Sickle Cell Anemia and Babesia Infection

Babesia is an intraerythrocytic, obligate Apicomplexan parasite that has, in the last century, been implicated in human infections via zoonosis and is now widespread, especially in parts of the USA and Europe. It is naturally transmitted by the bite of a tick, but transfused blood from infected donors has also proven to be a major source of transmission. When infected, most humans are clinically asymptomatic, but the parasite can prove to be lethal when it infects immunocompromised individuals. Hemolysis and anemia are two common symptoms that accompany many infectious diseases, and this is particularly true of parasitic diseases that target red cells. Clinically, this becomes an acute problem for subjects who are prone to hemolysis and depend on frequent transfusions, like patients with sickle cell anemia or thalassemia. Little is known about Babesia’s pathogenesis in these hemoglobinopathies, and most parallels are drawn from its evolutionarily related Plasmodium parasite which shares the same environmental niche, the RBCs, in the human host. In vitro as well as in vivo Babesia-infected mouse sickle cell disease (SCD) models support the inhibition of intra-erythrocytic parasite proliferation, but mechanisms driving the protection of such hemoglobinopathies against infection are not fully studied. This review provides an overview of our current knowledge of Babesia infection and hemoglobinopathies, focusing on possible mechanisms behind this parasite resistance and the clinical repercussions faced by Babesia-infected human hosts harboring mutations in their globin gene.

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Aroyl-Pyrrolyl-Hydroxy-Amides (APHAs), a Novel Family of Synthetic Histone Deacetylases Inhibitors, Are Potent Inducers of Human g-Globin Gene Expression.

Blood ◽

10.1182/blood.v104.11.1216.1216 ◽

2004 ◽

Vol 104 (11) ◽

pp. 1216-1216

Author(s):

Antonello Mai ◽

Silvio Massa ◽

Antonella Di Noia ◽

Katija Jelicic ◽

Elena Alfani ◽

...

Keyword(s):

Gene Expression ◽

Sickle Cell ◽

Sickle Cell Anemia ◽

Histone Deacetylases ◽

Globin Gene ◽

Renilla Luciferase ◽

In Vitro Assays ◽

Luciferase Reporter ◽

Globin Gene Expression

Abstract Post-natal pharmacological reactivation of HbF, by restoring the unbalanced α/non-α globin chain production in red cells of patients affected by β-thalassemia or sickle cell anemia, represents a potential cure for these diseases. Many classes of compounds have been identified capable to induce Hb F synthesis in vitro by acting at different levels of the globin gene expression regulatory machinery. One of these classes is represented by inhibitors of a family of enzymes, the histone deacetylases (HDACs), involved in chromatin remodelling and gene transcription regulation. HDACs act in multi-protein complexes that remove acetyl groups from lysine residues on several proteins, including histones and are divided into three distinct structural classes, depending on whether their catalytic activity is zinc (class I/II)- or NAD+ (class III)-dependent. The effects of the HDACs inhibitors identified so far on HbF synthesis is, however, modest and often associated with high toxicity. Therefore, the potential of their clinical use is unclear. We have recently described a new family of synthetic HDACs inhibitors, the Aroyl-pyrrolyl-hydroxy-amides (APHAs), that induce differentiation, growth arrest and/or apoptosis of transformed cell in culture [Mai A et al, J Med Chem2004;47:1098]. In this study, we investigate the capability of 10 different APHA compounds to induce Hb F in two in vitro assays. One assay is based on the ability of APHA compounds to activate either the human Aγ-driven Firefly (Aγ-F) or the β-promoter drives Renilla Luciferase (β-R) reporter in GM979 cells stably transfected with a Dual Luciferase Reporter construct. The second assay is represented by the induction of γ-globin expression (by quantitative RT-PCR) in primary adult erythroblasts obtained in HEMA cultures of mononuclear cells from normal donors. The majority of the compounds tested did not significantly increased the Aγ−F (Aγ−F+β−R) reporter ratio in GM979 cells. However, the compound MC1575 increased by 3-fold (from 0.09 to 0.30) the reporter ratio in GM979 cells at a concentration of 20 μM, with modest effects of the proliferation activity of GM979 cells over the three days of the assay. When MC1575 was added at a concentration of 2–10 μM in cultures of primary adult erythroblasts induced to differentiate in serum-free media for 4 days, it induced a three fold increase of the γ/(γ+β) globin ratio (from 0.04 to 0.12), with no apparent cellular toxicity. Among the HDAC inhibitors tested in this study, MC1575 was not the most potent inhibitor of total enzyme activity. However, it was the compound that most selectively inhibited the activity of the maize homologue of mammalian class IIa HDAC enzymes [Mai et al, J Med Chem2003;46:4826]. These results are consistent with the hypothesis that each class of histone deacetylases might have a specific biological function and indicate that those of class IIa might represent the enzymes most specifically involved in globin gene regulation. We suggest that, by targeting the chemical inhibitors toward the catalytic domain of this class of enzymes, it should be possible to identify more specific, more potent and less toxic compounds for pharmacological treatment of β-thalassemia or sickle cell anemia.

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Phytomedicines and Nutraceuticals: Alternative Therapeutics for Sickle Cell Anemia

The Scientific World JOURNAL ◽

10.1155/2013/269659 ◽

2013 ◽

Vol 2013 ◽

pp. 1-12 ◽

Cited By ~ 24

Author(s):

Ngozi Awa Imaga

Keyword(s):

Sickle Cell ◽

Sickle Cell Anemia ◽

Globin Gene ◽

Alternative Therapy ◽

Clinical Manifestations ◽

Base Substitution ◽

Beta Globin ◽

Inherited Disease ◽

Gene Encoding

Sickle cell anemia is a genetically inherited disease in which the “SS” individual possesses an abnormal beta globin gene. A single base substitution in the gene encoding the humanβ-globin subunit results in replacement ofβ6 glutamic acid by valine, leading to the devastating clinical manifestations of sickle cell disease. This substitution causes drastic reduction in the solubility of sickle cell hemoglobin (HbS) when deoxygenated. Under these conditions, the HbS molecules polymerize to form long crystalline intracellular mass of fibers which are responsible for the deformation of the biconcave disc shaped erythrocyte into a sickle shape. First-line clinical management of sickle cell anemia include, use of hydroxyurea, folic acid, amino acids supplementation, penicillinprophylaxis, and antimalarial prophylaxis to manage the condition and blood transfusions to stabilize the patient's hemoglobin level. These are quite expensive and have attendant risk factors. However, a bright ray of hope involving research into antisickling properties of medicinal plants has been rewarding. This alternative therapy using phytomedicines has proven to not only reduce crisis but also reverse sickling (in vitro). The immense benefits of phytomedicines and nutraceuticals used in the management of sickle cell anemia are discussed in this paper.

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A Library of Sickle Cell Anemia Induced Pluripotent Stem Cells of Diverse Haplotypes and Ethnicities

Blood ◽

10.1182/blood.v126.23.2354.2354 ◽

2015 ◽

Vol 126 (23) ◽

pp. 2354-2354

Author(s):

Seonmi Park ◽

Andreia Gianotti-Sommer ◽

David H.K. Chui ◽

Maria Stella Figueiredo ◽

Abdulrahman Alsultan ◽

...

Keyword(s):

Gene Expression ◽

Stem Cells ◽

Induced Pluripotent Stem Cells ◽

Sickle Cell ◽

Sickle Cell Anemia ◽

Pluripotent Stem Cells ◽

Globin Gene ◽

Genetic Modifiers ◽

Induced Pluripotent

Abstract The mutation causing sickle cell anemia (rs334, GAG-GTG, glu6val) had several independent origins in Africa, the Middle East and India and spread throughout parts of the world by wars, slave trading and population migrations. The genetic background upon which the HbS mutation occurred, or the β-globin gene (HBB) haplotype, is associated with differences in the phenotype of this disease and the ability of affected individuals to synthesize fetal hemoglobin (HbF). The main modifier of the disease phenotype is the level of HbF in the blood of affected individuals. HbF inhibits the polymerization of HbS, the proximate cause of disease pathophysiology. As part of the NHLBI NextGen consortium (U01HL107443) we established a library of induced pluripotent stem cells (iPSC) from patients with sickle cell anemia of diverse HBB haplotypes and HbF phenotypes. The purpose of establishing this library was to allow genetic studies of globin gene expression during the erythroid differentiation of iPSC of diverse genotypes. During these studies we have implemented an efficient and highly reproducible platform for the production of large numbers of sickle cell anemia-specific iPSC, derived and characterized a novel in vitro system for the production of an unlimited supply of erythroid lineage cells from the directed differentiation of normal and disease-specific iPSC and used this system to recapitulate erythroid-lineage ontogeny in vitro with the sequential development of primitive and definitive erythropoiesis, accompanied by the appropriate expression of stage-specific globin genes. We have recently finished whole genome DNA and RNA sequencing analysis in some of these lines aimed at identifying developmental gene expression profile differences between erythroid precursors that produce primarily HbF and those that produce primarily HbA or HbS as part of our search for novel HbF genetic modifiers associated with markedly elevated HbF levels found in sickle cell anemia patients naturally, or in response to hydroxyurea treatment. Furthermore, our labs are also focusing on using a CRISPR-based gene editing platform to study the effect of novel HbF genetic modifiers and explore globin switching. Cell lines established are shown in the table. Table 1. Number of subjects recruited to date 98 Number of subjects with iPSC lines established 56 Average number of iPSC lines per subject 3 (total of 158 lines generated) Quality control status of iPSC lines All lines are expanded and banked, mycoplasma free, express pluripotency markers Subjects with target cells differentiated (erythrocytes) 25 Samples have been collected on African American patients with sickle cell anemia with diverse HBB haplotypes, predominantly homozygotes and compound heterozygotes for the Benin and Bantu haplotypes, Saudi Arabian patients with the Arab-Indian haplotype and the Saudi Benin haplotype that is characterized by HbF levels about twice as high as in African Benin haplotype patients and from Brazilian patients who are predominantly homozygotes for the Bantu haplotype that typically is associated with the lowest HbF of all HBB haplotypes. This iPSC-based library and the data associated with it represents a valuable readily available resource for the sickle cell research community and all the generated lines will be available for distribution early in 2016 through WiCell. Disclosures No relevant conflicts of interest to declare.

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Sickling Cells, Cyclic Nucleotides, and Protein Kinases: The Pathophysiology of Urogenital Disorders in Sickle Cell Anemia

Anemia ◽

10.1155/2012/723520 ◽

2012 ◽

Vol 2012 ◽

pp. 1-13 ◽

Cited By ~ 4

Author(s):

Mário Angelo Claudino ◽

Kleber Yotsumoto Fertrin

Keyword(s):

Protein Kinases ◽

Sickle Cell ◽

Sickle Cell Anemia ◽

Cyclic Nucleotides ◽

Single Point ◽

Clinical Manifestations ◽

Single Point Mutation ◽

Tract Infections

Sickle cell anemia is one of the best studied inherited diseases, and despite being caused by a single point mutation in theHBBgene, multiple pleiotropic effects of the abnormal hemoglobin S production range from vaso-occlusive crisis, stroke, and pulmonary hypertension to osteonecrosis and leg ulcers. Urogenital function is not spared, and although priapism is most frequently remembered, other related clinical manifestations have been described, such as nocturia, enuresis, increased frequence of lower urinary tract infections, urinary incontinence, hypogonadism, and testicular infarction. Studies on sickle cell vaso-occlusion and priapism using bothin vitroandin vivomodels have shed light on the pathogenesis of some of these events. The authors review what is known about the deleterious effects of sickling on the genitourinary tract and how the role of cyclic nucleotides signaling and protein kinases may help understand the pathophysiology underlying these manifestations and develop novel therapies in the setting of urogenital disorders in sickle cell disease.

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Study on the Efficacy of a JAK Inhibitor Pharmacological Agent As Inducer of Fetal Hemoglobin Production in Cultured Erythroid Precursors from Sickle Cell and Beta-Thalassemia Patients

Blood ◽

10.1182/blood.v126.23.2048.2048 ◽

2015 ◽

Vol 126 (23) ◽

pp. 2048-2048

Author(s):

Alice Pecoraro ◽

Antonio Troia ◽

Angela Vitrano ◽

Rosario Di Maggio ◽

Massimiliano Sacco ◽

...

Keyword(s):

Gene Expression ◽

Sickle Cell ◽

Globin Gene ◽

Beta Thalassemia ◽

Globin Mrna ◽

Gamma Globin ◽

Long Term Treatment ◽

Globin Gene Expression

Abstract Phenotypic improvement of hemoglobinopathies such as sickle cell disease and beta-thalassemia (beta-thal) has been shown in patients with high levels of fetal hemoglobin (HbF). In sickle cell disease (SCD) the beneficial effects of HbF are due to the inhibition of HbS polymerization and to the dilution of HbS determining the reduction of sickling and vascular occlusion. Moreover, in beta-thal, high levels of gamma-chains combined with the redundant alpha-chains, lead to a reduction of dyserythropoiesis and of the requirement for blood transfusions. The only drug approved for the treatment of adult patients with SCD and that has been entered in clinical practice of patients affected by beta-thal is hydroxyurea (HU); however there is a great variability in the responses of patients to HU, in fact some patients are good responder, while others exhibit little or no change in HbF levels after HU treatment; moreover a decrease in the efficacy during long term treatment was observed. Other pharmacological compounds, including 5-azacytidine and thalidomide have been shown to increase HbF production. Due to concerns about the safety of this agents, their use was limited to severe cases for whom conventional therapy was unfeasible. For this reason the search of new inducers of HbF production is important. Ruxolitinib is a JAK inhibitor and decreases the phosphorilation of STAT (Signal transducers and activators of transcription) family proteins, in particular STAT5 and STAT3. Phosphorylation of STAT5 is essential for basal erythropoiesis and for its acceleration during stress erythropoiesis. STAT3 plays an essential role in regulating gene expression of several genes involved in cell growth and apoptosis, in particular it was demonstrated to inhibit gamma-globin gene expression. The decrease of STAT3 phoshorilation could decrease the inhibition of gamma-globin gene expression; for this reason we considered ruxolitinib a candidate as inducer of HbF production. In our laboratory an ex vivo system was developed predictive of the in vivo response to hydroxyurea treatment by using liquid erythroid cultures, an in vitro culture system that recapitulates the process of human erythropoiesis. To evaluate the efficacy of ruxolitinib in increasing gamma-globin gene expression we carried out a study in vitro using liquid erythroid cultures. In this study we developed and exposed to ruxolitinib liquid erythroid precursors from 4 SCD and 17 beta-thal intermedia (beta-TI) patients. The use of quantitative Real-Time-polymerase chain reaction allowed us to determine the increase in gamma-globin mRNA expression in human erythroid cells treated with ruxolitinib compared to untreated cells. The results are summarized in Table 1 and showed that ruxolitinib at 200nM is able to determine a significant increase of gamma-globin gene expression (3.4±0.1)compared to HU (2.0± 0.2). In conclusion our study suggests that ruxolitinib could be considered an inducer of HbF and could be used in vivo for the treatment of hemoglobinopathies, particularly in patients who do not respond to HU therapy or who show a decreased response after long-term treatment. Table 1. Fold increase of Gamma-globin gene expression in presence of Ruxolitinib in erythroid cultured cells. Patient Sex Genotype gamma-globin mRNA fold increasein the presence of ruxolitinib #1 M b039/aaa +1 #2 F b039/aaa +1.65 #3 F b039/b039 +1.9 #4 F b039/IVS1,110 +1.5 #5 M IVS1,1/aaa +2.5 #6 M IVS1,110/IVS1,1 +9.2 #7 M b039/bs +6 #8 F bs/b039 +1.6 #9 F b039/IVS1,6 +1.7 #10 M IVS1,6/frcd6 +3 #11 M IVS1,6/bs +2.5 #12 M IVS1,6/frcd6 +8 #13 F IVS1,6/b039 +9 #14 M IVS1,1/b039 +2.2 #15 M db/IVS1,110 +8 #16 F db/IVS1,110 +1.8 #17 F IVS2,1/aaa +3.9 #18 M b039/-101 +1.4 #19 M IVS1,6/b039 +1 #20 M bs/IVS1,110 +1.4 #21 M IVS1,6/IVS1,6 +1.9 Disclosures No relevant conflicts of interest to declare.

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Abnormal Cytokine Production By Mast Cell Cultures from Sickle Cell Anemia Patients in Response to Inflammatory Stimuli and to Co-Culture with Eosinophils

Blood ◽

10.1182/blood-2019-129509 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 3566-3566

Author(s):

Myriam Salazar-Terreros ◽

Kleber Yotsumoto Fertrin ◽

Nicolas Moreno Reyes ◽

Fernando Ferreira Costa ◽

Carla F. Franco-Penteado

Keyword(s):

Mast Cell ◽

Sickle Cell ◽

Sickle Cell Anemia ◽

Peripheral Blood ◽

Cytokine Production ◽

Fold Increase ◽

Individual Response ◽

Activated State

Mast cell function (MC) in pathologic states can be studied through their ability to secrete mediators in vitro depending on MC phenotype and the nature of the stimuli. Reports on MC mediators in sickle cell anemia (SCA) patients are scarce, but clinical signs of MC activation syndrome, such as increased plasma histamine in vaso‐occlusive crisis (VOC), and normal or slightly elevated serum tryptase have been reported. However, assessing the biological relevance of MC as a cytokine source is more challenging because it is unclear under which circumstances they secrete those products in vivo, or if the cytokines measured systemically stem from a different cell type. We aimed to investigate the profile of mediators involved in the inflammatory process produced by MC in SCA. Methods:The supernatant of 5-week old MC cultures (17 SCA, 8 HV) obtained from peripheral blood CD34+ cells from 29 SCA patients and 13 healthy volunteers (HV) was analyzed using a multiplex platform and colorimetric assays for endothelin-1 (ET-1) and substance P (SP) (10 SCA, 6 HV). A correlation matrix (Pearson correlations, R software, v. 3.6.1) was generated using laboratory and clinical data chosen based on their value as inflammatory or prognosis markers (hydroxyurea [HU] treatment, fetal hemoglobin [HbF], hemoglobin [Hb], vaso-occlusive crisis [VOC], percentage of peripheral blood neutrophils, eosinophils (Eos), basophils, erythroblasts, and reticulocytes), MC surface expression of CD117, CD48 and CD63, and the supernatant content of 11 cytokines. To investigate MC cytokine release, we tested the supernatants from Eos-MC co-cultures (3:1 ratio), and after stimulation with ET-1 (20 nM), SP (10 µM) and imatinib (20 µg/ml)(n=3 per treatment). Results: Out of 26 cytokines, we found elevated levels of the following in the supernatants of SCA-MC cultures (data represented as mean in pg/ml±SE): TNFα: SCA=88.7±18.4, HV=32.6±3.8; IFNγ: SCA=55.3±11.2, HV=15.7±1.8; MCP1: SCA=555.0±147.2, HV=145.3±35.2; RANTES: SCA=24.7±3.9, HV=10.7±1.8 (p<0.05). However, SCA-MC from patients treated with HU (n = 11) showed higher values of IL-1b, IL-4, IL-5, IL-9, IL-15, and FGF than HV (n=8) and HU-free patients (n=6) (p<0.05). Supernatants from SCA-MC had higher ET-1 production compared to HV-MC (SCA=16.3±1.2, HV=11.93±1.3, pg/ml, p=0.02) but SP production was similar (SCA=27.9±1.3; HV=31.49±0.7 pg/ml). ET-1 stimulation of MC cultures caused 2-fold increase in IL-1AR production on HV-MC, but failed to produce any effect on SCA-MC. Similarly, imatinib reduced FGF only in HV-MC samples (HV: 15.1±3.5, HV-HU: 4.0±1.6, pg/ml). No effect on cytokine production was observed with SP. Conversely, Eos-MC cocultures showed a 10- and 4-fold increase of IL-5 and IL-9, respectively, regardless of the origin of Eos (HV or SCA). SCA-MC/SCA-Eos co-cultures had elevated proinflammatory (IL-1b, IL-12, TNF-α) and angiogenic (FGF, VEGF) cytokines, RANTES, IL-7, IL-4, and IL1-RA compared with SCA-MC/HV-Eos and HV-MC/HV-Eos (p<0.05). Preliminary multiparametric analysis on data from SCA patients showed a strong negative correlation between HU therapy and VGEF production, and between HbF levels and CD63 expression (MC activation marker).We also found a positive correlation between history of VOC and eotaxin-1 produced by SCA-MC. Conclusions: We found that MC responses depend both on the origin of the cultured cell and the stimuli utilized. Despite differences between in vitro and in vivo MC populations, our data show that cultured SCA-MC have a sustained activated state and produce a repertoire of mediators that could contribute to a perivascular microenvironment in favor of leukocyte and endothelium activation. In terms of cytokine production, cultured SCA-MC were more sensitive to stimulation by SCA-Eos than by HV-Eos, which may be relevant to the pathophysiology of airway inflammation in SCA patients with asthma. Differences in cytokine production between SCA-MC cultures from patients treated or not with HU may reflect the variability in adherence to treatment, individual response to each compound, or epigenetic modifications during the MC differentiation process that affect the phenotype of the mature MC. These results support that mediators produced by MC can contribute to the chronic inflammatory state and may be implicated in exacerbated responses to eosinophil activation in SCA. Disclosures Fertrin: Agios Pharmaceuticals, Inc.: Research Funding.

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Metabolomic Characterization of Red Blood Cell Differentiation

Blood ◽

10.1182/blood-2020-137175 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 35-35

Author(s):

Kelsey Temprine ◽

Amanda Sankar ◽

Costas Lyssiotis ◽

Yatrik Shah

Keyword(s):

Amino Acid ◽

Sickle Cell ◽

Sickle Cell Anemia ◽

Sodium Butyrate ◽

Global Changes ◽

Ineffective Erythropoiesis ◽

Irreversible Loss ◽

In Vivo Models

Background: Erythropoiesis is the highly coordinated multi-step process by which multipotent hematopoietic stem cells differentiate into mature enucleated red blood cells (RBCs). As erythroid cells become more terminally differentiated, they undergo changes in morphology and gene expression, start synthesizing hemoglobin, commit to an irreversible loss of proliferation, and eventually expulse their nuclei and other cytoplasmic organelles. Thus, RBCs must rely on their proteome and metabolome for proper function. The RBC proteome is estimated to contain 2,800 proteins, including a variety of receptors and transporters that allow RBCs to uptake xenobiotics or endogenous metabolites as they circulate for ∼120 days. Furthermore, they are metabolically active with glycolysis, nucleotide catabolism, and glutathione metabolism as the major pathways supporting cell survival and function. However, it is unclear how the metabolome is altered during erythropoiesis, what role metabolites play in normal erythropoiesis, and if dysregulation of metabolites contributes to diseases of ineffective erythropoiesis, such as sickle cell anemia and thalassemia. Methods: Four models of erythropoiesis were used in this study. 1) Mice were treated with phenylhydrazine (Phz) to induce acute hemolysis followed by erythropoietic recovery, leading to an increase in circulating reticulocytes. 2) Mice were lethally irradiated and transplanted with wild-type or sickle cell bone marrow, leading to anemic profiles in sickle cell chimeras. 3) The mouse erythroleukemic (MEL) cell line was treated with DMSO to induce differentiation. 4) The human erythroleukemic (K562) cell line was treated with sodium butyrate to induce differentiation. For the in vivo mouse models, blood was collected from control and treated animals, and complete blood count (CBC) analysis was performed. For the in vitro cell culture models, the mRNA levels of β-globin were measured by Q-RT-PCR in control and differentiated cells, and the degree of hemoglobinization was determined visually and via staining for heme. In addition, metabolites were extracted from the collected RBCs and erythroleukemic cell lines, and a Snapshot LC/MS metabolomic platform was used to identify commonly altered metabolites. Results: We first validated our four models of erythropoiesis. Treatment with Phz decreased the number of total RBCs while increasing the RBC distribution width, indicating an increased number of reticulocytes (more immature RBCs) in circulation. Similar results were seen in the sickle cell chimeras. Treatment of MEL and K562 cells with DMSO and sodium butyrate, respectively, resulted in increased expression of β-globin, increased levels of heme, and increased red color. Then, using our Snapshot metabolomic platform, we identified global changes in RBC metabolism during erythropoiesis. Analyses of the commonly altered metabolites in the in vitro and in vivo models revealed an increase in amino acid, mitochondrial, and urea cycle metabolism during erythropoiesis. L-aspartate levels were particularly upregulated, especially in DMSO-treated MEL cells. We are now investigating the role of aspartate in the regulation of erythropoiesis. Conclusions: We defined how the metabolome was altered in multiple in vitro and in vivo models of erythropoiesis and identified global changes in RBC metabolism between the different models. Specifically, we found that L-aspartate was upregulated during RBC differentiation in all four models. Aspartate is an amino acid that plays a role in many processes in cells, including nucleotide biosynthesis, redox homeostasis, and amino acid biosynthesis. We hypothesize that aspartate metabolism is critical for RBC differentiation and that its dysregulation exacerbates disease of ineffective erythropoiesis, such as sickle cell anemia and β-thalassemia. We are currently testing its role in inducing hemoglobinization and in regulating the commitment of erythroid progenitor cells to an irreversible loss of proliferation. Overall, we believe that understanding the precise mechanisms by which cellular metabolism plays a role in proper RBC differentiation may lead to better therapies for diseases of ineffective erythropoiesis, such as sickle cell anemia and thalassemia. Disclosures No relevant conflicts of interest to declare.

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Prolongation of sickle cell survival by dimethyl adipimidate is compromised by immune sensitization

Blood ◽

10.1182/blood.v64.1.161.161 ◽

1984 ◽

Vol 64 (1) ◽

pp. 161-165 ◽

Cited By ~ 11

Author(s):

MS Guis ◽

WM Lande ◽

N Mohandas ◽

R Pennathur-Das ◽

H Preisler ◽

...

Keyword(s):

Sickle Cell Disease ◽

Sickle Cell ◽

Sickle Cell Anemia ◽

Initial Study ◽

Red Cells ◽

Adult Males ◽

Cell Disease ◽

Sickle Cells

Abstract The effect of dimethyl adipimidate (DMA), an amino-reactive crosslinking reagent with demonstrated antisickling properties in vitro, on the survival of 51Cr-labeled autologous sickle cells was evaluated in five adult males with sickle cell anemia. The survival of cells pretreated with 5 mmol/L DMA (pH 7.4), normal (t1/2 28–33 days) in four subjects and near-normal (t1/2 20 days) in the fifth, was considerably longer than that usually observed in sickle cell disease. In fact, the effect of DMA on the survival of sickle cells in vivo equals or exceeds that of any other agent tested to date. In three subjects, the survival of a second infusion of DMA-treated red cells was much shorter (t1/2 1.8, 3, 4.5 days) than in the initial study. An antibody was detected in the serum of these subjects that was directed to DMA-treated red cells. Modification of the immunogenicity of treated cells will be required before further consideration of DMA for use in the therapy of sickle cell anemia.

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THE PHYSICAL STATE OF HEMOGLOBIN IN SICKLE-CELL ANEMIA ERYTHROCYTES IN VIVO

Journal of Experimental Medicine ◽

10.1084/jem.127.4.711 ◽

1968 ◽

Vol 127 (4) ◽

pp. 711-716 ◽

Cited By ~ 43

Author(s):

Johanna Döbler ◽

John F. Bertles

Keyword(s):

Electron Microscopy ◽

Sickle Cell ◽

Sickle Cell Anemia ◽

Physical State ◽

Venous Blood ◽

Significant Loss ◽

Filamentous Form ◽

Hb S

Venous blood removed anaerobically from patients with sickle-cell anemia was transferred immediately into fixative, thus precluding significant loss or gain of oxygen by the cells. Electron microscopy demonstrated an intraerythrocytic fibrillar fine structure similar to that described in prior studies on erythrocytes sickled by deoxygenation in vitro. Observations reported here lead to these conclusions: (a) explanations of the sickling process derived from in vitro experimentation may with validity be applied to sickling in vivo; and (b) the term "sickled" must be used with caution: a sickle-shaped membrane does not necessarily endose Hb S in filamentous form.

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Prolongation of sickle cell survival by dimethyl adipimidate is compromised by immune sensitization

Blood ◽

10.1182/blood.v64.1.161.bloodjournal641161 ◽

1984 ◽

Vol 64 (1) ◽

pp. 161-165

Author(s):

MS Guis ◽

WM Lande ◽

N Mohandas ◽

R Pennathur-Das ◽

H Preisler ◽

...

Keyword(s):

Sickle Cell Disease ◽

Sickle Cell ◽

Sickle Cell Anemia ◽

Initial Study ◽

Red Cells ◽

Adult Males ◽

Cell Disease ◽

Sickle Cells

The effect of dimethyl adipimidate (DMA), an amino-reactive crosslinking reagent with demonstrated antisickling properties in vitro, on the survival of 51Cr-labeled autologous sickle cells was evaluated in five adult males with sickle cell anemia. The survival of cells pretreated with 5 mmol/L DMA (pH 7.4), normal (t1/2 28–33 days) in four subjects and near-normal (t1/2 20 days) in the fifth, was considerably longer than that usually observed in sickle cell disease. In fact, the effect of DMA on the survival of sickle cells in vivo equals or exceeds that of any other agent tested to date. In three subjects, the survival of a second infusion of DMA-treated red cells was much shorter (t1/2 1.8, 3, 4.5 days) than in the initial study. An antibody was detected in the serum of these subjects that was directed to DMA-treated red cells. Modification of the immunogenicity of treated cells will be required before further consideration of DMA for use in the therapy of sickle cell anemia.

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