ABSTRACT
A 6-year-old spayed female Scottish Fold cat presented with lethargy and anorexia. A complete blood cell count indicated severe anemia and mild thrombocytopenia. Examination of peripheral blood smears revealed marked changes in the erythroid lineage, including the presence of basophilic stippling and Howell-Jolly bodies as well as an increase in nucleated erythrocytes, polychromatophils, ovalocytes, and schistocytes. Additionally, some erythrocytes contained a ring or figure-eight shaped structure known as a Cabot ring, which were especially observed in polychromatophilic erythrocytes. Hemolytic diseases (Mycoplasma infection and IMHA) were diagnostically excluded, and the cat was treated through prednisolone administration, whole blood transfusion, and administration of vitamins (K2 and B12); however, the anemia progressively worsened. Cabot rings were observed until Day 22 and subsequently disappeared as the number of nucleated RBCs increased, and the erythrocyte lineage shifted to immature population. On Day 42, peripheral blood examination revealed further left shifting and appearance of many rubriblasts. The patient died at home on Day 43. Necropsy revealed neoplastic cells infiltrating the bone marrow and other organs, which were immunopositive to CD71 which is an erythroid lineage marker. In humans, Cabot rings have been observed in megaloblastic anemia, lead poisoning, myelodysplastic syndrome, and myelofibrosis; further, they are thought to be related to stressed bone marrow and dyserythropoiesis. This is the first case report of a cat with Cabot rings, which are suggestive of defects in erythroid lineage production.
Subject(s)
Cat Diseases , Myeloproliferative Disorders , Cats , Female , Cat Diseases/pathology , Cat Diseases/diagnosis , Animals , Myeloproliferative Disorders/veterinary , Myeloproliferative Disorders/pathology , Myeloproliferative Disorders/complications , Fatal Outcome , Erythrocytes, Abnormal/pathology , Anemia/veterinary , Anemia/pathology , Erythrocytes/pathologyABSTRACT
Schistocytes are fragmented red blood cells produced as a result of mechanical damage to erythrocytes, usually due to microangiopathic thrombotic diseases or mechanical factors. The early laboratory detection of schistocytes has a critical impact on the timely diagnosis, effective treatment, and positive prognosis of diseases such as thrombocytopenic purpura and hemolytic uremic syndrome. Due to the rapid development of science and technology, laboratory hematology has also advanced. The accuracy and efficiency of tests performed by fully automated hematology analyzers and fully automated morphology analyzers have been considerably improved. In recent years, substantial improvements in computing power and machine learning (ML) algorithm development have dramatically extended the limits of the potential of autonomous machines. The rapid development of machine learning and artificial intelligence (AI) has led to the iteration and upgrade of automated detection of schistocytes. However, along with significantly facilitated operation processes, AI has brought challenges. This review summarizes the progress in laboratory schistocyte detection, the relationship between schistocytes and clinical diseases, and the progress of AI in the detection of schistocytes. In addition, current challenges and possible solutions are discussed, as well as the great potential of AI techniques for schistocyte testing in peripheral blood.
Subject(s)
Artificial Intelligence , Erythrocytes, Abnormal , Humans , Erythrocytes, Abnormal/pathology , Machine LearningABSTRACT
BACKGROUND: The effectiveness of red cell transfusion in a given blood unit that relied on both quantity and quality of donated cells undoubtedly affects prognostic outcomes. OBJECTIVE: We aimed to determine the frequency of subclinical functional hemoglobin and red cell abnormalities in donated blood of Fayoum University Hospital in Egypt. Additionally, to assess the usefulness of reticulocyte mean hemoglobin content (RET-He) and immature reticulocyte fraction (IRF) as screening measures for such abnormalities. MATERIAL AND METHODS: This cross-sectional study enrolled 200 volunteer blood donors who met the national standard criterion of blood donation. Complete blood count with reticulocyte parameters, serum ferritin, sickling test, G6PD assay, Mentzer index, and naked-eye single tube red cell osmotic fragility test were carried out. RESULTS: Functional red cell abnormalities represented 44 % of this cohort. Out of them, 4.5 % had iron deficiency, 11 % had a positive sickling test, 19 % had G6PD deficiency, and 9.5 % had suspicious thalassemia. The sensitivity and specificity test for RET-He in selective identification of functional hemoglobin abnormalities in donated blood were 83.3 % and 61.2 %, respectively at a cutoff value of 26.9. Though there was no statistically significant effect of RET-He on the selective detection of G6PD deficiency, IRF had a statistically significant high level with a p-value of 0.04. CONCLUSION: Subclinical functional red cell abnormalities seem to be prevalent among blood donors. Reticulocyte/ erythrocyte indices could be useful screening tools for red cell abnormalities. Further studies are required for assessing the impact of transfusing such abnormalities to neonates and other critical recipients.
Subject(s)
Erythrocytes, Abnormal , Humans , Infant, Newborn , Anemia, Iron-Deficiency/diagnosis , Blood Donors , Cross-Sectional Studies , Egypt , Glucosephosphate Dehydrogenase Deficiency , Hemoglobins/analysis , Hospitals, University , Proto-Oncogene Proteins c-ret , Reticulocytes/chemistry , Reticulocytes/pathology , Erythrocytes, Abnormal/chemistry , Erythrocytes, Abnormal/pathologyABSTRACT
Sickle cell disease (SCD) is a genetic disorder that affects millions of individuals worldwide. Chronic anemia, hemolysis, and vasculopathy are associated with SCD, and their role has been well characterized. These symptoms stem from hemoglobin (Hb) polymerization, which is the primary event in the molecular pathogenesis of SCD and contributes to erythrocyte or red blood cell (RBC) sickling, stiffness, and vaso-occlusion. The disease is caused by a mutation at the sixth position of the ß-globin gene, coding for sickle Hb (HbS) instead of normal adult Hb (HbA), which under hypoxic conditions polymerizes into rigid fibers to distort the shapes of the RBCs. Only a few therapies are available, with the universal effectiveness of recently approved therapies still being monitored. In this review, we first focus on how sickle RBCs have altered metabolism and then highlight how this understanding reveals potential targets involved in the pathogenesis of the disease, which can be leveraged to create novel therapeutics for SCD.
Subject(s)
Anemia, Sickle Cell , Vascular Diseases , Anemia, Sickle Cell/drug therapy , Anemia, Sickle Cell/genetics , Drug Discovery , Erythrocytes, Abnormal/metabolism , Erythrocytes, Abnormal/pathology , Hemoglobin A/metabolism , Hemoglobin, Sickle/genetics , Hemoglobin, Sickle/metabolism , Humans , Vascular Diseases/etiologySubject(s)
Erythrocytes, Abnormal , Glucosephosphate Dehydrogenase Deficiency , Lymphoma, Large B-Cell, Diffuse , Adult , Erythrocytes, Abnormal/metabolism , Erythrocytes, Abnormal/pathology , Glucosephosphate Dehydrogenase Deficiency/blood , Glucosephosphate Dehydrogenase Deficiency/drug therapy , Glucosephosphate Dehydrogenase Deficiency/pathology , Humans , Lymphoma, Large B-Cell, Diffuse/blood , Lymphoma, Large B-Cell, Diffuse/drug therapy , Lymphoma, Large B-Cell, Diffuse/pathology , MaleABSTRACT
Red blood cell (RBC) populations are inherently heterogeneous, given mature RBC lack the transcriptional machinery to re-synthesize proteins affected during in vivo aging. Clearance of older, less functional cells thus aids in maintaining consistent hemorheological properties. Scenarios occur, however, where portions of mechanically impaired RBC are re-introduced into blood (e.g., damaged from circulatory support, blood transfusion) and may alter whole blood fluid behavior. Given such perturbations are associated with poor clinical outcomes, determining the tolerable level of abnormal RBC in blood is valuable. Thus, the current study aimed to define the critical threshold of blood fluid properties to re-infused physically-impaired RBC. Cell mechanics of RBC were impaired through membrane cross-linking (glutaraldehyde) or intracellular oxidation (phenazine methosulfate). Mechanically impaired RBC were progressively re-introduced into the native cell population. Negative alterations of cellular deformability and high shear blood viscosity were observed following additions of only 1-5% rigidified RBC. Low-shear blood viscosity was conversely decreased following addition of glutaraldehyde-treated cells; high-resolution microscopy of these mixed cell populations revealed decreased capacity to form reversible aggregates and decreased aggregate size. Mixed RBC populations, when exposed to supraphysiological shear, presented with compounded mechanical impairment. Collectively, key determinants of blood flow behavior are sensitive to mechanical perturbations in RBC, even when only 1-5% of the cell population is affected. Given this fraction is well-below the volume of rigidified RBC introduced during circulatory support or transfusion practice, it is plausible that some adverse events following surgery and/or transfusion may be related to impaired blood fluidity.
Subject(s)
Blood Viscosity , Erythrocyte Deformability , Erythrocytes, Abnormal/pathology , Blood Flow Velocity , Cross-Linking Reagents/toxicity , Erythrocyte Deformability/drug effects , Erythrocyte Transfusion , Erythrocytes, Abnormal/drug effects , Erythrocytes, Abnormal/metabolism , Glutaral/toxicity , Humans , Male , Methylphenazonium Methosulfate/toxicity , Models, Biological , Oxidative Stress , Stress, Mechanical , Superoxides/bloodABSTRACT
Red blood cell (RBC) morphology is, in general, the key diagnostic feature for hereditary spherocytosis (HS) and hereditary elliptocytosis (HE). However, in hereditary pyropoikilocytosis (HPP), the severe clinical form of HE, the morphological diagnosis is difficult due to the presence of a RBC morphological picture characterized by a mixture of elliptocytes, spherocytes, tear-drop cells, and fragmented cells. This difficulty increases in new-borns and/or patients requiring frequent transfusions, making impossible the prediction of the disease course or its severity. Recently, it has been demonstrated that the measurement of osmotic gradient ektacytometry (OGE), using a laser-assisted optical rotational ektacytometer LoRRca (MaxSis, RR Mechatronics), allows a clear differentiation between HS and HE, where the truncated osmoscan curve reflects the inability of the already elliptical cells to deform further under shear stress in the face of hypotonicity. In HPP, however, the RBCs appear to have a significantly decreased ability to maintain deformability in these conditions, and the classical trapezoidal profile of HE is less evident or indistinguishable from HS. Here, two unrelated patients with hereditary hemolytic anemia (HHA) due to HPP and HS, respectively, are described with the joint inheritance of a complex set of five genetic defects. Two of these defects are novel alpha-spectrin gene (SPTA1) variants, one is a microdeletion that removes the entire SPTA1 gene, and two are well-known low-expression polymorphic alleles: α-LELY and α-LEPRA. In the HPP patient (ID1), with many circulating spherocytes, the interactions between the two SPTA1 gene variants may lead, in addition to an elongation defect (elliptocytes), to a loss of membrane stability and vesiculation (spherocytes), and RBCs appear to have a significantly decreased ability to maintain deformability in hypotonic conditions. Due to this, the classical trapezoidal profile of HE may become less evident or indistinguishable from HS. The second patient (ID2) was a classical severe form of HS with the presence of more than 20% of spherocytes and few pincered cells. The severity of clinical manifestation is due to the coinheritance of a microdeletion of chromosome 1 that removes the entire SPTA1 gene with a LEPRA SPTA1 variant in trans. The diagnostic interest of both observations is discussed.
Subject(s)
Anemia, Hemolytic, Congenital/genetics , Elliptocytosis, Hereditary/genetics , Erythrocytes, Abnormal/pathology , Spectrin/genetics , Spherocytosis, Hereditary/genetics , Adult , Anemia, Hemolytic, Congenital/pathology , Chronic Disease , Elliptocytosis, Hereditary/pathology , Female , Genetic Variation , Humans , Infant , Male , Spherocytosis, Hereditary/pathologySubject(s)
Adenocarcinoma of Lung , Carbazoles , Erythrocytes, Abnormal , Lung Neoplasms , Piperidines , alpha-Thalassemia , Adenocarcinoma of Lung/blood , Adenocarcinoma of Lung/drug therapy , Carbazoles/administration & dosage , Carbazoles/adverse effects , Erythrocytes, Abnormal/metabolism , Erythrocytes, Abnormal/pathology , Humans , Lung Neoplasms/blood , Lung Neoplasms/drug therapy , Male , Middle Aged , Piperidines/administration & dosage , Piperidines/adverse effects , alpha-Thalassemia/blood , alpha-Thalassemia/drug therapy , alpha-Thalassemia/pathologySubject(s)
Elliptocytosis, Hereditary , Erythrocytes, Abnormal , Thrombocytosis , beta-Thalassemia , Adult , Elliptocytosis, Hereditary/genetics , Elliptocytosis, Hereditary/metabolism , Elliptocytosis, Hereditary/pathology , Erythrocytes, Abnormal/metabolism , Erythrocytes, Abnormal/pathology , Humans , Male , Thrombocytosis/genetics , Thrombocytosis/metabolism , Thrombocytosis/pathology , beta-Thalassemia/genetics , beta-Thalassemia/metabolism , beta-Thalassemia/pathologyABSTRACT
Sickle cell disease, a genetic disorder affecting a sizeable global demographic, manifests in sickle red blood cells (sRBCs) with altered shape and biomechanics. sRBCs show heightened adhesive interactions with inflamed endothelium, triggering painful vascular occlusion events. Numerous studies employ microfluidic-assay-based monitoring tools to quantify characteristics of adhered sRBCs from high resolution channel images. The current image analysis workflow relies on detailed morphological characterization and cell counting by a specially trained worker. This is time and labor intensive, and prone to user bias artifacts. Here we establish a morphology based classification scheme to identify two naturally arising sRBC subpopulations-deformable and non-deformable sRBCs-utilizing novel visual markers that link to underlying cell biomechanical properties and hold promise for clinically relevant insights. We then set up a standardized, reproducible, and fully automated image analysis workflow designed to carry out this classification. This relies on a two part deep neural network architecture that works in tandem for segmentation of channel images and classification of adhered cells into subtypes. Network training utilized an extensive data set of images generated by the SCD BioChip, a microfluidic assay which injects clinical whole blood samples into protein-functionalized microchannels, mimicking physiological conditions in the microvasculature. Here we carried out the assay with the sub-endothelial protein laminin. The machine learning approach segmented the resulting channel images with 99.1±0.3% mean IoU on the validation set across 5 k-folds, classified detected sRBCs with 96.0±0.3% mean accuracy on the validation set across 5 k-folds, and matched trained personnel in overall characterization of whole channel images with R2 = 0.992, 0.987 and 0.834 for total, deformable and non-deformable sRBC counts respectively. Average analysis time per channel image was also improved by two orders of magnitude (â¼ 2 minutes vs â¼ 2-3 hours) over manual characterization. Finally, the network results show an order of magnitude less variance in counts on repeat trials than humans. This kind of standardization is a prerequisite for the viability of any diagnostic technology, making our system suitable for affordable and high throughput disease monitoring.
Subject(s)
Anemia, Sickle Cell/blood , Deep Learning , Erythrocytes, Abnormal/classification , Microfluidics/statistics & numerical data , Anemia, Sickle Cell/diagnostic imaging , Biophysical Phenomena , Computational Biology , Diagnosis, Computer-Assisted/statistics & numerical data , Erythrocyte Deformability/physiology , Erythrocytes, Abnormal/pathology , Erythrocytes, Abnormal/physiology , Hemoglobin, Sickle/chemistry , Hemoglobin, Sickle/metabolism , High-Throughput Screening Assays/statistics & numerical data , Humans , Image Interpretation, Computer-Assisted/statistics & numerical data , In Vitro Techniques , Lab-On-A-Chip Devices/statistics & numerical data , Laminin/metabolism , Neural Networks, Computer , Protein MultimerizationABSTRACT
Schistocytosis is the morphological hallmark of the microangiopathic haemolytic anaemia of thrombotic microangiopathy (TMA). Consensus guidelines for manual schistocyte quantitation are available, but limited research has evaluated them. The 2012 International Council for Standardization in Haematology (ICSH) recommends a schistocyte quantitation of 1% as a robust cut-off for significance, with the quantitation including helmet, crescent, triangle and keratocyte poikilocytes; and microspherocytes only in the presence of helmets, crescents/triangles, and keratocytes. We aimed to evaluate the relative contribution of these different poikilocytes to schistocyte counting; compare the ICSH method with our proposed method which counts only cells most specific for red cell fragmentation (helmet, crescent and triangular schistocytes); and evaluate inter- and intra-observer agreement. Blood films were sourced from the Australian Snakebite Project, including non-envenomed and envenomed cases, with and without TMA. In blood films across the range of schistocytosis, the predominant poikilocytes present were helmets and crescents. Triangles, keratocytes and microspherocytes were typically only present when ICSH schistocyte count was >1%. With results dichotomised as <1.0% or ≥1.0%, our proposed new method versus the ICSH method showed almost perfect agreement [observed agreement 95%, Cohen's kappa (κ)=0.84, SE 0.04, 95% CI 0.76-0.92, p<0.005]. Inter-observer strength of agreement for our method was moderate (Fleiss' κ for comparisons between three non-unique microscopists κ=0.50, SE 0.05, 95% CI 0.41-0.59, p<0.005). Intra-observer reproducibility assessed in two microscopists ranged from substantial (Cohen's κ=0.71, SE 0.08, 95% CI 0.55-0.86, p<0.005) to borderline almost perfect agreement (Cohen's κ=0.81, SE 0.07, 95% CI 0.68-0.93, p<0.005). Schistocyte quantitation using our new method is simpler than the 2012 ICSH method and had almost perfect agreement. Our finding of moderate inter-observer agreement in quantitating helmet, triangle and crescent schistocytes is applicable to both the ICSH and our newly proposed method. This finding underscores the importance of clinicopathological correlation and repeated examinations in the context of a clinically suspected TMA.
Subject(s)
Erythrocyte Count/standards , Erythrocytes, Abnormal/pathology , Purpura, Thrombotic Thrombocytopenic/pathology , Thrombotic Microangiopathies/pathology , Cell Count/methods , Cell Count/standards , Erythrocyte Count/methods , Humans , Observer VariationSubject(s)
Acid-Base Imbalance/genetics , Anemia, Hemolytic, Congenital/genetics , Erythrocytes, Abnormal/pathology , Metabolism, Inborn Errors/genetics , Rh-Hr Blood-Group System/genetics , Acid-Base Imbalance/pathology , Adult , Anemia, Hemolytic, Congenital/pathology , Blood Proteins/genetics , Erythrocytes, Abnormal/metabolism , Female , Humans , Membrane Glycoproteins/genetics , Metabolism, Inborn Errors/pathology , PregnancyABSTRACT
Hereditary red blood cell (RBC) membranopathies are characterized by mutations in genes encoding skeletal proteins that alter the membrane complex structure. Hereditary spherocytosis (HS) is the most common inherited RBC membranopathy leading to hereditary hemolytic anemia with a worldwide distribution and an estimated prevalence, in Europe, of about 1:2000 individuals. The recent availability of targeted next generation sequencing (t-NGS) and its combination with RBC deformability measured with a laser-assisted optical rotational ektacytometer (LoRRca) has demonstrated to be the most powerful contribution to lower the percentage of hereditary hemolytic anemia undiagnosed cases. In order to know the kind and frequency of RBC membrane mutations in our geographical area (Catalonia) and to better understand their pathophysiology, 42 unrelated, non-transfusion-dependent (NTD) patients with hereditary hemolytic anemia have been studied by combining t-NGS and LoRRca. The osmoscan module of LoRRca provides three rheological profiles that reflect the maximal deformability (EImax), osmotic fragility (Omin), and hydration state (Ohyper) of RBCs and contribute to a better understanding of the contribution RBC rheology to the severity of anemia. From the 42 patients studied, 37 were suspected to be a RBC membrane defect due to phenotypic characteristics and abnormal RBC morphology and, from these, in 31 patients (83.8% of cases) the mutation was identified by t-NGS. No definite diagnosis was achieved in 11 patients (26.2% of cases), including 6 out of 37 cases, with suspected membranopathy, and 5 with unclassifiable HHA. In all these undiagnosed patients, the existence of hemoglobinopathy and/or enzymopathy was ruled out by conventional methods.
Subject(s)
Anemia, Hemolytic, Congenital/diagnosis , Anemia, Hemolytic, Congenital/etiology , Erythrocyte Deformability/genetics , Erythrocyte Membrane/genetics , Erythrocyte Membrane/metabolism , High-Throughput Nucleotide Sequencing , Osmotic Fragility/genetics , Adolescent , Adult , Aged , Alleles , Anemia, Hemolytic, Congenital/blood , Biomarkers , Child , Erythrocyte Membrane/pathology , Erythrocytes, Abnormal/pathology , Female , Genetic Association Studies , Genetic Predisposition to Disease , Humans , Male , Middle Aged , Mutation , Osmotic Pressure , Spherocytosis, Hereditary/blood , Spherocytosis, Hereditary/diagnosis , Spherocytosis, Hereditary/genetics , Spherocytosis, Hereditary/metabolism , Young AdultABSTRACT
OBJECTIVE: Hepatic and biliary tract diseases are common in sickle cell disease (SCD) patients, likely due to sickling, hemosiderosis, viral hepatitis, or cholelithiasis. Literature is lacking on associations between SCD, autoimmune hepatitis (AIH), and/or sclerosing cholangitis (SC)-together, autoimmune liver disease (AILD). We aimed to better understand the relationship of these diseases in pediatric patients. MATERIALS AND METHODS: A retrospective analysis of patients with SCD and AILD at the Children's Hospital of Philadelphia (January 2008 to August 2015). RESULTS: Seven patients, ages 8 to 23 years (3 males), were identified. Three had AIH, 2 SC, and 2 AIH/SC overlap, known as autoimmune SC. All patients with AIH treated with azathioprine significantly improved their liver enzymes. One patient with SC and inflammatory bowel disease underwent successful bone marrow transplant. Two SC patients died from SCD complications. CONCLUSIONS: In this cohort, there seems to be an association between SCD and AILD; SC in this population was severe. Physicians should be aware of this and evaluate patients with SCD and elevated liver enzymes for AILD.
Subject(s)
Anemia, Sickle Cell/complications , Erythrocytes, Abnormal/pathology , Hepatitis, Autoimmune/pathology , Adolescent , Adult , Child , Female , Follow-Up Studies , Hepatitis, Autoimmune/etiology , Humans , Male , Prognosis , Retrospective Studies , Young AdultABSTRACT
Infantile pyknocytosis is a rare and self-limiting cause of hemolytic anemia in neonates. It can result in severe anemia and hyperbilirubinemia. The pathogenesis is unknown: a genetic origin has been discussed; however, based on the current literature it is not clear which genetic mutations should be considered. We present a case of a premature twin, in whom genetic screening was performed. Genetic mutations in 46 genes associated with hereditary hemolytic anemia and dyserythropoietic anemia were tested. No mutations were found. In infantile pyknocytosis, a genetic defect in these genes is unlikely.