deficiency of Nmnat3 resulted in glycolysis stall and a shift to the pentose phosphate pathway. Our findings indicate the critical roles of Nmnat3 in maintenance of the NAD pool in mature erythrocytes and the physiological impacts at its absence in mice.

Gain-of-function Lyn induces anemia: appropriate Lyn activity is essential for normal erythropoiesis and Epo receptor signaling.

Lyn is involved in erythropoietin (Epo)-receptor signaling and erythroid homeostasis. Downstream pathways influenced following Lyn activation and their significance to erythropoiesis remain unclear. To address this, we assessed a gain-of-function Lyn mutation (Lyn(up/up)) on erythropoiesis and Epo receptor signaling. Adult Lyn(up/up) mice were anemic, with dysmorphic red cells (spherocyte-like, acanthocytes) in their circulation, indicative of hemolytic anemia and resembling the human disorder chorea acanthocytosis. Heterozygous Lyn(+/up) mice became increasingly anemic with age, indicating that the mutation was dominant. In an attempt to overcome this anemia, extramedullary erythropoiesis was activated. As the mice aged, the levels of different immature erythroid populations changed, indicating compensatory mechanisms to produce more erythrocytes were dynamic. Changes in Epo signaling were observed in Lyn(+/up) erythroid cell lines and primary CD71(+) Lyn(up/up) erythroblasts, including significant alterations to the phosphorylation of Lyn, the Epo receptor, Janus kinase 2, Signal Transducer and Action of Transcription-5, GRB2-associated-binding protein-2, Akt, and Forkhead box O3. As a consequence of altered Lyn signaling, Lyn(+/up) cells remained viable in the absence of Epo but displayed delayed Epo-induced differentiation. These data demonstrate that Lyn gene dosage and activity are critical for normal erythropoiesis; constitutively active Lyn alters Epo signaling, which in turn produces erythroid defects.

Effects of aluminum chloride on some trace elements and erythrocyte osmotic fragility in rats.

Aluminum (Al) is a nonessential, toxic element to which humans are constantly exposed as a result of an increase in industrialization and improving technology practices. The aim of the study was to investigate the effects of different durations and doses of Al exposure on serum and tissue element levels and erythrocyte osmotic fragility in rats. A total of 40 male Wistar Albino rats were divided into five groups: control, group I (3 weeks, 8 mg/kg), group II (6 weeks, 8 mg/kg), group III (3 weeks, 16 mg/kg), and group IV (6 weeks, 16 mg/kg). Al chloride (AlCl3) was injected intraperitoneally (i.p.) five times a week. At the end of the experimental period, levels of Al, iron (Fe), copper (Cu), and zinc (Zn) in serum, liver, and kidney tissues were measured using inductively coupled plasma optical emission spectrometry. Osmotic fragility was determined using a spectrophotometer. The results of the experiment indicate that Al induced a statistically significant increase in Al and Fe concentrations in liver and serum as well as in Cu in the kidney. The Fe concentration in serum and kidney tissues was significantly lower in all the groups. As a result of our study, it may be concluded that tissue Al accumulation may lead to an increase in osmotic fragility of erythrocytes and abnormal trace element levels.