Nucleated red blood cell distribution in critically ill patients with acute pancreatitis: a retrospective cohort study.

OBJECTIVES: This study examined the potential association between nucleated red blood cell (NRBC) levels and mortality in critically ill patients with acute pancreatitis (AP) in the intensive care unit, due to limited existing research on this correlation. METHODS: This retrospective cohort study utilized data from the MIMIC-IV v2.0 and MIMIC-III v1.4 databases to investigate the potential relationship between NRBC levels and patient outcomes. The study employed restricted cubic splines (RCS) regression analysis to explore non-linear associations. The impact of NRBC on prognosis was assessed using a generalized linear model (GLM) with a logit link, adjusted for potential confounders. Furthermore, four machine learning models, including Gradient Boosting Classifier (GBC), Random Forest, Gaussian Naive Bayes, and Decision Tree Classifier model, were constructed using NRBC data to generate risk scores and evaluate the potential of NRBC in predicting patient prognosis. RESULTS: A total of 354 patients were enrolled in the study, with 162 (45.8%) individuals aged 60 years or older and 204 (57.6%) males. RCS regression analysis demonstrated a non-linear relationship between NRBC levels and 90-day mortality. Receiver Operating Characteristic (ROC) analysis identified a 1.7% NRBC cutoff to distinguish survivor from non-survivor patients for 90-day mortality, yielding an Area Under the Curve (AUC) of 0.599, with a sensitivity of 0.475 and specificity of 0.711. Elevated NRBC levels were associated with increased risks of 90-day mortality in both unadjusted and adjusted models (all Odds Ratios > 1, P < 0.05). Assessment of various machine learning models with nine variables, including NRBC, Sex, Age, Simplified Acute Physiology Score II, Acute Physiology Score III, Congestive Heart Failure, Vasopressin, Norepinephrine, and Mean Arterial Pressure, indicated that the GBC model displayed the highest predictive accuracy for 90-day mortality, with an AUC of 0.982 (95% CI 0.970-0.994). Post hoc power analysis showed a statistical power of 0.880 in the study. CONCLUSIONS: Elevated levels of NRBC are linked to an increased mortality risk in critically ill patients with AP, suggesting its potential for predicting mortality.

Nucleated red blood cells as a biomarker for mortality in infants and neonates requiring veno-arterial extracorporeal membrane oxygenation for cardiac disease.

INTRODUCTION: Nucleated red blood cells (NRBC) are rare in the peripheral circulation of healthy individuals and their presence have been associated with mortality in adults and very low birth weight newborns, however, its value as a biomarker for mortality in infants requiring veno-arterial (VA) extracorporeal membrane oxygenation (ECMO) has yet to be studied. We sought to determine if NRBC can serve as a biomarker for ECMO mortality and inpatient mortality in infants requiring V-A ECMO. METHODS: A single-center retrospective chart review analyzing infants <1 year of age requiring VA ECMO due to myocardial dysfunction or post-cardiotomy between January 1, 2011 to June 30, 2020. RESULTS: One hundred two patients required VA ECMO. Sixty-five patients required ECMO post-cardiotomy, 19 for perioperative deterioration, and 18 for myocardial dysfunction. Fifty-one patients (50%) died (21 died on ECMO, 30 died post-ECMO decannulation). Multivariable analysis found Age <60 days (OR 13.0, 95% CI 1.9-89.6, p = 0.009), NRBC increase by >50% post-ECMO decannulation (OR 17.1, 95% CI 3.1-95.1, p = 0.001), Single Ventricle (OR 9.0, 95% CI 1.7-47.7, p = 0.01), and lactate at ECMO decannulation (OR 3.0, 95% CI 1.3-7.1, p = 0.011) to be independently associated with inpatient mortality. ROC curves evaluating NRBC pre-ECMO decannulation as a biomarker for mortality on ECMO (AUC 0.80, 95% CI 0.68-0.92, p ⩽ 0.001) and post-ECMO decannulation (AUC 0.75, 95% CI 0.65-0.84, p ⩽ 0.001) show NRBC to be an accurate biomarker for mortality. CONCLUSIONS: Greater than 50% increase in NRBC post-ECMO decannulation is associated with inpatient mortality. NRBC value pre-ECMO decannulation may be a useful biomarker for mortality while on ECMO and post-decannulation.

Nucleated red blood cells as a biomarker for mortality in neonates following cardiac surgery.

INTRODUCTION: Nucleated red blood cells (NRBCs) are immature red cells that under normal conditions are not present in the peripheral circulation. Several studies have suggested an association between elevated NRBC and poor outcome in critically ill adults and neonates. We sought to determine if elevations in NRBC value following cardiac surgery and following clinical events during the hospital stay can be used as a biomarker to monitor for mortality risk in neonates post-cardiac surgery. MATERIALS AND METHODS: We constructed a retrospective study of 264 neonates who underwent cardiac surgery at Children's Hospital, New Orleans between 2011 and 2020. Variables included mortality and NRBC value were recorded following cardiac surgery and following peri-operative clinical events. The study was approved by LSU Health IRB. Sensitivity, specificity, receiver operating characteristic (ROC) curves with area under the curve (AUC) and logistic regression analysis were performed. RESULTS: Thirty-six patients (13.6%) died, of which 32 had an NRBC value ≥10/100 white blood cell (WBC) during hospitalisation. Multi-variable analysis found extracorporeal membrane oxygenation use (OR 10, 95% CI 2.9-33, p=<0.001), NRBC ≥10/100 WBC (OR 16.1, CI 4.1-62.5, p ≤ 0.001) and peak NRBC in the 14-day period post-cardiac surgery (continuous variable, OR 1.05, 95% CI 1.0-1.09, p = 0.03), to be independently associated with mortality. Using a cut-off NRBC value of 10/100 WBC, there was an 88.9% sensitivity and a 90.8% specificity, with ROC curve showing an AUC of 0.9 and 0.914 for peak NRBC value in 14 days post-surgery and entire hospitalisation, respectively. CONCLUSIONS: NRBC ≥10/100 WBC post-cardiac surgery is strongly associated with mortality. Additionally, NRBC trend appears to show promise as an accurate biomarker for mortality.

Isolation and characterization of fetal nucleated red blood cells from maternal blood as a target for single cell sequencing-based non-invasive genetic testing.

PURPOSE: Although non-invasive prenatal testing (NIPT) based on cell-free DNA (cfDNA) in maternal plasma has been prevailing worldwide, low levels of fetal DNA fraction may lead to false-negative results. Since fetal cells in maternal blood provide a pure source of fetal genomic DNA, we aimed to establish a workflow to isolate and sequence fetal nucleated red blood cells (fNRBCs) individually as a target for NIPT. METHODS: Using male-bearing pregnancy cases, we isolated fNRBCs individually from maternal blood by FACS, and obtained their genomic sequence data through PCR screening with a Y-chromosome marker and whole-genome amplification (WGA)-based whole-genome sequencing. RESULTS: The PCR and WGA efficiencies of fNRBC candidates were consistently lower than those of control cells. Sequencing data analyses revealed that although the majority of the fNRBC candidates were confirmed to be of fetal origin, many of the WGA-based genomic libraries from fNRBCs were considered to have been amplified from a portion of genomic DNA. CONCLUSIONS: We established a workflow to isolate and sequence fNRBCs individually. However, our results demonstrated that, to make cell-based NIPT targeting fNRBCs feasible, cell isolation procedures need to be further refined such that the nuclei of fNRBCs are kept intact.

High-throughput isolation of fetal nucleated red blood cells by multifunctional microsphere-assisted inertial microfluidics.

Being easy, safe and reliable, non-invasive prenatal diagnosis (NIPD) has been greatly pursued in recent years. Holding the complete genetic information of the fetus, fetal nucleated red blood cells (fNRBCs) are viewed as a suitable target for NIPD application. However, effective separating fNRBCs from maternal peripheral blood for clinic use still faces great challenges, given that fNRBCs are extremely rare in maternal blood circulation. Here, by combining the high-throughput inertial microfluidic chip with multifunctional microspheres as size amplification, we develop a novel method to isolate fNRBCs with high performance. To enlarge the size difference between fNRBCs and normal blood cells, we use the gelatin coated microspheres to capture fNRBCs with anti-CD147 as specific recognizer at first. The size difference between fNRBCs captured by the microspheres and normal blood cells makes it easy to purify the captured fNRBCs through the spiral microfluidic chip. Finally, the purified fNRBCs are mildly released from the microspheres by enzymatically degrading the gelatin coating. Cell capture efficiency about 81%, high purity of 83%, as well as cell release viability over 80% were achieved using spiked samples by this approach. Additionally, fNRBCs were successfully detected from peripheral blood of pregnant women with an average of 24 fNRBCs per mL, suggesting the great potential of this method for clinical non-invasive prenatal diagnosis.

Unique monoclonal antibodies specifically bind surface structures on human fetal erythroid blood cells.

BACKGROUND: Continuing efforts in development of non-invasive prenatal genetic tests have focused on the isolation of fetal nucleated red blood cells (NRBCs) from maternal blood for decades. Because no fetal cell-specific antibody has been described so far, the present study focused on the development of monoclonal antibodies (mAbs) to antigens that are expressed exclusively on fetal NRBCs. METHODS: Mice were immunized with fetal erythroid cell membranes and hybridomas screened for Abs using a multi-parameter fluorescence-activated cell sorting (FACS). Selected mAbs were evaluated by comparative FACS analysis involving Abs known to bind erythroid cell surface markers (CD71, CD36, CD34), antigen-i, galactose, or glycophorin-A (GPA). Specificity was further confirmed by extensive immunohistological and immunocytological analyses of NRBCs from umbilical cord blood and fetal and adult cells from liver, bone marrow, peripheral blood, and lymphoid tissues. RESULTS: Screening of 690 hybridomas yielded three clones of which Abs from 4B8 and 4B9 clones demonstrated the desired specificity for a novel antigenic structure expressed on fetal erythroblast cell membranes. The antigenic structure identified is different from known surface markers (CD36, CD71, GPA, antigen-i, and galactose), and is not present on circulating adult erythroid cells, except for occasional detectability in adult bone marrow cells. CONCLUSIONS: The new mAbs specifically bind the same or highly overlapping epitopes of a surface antigen that is almost exclusively expressed on fetal erythroid cells. The high specificity of the mAbs should facilitate development of simple methods for reliable isolation of fetal NRBCs and their use in non-invasive prenatal diagnosis of fetal genetic status.

Placental pathology in full-term infants with hypoxic-ischemic neonatal encephalopathy and association with magnetic resonance imaging pattern of brain injury.

OBJECTIVE: To investigate the relationship between placental pathology and pattern of brain injury in full-term infants with neonatal encephalopathy after a presumed hypoxic-ischemic insult. STUDY DESIGN: The study group comprised full-term infants with neonatal encephalopathy subsequent to presumed hypoxia-ischemia with available placenta for analysis who underwent cerebral magnetic resonance imaging (MRI) within the first 15 days after birth. Macroscopic and microscopic characteristics of the placenta were assessed. The infants were classified according to the predominant pattern of brain injury detected on MRI: no injury, predominant white matter/watershed injury, predominant basal ganglia and thalami (BGT) injury, or white matter/watershed injury with BGT involvement. Maternal and perinatal clinical factors were recorded. RESULTS: Placental tissue was available for analysis in 95 of 171 infants evaluated (56%). Among these 95 infants, 34 had no cerebral abnormalities on MRI, 27 had white matter/watershed injury, 18 had BGT injury, and 16 had white matter/watershed injury with BGT involvement. Chorioamnionitis was a common placental finding in both the infants without injury (59%) and those with white matter/BGT injury (56%). On multinomial logistic regression analysis, white matter/watershed injury with and without BGT involvement was associated with decreased placental maturation. Hypoglycemia was associated with an increased risk of the white matter/BGT injury pattern (OR,5.4; 95% CI, 1.4-21.4). The BGT injury pattern was associated with chronic villitis (OR, 12.7; 95% CI, 2.4-68.7). A placental weight <10th percentile appeared to be protective against brain injury, especially for the BGT pattern (OR, 0.1; 95% CI, 0.01-0.7). CONCLUSION: Placental weight <10th percentile was mainly associated with normal cerebral MRI findings. Decreased placental maturation and hypoglycemia <2.0 mmol/L were associated with increased risk of white matter/watershed injury with or without BGT involvement. Chronic villitis was associated with BGT injury irrespective of white matter injury.

Nucleated red blood cells as a novel biomarker in the diagnosis and prediction of sepsis severity in children.

Introduction: Sepsis is a vitally serious disease leading to high mortality. Nucleated red blood cells (NRBCs) are present in some noninfectious diseases, but the relationship between NRBCs and sepsis in children remains unknown. The purpose of this study was to compare the clinical characteristics and outcomes of sepsis with positive NRBCs and negative NRBCs in children, and to further explore whether the count of NRBCs has a relationship with the severity of sepsis. Methods: We enrolled children with sepsis who were admitted to the Children's Hospital of Chongqing Medical University between January 2020 and December 2022. The children's clinical data, laboratory data and outcomes were recorded and analyzed. Results: One hundred and fifteen children met the inclusion criteria in our study. Compared to negative NRBCs patients, the C-reactive protein, alanine transaminase, urea nitrogen values, mortality rate and length of hospitalization were found to be significantly increased, while platelet counts, and hemoglobin were significantly decreased in sepsis patients with positive NRBC (P < 0.05). Receiver operating characteristic (ROC) curves analysis showed that the optimal cutoff value of the NRBC count in the diagnosis of severe sepsis was 3, with a sensitivity of 87.5% and specificity of 94.9%. The area under the ROC curve was 0.877 (95% CI: 0.798-0.957). Discussion: These findings demonstrated that NRBC count has the potential to be a biomarker for the diagnosis of sepsis in children, especially an NRBC count greater than 3, which may predict the severity and poor prognosis in children suffering from sepsis.

Frequency of Nucleated Red Blood Cells in the Peripheral Blood of ICU-Admitted Patients.

Background Nucleated red blood cells (NRBCs) are not normally found in the peripheral blood of normal healthy individuals. The presence of NRBCs on an adult peripheral blood smear indicates that there is an extremely high demand for the bone marrow to manufacture RBCs and that immature red blood cells are being released into the bloodstream. Anemia, myelofibrosis, thalassemia, miliary tuberculosis, malignancies of the bone marrow (myelomas, leukemias, lymphomas), and prolonged hypoxemia are a few possible pathogenic reasons. Critically ill patients who have NRBCs have a high mortality rate and a worse prognosis. OBJECTIVE: To evaluate the clinical significance of NRBCs in the peripheral blood of critically ill patients admitted to the ICU to find a cut-off to predict mortality. MATERIALS AND METHODS: A cross-sectional study was carried out over a period of six months September 1, 2020, to March 31, 2021, in Lahore, Pakistan. A total of 800 critically ill patients of both sexes in the age group of 18-70 years were included. Patients younger than 18 years and patients who underwent surgery were excluded. A quantity of 3 ml of whole blood sample in an ethylenediamine tetraacetic acid (EDTA) vial from each patient was run on SYSMEX XN-9000 (Sysmex Corporation, Kobe, Hyogo, Japan) and the results were reviewed on peripheral smears. RESULTS: The incidence of NRBCs in ICU-admitted patients was 62.5% (500/800). The total number of NRBC-positive patients recovering after the treatment was 364 (72.8%). The overall mortality of NRBC-positive patients was 30% (150/500). It was significantly higher (p<0.001) than that of NRBC-negative patients (14%; 44/300). During treatment, the highest mortality rate was seen in patients due to malignancy (100%), followed by sepsis (58.8%). It was observed that the disease pattern and number of NRBCs were significantly different (p<0.001) among all disease groups. However, there was no statistically significant difference in NRBCs on the basis of gender (p >0.05). In our study, a cutoff of NRBCs of 2.50 showed a high risk of mortality with a sensitivity of 91%. CONCLUSION: The presence of NRBCs may predict mortality in critically ill ICU-admitted patients. Their presence in the blood may be regarded as a marker of severity suggesting a high risk of ICU death.

Nucleated red blood cell: a feasible quality parameter of cord blood units.

INTRODUCTION: Umbilical cord blood is an alternative source of hematopoietic progenitor cells for bone marrow transplantation; however, it is associated with a higher graft failure rate. The presence of a high rate of nucleated red blood cells (NRBCs) seems to be related to a greater capacity for engraftment, although is also associated with fetal distress conditions. We analyzed the correlation of the NRBC with quality parameters and its association with the utilization score of a cord blood unit. STUDY DESIGN AND METHOD: Data of 3346 units collected in a public cord blood bank from May 2010 to December 2017 were analyzed, retrospectively, to identify factors associated with an increased number of nucleated red blood cells and its correlation with the engraftment capacity measured through total nucleated cells (TNCs) and CD34 positive cells. We also evaluated the utilization score of these units and identified an NRBC cutoff associated with a higher score. RESULTS: The median volume collected was 104 mL (42-255), the pre-processing TNC count was 144.77 × 107 (95.46-477.18), the post-processing TNC count was 119.44 × 107 (42.7-477.18), the CD34 count was 4.67 × 106 (0.31-48.01), the NRBC count was 5 (0-202) and the utilization score was 0.0228 (0.00143-0.9740). The NRBC showed a correlation with the collected volume, TNC and CD34 positive cells and a higher utilization score and the receiver operating characteristic (ROC) curve analysis identified the five NRBC/100 leukocytes cutoff that correlates better with the probability of use. No association with pathological conditions and the NRBC rate was observed. CONCLUSIONS: The NRBC is a feasible parameter for the screening of the cord blood unit (CBU) and the minimum cutoff of five NRBC/100 leukocytes can be a strategy in conjunction with the TNC to identify better units for cord blood bank sustainability.

Nucleated Red Blood Cell Counts of Neonates Born Emergently 1-4 h after a Maternal Cardiac Arrest.

A high nucleated red blood cell (NRBC) count in a newborn infant at birth is sometimes used to imply that fetal hypoxia occurred. However, it is debated whether many hours are required between fetal hypoxia and the appearance of high NRBC or alternatively, whether this can occur very quickly, with fetal hypoxia within minutes to a few hours before birth. We sought relevant information from four unfortunate cases, where during a previously healthy pregnancy, the mother had a sudden cardiac arrest, with cardiopulmonary resuscitation begun at the incident scene and continued through emergent cesarean section delivery.

Epigenome-wide association study of bronchopulmonary dysplasia in preterm infants: results from the discovery-BPD program.

BACKGROUND: Bronchopulmonary dysplasia (BPD) is a lung disease in premature infants caused by therapeutic oxygen supplemental and characterized by impaired pulmonary development which persists into later life. While advances in neonatal care have improved survival rates of premature infants, cases of BPD have been increasing with limited therapeutic options for prevention and treatment. This study was designed to explore the relationship between gestational age (GA), birth weight, and estimated blood cell-type composition in premature infants and to elucidate early epigenetic biomarkers associated with BPD. METHODS: Cord blood DNA from preterm neonates that went on to develop BPD (n = 14) or not (non-BPD, n = 93) was applied to Illumina 450 K methylation arrays. Blood cell-type compositions were estimated using DNA methylation profiles. Multivariable robust regression analysis elucidated CpGs associated with BPD risk. cDNA microarray analysis of cord blood RNA identified differentially expressed genes in neonates who later developed BPD. RESULTS: The development of BPD and the need for oxygen supplementation were strongly associated with GA (BPD, p < 1.0E-04; O2 supplementation, p < 1.0E-09) and birth weight (BPD, p < 1.0E-02; O2 supplementation, p < 1.0E-07). The estimated nucleated red blood cell (NRBC) percent was negatively associated with birth weight and GA, positively associated with hypomethylation of the tobacco smoke exposure biomarker cg05575921, and high-NRBC blood samples displayed a hypomethylation profile. Epigenome-wide association study (EWAS) identified 38 (Bonferroni) and 275 (false discovery rate 1%) differentially methylated CpGs associated with BPD. BPD-associated CpGs in cord blood were enriched for lung maturation and hematopoiesis pathways. Stochastic epigenetic mutation burden at birth was significantly elevated among those who developed BPD (adjusted p = 0.02). Transcriptome changes in cord blood cells reflected cell cycle, development, and pulmonary disorder events in BPD. CONCLUSIONS: While results must be interpreted with caution because of the small size of this study, NRBC content strongly impacted DNA methylation profiles in preterm cord blood and EWAS analysis revealed potential insights into biological pathways involved in BPD pathogenesis.

Comparison of transcriptome profiles of nucleated red blood cells in cord blood between preterm and full-term neonates.

BACKGROUND: The reactivation of fetal γ-globin expression is an effective strategy for ameliorating the clinical symptoms of ß-hemoglobinopathies. However, the mechanism of globin switching, especially the roles of long non-coding RNAs (lncRNAs) in this process, remains elusive. METHODS: We compared the in vivo transcriptome profiles of nucleated red blood cells (NRBCs) isolated from the umbilical cord blood of preterm and full-term newborns. We collected 75 umbilical cord blood samples and performed qPCR of the candidate genes. RESULTS: In this study, we identified 7,166 differentially expressed protein-coding genes, 3,243 differentially expressed lncRNAs, and 79 differentially expressed microRNAs. Our data show that the Fanconi anemia pathway and the H19/let-7/LIN28B axis may be involved in γ- to ß-globin gene switching. Moreover, we constructed the hub gene network of the differentially expressed transcription factors. Based on qPCR, we found that BCL11A was differentially expressed based on biological sex. We also confirmed that H19 is differentially expressed and established the H19-related network to reveal the potential regulatory mechanisms. CONCLUSION: We present the profiles of the in vivo transcriptome differences of NRBCs between preterm and full-term neonates for the first time, and provide novel research targets for ß-hemoglobinopathies.

Improving the efficiency of the autoverification workflow for nucleated red blood cell reporting in the hematology laboratory.

OBJECTIVE: Although the microscopic manual count is considered the standard method for NRBC enumeration, modern hematology analyzers can perform this task automatically with reliable accuracy and efficiency. This study aims to evaluate the diagnostic performance of the Sysmex XN hematology analyzer and to construct the optimal workflow for accurate and efficient NRBC reporting. METHODS: Specimens containing different levels of NRBC were included. Analytical performance was evaluated via method comparison with flow cytometry (FCM) and manual count (MC). Clinical sensitivity was analyzed by ROC analysis using manual count as the standard method. RESULTS: Correlation study of %NRBC with FCM and MC demonstrated an r value of 0.925 (95% CI 0.905 to 0.942) and 0.990 (95% CI 0.987 to 0.992) with a mean difference of -0.8 (95%CI: -6.7 to +5.0) and +0.50 (95% CI: -6.7 to +7.7), respectively. When the automated NRBC count was equal to zero and >0.07 × 109 /L, the false-negative rate and false-positive rate were 100%, respectively; hence, manual slide review could be omitted. A false-positive rate of 72.7% was noted in specimens containing NRBC count less than 0.07 × 109 /L. CONCLUSION: The Sysmex XN can help improve the efficiency of NRBC enumeration owing to its accuracy, rapidity, and automation. However, further studies are required to improve the accuracy of detection in specimens containing a very low level of NRBC.

Nucleated red blood cells participate in myocardial regeneration in the toad Bufo Gargarizan Gargarizan.

Heart regeneration is negligible in humans and mammals but remarkable in some ectotherms. Humans and mammals lack nucleated red blood cells (NRBCs), while ectotherms have sufficient NRBCs. This study used Bufo gargarizan gargarizan, a Chinese toad subspecies, as a model animal to verify our hypothesis that NRBCs participate in myocardial regeneration. NRBC infiltration into myocardium was seen in the healthy toad hearts. Heart needle-injury was used as an enlarged model of physiological cardiomyocyte loss. It recovered quickly and scarlessly. NRBC infiltration increased during the recovery. Transwell assay was done to in vitro explore effects of myocardial injury on NRBCs. In the transwell system, NRBCs could infiltrate into cardiac pieces and could transdifferentiate toward cardiomyocytes. Heart apex cautery caused approximately 5% of the ventricle to be injured to varying degrees. In the mildly to moderately injured regions, NRBC infiltration increased and myocardial regeneration started soon after the inflammatory response; the severely damaged region underwent inflammation, scarring, and vascularity before NRBC infiltration and myocardial regeneration, and recovered scarlessly in four months. NRBCs were seen in the newly formed myocardium. Enzyme-linked immunosorbent assay and Western blotting showed that the levels of tumor necrosis factor-α, interleukin- 1ß, 6, and11, cardiotrophin-1, vascular endothelial growth factor, erythropoietin, matrix metalloproteinase- 2 and 9 in the serum and/or cardiac tissues fluctuated in different patterns during the cardiac injury-regeneration. Cardiotrophin-1 could induce toad NRBC transdifferentiation toward cardiomyocytes in vitro. Taken together, the results suggest that the NRBC is a cell source for cardiomyocyte renewal/regeneration in the toad; cardiomyocyte loss triggers a series of biological processes, facilitating NRBC infiltration and transition to cardiomyocytes. This finding may guide a new direction for improving human myocardial regeneration.

Prognostic impacts of peripheral blood erythroblasts after single-unit cord blood transplantation.

INTRODUCTION: The appearance of erythroblasts (EBLs) in peripheral blood occurs in a variety of serious conditions and has been associated with mortality in critically ill patients. However, the incidence, risk factor, and outcomes of EBLs after cord blood transplantation (CBT) remain unclear. METHODS: We have investigated the impact of EBLs on transplant outcomes on 225 adult patients who underwent single-unit CBT at our single institute. RESULTS: The cumulative incidences of EBL ≥200 × 106 /L and EBL ≥1000 × 106 /L at 60 days after CBT were 17% and 4%, respectively, detected after a median of 35 days and 36.5 days. Multivariate analysis using erythroblastosis as time-dependent covariates demonstrated the significant association of EBL ≥1000 × 106 /L, but not EBL ≥200 × 106 /L, with the development of grade III-IV acute graft-versus-host disease (GVHD, hazard ratio [HR]: 18.56; P < .001), higher nonrelapse mortality (HR: 13.38; P < .001), and overall mortality (HR: 4.97; P = .001). CONCLUSION: These data suggested that higher levels of EBLs were recognized as a significant risk factor for severe acute GVHD and mortality after single-unit CBT. Higher levels of EBLs may serve as a surrogate marker for poor single CBT outcomes.

Increased Nucleated RBCs in Cord Blood: Not an Exclusion Criterion but a Quality Indicator for Hematopoietic Progenitor Cell Transplantation.

Increased nucleated red blood cell (NRBC) counts have been reported to be associated with adverse fetal outcomes, and cord blood units (CBUs) with increased NRBC counts require a 2nd questionnaire to determine their suitability for transplantation. However, a recent study demonstrated a positive correlation of NRBCs with CD34+ cells and total nucleated cells (TNCs). We evaluated the association between the NRBC count and hematopoietic progenitor cell (HPC) content (TNC and CD34+ cell counts) in Korean full-term newborn CBUs. In addition, we assessed whether an increased NRBC count is associated with newborn health problems that impair CBU safety. Among the 32,876 units processed from May 2006 to December 2018, a total of 23,385 CBUs with a TNC count ≥ 7 × 108 and reliable perinatal information were analyzed to assess the association of the NRBC count with CBU parameters, and the newborns associated with 457 CBUs that required the 2nd questionnaire due to an increased NRBC (≥ 15 NRBCs/100 WBCs) were assessed at one year for health problems that threatened CBU safety. The majority of the CBUs that required the 2nd questionnaire due to an increased NRBC count (96.9%) were determined to be suitable for transplantation. Those with an increased NRBC count showed significantly higher CD34+ cell and TNC counts and a higher rate of transplantation (P < 0.001, < 0.001 and 0.025, respectively). NRBCs showed a significant positive correlation with TNCs and CD34+ cells and a significant negative correlation with birth weight (all P < 0.001; adjusted r = 0.185, 0.369 and - 0.029, respectively). In the multiple linear regression analysis, NRBCs showed independent and positive correlations with TNCs and CD34+ cells after adjustments for birth weight and gestational age (all P < 0.001; ß = 0.182, adjusted R2 = 0.053 and ß = 0.367, adjusted R2 = 0.418). An increased NRBC count in full-term normal delivery is a surrogate marker of HPCs in CBUs rather than an exclusion criterion for CBU safety. Moreover, providing the NRBC count together with the NRBC-corrected TNC count will be useful for clinicians to select CBUs for transplantation.

Association of nucleated red blood cell count with mortality among neonatal intensive care unit patients.

BACKGROUND: Nucleated red blood cells (nRBCs) are associated with adverse outcomes for pediatric and adult intensive care patients. METHODS: The association between nRBC count and mortality was examined in an observational cohort of patients admitted to the neonatal intensive care unit from December 2015-December 2018. RESULTS: Among the 1059 patients with at least one nRBC count obtained, 45 infants (4.2%) experienced in-hospital mortality prior to NICU discharge, the primary outcome measured in this study. Infants with any nRBC count >0 had a significantly higher risk of mortality (5.3% [45/849] vs. 0% [0/351], p < 0.001 by Fisher exact), and time to mortality decreased with higher nRBC counts (Spearman correlation -0.59, p < 0.001). The association between nRBC count and mortality remained significant even when restricting only to infants who were older than 7 days at time of nRBC count. CONCLUSION: Among neonatal intensive care unit patients, including those >7 days old, nRBCs are associated with significantly elevated mortality risk. A prospective study to better characterize clinical co-variants is necessary to better establish the use of nRBCs as a predictor of mortality.

Single-cell transcriptomic landscape of nucleated cells in umbilical cord blood.

BACKGROUND: For both pediatric and adult patients, umbilical cord blood (UCB) transplant is a therapeutic option for a variety of hematologic diseases, such as blood cancers, myeloproliferative disorders, genetic diseases, and metabolic disorders. However, the level of cellular heterogeneity and diversity of nucleated cells in UCB has not yet been assessed in an unbiased and systemic fashion. In the present study, nucleated cells from UCB were subjected to single-cell RNA sequencing to simultaneously profile the gene expression signatures of thousands of cells, generating a rich resource for further functional studies. Here, we report the transcriptomes of 17,637 UCB cells, covering 12 major cell types, many of which can be further divided into distinct subpopulations. RESULTS: Pseudotemporal ordering of nucleated red blood cells identifies wave-like activation and suppression of transcription regulators, leading to a polarized cellular state, which may reflect nucleated red blood cell maturation. Progenitor cells in UCB also comprise 2 subpopulations with activation of divergent transcription programs, leading to specific cell fate commitment. Detailed profiling of cytotoxic cell populations unveiled granzymes B and K signatures in natural killer and natural killer T-cell types in UCB. CONCLUSIONS: Taken together, our data form a comprehensive single-cell transcriptomic landscape that reveals previously unrecognized cell types, pathways, and mechanisms of gene expression regulation. These data may contribute to the efficacy and outcome of UCB transplant, broadening the scope of research and clinical innovations.

Noninvasive Prenatal Diagnostics: Recent Developments Using Circulating Fetal Nucleated Cells.

PURPOSE OF REVIEW: The purpose of this review is to highlight recent research advances in noninvasive prenatal diagnostic methods. RECENT FINDINGS: Recent studies developing noninvasive prenatal diagnostic (NIPD) methods have been focused on either fetal nucleated red blood cells (fNRBCs) or circulating trophoblasts (cTBs). Enriched cTBs were successfully utilized for whole genome profiling and short tandem repeat (STR) identification to confirm feto-maternal relationship. However, further analysis of isolated fNRBCs remains confined to examining fetal cytogenetics. SUMMARY: Invasive prenatal diagnostic procedures, amniocentesis and chorionic villus sampling, are the gold standard for the diagnosis of fetal chromosomal abnormalities and genetic disorders. Meanwhile, noninvasive techniques of analyzing circulating cell-free fetal DNA (cffDNA) have been limited to screening tools and are highly fragmented and confounded by maternal DNA. By detecting circulating fetal nucleated cells (CFNCs) we are able to noninvasively confirm fetal chromosomal abnormalities, truly realizing the concept of "noninvasive prenatal diagnostics". The primary technical challenge is the enrichment of the low abundance of CFNCs in maternal peripheral blood. For any cell-based NIPD method, both fetal whole genome profiling and confirmation of the feto-parental relationship are essential. This has been successfully performed using enriched and isolated cTBs, making cTB a better candidate for NIPD. cTB enumeration also correlates with abnormal fetal or placental development. On the other hand, downstream analysis of fNRBCs remains limited to examining fetal sex and aneuploidies. Furthermore, trophoblast-based NIPD via an endocervical sample is also promising because of reduced dilution from hematologic cells.