Search

Based on the relationship between the intracellular concentration of sickle hemoglobin S (HbS) and the delay that occurs prior to the onset of sickling following deoxygenation, targeting the intracellular HbS concentration is a recognized therapeutic approach for sickle cell disease (SCD). We and others have shown that restricting iron by dietary or pharmacologic means improves hematologic parameters, inflammation, and organ damage in mouse models of SCD. Clinical evidence corroborating these findings is confined to case reports and small case series studies, none of which account for treatment or α‐thalassemia. We hypothesize that increased transferrin saturation is associated with increased mean cellular hemoglobin concentration (MCHC) which in turn is associated with decreased red cell counts and worsening anemia. To investigate this hypothesis, we examined the relationships between transferrin saturation and MCHC with each of the parameters that define MCHC in sickle patients (HbSS without α‐thalassemia) and healthy volunteers (HVs). Results indicate that transferrin saturation and MCHC are positively correlated with each other in sickle patients and HV. In patients with SCD, MCHC and transferrin saturation are negatively correlated with RBC count and are not correlated with hemoglobin, whereas each is positively associated with HV. Transferrin saturation and MCHC are each positively correlated with the hemolysis marker, lactate dehydrogenase. These observations support a model where increased transferrin saturation contributes to higher intracellular HbS concentrations with subsequent increases in sickling and hemolysis in sickle patients, suggesting that pharmacologic approaches to decrease serum iron may provide a therapeutic approach for patients with SCD. Trial Registration: This study was registered with ClinicalTrials.gov identifiers: NCT00011648, NCT00081523, and NCT04817670. [ABSTRACT FROM AUTHOR]

Introduction: Reticulocyte count and novel derived parameters provide insight into the effectiveness of erythropoiesis and may be useful tools in the classification and diagnosis of anemias. However, there is no standardisation, so we consider it necessary that each laboratory evaluates the parameters according to its own methodology and instrumentation and establishes its own reference ranges. Our aim was to establish the reference intervals (RIs) of reticulocyte profile provided by the Beckman Coulter DxH 900 haematological autoanalyzer in our reference population. Methods: One hundred and seventy‐five healthy adults (18 to 62 years) were included. Subjects were collected from the blood donation centre of the Hospital Clínico San Carlos (Madrid, Spain) upon informed consent. Whole blood was collected and assayed for 14 haematological parameters on the Beckman Coulter DxH 900 analyzer in the haematology laboratory of the Clinical Analysis Department. RIs were established as per Clinical and Laboratory Standards Institute EP28‐A3c guidelines using three different statistical approaches. Results: RIs estimated using the non‐parametric method and the Harrell‐Davis bootstrap method were very similar. RIs estimated by the robust method were narrower. Gender partitioning was required for two haematological parameters (low haemoglobin density (LHD) and microcytic anaemia factor (MAF)). The rest of the parameters did not need to be partitioned according to Lahti's method. Conclusion: RIs have been established for 14 hematologic parameters of the reticulocyte profile for the Beckman Coulter DxH 900 haematology analyzer using a healthy cohort of adult subjects. [ABSTRACT FROM AUTHOR]

ABSTRACT Introduction Material and Methods Results Conclusions Hereditary spherocytosis (HS) is a congenital haemolytic disorder, resulting from plasma membrane protein deficiency of red blood cells (RBCs). Typical pathological signs are anemia, jaundice, and splenomegaly; in newborns, jaundice is the main symptom.This study focused on the state of art about the HS diagnosis, from traditional to innovative methods, including diagnostic algorithms that can be applied for pediatric and adult patients, for different laboratory diagnostic levels.The first erythrocyte parameters used for HS diagnosis were the mean corpuscular hemoglobin concentration (MCHC), mean corpuscular volume (MCV), and red blood cell distribution width (RDW); nowadays new parameters are used in blood cell counter. Advia analyzers (Siemens Medical Solutions) supply the hyper‐dense cell percentage (% Hyper), which reflects the red blood cells hyperchromia. Sysmex instruments (i.e. XT‐4000i, XE‐5000, XN‐Series) provide the MicroR, that is the percentage of erythrocytes smaller than 60 fL, Hypo‐He, which is the percentage of erythrocytes with a content of hemoglobin less than 17 pg and % Hyper‐He, which represents the percentage of RBC with cellular hemoglobin content higher than 49 pg. CELL‐DYN Sapphire (Abbott Diagnostics) introduced the HPR parameter (% HPR), which represents the erythrocytes with hemoglobin > 410 g/L. Beckman Coulter instruments supply the mean sphered corpuscular volume (MSCV), which is the average volume of all erythrocytes, including mature erythrocytes and reticulocytes. Other reference tests for screening and diagnosis of HS are the acidified glycerol lysis test (AGLT), the eosin‐5‐maleimide (EMA) binding test and genetic testing by next‐generation sequencing.The diagnostic workup of hereditary spherocytosis could be improved thanks to all the available tests, including new molecular tools. However, it requires synergy between clinicians and laboratory staff, evaluating clinical manifestations, all available data related to the disease and the prognosis to fill the diagnostic gaps in the near future. [ABSTRACT FROM AUTHOR]

Therapeutic neutralization of Oncostatin M (OSM) causes mechanism‐driven anemia and thrombocytopenia, which narrows the therapeutic window complicating the selection of doses (and dosing intervals) that optimize efficacy and safety. We utilized clinical data from studies of an anti‐OSM monoclonal antibody (GSK2330811) in healthy volunteers (n = 49) and systemic sclerosis patients (n = 35), to quantitatively determine the link between OSM and alterations in red blood cell (RBC) and platelet production. Longitudinal changes in hematopoietic variables (including RBCs, reticulocytes, platelets, erythropoietin, and thrombopoietin) were linked in a physiology‐based model, to capture the long‐term effects and variability of therapeutic OSM neutralization on human hematopoiesis. Free serum OSM stimulated precursor cell production through sigmoidal relations, with higher maximum suppression (Imax) and OSM concentration for 50% suppression (IC50) for platelets (89.1% [95% confidence interval: 83.4–93.0], 6.03 pg/mL [4.41–8.26]) than RBCs (57.0% [49.7–64.0], 2.93 pg/mL [2.55–3.36]). Reduction in hemoglobin and platelets increased erythro‐ and thrombopoietin, respectively, prompting reticulocytosis and (partially) alleviating OSM‐restricted hematopoiesis. The physiology‐based model was substantiated by preclinical data and utilized in exploration of once‐weekly or every other week dosing regimens. Predictions revealed an (for the indication) unacceptable occurrence of grade 2 (67% [58–76], 29% [20–38]) and grade 3 (17% [10–25], 3% [0–7]) anemias, with limited thrombocytopenia. Individual extent of RBC precursor modulation was moderately correlated to skin mRNA gene expression changes. The physiological basis and consideration of interplay among hematopoietic variables makes the model generalizable to other drug and nondrug scenarios, with adaptations for patient populations, diseases, and therapeutics that modulate hematopoiesis or exhibit risk of anemia and/or thrombocytopenia. [ABSTRACT FROM AUTHOR]

Introduction: The International Council for Standardization in Haematology convened a working group to assess and propose improvements upon the state of standardization and harmonization of reticulocyte parameters among commercial hematology analyzers. Methods: An international group of laboratory hematologists prospectively collected and analyzed clinical samples using locally available IVD commercial hematology analyzers. Eight hundred and fifty‐five total samples were collected at 6 sites using 9 distinct analyzer types. Samples were assessed for reticulocyte percent (RET%), immature reticulocyte fraction (IRF), and reticulocyte hemoglobin content (RHC). Method comparison and regression statistics were calculated. These analyses were used to determine whether statistical recalibration offered a potential avenue for increasing comparability between these methods. Results: While methods producing reticulocyte percent were the most comparable in this study, the state of harmonization for the IRF and RHC was reduced with pearson correlation coefficients ranging from 0.955 to 0.77 and 0.927 and 0.680, respectively. Nevertheless, use of parameters from the Passing Bablok regression substantially improved the comparability of the results. In addition, precision data was derived which also demonstrated substantial differences between analyzer systems. Conclusion: While reticulocyte counting is correlated between the automated methods evaluated in this study, the current state of harmonization of other reticulocyte parameters is not as strong. A major challenge in moving this field forward is the need for commutable materials to facilitate comparisons between analyzers not co‐located. A potential alternate approach to improve the current state would be instrument re‐calibration. However, this is challenging both technically and due to national regulatory frameworks. [ABSTRACT FROM AUTHOR]

Introduction: Detection of iron deficiency (ID) remains challenging. We aimed to evaluate the performance of reticulocyte hemoglobin equivalent (Ret‐He) as a potential diagnostic marker to assess ID and iron deficiency anemia (IDA) in a large pediatric cohort. Methods: A total of 3158 patients (aged 15 days to 19 years with a median age of 8.5 years; 60.2% female) were retrospectively studied. Statistical analysis was performed (a) to evaluate relationship of Ret‐He with other relevant complete blood count and iron panel parameters; (b) to compare the levels of Ret‐He in ID and IDA groups to a control group; and (c) to assess sensitivity and specificity of Ret‐He in ID, IDA, and anemia without ID groups. Results: Ret‐He values were significantly positively correlated to ferritin and transferrin saturation (TSAT). The median Ret‐He was significantly lower in ID. A Ret‐He cutoff of ≤30.0 pg distinguished cases of ID from the control group with a sensitivity of 90.2%, specificity of 59.5%, and area under curve (AUC) of 0.88. Ret‐He showed better diagnostic performance in the IDA group and acceptable performance for ID without anemia. The sensitivity, specificity, and AUC were 90.1%, 80.9%, and 0.93 for IDA at cutoff value of ≤27.4 pg, and 80.8%, 51.1%, and 0.70 for ID without anemia at cutoff value of ≤30.8 pg, respectively. Conclusion: Our large pediatric tertiary care hospital study demonstrates that Ret‐He is a reliable marker to help confirm IDA in pediatric population. However, further studies are needed for its use to capture the early stages of ID. [ABSTRACT FROM AUTHOR]

Introduction: The Atellica Hema (Siemens Healthineers, Tarrytown, NY, USA) is a new generation multi‐parameter analyzer for full blood count, 6‐part differential and reticulocyte testing by impedance variation and fluorescence flow cytometry. In this study, we verified the whole blood and limited body fluid modes of the Atellica Hema 580. Methods: We evaluated precision, linearity, carry‐over, throughput and performed a method comparison to assess the performance of the Atellica Hema 580. For comparison of the Atellica Hema 580 with the Sysmex XN‐1000 (Sysmex, Kobe, Japan), 140 samples from adult and pediatric patients including both normal and abnormal hematology profiles were analyzed in parallel. Results: The Atellica Hema 580 demonstrated acceptable imprecision within the manufacturer's specifications for whole blood and body fluid modes, good linearity for high and low ranges and no significant carryover. The full blood count, differential and reticulocyte correlated well with the Sysmex XN‐1000, except for mean cell hemoglobin concentration, basophil and large immature cells. The optical platelet count, reflexed in 34 samples with a platelet count <150 × 109/l, showed a strong correlation with the fluorescent platelet count on the Sysmex XN‐1000. The morphology flagging efficiency was 92% for white blood cells, 95% for red blood cells and 87% for platelets. Conclusion: The Atellica Hema 580 showed good analytical performance and workflow efficiency for a wide range of patient samples. [ABSTRACT FROM AUTHOR]

Blood doping is prohibited for athletes but has been a well‐described practice within endurance sports throughout the years. With improved direct and indirect detection methods, the practice has allegedly moved towards micro‐dosing, that is, reducing the blood doping regime amplitude. This narrative review evaluates whether blood doping, specifically recombinant human erythropoietin (rhEpo) treatment and blood transfusions are performance‐enhancing, the responsible mechanism as well as detection possibilities with a special emphasis on micro‐dosing. In general, studies evaluating micro‐doses of blood doping are limited. However, in randomized, double‐blinded, placebo‐controlled trials, three studies find that infusing as little as 130 ml red blood cells or injecting 9 IU × kg bw−1 rhEpo three times per week for 4 weeks improve endurance performance ~4%–6%. The responsible mechanism for a performance‐enhancing effect following rhEpo or blood transfusions appear to be increased O2‐carrying capacity, which is accompanied by an increased muscular O2 extraction and likely increased blood flow to the working muscles, enabling the ability to sustain a higher exercise intensity for a given period. Blood doping in micro‐doses challenges indirect detection by the Athlete Biological Passport, albeit it can identify ~20%–60% of the individuals depending on the sample timing. However, novel biomarkers are emerging, and some may provide additive value for detection of micro blood doping such as the immature reticulocytes or the iron regulatory hormones hepcidin and erythroferrone. Future studies should attempt to validate these biomarkers for implementation in real‐world anti‐doping efforts and continue the biomarker discovery. [ABSTRACT FROM AUTHOR]

Reticulocyte indices are used to characterize anemia, including the identification of regeneration. In people, the immature reticulocyte fraction (IRF), percentage of hypochromic red blood cells (%HYPO‐RBC), and other reticulocyte indices have been used as earlier indicators of erythropoiesis and as valuable monitoring tools in the assessment of various therapies. The reference intervals (RI) of the IRF and %HYPO‐RBC have not been reported in dogs. The objective of this study was to establish RIs for novel variables (IRF, %HYPO‐RBC, and CH‐delta) and assess RIs for more commonly reported reticulocyte indices in healthy dogs. RIs were calculated from blood results retrospectively collected from 106 client‐owned healthy dogs at the time of induction into a blood donor program using the ADVIA 2120 hematology analyzer (Siemens Healthcare Diagnostics). For the calculation of RIs, appropriate tests were applied for outlier detection and normality assessment. For variables normally distributed, RIs and their respective 90% confidence intervals (CIs) were calculated using parametric methods, while for variables not normally distributed, robust methods were used and bootstrapping for calculating the 90% CIs. The following RIs were established: reticulocyte hemoglobin content (CHr) 24.5‐28 pg, mean reticulocyte volume (MCVr) 85.9‐99.3 fL, mean corpuscular hemoglobin concentration of reticulocytes (CHCMr) 271.0‐306.3 g/L, IRF 10.4%‐43.5%, CH‐delta 0.5‐4.3 pg, and percentage of hypochromic red blood cells (%HYPO‐RBC) 0.10%‐0.80%. The results of this study provide RIs for novel reticulocyte variables. Further studies are required to determine the clinical utility of IRF, %HYPO‐RBC, and CH delta as early indicators of erythropoietic activity in canine patients. [ABSTRACT FROM AUTHOR]

The article presents letter to the editor focuses on the advantages of incorporating Patient-Based Real-Time Quality Control (PBRTQC) as a complementary tool in modern quality assurance strategies. Topics include the role of PBRTQC in enhancing error detection and patient safety, the optimization of laboratory-specific settings for improved performance, and the challenges of implementation due to software limitations and sample variability.

Introduction: This study assessed the comparability of complete blood count (CBC) parameters between capillary and venous samples, and extended previous research by examining the influence of different storage temperatures on CBC stability up to 7 days after sample collection. Methods: Venous and capillary blood samples were collected from 93 adult patients. Hemoglobin (Hb), hematocrit (Ht), mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC), mean platelet volume (MPV), leukocytes, lymphocytes, basophils, eosinophils, erythrocytes, red cell distribution width (RDW), immature granulocytes (IG), immature reticulocyte fraction (IRF), monocytes, neutrophils, platelets, and reticulocytes were measured. Deming regression and mean relative differences between venous and capillary measurements were contrasted with desirable total allowable error (TEa). Stability was assessed in 20–27 venous blood samples stored at 4, 21–22, or 30°C, and analyzed at 0, 24, 48, 72, 96, 120, 144, and 168 h. Mean relative change with respect to baseline measurements was compared to the desirable TEa to determine acceptable stability. Results: Deming regression demonstrated strong linear correlations and acceptable variation between venous and capillary measurements. Erythrocytes, Hb, Ht, MCH, MCV, RDW, reticulocytes, and platelets showed acceptable stability for at least 96 h at 4°C. Mean relative change exceeded desirable TEa after 24 h at 30°C for all parameters, except erythrocytes, Hb, leukocytes, and MCH. Conclusion: Clinical laboratory specialists and clinicians should be aware of potential differences between venous and capillary measurements, and the influence of storage conditions. Clinical validity of delayed CBC analysis depends on the clinical situation and required precision of the result. [ABSTRACT FROM AUTHOR]

Background: The regenerative response following Babesia rossi infection in dogs is mild, despite severe hemolytic anemia. Objective: We aimed to compare the admission absolute reticulocyte count (ARC) and reticulocyte indices in 103 dogs naturally infected with B. rossi with 10 dogs suffering from immune‐mediated hemolytic anemia (IMHA) and 14 healthy control dogs. The regenerative response was also evaluated in five dogs experimentally infected with B. rossi. Methods: This is a retrospective observational study of records generated on the ADVIA 2120 hematology analyzer. Results: The median hematocrits (HCT) of the B. rossi and IMHA groups were significantly lower than the control group (p <.001 for both); however, no differences were seen between the B. rossi and IMHA groups. Compared with the control group, the median ARC was significantly higher in the B. rossi (p =.006) and IMHA (p =.019) groups but significantly lower in the B. rossi group than the IMHA group (p =.041). In the experimentally infected dogs, there was a sudden decrease in the ARC approximately 48 h after the detection of peripheral parasitemia, which was followed by an increase after treatment. Reticulocytes of naturally infected B. rossi dogs were larger, with more variation in cellular volume. The reticulocytes of the experimentally infected dogs decreased in size with decreasing hemoglobin concentrations as the study progressed. Conclusions: The regenerative response in dogs naturally infected with B. rossi is inadequate, given the severity of the anemia observed, and it might be a result of direct suppressive action by the parasite or host response on the bone marrow. [ABSTRACT FROM AUTHOR]

Background: Hematological parameters vary significantly throughout growth and development due to physiological processes such as fetal‐to‐adult erythropoiesis and puberty. Pediatric age‐ and sex‐specific reference intervals (RIs) are thus essential for appropriate clinical decision‐making. The current study aimed to establish RIs for both common and novel hematology parameters on the Mindray BC‐6800Plus system. Methods: Six hundred and eighty‐seven healthy children and adolescents (30 days to 18 years) were enrolled. Participants were recruited as part of the Canadian Laboratory Initiative on Pediatric Reference Intervals Program upon informed consent or identified from apparently healthy outpatient clinics. Whole blood was collected and assayed for 79 hematology parameters on the BC‐6800Plus system (Mindray). Age‐ and sex‐specific RIs were established as per Clinical and Laboratory Standards Institute EP28‐A3c guidelines. Results: Dynamic reference value distributions were observed for several hematology parameters, including erythrocytes, leukocytes, platelets, reticulocytes, and research‐use‐only markers. Age partitioning was required for 52 parameters, demonstrating changes in infancy and puberty. Sex partitioning was required for 11 erythrocyte parameters (i.e., red blood cell (RBC), hemoglobin, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin concentration, RBC distribution width coefficient of variation, hemoglobin distribution width, macrocyte count, macrocyte percentage, RBC (optical), and reticulocyte production index). Few parameters had undetectable levels in our healthy cohort (i.e., nucleated RBC count and immature granulocyte count). Conclusions: The current study completed hematological profiling for 79 parameters on the BC‐6800Plus system in a healthy cohort of Canadian children and adolescents. These data emphasize the complex biological patterns of hematology parameters in childhood, particularly at the onset of puberty, and support the need for age‐ and sex‐specific RIs for clinical interpretation. [ABSTRACT FROM AUTHOR]

Objective: The study aimed to determine the impact of Red Blood Cells (RBCs) generated from peripheral blood mononuclear cells (PBMCs) on T cell proliferation and host response following whole blood stimulation. Background: Culturing RBCs is a potential solution for donor shortage. The impact of immature cultured RBCs which express CD71+ on host immune response is not known. Methods/Materials: PBMCs were seeded in an erythroid expansion medium. CD71+ cells were isolated at days 14 and 21 of culture and incubated with either purified T cells or with LPS‐stimulated whole blood. Controls were incubated with medium. Results: At day 9, the percentage of cells that expressed CD45 and CD71 reached to the highest level (32.9%, IQR; 26.2–39.05) while the percentage of cells that expressed CD71 and CD235a reached to the highest level on day 17 (70.2%, IQR; 66.1–72.8). Incubation of T cells with days 14 CD71+ cells and day 21 CD71+ cells increased T cell proliferation. In a whole blood stimulation assay, day 21 CD71+ cells, but not day 14 CD71+ cells, inhibited the production of IL‐6 and TNFα. Conclusion: Cultured erythroid cells can modulate the immune response by promoting T cell proliferation and inhibiting cytokine secretions following whole blood stimulation. [ABSTRACT FROM AUTHOR]

Background: There are few reports in dogs that have evaluated the utility of semi‐quantitative scoring of bone marrow iron stores in conjunction with reticulocyte hemoglobin (CHr) to identify iron‐restricted erythropoiesis due to absolute iron deficiency or iron sequestration. Objectives: An established system for scoring iron stores in human bone marrow samples was applied to dogs. The objectives were to evaluate interobserver agreement (Κω), determine marrow iron scores in dogs without detectable hematologic abnormalities, and assess combined interpretation of iron scores and CHr to evaluate for iron‐restricted erythropoiesis. Methods: Four blinded observers independently scored iron in 139 Prussian blue‐stained canine marrow samples from 0 (none) to 6 (very heavy), including healthy controls (n = 12), clinically ill dogs with (n = 100) and without (n = 16) detectable hematologic abnormalities, and dogs with experimental nutritional iron deficiency (n = 11). Additional medical record data were available for 118 dogs to evaluate for other evidence of iron deficiency (abnormal CHr, RBC indices, serum iron variables, external blood loss, or nutritional deficiencies). Results: Mean Κω was 0.69 (substantial agreement) for all samples but was 0.44 (moderate agreement) for samples with iron scores <3, indicating distinguishing scores 0–2 may not be reliable. Dogs without detectable hematologic abnormalities had scores from 3–5. Dogs with scores <3 and decreased CHr often had more indicators of iron deficiency vs dogs only having low iron scores or low CHr. Conclusions: Evaluation of dogs with marrow iron score <3 for external blood loss or nutritional deficiencies is likely clinically worthwhile, particularly if there is also decreased CHr. [ABSTRACT FROM AUTHOR]

The article discusses a rare case of neonatal-onset congenital dyserythropoietic anemia type II (CDA II) diagnosed in a two-month-old male, confirmed through bone marrow cytological analysis and genetic testing identifying pathogenic secretory pathway component (SEC)23B mutations. Topics include the clinical presentation and diagnostic features of CDA II, the genetic basis involving SEC23B mutations, and potential management options such as transfusions.

Background: Evaluating regeneration is essential for the classification and differential diagnosis of anemia in dogs. Early detection of regeneration is challenging in anemic dogs. Objectives: This study assessed the value of immature reticulocyte fraction (IRF) in differentiating preregenerative anemia (PRA) from nonregenerative anemia (NRA) in dogs. Animals: Ninety‐four dogs: 49 controls and 45 with anemia. Methods: Case‐control study. Fractions of low‐, medium‐ (MFR), and high‐fluorescence reticulocytes (HFR), were measured using the ADVIA 2120i hematology analyzer. The IRF was calculated as the sum of percentages of MFR and HFR. Data from dogs with regenerative anemia (RA, n = 19), PRA (n = 11), and NRA (n = 15) were retrospectively analyzed. The value of IRF was compared with reticulocyte production index (RPI) using the receiver operating characteristic (ROC) curve. Results: The median of IRF was significantly higher in dogs with RA (46.5%; range, 40.9‐53.6%; P <.001) and PRA (40.6%; range, 27.7‐47.1%; P =.01) than in controls (22.1%; range, 16.9‐29.3%). The IRF in dogs with PRA showed no difference compared to dogs with RA (P >.99) but was higher than dogs with NRA (18.7%; range, 8.8‐24%; P =.00). The area under the ROC curve of IRF was superior to that of RPI (0.897 vs 0.818, P =.00) in differentiating dogs with PRA from NRA. Conclusions and Clinical Importance: The IRF is a reliable variable for detecting early regeneration in anemic dogs without reticulocytosis. The study suggests that the measurement of IRF could be useful in classifying anemic dogs. [ABSTRACT FROM AUTHOR]

Introduction: The Thomas‐plot has proven to be a helpful tool to discriminate between different types of anemia. This plot combines the reticulocyte hemoglobin content (Ret‐He) with the soluble transferrin receptor (sTfR)/log ferritin (fer) ratio. In this study, we designed an alternative Thomas‐plot in which Ret‐He is combined with the transferrin (Tf)/log ferritin ratio. We validated both Thomas‐plots in a population of anemic patients and compared the performance to the current laboratory diagnostics of anemia. Methods: A total of 536 anemic patients were included. The first 188 patients were used to generate ROC curves to define the optimal cut‐off values for both Thomas‐plots. With the following 348 patients included we studied the performance of the alternative and classical Thomas‐plots compared to current anemia diagnostics. Results: Cut‐off values were defined (Ret‐He: 31.2 pg, sTfR/log(fer): 0.91, and Tf/log(fer): 1.71). With both Thomas‐plots the amount of e causa ignota (ECI) cases dropped from 39% to 27%. A more in depth analysis on the iron status of anemia of chronic disease (ACD) patients and a subdivision between latent and classical iron deficiencies could be made with the help of both plots. A shift from classical iron deficiency anemia (IDA) cases according to the classical Thomas‐plot toward functional IDA according to the alternative Thomas‐plot was observed. Conclusion: The alternative Thomas‐plot is an effective tool that gives a more in depth view on the iron status of anemic patients. In addition, it is easier to implement due to the use of transferrin rather than the soluble transferrin receptor. [ABSTRACT FROM AUTHOR]

Aims: Falsely lower or even negative phosphatidylethanol (PEth) levels may theoretically be seen in patients with haemolytic diseases, and the present study aimed to elucidate this hypothesis. Methods: PEth and carbohydrate‐deficient transferrin (CDT) from 9893 serum and whole blood samples were included along with markers of haemolysis (i.e. haptoglobin, HbA1c, reticulocytes, LD and Hb). Cases showing discrepancy between PEth and CDT, that is, a low PEth value and a high CDT value, were considered to be possibly caused by falsely lowered PEth despite high alcohol consumption. These cases (N = 233) were compared to the control group without PEth and CDT mismatch. Results: The levels of haptoglobin were significantly lower in the cases showing low PEth and high CDT (estimate = −0.62, p = 0.002). The levels of HbA1c (estimate = −3.26, p = 0.001) and Hb (estimate = −0.507, p < 0.001) were also significantly lower in this group. These findings indicate haemolytic diseases in the low PEth/high CDT group. There were no significant differences for reticulocytes and LD concentrations between the low PEth/high CDT group and the control group. Conclusions: These results indicate that falsely low PEth values could be associated with markers of haemolytic diseases, although more research is needed to highlight this further. [ABSTRACT FROM AUTHOR]

Introduction: Established parameters for the investigation of suspected iron deficiency have recognized limitations that affect their sensitivity and specificity. Reticulocyte haemoglobin content (RHC) is an early biomarker of iron deficiency or restriction, which also demonstrates response to iron therapy. RHC parameters are offered on all major automated haematology analysers. There is no external quality assessment (EQA) for RHC in the UK, restricting its wider clinical application. The UK National External Quality Assessment Scheme (UKNEQAS) for Haematology has completed a pilot study on the use of stabilized blood for RHC EQA. Methods: Blood was obtained from two JAK2 V617F mutation-positive, polycythaemia vera patients undergoing regular venesection, and from a healthy volunteer donor. Aliquots of each donation were distributed to users of Abbott, Horiba, Siemens and Sysmex automated analysers for RHC analysis on days 1, 3, 5 and 7. Results: Results were received from 20 laboratories using four different platforms. The daily mean and standard deviation (SD) were calculated for each aliquot, by day of analysis and platform. With RHC there was no significant within-platform difference (p > 0.5) between testing days, although there were statistically significant differences (p < 0.001) between different platforms. The greatest difference was seen between Abbott MCHr and Horiba RhCc (-6.14 ± 1.25 pg). Despite inter-platform differences, it was possible to define iron deficient, borderline, and normal cut-offs to classify 95% of results correctly. Conclusions: The results demonstrate that it is possible to provide RHC EQA material using UKNEQAS standard procedures. The results are clinically relevant but indicate a requirement for inter-user comparison. [ABSTRACT FROM AUTHOR]

Introduction: Assessing the percentage of reticulocytes (%Retic) is useful for diagnosing and treating blood diseases that present with anaemia. The Celltac G+™ hematology analyzer (HA) uses a novel reticulocyte identification method that involves metachromatic nucleic acid staining with acridine orange and crossover analysis of emission light of DNA/RNA (determination of red cells, nucleic acid-containing cells, and platelets, RNP Determination™). The red and green fluorescence generated by stained singlestranded RNA and double-stranded DNA express immaturity and morphological abnormality of erythrocytes by detecting erythrocyte RNA and DNA content. Methods: The basic performance of the test automated analyzer (TAA) Celltac G+ was evaluated and compared with the flow cytometry reference method and the comparative automated analyzer (CAA) XN-1000/2000™. In addition, its precision, limit of quantity (LoQ), linearity, analytical measurement interval (AMI), accuracy, and comparability and the effects of interfering substances were evaluated. Results: Evaluation of %Retic by the TAA demonstrated good precision and linearity. The AMI was confirmed from 0.02 to 8.23, and the LoQ in %Retic as the coefficient of variation within an 11% limit (SD, within a 0.01 limit) was 0.14. TAA correlated well with the reference method and routine HA (CAA). Some deviations were found between TAA and CAA in DNA measurements of erythrocytes from abnormal samples. Conclusion: Celltac G+ uses a novel measurement principle and can assess erythrocyte immaturity independent of DNA contents. It represents a new HA that provides novel, useful information on immaturity and morphological abnormality of erythrocytes. [ABSTRACT FROM AUTHOR]

Background: Clinical pathology and body weight information for the cynomolgus monkey in the literature is primarily derived from a small number of animals with limited age ranges, varying geographic origins, and mixed genders. Objectives: This study aimed to summarize the age‐ and sex‐related changes in clinical pathology analytes and body weights in cynomolgus monkeys of Mauritian origin. Methods: Pre‐study age and body weight data were reviewed in 1819 animals, and pre‐study hematologic, coagulation, and serum biochemical analytes were reviewed in 1664 animals. Results: Body weights were statistically higher (P < 0.01) in males than females in all age groups (2–10 years). These measurements became prominent after 4 years of age and peaked at 7 to 8 years of age in both sexes. Sex‐related differences were noted in reticulocyte (RETIC) counts, creatinine, cholesterol, and triglyceride concentrations, and alkaline phosphatase (ALP) and gamma‐glutamyl transferase (GGT) activities. Age‐related differences were noted in RETIC and lymphocyte counts, creatinine, triglyceride, phosphorus, and globulin concentrations, and ALP and GGT activities. The youngest (2 to <3 year) age group had the fewest number of clinical pathologic analyte differences including ALP and GGT activity differences which occurred in all age groups from 2 to 10 years; they also had age‐related lower globulin concentrations. There were no age‐ or sex‐related differences in coagulation measurands. Conclusions: Sexual dimorphism in body weight was apparent for all ages from 2 to 10 years of age. The only difference in clinical pathology analytes unique to the 2 to <3 years of age group were age‐related lower globulin levels. [ABSTRACT FROM AUTHOR]

Objectives: Large‐scale generation of universal red blood cells (RBCs) from O‐negative (O‐ve) human induced pluripotent stem cells (hiPSCs) holds the potential to alleviate worldwide shortages of blood and provide a safe and secure year‐round supply. Mature RBCs and reticulocytes, the immature counterparts of RBCs generated during erythropoiesis, could also find important applications in research, for example in malaria parasite infection studies. However, one major challenge is the lack of a high‐density culture platform for large‐scale generation of RBCs in vitro. Materials and Methods: We generated 10 O‐ve hiPSC clones and evaluated their potential for mesoderm formation and erythroid differentiation. We then used a perfusion bioreactor system to perform studies with high‐density cultures of erythroblasts in vitro. Results: Based on their tri‐lineage (and specifically mesoderm) differentiation potential, we isolated six hiPSC clones capable of producing functional erythroblasts. Using the best performing clone, we demonstrated the small‐scale generation of high‐density cultures of erythroblasts in a perfusion bioreactor system. After process optimization, we were able to achieve a peak cell density of 34.7 million cells/ml with 92.2% viability in the stirred bioreactor. The cells expressed high levels of erythroblast markers, showed oxygen carrying capacity, and were able to undergo enucleation. Conclusions: This study demonstrated a scalable platform for the production of functional RBCs from hiPSCs. The perfusion culture platform we describe here could pave the way for large volume‐controlled bioreactor culture for the industrial generation of high cell density erythroblasts and RBCs. [ABSTRACT FROM AUTHOR]

Red blood cells are produced by terminal erythroid differentiation, which involves the dramatic morphological transformation of erythroblasts into enucleated reticulocytes. Microtubules are important for enucleation, but it is not known if the centrosome, a key microtubule‐organizing center, is required as well. Mice lacking the conserved centrosome component, CDK5RAP2, are likely to have defective erythroid differentiation because they develop macrocytic anemia. Here, we show that fetal liver‐derived, CDK5RAP2‐deficient erythroid progenitors generate fewer and larger reticulocytes, hence recapitulating features of macrocytic anemia. In erythroblasts, but not in embryonic fibroblasts, loss of CDK5RAP2 or pharmacological depletion of centrosomes leads to highly aberrant spindle morphologies. Consistent with such cells exiting mitosis without chromosome segregation, tetraploidy is frequent in late‐stage erythroblasts, thereby giving rise to fewer but larger reticulocytes than normal. Our results define a critical role for CDK5RAP2 and centrosomes in spindle formation specifically during blood production. We propose that disruption of centrosome and spindle function could contribute to the emergence of macrocytic anemias, for instance, due to nutritional deficiency or exposure to chemotherapy. Synopsis: Immature erythroid cells differentiate into red blood cells by undergoing a defined number of cell divisions followed by ejection of their nuclei. This study investigates the role of centrosomes in this process using ex vivo differentiation of fetal liver‐derived erythroid progenitors in mice. The conserved centrosomal protein CDK5RAP2 facilitates erythroid differentiation both ex vivo and in vivoCentrosomes and CDK5RAP2 are crucial for normal spindle assembly in erythroblasts but not in embryonic fibroblastsErythroblasts lacking CDK5RAP2 or centrosomes develop tetraploidyTP53 is activated in erythroblasts lacking CDK5RAP2 or centrosomes but is not responsible for the observed defects in erythropoiesisUnlike embryonic fibroblasts, erythroblasts do not expand their centrosomes in mitosis [ABSTRACT FROM AUTHOR]

Iron overload is directly associated with diabetes mellitus, loss of islet beta cell, and insulin resistance. Likewise, long noncoding RNA (lncRNA) is associated with type 2 diabetes (T2D). Moreover, lncRNAs could be induced by iron overload. Therefore, we are going to explore the molecular mechanism of lncRNA XIST in iron overload-related T2D. Real-time quantitative PCR and Western blot were used to detect gene and protein levels, respectively. TUNEL and MTT assay were performed to examine cell survival. The glucose test strip, colorimetric analysis kit, ferritin ELISA kit, and insulin ELISA kit were performed to examine the levels of glycolic, iron, and total iron-binding capacity, ferritin, and insulin in serum. Fluorospectrophotometry assay was used to examine labile iron pool level. XIST was higher expressed in T2D and iron overload-related T2D rat tissues and cells, and iron overload-induced promoted XIST expression in T2D. Higher XIST expression was associated with iron overload in patients with T2D. Knockdown of XIST alleviated iron overload and iron overload-induced INS-1 cells injury. Further, we found that XIST can sponge miR-130a-3p to trigger receptor-like kinase 2 (ALK2) expression. Moreover, knockdown of ALK2 alleviated iron overload and iron overload-induced INS-1 cells injury by inhibiting bone morphogenetic protein 6 (BMP6)/ALK2/SMAD1/5/8 axis but reversed with XIST upregulation, which was terminally boosted by overexpression of miR-130a-3p. XIST has the capacity to promote iron overload and iron overload-related T2D initiation and development through inhibition of ALK2 expression by sponging miR-130a-3p, and that targeting this axis may be an effective strategy for treating patients with T2D. [ABSTRACT FROM AUTHOR]

Introduction: Reticulocytes are erythroid precursors that develop into mature erythrocytes, and they are an important tool to assess erythropoietic activity, as their count indicates the balance between the cells released from the bone marrow, their stage of maturity, and their rate of development into mature erythrocytes. Considering the described biological variability of the absolute reticulocyte count (ARC) and the immature reticulocyte fraction (IRF) and the limited information available on these hematological parameters in children, this study determined the reference intervals (RIs) of these parameters in a healthy pediatric population. Methods: A retrospective, observational, and analytical study was designed to establish RIs for the ARC and the IRF according to age and sex. An indirect sampling method was applied to a mixed database of complete blood counts from children aged 2 months to 18 years, using the truncated maximum likelihood indirect method for reference interval estimation. Percentiles were calculated to obtain bimodal RIs. Results: From a total of 190,812 samples, 6,814 were selected. Gender stratification was not necessary for the ARC and the IRF but they required partitioning into six and two age groups, respectively. Conclusion: This study determined, by an indirect sampling method, RIs for the ARC and the IRF in a pediatric population according to age and sex. [ABSTRACT FROM AUTHOR]

The haematological module of the athlete biological passport (ABP) monitors longitudinal haematological variations that could be indicative of blood manipulation. This study applied a multi‐parametric model previously validated in elite cyclists to compare inferred and actual PV variations, whereas the potential influence of the oral contraceptive pill (OCP) cycle on the ABP blood biomarkers and plasma volume (PV) in 14 physically active women taking OCPs was also investigated. Blood and serum samples were collected each week for 8 weeks, and the ABP haematological variables were determined according to the World Anti‐Doping Agency guidelines. Transferrin (sTFN), ferritin (FERR), albumin (ALB), calcium (Ca), creatinine (CRE), total protein (TP) and low‐density lipoprotein (LDL) were additionally computed as 'volume‐sensitive' variables in a multivariate analysis to determine individual estimations of PV variations. Actual PV variations were indirectly measured using a validated carbon monoxide rebreathing method. We hypothesised ABP markers to be stable during a standard OCP cycle and estimated PV variations similar to measured PV variations. Measured PV variations were in good agreement with the predictions and allowed to explain an atypical passport finding (ATPF). The ABP biomarkers, Hbmass and PV were stable over 8 weeks. Significant differences occurred only between Week 7 and Week 1, with lower levels of haemoglobin concentration ([Hb]), haematocrit (HCT) and red blood cell count (RBC)(−4.4%, p < 0.01; −5.1%, p < 0.01; −5.2%, p < 0.01) and higher levels of PV at week 7 (+9%, p = 0.05). We thus concluded that estimating PV variations may help interpret individual ABP haematological profiles in women. [ABSTRACT FROM AUTHOR]

Whole organismal declines in metabolic performance due to ageing are known to occur in virtually all higher vertebrates, the causes of such declines, however, are not fully understood. In mammals, haemoglobin concentration, haematocrit and red blood cell count begin to decrease in later life. In birds however, such knowledge on changes in haematological variables with increasing age is still scarce and existing data largely focuses on nestlings or differences between juveniles and adults. Here, forty‐five male zebra finches Taeniopygia guttata, aged between 15 and 84 months were brought to their metabolic peak through exercise and then sampled for a full suite of haematological variables, including four directly measured primary haematological variables which were modelled against age as a continuous factor. We found a significant decline in haematocrit due to ageing in the birds (p = 0.029), a phenomenon observed in several other avian species but still unexplained. Haemoglobin did not change significantly (p = 0.156), while neither erythrocyte size or number showed any trends with age (p = 0.898 and p = 0.922 respectively) indicating that declining haematocrit is due to mechanisms other than decreasing cells size or number. We considered reticulocytes, being generally larger than mature erythrocytes as an explanation for declining haematocrit, however an a priori examination revealed an opposite result and increased with age (p = 0.016), leading us to believe that the decline in haematocrit with increasing age relates to differences in water homeostasis potential in older birds. The age dependence of haematological variables appears less clear in birds than in mammals. Besides oxygen carrying capacity, haematocrit influences the viscoelastic properties of blood, and it remains to be shown if this decline in haematocrit in birds can explain declining aerobic metabolism. [ABSTRACT FROM AUTHOR]

Background: Reticulocyte indices have been suggested as alternatives to transferrin saturation (TSAT) for iron status assessment in humans and dogs but they have not been evaluated thoroughly in cats. Objectives: To assess the value of the reticulocyte indices for the diagnosis of iron deficiency in cats with chronic kidney disease (CKD) and chronic hematuria associated with subcutaneous ureteral bypasses (SUBs). Animals: Sixty‐four cats: 16 healthy, 14 CKD without SUB, and 34 CKD with SUB. Methods: Prospective observational cross‐sectional study of cats presented for routine nephrology visits. Primary outcomes included assessment of the diagnostic values of erythrocyte indices (mean corpuscular volume, hemoglobin, and hemoglobin concentration: MCV, MCH, and MCHC) and reticulocyte indices (mean corpuscular volume, MCVr; corpuscular hemoglobin, CHr), using TSAT as reference. Results: Iron deficiency was diagnosed in 9/64 cats (14%). A receiver‐operating characteristic curve analysis yielded a moderate discriminatory value for CHr in this diagnosis: area under the curve [AUC] =.75 (95% confidence interval, 0.48‐0.89); P =.006; sensitivity 67%, specificity 82% for a cutoff of 15.9 pg. This compared favorably to MCVr (AUC =.63; P =.29), MCV (AUC =.58; P =.45), MCH (AUC =.64; P =.19), and MCHC (AUC =.7; P =.03). Conclusion and Clinical Importance: CHr added moderate value to the diagnosis of iron deficiency in cats with CKD. [ABSTRACT FROM AUTHOR]

Introduction: The athlete biological passport monitors blood variables over time to uncover blood doping. With the phasing in of a new series of blood analyzers, the Sysmex XN series, it was necessary to examine the comparability of results with the previously employed XT/XE series. A previous comparison between XN and XT/XE series suggested a small but significant bias between the two instruments in the measurements of RET%. Here, we examined the comparability of RET% on the XN and XT/XE platform using data collected over the first year since the transition. Methods: The comparability of results obtained from XN and XT/XE instruments was assessed using three datasets: (i) 767 blood samples measured on both instrument series in 22 WADA‐accredited laboratories, (ii) 27 323 samples measured on either instrument across 31 laboratories, and (iii) 119 clinical samples and 110 anti‐doping samples measured on both instruments in a single laboratory. Results: Analysis of the three datasets confirms the previous observation of a bias toward higher RET% values for samples measured on Sysmex XN instruments compared with the XT/XE series. Using data across a larger number of XN instruments and a larger athlete population, the current work suggests that the bias is proportional and slightly higher than previously observed across most of the range RET% values. Conclusion: A model is proposed for the comparison of data across XN and XT/XE technologies whereby the instrument bias increases proportionally with RET% measured on Sysmex XN Series, but where the rate of increase is negatively related to IRF%. [ABSTRACT FROM AUTHOR]

Introduction: Iron deficiency anaemia (IDA) and anaemia of chronic disease (ACD) are common causes of anaemia with similar clinical and laboratory features. IDA is caused by low iron stores while ACD is due to iron‐restricted erythropoiesis occurring in inflammatory states. Differential diagnosis requires analysis of multiple biochemical and haematological parameters. IDA can occur simultaneously to ACD (mixed aetiology). It is essential that true iron deficiency is identified, as these patients will require iron therapy. This preliminary study investigated whether hepcidin, the master regulator of iron homeostasis, in conjunction with reticulocyte haemoglobin equivalent (RetHe) has the potential to differentiate IDA from ACD, and to exclude IDA in patients with mixed aetiology. Methods: Hepcidin concentration (measured using a commercially available ELISA method), RetHe, and iron parameters along with C‐reactive protein (CRP) were analysed in 77 Gastroenterology patients with anaemia in a secondary care setting. Results: Receiver operator characteristic (ROC) analysis showed that hepcidin at an optimal cut‐off concentration of <6ng/ml could identify IDA with a sensitivity and specificity of 88.9% and 90.6% respectively and could distinguish ACD from IDA with both a sensitivity and specificity of 100% at a cut‐off of >46ng/ml. Identifying true IDA in mixed aetiology patients could be achieved by RetHe analysis and applying an optimal cut‐off of <30pg. Conclusion: Hepcidin, in conjunction with RetHe, offers a new simplified diagnostic pathway for differential diagnosis of IDA and ACD, thereby reducing the diagnostic turnaround time and allowing appropriate treatment of patients with a true iron deficiency. [ABSTRACT FROM AUTHOR]

Introduction: The objective of the study was to evaluate the performance of the Abbott Alinity hq advanced multi angle polarized scatter separation (MAPSSTM)‐based optical RBC technology, for the differentiation between iron deficiency anemia (IDA) and β‐thalassemia carrier status. Methods: Four hundred and sixty‐four samples were analyzed. 228 were healthy controls, 30 were β‐thalassemia carriers, and 40 were IDA. Receiver operating characteristics analysis evaluated the performance of red cell parameters and mathematical formulas. Results: RBC concentration was the most efficient discriminant (area under the curve; AUC of 0.963, Youden Index of 0.88) followed by red blood cell distribution width in size distribution (AUC of 0.960 and YI of 0.86), and red blood cell distribution width coefficient of variation (AUC of 0.924, and YI of 0.74). The absolute reticulocyte concentration showed good diagnostic efficiency, with AUC of 0.808. Hemoglobin distribution width, the %CV of directly measured cellular hemoglobin concentration, and CHCr, the average hemoglobin concentration of reticulocytes have emerged as novel discriminating parameters, with AUC of 0.749 and 0.785, respectively. The England and Fraser index was the best discriminating mathematical formula based on Youden Index of 0.91. The Ricerca, red blood cell distribution width index, Green and King, and Mentzner Index formulas also showed strong discriminative power. The Shine and Lal index, together with the recent mathematical formula M/H, (ratio of percent microcytic and hypochromic red blood cells) demonstrated moderate performance with AUC of 0.796 and 0.740, respectively. Conclusion: Extended red cell analysis delivered by the advanced optical technology on the Alinity hq hematology analyzer has efficient diagnostic utility in the initial discrimination of the two most common microcytic anemias: IDA and β‐thalassemia trait. [ABSTRACT FROM AUTHOR]

Mitophagy is a form of autophagy specialized to selectively remove mitochondria. Although the PINK1/Parkin pathway is the best described mitophagy of damaged mitochondria, receptor/mediated mitophagy seems to have a pivotal role in cellular development and specialization. The most studied mitophagy receptor BCL2/adenovirus E1B 19‐kDa‐interacting protein 3‐like (BNIP3L/NIX) is shown to be important for the programmed removal of healthy mitochondria during terminal differentiation of erythrocytes, but its role has been proven in various cell types. Despite recent advances in our understanding of its regulation by phosphorylation and dimerization, there remain numerous questions on how BNIP3L/NIX tightly balances between cellular life and death decisions. This brief review intends to summarize ongoing dilemmas related to BNIP3L/NIX. [ABSTRACT FROM AUTHOR]

Introduction: The aim of our study was to examine the relationship of hepcidin‐25 with red blood cell and reticulocyte indices and to evaluate the diagnostic properties of hepcidin‐25 in the assessment of positive iron balance in end‐stage renal disease (ESRD) patients. Methods: Eighty anemic ESRD patients (hemoglobin < 110 g/L) were classified as having iron deficiency (ID, N = 20), iron sufficiency (IS, N = 29), and positive iron balance (PB, N = 31) using the conventional biomarkers for iron status evaluation. Hepcidin‐25 was determined by a chemiluminescent direct ELISA. Results: Hepcidin‐25 was significantly negatively correlated with the proportion of hypochromic erythrocytes (%HYPO) (P =.034) and immature reticulocyte fraction (P =.010) in ID and with the absolute reticulocyte concentration in ID (P =.048) and PB (P =.040). In multivariate models, hepcidin‐25 was independently negatively associated with the mean reticulocyte hemoglobin content (CHr; β = −0.493, P =.004) and red blood cell size factor (RSf) (β = −0.334, P =.036) only in the PB group. The best hepcidin‐25 value to exclude PB was 66.13 µg/L, showing a sensitivity of 61.3%, a specificity of 75.5%, and an AUC of 0.808. Conclusion: Our results suggest that hepcidin‐25 levels are independently negatively associated with the iron demand for the most recent erythropoiesis only in PB. Hepcidin‐25 performed acceptable in discriminating anemic ESRD patients with positive iron balance and may prove to be a useful additional tool in the evaluation of iron status. [ABSTRACT FROM AUTHOR]

Introduction: Reticulocytes (RET) are immature red blood cells, and RET enumeration in peripheral blood has important clinical value in diagnosis, treatment efficacy observation, and prognosis of anemic diseases. For RET enumeration, flow cytometric reference method has shown to be more precise than the manual method by light microscopy. However, flow cytometric method generates occasionally spurious RET counts in some situations. The manual method, which is subjective, imprecise, and tedious, currently remains as an accepted reference method. As a result, there is a need for manual method to be more objective, precise, and rapid. Methods: 40 EDTA‐K2 anticoagulated whole blood samples were randomly selected for the study. 784 microscopic images were taken from blood slides as dataset, and all mature RBCs and RETs in these images were located and labeled by experienced experts. Then, we leverage a Faster R‐CNN deep neural network to train a RET detection model and evaluate the model. Results: Both the recall and precision rate of the model are more than 97%, and average analysis time of a single image is 0.21 seconds. Conclusion: The deep learning method shows outstanding performance including high accuracy and fast speed. The experimental results show that the deep learning method holds the potential to act as a rapid computer‐aid method for manual RET enumeration for cytological examiners. [ABSTRACT FROM AUTHOR]

Introduction: UniCel DxH900 (Beckman Coulter, Miami, Florida, USA) is a quantitative, multi‐parameter, automated hematology analyzer for in vitro diagnostic use in clinical laboratories. The aim of this study was to evaluate the analytical performance of the new DxH900 analyzer to verify its diagnostic and clinical utility in the hematology laboratory of a tertiary care hospital in Spain. The most important and novel feature offered by DxH900 analyzer is providing MDW (monocyte distribution width), a new hematologic parameter which is being clinically validated as an early sepsis indicator with promising results. Methods: We evaluated imprecision (including MDW), linearity, and carryover of DxH900. Method comparison for cell blood count (CBC) was performed in relation to DxH800 with 100 samples. We compared leukocyte differential (DIFF) from DxH900 with manual 400‐cell differential. 390 samples were assessed for flag performance. Results: Results obtained for between days and within‐run imprecision were good. DxH900 showed excellent linearity (R = 1.00) over analytical range for white blood cells, red blood cells, hemoglobin, platelets, and reticulocyte count (RET) (R = 0.96) and no significant carryover effect. CBC and RET on the DxH900 correlated well with DxH800 (R ≥ 0.99). Comparison with manual differential showed excellent correlation (R ≥ 0.88), except for basophils. Flagging performance exhibited sensitivity over 90% for majority of alarm messages and very high negative predictive value (over 95%). Conclusion: UniCel DxH900 Coulter analyzer provides reliable results and fully comparable to DxH800. DxH900 is an accurate, highly precise analyzer with good analytical performances to be used effectively in high‐volume laboratories. [ABSTRACT FROM AUTHOR]

Aims: Total haemoglobin mass (tot-Hb) increases during high-altitude acclimatization. Normalization of tot-Hb upon descent is thought to occur via neocytolysis, the selective destruction of newly formed erythrocytes. Because convincing experimental proof of neocytolysis is lacking, we performed a prospective study on erythrocyte survival after a stay at the Jungfraujoch Research Station (JFJRS; 3450 m). Methods: Newly formed erythrocytes of 12 male subjects (mean age 23.3 years) were age cohort labelled in normoxia (110 m) and during a 19-day high-altitude sojourn by ingestion of 13C2-and 15N-labelled glycine respectively. Elimination dynamics for erythrocytes produced in normoxia and at high altitude were measured by isotope ratio mass spectrometry of haem, by determining tot-Hb, reticulocyte counts, erythrocyte membrane protein 4.1a/4.1b ratio and by mathematical modelling. Results: Tot-Hb increased by 4.7% ± 2.7% at high altitude and returned to pre-altitude values within 11 days after descent. Elimination of 13C-(normoxia) and 15N-(high altitude) labelled erythrocytes was not different. Erythropoietin levels and counts of CD71-positive reticulocytes decreased rapidly after descent. The band 4.1a/4.1b ratio decreased at altitude and remained low for 3-4 days after descent and normalized slowly. There was no indication of haemolysis. Conclusion: We confirm a rapid normalization of tot-Hb upon descent. Based on the lack of accelerated removal of age cohorts of erythrocytes labelled at high altitude, on patterns of changes in reticulocyte counts and of the band 4.1a/4.1b ratio and on modelling, this decrease did not occur via neocytolysis, but by a reduced rate of erythropoiesis along with normal clearance of senescent erythrocytes. [ABSTRACT FROM AUTHOR]

Introduction: In the hub and spoke laboratory network, the number of hematology analyzers (HAs) within each core center has increased, and the control of HAs alignment is becoming necessary requirement to ensure analytical quality. In this scenario, HA alignment can be assessed by analyzing the same control material used for internal quality control on multiple HAs, assuming its commutability. The aim of the study was to verify the applicability of a protocol for the alignment of HAs based on control material rather than on fresh whole‐blood samples. Methods: The alignment of five HAs was evaluated for red (RBC, Hb, MCV, RET), white (WBC, NE, LY, MO, EO, BA, IG), and platelet (PLT) series parameters, following a protocol by SIBioC, using human sample (HS) and quality control material (QC), after the verification of commutability, according to the IFCC protocol. Maximum bias was derived from biological variation data. Results: A complete alignment between instruments was confirmed for the majority of the parameters investigated both for HS and QC material. Partial misalignments or inconcludent results were instead evident for MCV, MO, EO, BA, and IG. Interestingly, QC material was found to be not commutable for LY, MO, and BA. Conclusion: The alignment of hematologic analyzers for main cell population parameters may be verified with both QC and HS, displaying consistent results and interpretation. The evaluation for some white series parameters (EO, BA, and IG) is critical, and particular attention must be paid to the values of the material used for the alignment. [ABSTRACT FROM AUTHOR]

The widespread use of tris(1,3‐dichloro‐2‐propyl) phosphate (TDCIPP) as a flame retardant has led to its release to the environment. Thus, the toxicological effects of TDCIPP on humans and animals are of importance. For better understanding of its potential toxicities, TDCIPP (250, 500, or 650 mg/kg/day) or vehicle control was administrated orally to adult male Wistar‐Imamichi rats for 7 days. After the final administration of compounds, organ weights, histopathology, blood biochemistry, and hematology were examined. Hepatic toxicity was observed at doses ≥ 500 mg/kg/day of TDCIPP, and renal toxicity was observed at 650 mg/kg/day. The anti‐androgenic activity of TDCIPP was previously confirmed in vitro and in vivo, but weights of epididymis, an androgen‐dependent organ, were not affected by TDCIPP treatment in adults. Serum alkaline phosphatase activity was significantly decreased in all TDCIPP‐treated rats independent of dose. Hemoglobin concentration, hematocrit, red blood cell count, and reticulocyte count were decreased in all TDCIPP‐treated rats, but mean corpuscular volume, total iron‐binding capacity, and serum iron were normal, suggesting that renal anemia was caused by TDCIPP. Together with previous reports on effects of anti‐androgenic substances on red blood cell indices, anemia caused by TDCIPP could be due to its anti‐androgenic activity. These considerations will contribute to further assessment of the toxicity of the compound. Tris(1,3‐dichloro‐2‐propyl) phosphate (TDCIPP) was administrated orally to adult male Wistar‐Imamichi rats for 7 days. Serum ALP activity was significantly decreased in all TDCIPP‐treated rats independent of dose. Hemoglobin concentration, hematocrit, red blood cell count, and reticulocyte count were decreased by TDCIPP, suggesting that renal anemia is possibly caused by its anti‐androgenic activity. These considerations will contribute to further assessment of the toxicity of the compound. [ABSTRACT FROM AUTHOR]

Background: Recent case reports have described the efficacy of daratumumab to treat refractory pure red cell aplasia (PRCA) following major ABO mismatched allogeneic hematopoietic stem cell transplantation (HSCT). In this report, we describe the use of daratumumab as a first-line agent for treatment of delayed red blood cell (RBC) engraftment following a major ABO mismatched pediatric HSCT and provide a review of the literature.Study Design and Materials: We report on a 14-year-old with DOCK8 deficiency who underwent a myeloablative, haploidentical bone marrow transplant from her major ABO mismatched sister (recipient O+, donor A+) for treatment of her primary immunodeficiency. Despite achieving full donor chimerism, she had delayed RBC engraftment requiring ongoing transfusions. Due to iron deposition, symptomatic anemia, and persistence of anti-A iso-hemagglutinins despite discontinuation of immunosuppression, treatment for delayed RBC engraftment with the CD38-targeted monoclonal antibody daratumumab was selected as a less immunosuppressive agent that could more selectively target iso-hemagglutinin producing plasma cells without causing broad B-cell aplasia.Results: Clinical effect with daratumumab was demonstrated by reduced iso-hemagglutinin titer, increased reticulocytosis, normalization of her hemoglobin, and transfusion independence. In the 11-month follow-up period to date, no additional transfusions or immunosuppression have been necessary, despite persistence of low-level anti-A iso-hemagglutinin.Conclusion: Our experience suggests that daratumumab was an effective first-line therapy for delayed RBC engraftment and that earlier consideration for daratumumab in treatment of delayed RBC engraftment may be warranted. [ABSTRACT FROM AUTHOR]

The micronucleus test (MNT) is the most widely applied short‐term assay to detect clastogens or spindle disruptors. The use of flow cytometry (FCM) has been reported for micronucleated erythrocytes scoring in peripheral blood. The aim of this study was to develop a novel and practical protocol for MNT in rat peripheral blood by FCM, with the method validation. CD71‐fluorescein isothiocyanate and DRAQ5 were adopted for the fluorescent staining of proteins and DNA, respectively, to detect micronuclei. To validate the method, groups of male Sprague–Dawley rats (five per group) received two oral gavage doses at 0 and 24 h of six chemicals (four positive mutagens: ethyl methanesulphonate [EMS], cyclophosphamide [CP], colchicine [COL], and ethyl nitrosourea [ENU]; two nongenotoxic chemicals: sodium saccharin and eugenol). Blood samples were collected from the tail vein before and on the five continuous days after treatments; all of which were analyzed for micronuclei presence by both the manual (Giemsa staining) and FCM methods. The FCM‐based method consistently demonstrated highly sensitive responses for micronucleus detection at all concentrations and all time points for EMS, CP, COL, and ENU. Sodium saccharin and eugenol could be identified as negative in this protocol. Results obtained with the FCM‐based method correlated well with the micronucleus frequencies (r = 0.659–0.952), and the proportion of immature erythrocytes (r = 0.915–0.981) tested by Giemsa staining. The method reported here, with easy operation, low background, and requirement for a regular FCM, could be an efficient system for micronucleus scoring. The objective of this study was to develop a novel and practical protocol for the micronucleus test in rat peripheral blood by flow cytometry (FCM), followed by method validation. Male Sprague–Dawley rats were treated at 24‐h intervals by gavage with six chemicals (ethyl methanesulphonate [EMS], cyclophosphamide [CP], colchicine [COL], ethyl nitrosourea [ENU], sodium saccharin, and eugenol). The method reported here, with easy operation, low background, and requirement for a regular FCM, could be an efficient system for micronucleus scoring. [ABSTRACT FROM AUTHOR]

The article discusses research which investigated red blood cell (RBC) and reticulocyte-related parameters in adult transfusion-dependent thalassemia (TDT) and non-transfusion-dependent thalassemia (NTDT) patients with beta thalassemia. Topics explored include the analysis of RBC transfusion history of patients involved in the study, the results of the whole blood analysis and complete blood count performed, and the correlation between RBC parameters and pretransfusion hemoglobin levels.

The article explores Sideroblastic anemias (SA) are characterized by the presence of ring sideroblasts (RS) in the bone marrow secondary to impaired heme synthesis. RS result from the accumulation of iron in the mitochondria surrounding the nuclear membrane of erythroid precursors. Acquired causes of SA include nonclonal (alcohol consumption, hypothermia, copper deficiency, zinc toxicity, isoniazid, and other drugs) and clonal disorders.

Background: Vel expression on erythrocytes is variable due to polymorphisms, complicating Vel typing. Weak Vel expression can be caused by mutations within SMIM1 in a heterozygous setting, suggesting a dominant negative effect of SMIM1 mutants on wild type (wt)SMIM1 expression. Here we report how SMIM1 expression is regulated during erythropoiesis, to understand its variable expression on erythrocytes. Study Design and Methods: Peripheral blood reticulocytes at different stages, cultured erythroid precursors and HEK293T cells were used to investigate expression and putative competition between wtSMIM1 and mutated SMIM1 VEL*01W.01, (c.152T>A (p.Met51Lys)), VEL*01W.02 (c.152T>G (p.Met51Arg)), and VEL*01W.03 (c.161T>C (p.Leu54Pro)). Results: Depending on the mutations in SMIM1 an effect on total and membrane expression of SMIM1 was observed in transfected HEK293T cells, but co‐expression of wtSMIM1 and mutatedSMIM1 did not have an effect on wtSMIM1 membrane expression. During differentiation of donors expressing VEL*01W.01, VEL*01W.03, Vel‐positive, Vel‐negative (homozygote SMIM1*64_80del), and Vel‐heterozygote SMIM1*64_80del primary human erythroblasts no overt defect was found in Vel expression dynamics or total SMIM1 expression levels when compared with wtSMIM1 erythroblasts. However, during enucleation, total Vel expression was significantly lower on reticulocytes of Vel‐weak donors expressing heterozygote mutated SMIM1 compared to Vel‐positive or Vel‐heterozygote SMIM1*64_80del donors, while Vel expression on extruded nuclei was maintained. In addition, reticulocyte maturation in vivo showed further loss of Vel expression in these individuals and nearly absent on erythrocytes. Conclusion: These results suggest that SMIM1 mutations exert a dominant negative effect on wtSMIM1 probably by affecting SMIM1 multimerization and thereby Vel epitope presentation at the latest stages of erythroid differentiation. [ABSTRACT FROM AUTHOR]

Arachidonic acid 15‐lipoxygenases (ALOX15) are lipid peroxidizing enzymes, which has previously been implicated in the maturational breakdown of intracellular organelles and plasma membrane remodeling during reticulocyte‐erythrocyte transition. Conventional Alox15−/− mice are viable, develop normally but do not exhibit a major defective erythropoietic phenotype. To characterize the putative in vivo relevance of Alox15 for red blood cell development, we explored the impact of systemic inactivation of the Alox15 gene on mouse erythropoiesis. We found that Alox15−/− mice exhibited reduced erythrocyte counts, elevated reticulocyte counts and red cell hyperchromia. The structure of the plasma membrane of Alox15−/− erythrocytes is altered and a significant share of the red cells was present as echinocytes and/or acanthocytes. An increased share of the Alox15−/− erythrocytes cells were annexin V positive, which indicates a loss of plasma membrane asymmetry. Erythrocytes of Alox15−/− mice were more susceptible to osmotic hemolysis and exhibited a reduced ex vivo life span. When we transgenically expressed human ALOX15 in Alox15−/− mice under the control of the aP2 promoter the defective erythropoietic system was rescued and the impaired osmotic resistance was normalized. Together these data suggest the involvement Alox15 in the maturational remodeling of the plasma membrane during red cell development. [ABSTRACT FROM AUTHOR]

The aim of the present in vivo study was to determine the kinetics of the genotoxic and cytotoxic activities of cladribine and clofarabine in mouse normoblasts using flow cytometry. Mice in groups of five were treated with cladribine or clofarabine. Blood samples were obtained from the mouse tails before treatment and every 8 hr posttreatment for 72 hr. These samples were cytometrically scored for micronucleated reticulocytes (RETs), and the percentage of RETs was determined. The results showed that clofarabine and cladribine have early cytotoxic effects that are related to the genotoxic effects reported in previous studies; the drugs have both complex long‐lasting genotoxic and cytotoxic kinetic activity, with similar profiles that suggest a relationship between the genotoxic and cytotoxic parameters. The initial genotoxkinetics timing of clofarabine is equivalent to those of difluorodeoxycytidine, likely because both agents inhibit DNA polymerase. Clofarabine shows a higher genotoxic and cytotoxic efficiency than cladribine, in agreement with previous results. [ABSTRACT FROM AUTHOR]

We previously described flow cytometry‐based methods for scoring the incidence of micronucleated reticulocytes (MN‐RET) and PIG‐A mutant phenotype reticulocytes (MUT RET) in rodent and human blood samples. The current report describes important methodological improvements for human blood analyses, including immunomagnetic enrichment of CD71‐positive reticulocytes prior to MN‐RET scoring, and procedures for storing frozen blood for later PIG‐A analysis. Technical replicate variability in MN‐RET and MUT RET frequencies based on blood specimens from 14 subjects, intra‐subject variability based on serial blood draws from 6 subjects, and inter‐subject variation based on up to 344 subjects age 0 to 73 years were quantified. Inter‐subject variation explained most of the variability observed for both endpoints (≥77%), with much lower intra‐subject and technical replicate variability. The relatively large degree of inter‐subject variation is apparent from mean and standard deviation values for MN‐RET (0.15 ± 0.10%) and MUT RET (4.7 ± 5.0 per million, after omission of two extreme outliers). The influences of age and sex on inter‐subject variation were investigated, and neither factor affected MN‐RET whereas both influenced MUT RET frequency. The lowest MUT RET values were observed for subjects <11 years old, and males had moderately higher frequencies than females. These results indicate that MN‐RET and MUT RET are automation‐compatible biomarkers of genotoxicity that bridge species of toxicological interest to include human populations. These data will be useful for appropriately designing future human studies that include these biomarkers of genotoxicity, and highlight the need for additional work aimed at identifying the sources of inter‐individual variability reported herein. [ABSTRACT FROM AUTHOR]