BMP6 and VDR gene polymorphisms are associated with osteonecrosis in a sickle cell anaemia cohort.

The occurrence and severity of osteonecrosis in sickle cell anaemia (SCA) vary due to risk factors, including genetic modifiers. Bone morphogenetic proteins (BMPs), particularly BMP6, and the vitamin D receptor (VDR) play key roles in cartilage and bone metabolism, making them potential contributors to orthopaedic outcomes in SCA. Here, we evaluated the association of polymorphisms in BMP6 (rs3812163, rs270393 and rs449853) and VDR (FokI rs2228570 and Cdx2 rs11568820) genes with osteonecrosis risk in a Brazilian SCA cohort. A total of 177 unrelated SCA patients were selected. The AA genotype of BMP6 rs3812163 was independently associated with a lower osteonecrosis risk (p = 0.015; odds ratio (OR): 0.38; 95% confidence interval (CI): 0.18-0.83) and with the long-term cumulative incidence of osteonecrosis (p = 0.029; hazard ratio: 0.56, 95% CI: 0.34-0.94). The VDR rs2228570 TT genotype was independently associated with a lower osteonecrosis risk (p = 0.039; OR: 0.14; 95% CI: 0.02-0.90). In summary, our results provide evidence that BMP6 rs3812163 and the VDR rs2228570 might be implicated in osteonecrosis pathophysiology in SCA and might help identify individuals at high risk.

Comparative transcriptomic analysis of circulating endothelial cells in sickle cell stroke.

Ischemic stroke (IS) is one of the most impairing complications of sickle cell anemia (SCA), responsible for 20% of mortality in patients. Rheological alterations, adhesive properties of sickle reticulocytes, leukocyte adhesion, inflammation and endothelial dysfunction are related to the vasculopathy observed prior to ischemic events. The role of the vascular endothelium in this complex cascade of mechanisms is emphasized, as well as in the process of ischemia-induced repair and neovascularization. The aim of the present study was to perform a comparative transcriptomic analysis of endothelial colony-forming cells (ECFCs) from SCA patients with and without IS. Next, to gain further insights of the biological relevance of differentially expressed genes (DEGs), functional enrichment analysis, protein-protein interaction network (PPI) construction and in silico prediction of regulatory factors were performed. Among the 2469 DEGs, genes related to cell proliferation (AKT1, E2F1, CDCA5, EGFL7), migration (AKT1, HRAS), angiogenesis (AKT1, EGFL7) and defense response pathways (HRAS, IRF3, TGFB1), important endothelial cell molecular mechanisms in post ischemia repair were identified. Despite the severity of IS in SCA, widely accepted molecular targets are still lacking, especially related to stroke outcome. The comparative analysis of the gene expression profile of ECFCs from IS patients versus controls seems to indicate that there is a persistent angiogenic process even after a long time this complication has occurred. Thus, this is an original study which may lead to new insights into the molecular basis of SCA stroke and contribute to a better understanding of the role of endothelial cells in stroke recovery.

Sickle Cell Disease in Brazil: Current Management.

Sickle cell disease (SCD) comprises inherited red blood cell disorders due to a mutation in the ß-globin gene (c20A > T, pGlu6Val) and is characterized by the presence of abnormal hemoglobin, hemoglobin S, hemolysis, and vaso-occlusion. This mutation, either in a homozygous configuration or in compound states with other ß-globin mutations, leads to polymerization of hemoglobin S in deoxygenated conditions, causing modifications in red blood cell shape, particularly sickling. Vaso-occlusive crisis (VOC) is the hallmark of the disease, but other severe complications may arise from repeated bouts of VOCs. SCD is considered a global health problem, and its incidence has increased in some areas of the world, particularly the Americas and Africa. Management of the disease varies according to the region of the world, mainly due to local resources and socioeconomic status. This review aimed to describe more recent data on SCD regarding available treatment options, especially in Brazil. New treatment options are expected to be available to all patients, particularly crizanlizumab, which is already approved in the country.

Piezo1 activation augments sickling propensity and the adhesive properties of sickle red blood cells in a calcium-dependent manner.

Haemoglobin S polymerization in the red blood cells (RBCs) of individuals with sickle cell anaemia (SCA) can cause RBC sickling and cellular alterations. Piezo1 is a mechanosensitive protein that modulates intracellular calcium (Ca2+ ) influx, and its activation has been associated with increased RBC surface membrane phosphatidylserine (PS) exposure. Hypothesizing that Piezo1 activation, and ensuing Gárdos channel activity, alter sickle RBC properties, RBCs from patients with SCA were incubated with the Piezo1 agonist, Yoda1 (0.1-10 µM). Oxygen-gradient ektacytometry and membrane potential measurement showed that Piezo1 activation significantly decreased sickle RBC deformability, augmented sickling propensity, and triggered pronounced membrane hyperpolarization, in association with Gárdos channel activation and Ca2+ influx. Yoda1 induced Ca2+ -dependent adhesion of sickle RBCs to laminin, in microfluidic assays, mediated by increased BCAM binding affinity. Furthermore, RBCs from SCA patients that were homo-/heterozygous for the rs59446030 gain-of-function Piezo1 variant demonstrated enhanced sickling under deoxygenation and increased PS exposure. Thus, Piezo1 stimulation decreases sickle RBC deformability, and increases the propensities of these cells to sickle upon deoxygenation and adhere to laminin. Results support a role of Piezo1 in some of the RBC properties that contribute to SCA vaso-occlusion, indicating that Piezo1 may represent a potential therapeutic target molecule for this disease.

Filter cake extract from the beet sugar industry as an economic growth medium for the production of Spirulina platensis as a microbial cell factory for protein.

BACKGROUND: Beet filter cake (BFC) is a by-product of sugar beet processing, which is difficult to dispose of and involves severe environmental concerns. Spirulina platensis is a microalga with a high protein content essential for human and animal nutrition. The present study aimed to utilize the beet filter cake extract (BFCE) to produce Spirulina platensis commercially. However, the cultivation of S. platensis on BFCE to produce economically single-cell protein has not been reported previously. RESULTS: The batch experiment revealed the maximum dry weight at Zarrouk's medium (0.4 g/L) followed by 0.34 g/L in the treatment of 75% BFCE. The highest protein content was 50% in Zarrouk's medium, followed by 46.5% in 25% BFCE. However, adding a higher concentration of 100% BFCE led to a protein content of 31.1%. In the adaption experiment, S platensis showed an increase in dry cell weight and protein content from 25 to 75% BFCE (0.69 g/L to 1.12 g/L and 47.0% to 52.54%, respectively) with an insignificant variation compared to Zarrouk's medium (p ≤ 0.05), indicating that S. platensis can be economically produced when cultivated on 75% BFCE The predicated parameters from response surface methodology were NaNO3 (2.5 g/L), NaHCO3 (0.67 g/L), BFCE (33%) and pH = 8, which resulted in biomass yield and protein content (0.56 g/L and 52.5%, respectively) closer to that achieved using the standard Zarrouk's medium (0.6 g/L and 55.11%). Moreover, the total essential amino acid content was slightly higher in the optimized medium (38.73%) than SZM (36.98%). CONCLUSIONS: Therefore, BFCE supplemented medium could be used as a novel low-cost alternative growth medium for producing a single-cell protein with acceptable quantity and quality compared to the standard Zarrouk's medium.

Novel Insights into the Pathophysiology and Treatment of Sickle Cell Disease.

The polymerization of hemoglobin under deoxygenation is the main pathophysiological event in sickle cell diseases, described more than 70 years ago. The last two decades have seen a major increase in knowledge about the cascade of events that follow the polymerization of hemoglobin and the ensuing sickling of red blood cells. Several distinctive therapeutic targets have been discovered as a result, and a few drugs with innovative mechanisms of action are already on the market, while several others are the focus of ongoing trials. The aim of this narrative review is to describe some of the more recent data in the SCD literature regarding pathophysiology and novel treatments.

Novel inhibitor of hematopoietic cell kinase as a potential therapeutic agent for acute myeloid leukemia.

Myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML) are characterized by risk of relapses, poor survival, unwanted side effects and high toxicity with the current therapies. In light of these facts, there are efforts to develop new drugs specific for deregulated molecules that participate in leukemia pathogenesis. Hematopoietic cell kinase (HCK), an Src kinase family member, is overexpressed on hematopoietic stem cells of MDS and de novo AML patients and involved in the oncogenic process. Thus, we investigated in vitro, ex vivo and in vivo effects of a novel chemical compound targeting HCK inhibition (iHCK-37), in combination with the most used drugs for the treatment of MDS and de novo AML, 5-Azacytidine and Cytarabine. Herein, the combination treatment with iHCK-37 and 5-Azacytidine or Cytarabine demonstrated additive effects against leukemia cells, compared to either drug alone. iHCK-37 plus 5-Azacytidine or Cytarabine treatment was able to reduce the activation of oncogenic pathways, MAPK/ERK and PI3K/AKT, leading to reduction of ERK and AKT phosphorylation, and increased BAX and decreased BCL-XL protein expression. Moreover, treatment with iHCK-37 reduced MDS and AML CD34-positive cell numbers inside a 3D-structure but did not affect normal CD34-positive cell numbers. In vivo analysis showed that leukemic mice treated with iHCK-37 had reduced ERK and AKT proteins phosphorylation levels and leukocyte numbers. In conclusion, the iHCK-37 inhibitor has anti-neoplastic activity in leukemia cells without altering apoptosis and survival rate of normal cells, suggesting on-target malignant cell killing activity as a single agent or in combination with 5-Azacytidine or Cytarabine.

Rac GTPases in acute myeloid leukemia cells: Expression profile and biological effects of pharmacological inhibition.

Acute myeloid leukemia (AML) is a highly heterogeneous hematological neoplasm with low survival rates. Thus, the investigation of new therapeutic targets is essential. The Rac subfamily of GTPase proteins has been shown to participate in the physiopathology of hematological malignancies. However, their expression and function in AML remain unclear. In this study, we evaluated Rac1, Rac2 and Rac3 gene expressions in AML and their impact on clinical outcomes. We further investigated the effects of the in vitro treatment with a Rac inhibitor (EHT-1864) on AML cell lines. Rac3 expression was increased in AML derived from myelodysplastic syndromes compared to healthy donors. Rac2 expression did not differ between AML patients and healthy donors, but de novo AML patients with higher Rac2 presented lower overall survival. Oncogenic pathway gene-sets related to AKT/mTOR were identified as associated with Rac1, Rac2 and Rac3 expressions. EHT-1864 treatment reduced the viability of OCI-AML3, KG1 and Kasumi-1 cells in a time and dose-dependent manner. In OCI-AML3 cells, treatment with EHT-1864 induced apoptosis, autophagy, and led to the accumulation of cells in the G1 phase of the cell cycle. These changes were concomitant with alterations in p53 and cyclins. Dowregulation of the PI3K/AKT/mTOR pathway was also observed. Interestingly, the combined treatment of EHT-1864 and low doses of daunorubicin enhanced OCI-AML3 cell apoptosis. In conclusion, Rac2 expression is a prognostic factor in AML and our preclinical results suggest that Rac inhibition may be an attractive mechanism to compose the antineoplastic strategy for this disease.

Selection of plasma donors for the production of anti-SARS-CoV-2 immunoglobulin-based therapies: Strategies for quantitative antibody measurements.

Even after two years of the pandemic, a completely effective treatment against SARS-CoV-2 has not yet been established. Considering this fact and the emergence of successive new viral variants, the development of therapies based on natural polyclonal antibodies recovered from convalescent plasma remains relevant. This study presents a comparison between different methods of screening antibodies in samples of 41 individuals previously diagnosed with COVID-19. We found a significant correlation between Abbot Architect anti-SARS-CoV-2 IgG and Abbott Allinity SARS-CoV-2 IgG II Quantitative assay intensity of reactivity and neutralizing antibody (nAb) titers. Thus, we propose an initial antibody screening with IgG anti-N Abbott Architect test, with an index of, for example, > 3.25 or SARS-CoV-2 IgG II Quantitative Abbott Allinity assay > 137.65 AU/mL as good predictors of Nab ≥ 1:80. For the quantitative method, this threshold demonstrated a 100 % sensitivity and 80 % specificity, with 97.3 % accuracy. An interesting observation was the increase in the neutralizing activity of the anti-SARS-CoV-2 antibodies with the longest interval between the end of the symptoms and the collection, demonstrating that the delay in plasma collection does not affect the achievement of adequate nAbs levels. These results demonstrate the possibility of using faster and more widely available commercial serological tests with a good correlation with viral neutralization tests in culture, allowing for optimized large-scale donor selection, which will be of utmost importance for the development of therapies such as hyperimmune immunoglobulin.

New germline GATA1 variant in females with anemia and thrombocytopenia.

Familial forms of bone marrow defects are rare disorders and description of new cases are valuable opportunities to clarify the molecular machinery that triggers hematopoiesis and blood formation, as well as risk to malignant transformation. We investigated the genetic scenario and possible patterns of transmission in a rare case of familial myeloid disorder with a history of exposure to pesticides. Blood counts of two proband sisters, age 41 and 42, revealed mild anemia, neutrophilia and thrombocytopenia with bone marrow finding mimicking primary myelofibrosis in the cellular phase. We analyzed the coding regions of 78 myeloid neoplasms-related genes and 16 encoding xenobiotic metabolizing genes using Next-Generation Sequencing. The GATA1 variant c.788C > T, p.T263M, located in the C-terminal zinc finger domain of GATA1, was detected in the DNA of the two sisters. The screening of the other kindreds also revealed the p.T263M variant in the mother and two daughters with the same bone marrow disorder. This is the first report of an alteration in the GATA1 CF domain causing anemia, thrombocytopenia and megakaryocyte proliferation with mild myelofibrosis, correlating a new GATA1 germline variant with myeloid disorder.

NT157, an IGF1R-IRS1/2 inhibitor, exhibits antineoplastic effects in pre-clinical models of chronic myeloid leukemia.

Chronic myeloid leukemia (CML) is successfully treated with BCR-ABL1 tyrosine kinase inhibitors, but a significant percentage of patients develop resistance. Insulin receptor substrate 1 (IRS1) has been shown to constitutively associate with BCR-ABL1, and IRS1-specific silencing leads to antineoplastic effects in CML cell lines. Here, we characterized the efficacy of NT157, a pharmacological inhibitor of IGF1R-IRS1/2, in CML cells and observed significantly reduced cell viability and proliferation, accompanied by induction of apoptosis. In human K562 cells and in murine Ba/F3 cells, engineered to express either wild-type BCR-ABL1 or the imatinib-resistant BCR-ABL1T315I mutant, NT157 inhibited BCR-ABL1, IGF1R, IRS1/2, PI3K/AKT/mTOR, and STAT3/5 signaling, increased CDKN1A, FOS and JUN tumor suppressor gene expression, and reduced MYC and BCL2 oncogenes. NT157 significantly reduced colony formation of human primary CML cells with minimal effect on normal hematopoietic cells. Exposure of primary CML cells harboring BCR-ABL1T315I to NT157 resulted in increased apoptosis, reduced cell proliferation and decreased phospho-CRKL levels. In conclusion, NT157 has antineoplastic effects on BCR-ABL1 leukemogenesis, independent of T315I mutational status.

Evaluation and optimization of prescribed concomitant antiplatelet and anticoagulation therapy centrally managed by an anticoagulation management service.

Patients on long-term anticoagulation combined with antiplatelet therapy have an increased risk of bleeding compared to patients on anticoagulation alone. The aim of this study was to evaluate the appropriateness of antiplatelet therapy in patients who are on long-term warfarin therapy and are managed by Brigham and Women's Hospital Anticoagulation Management Service (BWH AMS). This was a single-center, prospective chart review of patients managed by BWH AMS who were on long-term warfarin therapy plus full-dose aspirin (325 mg), an oral P2Y12 inhibitor (clopidogrel, prasugrel or ticagrelor) and/or acetylsalicylic acid/dipyridamole. Patients' cardiovascular (CV) benefit and risk of bleeding were assessed according to clinical guidelines. The major objective of the study was to determine the proportion of patients on dual antithrombotic therapy (DAT) or triple antithrombotic therapy (TAT) whose risk of bleeding outweighed CV benefit. Of the 2677 patients evaluated for inclusion,145 were on concomitant long-term warfarin therapy plus aspirin (325 mg), an oral P2Y12 inhibitor and/or acetylsalicylic acid/dipyridamole. A total of 85 patients (58.6%) had no clear indication for DAT or TAT per guideline recommendations and were categorized as bleeding risk outweighing CV benefit. The remaining 60 patients (41.4%) had an appropriate indication for DAT or TAT per guidelines and were categorized as CV benefit outweighing bleeding risk. BWH AMS pharmacists made 33 (22.9%) recommendations to providers to discontinue or de-escalate antiplatelet therapy. Interventions were accepted for 10 (30.3%) patients. Pharmacist involvement in the management of patients' antithrombotic regimens can optimize guideline-directed medical therapy and mitigate the potential risk of bleeding.

Artesunate Switches Monocytes to an Inflammatory Phenotype with the Ability to Kill Leukemic Cells.

Monocytes are components of the tumor microenvironment related to cancer progression and immune escape. Therapeutic strategies for reprogramming monocytes from a tumor-supporting phenotype towards a tumoricidal phenotype are of great interest. Artesunate (ART) may be an interesting option for cancer treatment; however, the role of ART in regulating the inflammatory tumor microenvironment has not yet been investigated. Our aim is to evaluate the immunomodulatory potential of ART in vitro in human primary monocytes. ART treatment induced an increase in inflammatory monocytes (CD14highCD16-) with HLA-DR high expression and MCP-1/IL-1ß release. On the other hand, ART treatment reduced CD206 and CD163 expression, and abolished the monocyte population known as non-classical and intermediate. Leukemia cells in contact with monocytes programmed with ART presented enhanced in vitro apoptosis suggesting that monocytes acquired the ability to kill leukemic cells. ART induced changes in the monocyte phenotype were mediated by JAK2/STAT3 downregulation. The induction of immunosuppressive environment is an important step for cancer progression. ART showed an immunomodulatory activity, leading immune cells to an antitumor phenotype and could be a candidate for immunotherapy in cancer patients.

Polyphenolic Flavonoid Compound Quercetin Effects in the Treatment of Acute Myeloid Leukemia and Myelodysplastic Syndromes.

Flavonoids are ubiquitous groups of polyphenolic compounds present in most natural products and plants. These substances have been shown to have promising chemopreventive and chemotherapeutic properties with multiple target interactions and multiple pathway regulations against various human cancers. Polyphenolic flavonoid compounds can block the initiation or reverse the promotion stage of multistep carcinogenesis. Quercetin is one of the most abundant flavonoids found in fruits and vegetables and has been shown to have multiple properties capable of reducing cell growth in cancer cells. Acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS) therapy remains a challenge for hematologists worldwide, and the outcomes for patients with both disorders continue to be poor. This scenario indicates the increasing demand for innovative drugs and rational combinative therapies. Herein, we discuss the multitarget effects of the flavonoid quercetin, a naturally occurring flavonol, on AML and MDS.

ANKHD1 is an S phase protein required for histone synthesis and DNA repair in multiple myeloma cells.

ANKHD1 is highly expressed in various cancers such as leukemia and multiple myeloma. Silencing of ANKHD1 expression leads to decreased cell proliferation and accumulation of cells at the S phase. In this study we found ANKHD1 expression to be higher at the S phase, suggesting it to be an S phase protein. We observed that ANKHD1 interacts with histone promoter regions and its inhibition downregulates expression of all core histones, implying a role in histone synthesis. Since histone synthesis occurs in parallel with DNA replication at S phase, we evaluated PCNA (Proliferating Cell Nuclear Antigen) expression, a protein involved in DNA replication and repair. PCNA expression was found to be significantly decreased in ANKHD1 silenced cells. We further observed accumulation γH2AX, a marker for DNA double stranded breaks and an early sign of DNA damage induced by replication stress, upon ANKHD1 silencing. The expressions of several genes implicated in DNA repair were also modulated in ANKHD1 silenced cells, confirming the role of ANKHD1 in DNA repair. Based on this study we speculate that ANKHD1 is an S phase protein required for histone synthesis and DNA repair. These results however, are preliminary and require thorough investigation.

LEF1-AS1, long non-coding RNA, inhibits proliferation in myeloid malignancy.

LEF1 antisense RNA 1 (LEF1-AS1) is an antisense long non-coding RNA encoded in the lymphoid enhancer-binding factor 1 (LEF1) locus. LEF1-AS1 is a conserved transcript dysregulated in hematopoiesis. This study aimed to functionally characterize the role of this transcript in myeloid malignancy and explore a possible regulatory effect of LEF1-AS1 upon LEF1. We show that LEF1-AS1 is highly expressed in normal hematopoietic stem cells but barely detectable in myeloid malignant cell lines. Additionally, bone marrow cells from myelodysplastic syndrome (n=12) and acute myeloid malignancy patients (n=28) expressed significantly reduced levels of LEF1-AS1 compared to healthy controls (n=15). Artificial LEF1-AS1 over-expression inhibited proliferation in HL60 and led to an upregulation of tumor suppressors p21 and p27, and reduced ERK1/2 activation. Unexpectedly, no underlying modulation of LEF1 was detected. Ectopic expression of LEF1-AS1 also inhibited proliferation in HELA, a cell line lacking endogenous expression of LEF1, supporting a LEF1-independent mechanism. Additionally, transient over-expression of LEF1-AS1 in AML patient cells also led to reduced proliferation and colony formation capacity. We used a mass spectrometry-based proteomics approach. Proteomic quantification identified the modulation of an important metabolic regulator, Fumarase, and concomitant accumulation of the metabolite fumarate.

Up-regulation of SPINT2/HAI-2 by Azacytidine in bone marrow mesenchymal stromal cells affects leukemic stem cell survival and adhesion.

The role of tumour microenvironment in neoplasm initiation and malignant evolution has been increasingly recognized. However, the bone marrow mesenchymal stromal cell (BMMSC) contribution to disease progression remains poorly explored. We previously reported that the expression of serine protease inhibitor kunitz-type2 (SPINT2/HAI-2), an inhibitor of hepatocyte growth factor (HGF) activation, is significantly lower in BMMSC from myelodysplastic syndromes (MDS) patients compared to healthy donors (HD). Thus, to investigate whether this loss of expression was due to SPINT2/HAI-2 methylation, BMMSC from MDS and de novo acute myeloid leukaemia (de novo AML) patients were treated with 5-Azacitidine (Aza), a DNA methyltransferase inhibitor. In MDS- and de novo AML-BMMSC, Aza treatment resulted in a pronounced SPINT2/HAI-2 levels up-regulation. Moreover, Aza treatment of HD-BMMSC did not improve SPINT2/HAI-2 levels. To understand the role of SPINT2/HAI-2 down-regulation in BMMSC physiology, SPINT2/HAI-2 expression was inhibited by lentivirus. SPINT2 underexpression resulted in an increased production of HGF by HS-5 stromal cells and improved survival of CD34+ de novo AML cells. We also observed an increased adhesion of de novo AML hematopoietic cells to SPINT2/HAI-2 silenced cells. Interestingly, BMMSC isolated from MDS and de novo AML patients had increased expression of the integrins CD49b, CD49d, and CD49e. Thus, SPINT2/HAI-2 may contribute to functional and morphological abnormalities of the microenvironment niche and to stem/progenitor cancer cell progression. Hence, down-regulation in SPINT2/HAI-2 gene expression, due to methylation in MDS-BMMSC and de novo AML-BMMSC, provides novel insights into the pathogenic role of the leukemic bone marrow microenvironment.

Hypocholesterolemia and dysregulated production of angiopoietin-like proteins in sickle cell anemia patients.

Angiopoietin-like proteins (ANGPTL) are responsible for inhibiting lipoprotein lipase activity, and ANGPTL3 and ANGPTL4 deficiencies have been shown to lower lipoprotein levels in animal models and in humans carrying loss-of-function mutations. Sickle cell anemia (SCA) is a hereditary hemolytic anemia characterized by vaso-occlusive crises and end-organ damage, which is curiously associated with hypocholesterolemia and a low incidence of atherosclerosis, whose underlying mechanisms are unclear. We hypothesized that ANGPTL3 and ANGPTL4 dysregulation is responsible for the hypolipidemic phenotype in SCA. We measured circulating concentrations of ANGPTL3 and ANGPTL4 and correlated them with hemolytic biomarkers and lipoproteins in 40 patients with SCA and 30 control individuals. The association between hemolysis and low cholesterol levels in SCA was confirmed along with surprisingly higher levels of ANGPTL3 and ANGPTL4 in SCA patients than in controls. ANGPTL3 correlated with hemolysis markers LDH and reticulocyte counts, while ANGPTL4 did not. Our data show a paradoxical increase in production of ANGPTL3 and ANGPTL4 in SCA, which would be expected to cause hyperlipidemia, due to increased inhibition of lipoprotein lipase. ANGPTL3, exclusively produced by the liver, correlated with hemolysis markers, suggesting a possible hepatic response to hemolysis. Further functional studies and replication in larger cohorts are warranted to investigate the dysregulation of lipid metabolism in SCA.

Red blood cells microparticles are associated with hemolysis markers and may contribute to clinical events among sickle cell disease patients.

Microparticles are sub-micron vesicles possessing protein and other materials derived from the plasma membrane of their parent cells, and literature suggests that they may have a role in the pathophysiology and downstream manifestations of sickle cell disease (SCD). The contributions of red blood cells microparticles (RMP) to the pathogenic mechanisms and clinical phenotypes of SCD are largely unknown. There is a controversy as to whether the proportions of intravascular hemolysis (approximately ≤ 30% of total hemolysis) would be enough to explain some complications seen in patients with SCD. We investigated RMP among 138 SCD patients and 39 HbAA individuals. Plasma RMPs were quantified by flow cytometry, plasma hemoglobin and heme by colorimetric assays, and haptoglobin and hemopexin by ELISA. The patients had higher RMP, plasma hemoglobin, and heme compared to the controls. On the contrary, haptoglobin and hemopexin were depleted in the patients. The RMP correlated positively with heme, lactate dehydrogenase, plasma hemoglobin, serum bilirubin, reticulocyte counts, and tricuspid regurgitant jet velocity of the patients. Contrarily, it correlated negatively with HbF, hemopexin, red blood cells counts, hemoglobin concentration, and haptoglobin. Although patients treated with hydroxyurea had lower RMP, this did not attain statistical significance. Patients with sickle leg ulcer and elevated tricuspid regurgitant jet velocity had higher levels of RMP. In conclusion, these data suggest that RMPs are associated with hemolysis and may have important roles in the pathophysiology and downstream complications of SCD.

Recombinant erythropoietin as alternative to red cell transfusion in sickle cell disease.

Disturbances in the physiological regulation of erythropoietin (EPO) in patients with sickle cell disease (SCD) may contribute to worsening anaemia and increased transfusion requirements, but the use of recombinant EPO in this group of patients is controversial. The objective of this study was to evaluate the use of this drug in adult patients with SCD and its effects on haemoglobin levels and transfusion requirements. We conducted a retrospective analysis at the University of Campinas, with nineteen adults with a diagnosis of SCD (HbSS and HbS/ß+ thalassaemia), who had received at least 1 year of EPO therapy between 2007 and 2014. Haemoglobin concentrations and trends of variation in transfused RBC volumes were compared before and after EPO administration. We observed that seven patients had a good response to treatment (Hb increment higher than 1·5 g/dl) and nine had a partial response (0·5-1·5 g/dl increment) and there was a significant decrease in the need for transfusion amongst those who usually required regular transfusions. There were no increases in the rates of vaso-occlusive crisis or venous thromboembolism in comparison to the year before the onset of the therapy. Erythropoietin therapy led to a marked increase in haemoglobin concentration with a concomitant decrease in the demand for transfusion. Considering all complications related to allogeneic transfusion, we believe that EPO therapy represents an important therapeutic tool in sickle cell anaemia.