Machine Learning-Based Prediction of Hemoglobinopathies Using Complete Blood Count Data.

BACKGROUND: Hemoglobinopathies, the most common inherited blood disorder, are frequently underdiagnosed. Early identification of carriers is important for genetic counseling of couples at risk. The aim of this study was to develop and validate a novel machine learning model on a multicenter data set, covering a wide spectrum of hemoglobinopathies based on routine complete blood count (CBC) testing. METHODS: Hemoglobinopathy test results from 10 322 adults were extracted retrospectively from 8 Dutch laboratories. eXtreme Gradient Boosting (XGB) and logistic regression models were developed to differentiate negative from positive hemoglobinopathy cases, using 7 routine CBC parameters. External validation was conducted on a data set from an independent Dutch laboratory, with an additional external validation on a Spanish data set (n = 2629) specifically for differentiating thalassemia from iron deficiency anemia (IDA). RESULTS: The XGB and logistic regression models achieved an area under the receiver operating characteristic (AUROC) of 0.88 and 0.84, respectively, in distinguishing negative from positive hemoglobinopathy cases in the independent external validation set. Subclass analysis showed that the XGB model reached an AUROC of 0.97 for ß-thalassemia, 0.98 for α0-thalassemia, 0.95 for homozygous α+-thalassemia, 0.78 for heterozygous α+-thalassemia, and 0.94 for the structural hemoglobin variants Hemoglobin C, Hemoglobin D, Hemoglobin E. Both models attained AUROCs of 0.95 in differentiating IDA from thalassemia. CONCLUSIONS: Both the XGB and logistic regression model demonstrate high accuracy in predicting a broad range of hemoglobinopathies and are effective in differentiating hemoglobinopathies from IDA. Integration of these models into the laboratory information system facilitates automated hemoglobinopathy detection using routine CBC parameters.

Interference of bovine hemoglobin-based oxygen carrier-201 (Hemopure) on four hematology analyzers.

INTRODUCTION: Hemoglobin-based oxygen carriers, for example HBOC-201 (Hemopure), are aimed to bridge acute anemia when blood transfusion is not available or refused by the patient. However, since HBOC-201 appears free in plasma, it interferes with laboratory tests. This study presents an overview of HBOC-201 interference on four commonly used hematology analyzers and suggests treatment monitoring possibilities. METHODS: Blood samples were spiked with therapeutic doses of HBOC-201 and nine hematology parameters were measured with the Sysmex XN-20, Siemens Advia 2120i, Abbott Alinity Hq and Abbot Cell Dyn Sapphire hematology analyzers. The results were compared to control samples and the bias was determined. RESULTS: Most parameters, including all cell counts, hematocrit and MCV, showed a non-significant bias compared to control. However, the standard, total hemoglobin (Hb) measurement as well as MCH and MCHC showed poor agreement with control, as HBOC-201 was included in this measurement. Yet, the flow cytometry-based Hb method quantified intracellular Hb in spiked samples, excluding HBOC-201. CONCLUSION: Of all included hematology parameters, only total Hb and the associated MCH and MCHC suffered from interference. In contrast, the flow cytometry-based Hb measurement provided an accurate measure of intracellular Hb. The difference between total Hb and cellular Hb represents the HBOC-201 concentration and can be used to monitor HBOC-201 treatment.

[Normocytic anaemia due to multiple causes]. / Een normocytaire anemie met meerdere oorzaken.

Anaemia can be subdivided into microcytic, normocytic and macrocytic categories, which give direction to the investigation to identify the cause of the anaemia. However, this is not always straightforward, as this case illustrates. A 73-year-old man with normocytic anaemia appeared to have both beta thalassemia trait (microcytic) and myelodysplastic syndrome (usually macrocytic). This led to a mixture of red blood cells of different sizes and an average normal MCV. The manual differentiation of the blood smear was helpful to find the right diagnosis.

[A patient with erythroderma and pruritus: Sézary syndrome]. / Een man met jeuk en roodheid van de huid.

BACKGROUND: Erythroderma could be the first sign of a cutaneous T-cell lymphoma (CTCL), such as Sézary syndrome. Causes of erythroderma include inflammatory dermatosis, toxicoderma, paraneoplastic erytroderma, and CTCL. Hence, diagnosing Sézary syndrome can be difficult. Sézary syndrome is a rare, aggressive disease characterized by erythroderma, generalized lymphadenopathy and the presence of clonally related neoplastic T-cells in skin, peripheral blood, and lymph nodes. Treatment consists of photochemotherapy (PUVA), radiotherapy, immunomodulatory agents, low dose cytotoxic agents, and intensive chemotherapy. Immunotherapy directed against CCR4 and PD1 are new, promising developments. CASE DESCRIPTION: A 51-year-old man presented with a 1-year history of progressive, itchy erythroderma and lymphocytosis. After extensive cytomorphological, histopathological and molecular examination the diagnosis of Sézary syndrome could be established. Combination treatment of interferon and photochemotherapy (PUVA) was started. CONCLUSION: Diagnostic delay in Sézary syndrome is common. Integrated cytomorphological, immunological, and molecular evaluation of peripheral blood in patients with unexplained erythroderma non-responsive to (topical) treatment is warranted.

Transition from fresh frozen plasma to solvent/detergent plasma in the Netherlands: comparing clinical use and transfusion reaction risks.

Plasma transfusion is indicated for replenishment of coagulative proteins to stop or prevent bleeding. In 2014, the Netherlands switched from using ~300mL fresh frozen plasma (FFP) units to using 200mL Omniplasma, a solvent/detergent treated pooled plasma (SD plasma), units. We evaluated the effect of the introduction of SD plasma on clinical plasma use, associated bleeding, and transfusion reaction incidences. Using diagnostic data from six Dutch hospitals, national blood bank data, and national hemovigilance data for 2011 to 2017, we compared the plasma/red blood cell (RBC) units ratio (f) and the mean number of plasma and RBC units transfused for FFP (~300mL) and SD plasma (200mL) for various patient groups, and calculated odds ratios comparing their associated transfusion reaction risks. Analyzing 13,910 transfusion episodes, the difference (Δf = fSD - fFFP) in mean plasma/RBC ratio (f) was negligible (Δfentire_cohort = 0.01 [95% confidence interval (CI): -0.02 - 0.05]; P=0.48). SD plasma was associated with fewer RBC units transfused per episode in gynecological (difference of mean number of units -1.66 [95% CI: -2.72, -0.61]) and aneurysm (-0.97 [-1.59, -0.35]) patients. SD plasma was further associated with fewer anaphylactic reactions than FFP (odds ratio 0.37 [0.18, 0.77; P<0.01]) while the differences for most transfusion reactions were not statistically significant. SD plasma units, despite being one third smaller in volume than FFP units, are not associated with a higher plasma/RBC ratio. SD plasma is associated with fewer anaphylactic reactions than FFP plasma/RBC units ratio.

Fatal Acute Hemolytic Transfusion Reaction due to Anti-Wra.

BACKGROUND: The Wra blood group antigen is a low-frequency antigen. Antibody screening sets used in pretransfusion laboratory investigations usually do not contain a Wr(a+) cell. If subsequent cross-matching is performed without indirect antiglobulin test (IAT), Wra antibodies reacting with donor red blood cells (RBCs) will be missed. For reasonable economic and time-saving arguments the risk of missing the detection of a potential clinically relevant antibody is worldwide accepted. CASE REPORT: A 66-year-old women with a negative antibody screen rapidly deteriorated after she received two units of RBCs for symptomatic anemia after hip surgery. Diagnosis of a transfusion reaction was obscured by pre-existing and nonspecific symptoms. Laboratory investigation indicated acute hemolysis. Cross-matching in IAT was positive for the first unit, and an extended antibody identification panel showed reactivity with Wr(a+) cells. The patient did not respond to supportive therapy and died within 48 h after the start of transfusion. CONCLUSION: This dramatic case provides further evidence on the clinical relevance of Wra blood group antibodies. In addition, it underlines the clinical importance of risk awareness in the blood transfusion chain and the possible complexity in relation to patient monitoring in daily transfusion practice.

Age of platelet concentrates and time to the next transfusion.

BACKGROUND: Storage time of platelet (PLT) concentrates has been negatively associated with clinical efficacy outcomes. The aim of this study was to quantify the association between storage time of PLT concentrates and interval to the next PLT transfusion for different types of PLT components, stored for up to 7 days and transfused to transfusion-dependent hematooncology patients with thrombocytopenia. STUDY DESIGN AND METHODS: From a cohort of patients from 10 major Dutch hospitals, patients were selected whose transfusion patterns were compatible with PLT transfusion dependency due to hematooncologic disease. Mean time to the next transfusion and mean differences in time to the next transfusion for different storage time categories (i.e., fresh, <4 days; intermediate, 4-5 days; and old, >5 days) were estimated, per component type, using multilevel mixed-effects linear models. RESULTS: Among a cohort of 29,761 patients who received 140,896 PLT transfusions we selected 4441 hematooncology patients who had received 12,724 PLT transfusions during periods of PLT transfusion dependency. Transfusion of fresh, compared to old, buffy coat-derived PLTs in plasma was associated with a delay to the next transfusion of 6.2 hours (95% confidence interval [CI], 4.5-8.0 hr). For buffy coat-derived PLTs in PAS-B and -C this difference was 7.7 hours (95% CI, 2.2-13.3 hr) and 3.9 hours (95% CI, -2.1 to 9.9 hr) while for apheresis PLTs in plasma it was only 1.8 hours (95% CI, -3.5 to 7.1 hr). CONCLUSION: Our results indicate that the time to the next transfusion shortens with increasing age of transfused buffy coat-derived PLT concentrates. This association was not observed for apheresis PLTs.

Validation of the Feverkidstool and procalcitonin for detecting serious bacterial infections in febrile children.

BackgroundTo validate the Feverkidstool, a prediction model consisting of clinical signs and symptoms and C-reactive protein (CRP) to identify serious bacterial infections (SBIs) in febrile children, and to determine the incremental diagnostic value of procalcitonin.MethodsThis prospective observational study that was carried out at two Dutch emergency departments included children with fever, aged 1 month to 16 years. The prediction models were developed with polytomous logistic regression differentiating "pneumonia" and "other SBIs" from "non-SBIs" using standardized, routinely collected data on clinical signs and symptoms, CRP, and procalcitonin.ResultsA total of 1,085 children were included with a median age of 1.6 years (interquartile range 0.8-3.4); 73 children (7%) had pneumonia and 98 children (9%) had other SBIs. The Feverkidstool showed good discriminative ability in this new population. After adding procalcitonin to the Feverkidstool, c-statistic for "pneumonia" increased from 0.85 (95% confidence interval (CI) 0.76-0.94) to 0.86 (0.77-0.94) and for "other SBI" from 0.81 (0.73-0.90) to 0.83 (0.75- 0.91). A model with clinical features and procalcitonin performed similar to the Feverkidstool.ConclusionThis study confirms the external validity of the Feverkidstool, with CRP and procalcitonin being equally valuable for predicting SBI in our population of febrile children. Our findings do not support routine dual use of CRP and procalcitonin.

Storage time of platelet concentrates and all-cause bacteremia in hematologic patients.

BACKGROUND: Extension of storage time of platelet (PLT) concentrates may result in an increased risk of bacteremia, directly via transfusion of contaminated products or indirectly via transfusion-related immunomodulation. We aimed to quantify the association of storage time of PLT concentrates and all-cause bacteremia in hematologic patients. STUDY DESIGN AND METHODS: We established a cohort of hematologic patients who received a PLT transfusion between 2005 and 2015. Cases were defined as patients with a bacteremia the day after transfusion and matched to as many controls as possible. A conditional logistic regression was performed, stratified by storage medium. RESULTS: Among 3514 patients receiving 36,032 PLT concentrates stored in plasma, 613 cases of bacteremia were found. The relative risk of all-cause bacteremia the day after transfusion was 0.80 (95% confidence interval [CI], 0.58-1.12) for PLT concentrates stored 3 to 4 days and 0.67 (95% CI, 0.49-0.92) for at least 5 days, compared to no more than 2 days. Among 1527 patients receiving 11,822 PLT concentrates stored in PLT additive solution, 182 cases of bacteremia were found. The relative risk of all-cause bacteremia was 1.14 (95% CI, 0.70-1.84) for PLT concentrates stored for 3 to 4 days and 1.19 (95% CI, 0.70-2.01) for at least 5 days, compared to not more than 2 days. CONCLUSION: Storage time of PLT concentrates was not associated with increased occurrence of all-cause bacteremia the day after transfusion. If anything, fewer cases of bacteremia occurred with increasing storage time of PLT concentrates in plasma. These bacteremias are not directly caused by transfusion of a contaminated product and the underlying mechanism warrants further research.

C-reactive protein, procalcitonin and the lab-score for detecting serious bacterial infections in febrile children at the emergency department: a prospective observational study.

BACKGROUND: C-reactive protein (CRP) and procalcitonin (PCT) are useful diagnostic tools to estimate the risk of serious bacterial infection (SBI) in febrile children at the emergency department (ED). The Lab-score combines these 2 biomarkers with urinalysis in an easy to use validated model. Kinetics of inflammatory markers suggests a differentiating role of duration of disease. AIM: : Appraisal of the diagnostic role of CRP and PCT in febrile children at risk of SBI, determining the differentiating value of duration of fever, and validating and updating the Lab-score. METHODS: In this prospective observational study previously healthy children with fever, 1 month to 16 years of age, attending the EDs of a university hospital and a teaching hospital (Rotterdam, the Netherlands) between 2009 and 2012 were included. Standardized information on clinical signs and symptoms, CRP, PCT and urinalysis were collected prospectively. Logistic multivariable regression analysis was used to assess diagnostic performance. The original Lab-score included CRP, PCT and urinalysis and the total score ranged 0-9 points. RESULTS: One thousand eighty-four children were included, median age was 1.6 years (interquartile range: 0.8-3.5), 170 children (16%) had SBI. CRP [receiver operating characteristic (ROC)-area 0.77 (95% confidence interval [CI]: 0.69-0.85)] and PCT [ROC-area 0.75 (95% CI: 0.67-0.83)] were both strong predictors of SBI. Duration of fever had no added diagnostic value to CRP and PCT. The Lab-score performed well [ROC area 0.79 (95% CI: 0.72-0.87)], but threshold values performed similar to often used cutoffs of single biomarkers. An updated Lab-score improved only moderately [ROC area 0.83 (95% CI: 0.76-0.90)]. PCT did not alter post-test probabilities for SBI substantially in patients with low (<20 mg/L) or elevated CRP (≥ 100 mg/L) levels (67% of population). CONCLUSION: CRP and PCT were both strong predictors of SBI. The original and updated Lab-score performed well, but thresholds values lacked diagnostic value for ruling out SBI. Depending on clinical risk thresholds, diagnostic testing can be limited to CRP or PCT, rather than both, in many febrile children.

Wnt3a deficiency irreversibly impairs hematopoietic stem cell self-renewal and leads to defects in progenitor cell differentiation.

Canonical Wnt signaling has been implicated in various aspects of hematopoiesis. Its role is controversial due to different outcomes between various inducible Wnt-signaling loss-of-function models and also compared with gain-of-function systems. We therefore studied a mouse deficient for a Wnt gene that seemed to play a nonredundant role in hematopoiesis. Mice lacking Wnt3a die prenatally around embryonic day (E) 12.5, allowing fetal hematopoiesis to be studied using in vitro assays and transplantation into irradiated recipient mice. Here we show that Wnt3a deficiency leads to a reduction in the numbers of hematopoietic stem cells (HSCs) and progenitor cells in the fetal liver (FL) and to severely reduced reconstitution capacity as measured in secondary transplantation assays. This deficiency is irreversible and cannot be restored by transplantation into Wnt3a competent mice. The impaired long-term repopulation capacity of Wnt3a(-/-) HSCs could not be explained by altered cell cycle or survival of primitive progenitors. Moreover, Wnt3a deficiency affected myeloid but not B-lymphoid development at the progenitor level, and affected immature thymocyte differentiation. Our results show that Wnt3a signaling not only provides proliferative stimuli, such as for immature thymocytes, but also regulates cell fate decisions of HSC during hematopoiesis.

Gene therapy: is IL2RG oncogenic in T-cell development?

The gene IL2RG encodes the gamma-chain of the interleukin-2 receptor and is mutated in patients with X-linked severe combined immune deficiency (X-SCID). Woods et al. report the development of thymus tumours in a mouse model of X-SCID after correction by lentiviral overexpression of IL2RG and claim that these were caused by IL2RG itself. Here we find that retroviral overexpression of IL2RG in human CD34+ cells has no effect on T-cell development, whereas overexpression of the T-cell acute lymphoblastic leukaemia (T-ALL) oncogene LMO2 leads to severe abnormalities. Retroviral expression of IL2RG may therefore not be directly oncogenic--rather, the restoration of normal signalling by the interleukin-7 receptor to X-SCID precursor cells allows progression of T-cell development to stages that are permissive for the pro-leukaemic effects of ectopic LMO2.

Wnt signaling in the thymus is regulated by differential expression of intracellular signaling molecules.

Wnt signaling is essential for T cell development in the thymus, but the stages in which it occurs and the molecular mechanisms underlying Wnt responsiveness have remained elusive. Here we examined Wnt signaling activity in both human and murine thymocyte populations by determining beta-catenin levels, Tcf-reporter activation and expression of Wnt-target genes. We demonstrate that Wnt signaling occurs in all thymocyte subsets, including the more mature populations, but most prominently in the double negative (DN) subsets. This differential sensitivity to Wnt signaling was not caused by differences in the presence of Wnts or Wnt receptors, as these appeared to be expressed at comparable levels in all thymocyte subsets. Rather, it can be explained by high expression of activating signaling molecules in DN cells, e.g., beta-catenin, plakoglobin, and long forms of Tcf-1, and by low levels of inhibitory molecules. By blocking Wnt signaling from the earliest stage onwards using overexpression of Dickkopf, we show that inhibition of the canonical Wnt pathway blocks development at the most immature DN1 stage. Thus, responsiveness to developmental signals can be regulated by differential expression of intracellular mediators rather than by abundance of receptors or ligands.

T-sing progenitors to commit.

T-cell development in the thymus is a complex and highly regulated process. During the process of differentiation from multipotent progenitor cells to mature T cells, proliferation, restriction of lineage potential, TCR gene rearrangements and selection events occur, all accompanied by changes in gene expression. A comprehensive understanding of thymocyte differentiation remains to be established. Two related, key issues have received much attention recently: the nature of the thymus seeding cell and the regulation of T-cell lineage commitment. Here we review the perspectives of different researchers working both on murine and human T-cell development and argue that a true T-cell commitment factor might not be required because of the unique properties of the thymus.

Effects of Delta1 and Jagged1 on early human hematopoiesis: correlation with expression of notch signaling-related genes in CD34+ cells.

It has been shown that Notch signaling mediated by ligands of both Jagged and Delta families expands the hematopoietic stem cell compartment while blocking or delaying terminal myeloid differentiation. Here we show that Delta1- and Jagged1-expressing stromal cells have distinct effects on the clonogenic and differentiation capacities of human CD34(+) CD38(+) cells. Jagged1 increases the number of bipotent colony-forming unit-granulocyte macrophage (CFU-GM) and unipotent progenitors (CFU-granulocytes and CFU-macrophages), without quantitatively affecting terminal cell differentiation, whereas Delta1 reduces the number of CFU-GM and differentiated monocytic cells. Expression analysis of genes coding for Notch receptors, Notch targets, and Notch signaling modulators in supernatant CD34(+) cells arising upon contact with Jagged1 and Delta1 shows dynamic and differential gene expression profiles over time. At early time points, modest upregulation of Notch1, Notch3, and Hes1 was observed in Jagged1-CD34(+) cells, whereas those in contact with Delta1 strikingly upregulated Notch3 and Hes1. Later, myeloid progenitors with strong clonogenic potential emerging upon contact with Jagged1 upregulated Notch1 and Deltex and downregulated Notch signaling modulators, whereas T/NK progenitors originated by Delta1 strikingly upregulated Notch3 and Deltex and, to a lesser extent, Hes1, Lunatic Fringe, and Numb. Together, the data unravel previously unrecognized expression patterns of Notch signaling-related genes in CD34(+) CD38(+) cells as they develop in Jagged1- or Delta1-stromal cell environments, which appear to reflect sequential maturational stages of CD34(+) cells into distinct cell lineages.