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Abstract Objectives: The purpose of the study was to compare the performance of GloCyte (Advanced Instruments, Norwood, MA), a new semiautomated instrument for cerebrospinal fluid cell counting, with the manual hemocytometer method and the automated Sysmex XN (Sysmex, Kobe, Japan) body fluid mode. The clinical impact of replacing the manual method with either automated method was determined. Methods: Fifty-seven samples from 38 patients were analyzed by all three methods. Pearson correlation and Passing-Bablok regression were used to compare methods. Cytospin smears were reviewed on all samples, and clinical histories were obtained. Results: There was a strong linear relationship between the manual and automated methods for WBC counts (R = 0.988 for GloCyte; R = 0.980 for Sysmex XN). Positive bias was absent or negligible for WBC counts less than 30/μL. GloCyte and manual RBC counts were equivalent. There were no samples for which replacement of manual WBC counts by automated counts would have changed the diagnosis. Both automated methods showed improved precision for WBC counts compared with the manual method. Conclusions: Replacing manual WBC counts by GloCyte or Sysmex XN WBC counts would improve consistency of results without compromising diagnostic accuracy.

Background Umbilical cord blood (UCB) has been proposed as the potential source of haematopoietic stem cells (HSC) for allogeneic transplantation. However, few studies have shown that a common disease in pregnancy such as preeclampsia would affect the quality of UCB-HSC. Total nucleated cell count (TNC) is an important parameter that can be used to predict engraftment including UCB banking. Colony forming unit (CFU) assay is widely used as an indicator to predict the success of engraftment, since direct quantitative assay for HSC proliferation is unavailable. The aim of this study is to investigate the effects of preeclampsia in pregnancy on the stemness and differentiation potency of UCB-HSC. Methods Mononuclear cells (MNC) were isolated from UCB and further enriched for CD34+ cells using immune-magnetic method followed by CFU assay. A panel of HSC markers including differentiated haematopoietic markers were used to confirm the differentiation ability of UCB-HSC by flow cytometry analysis. Results/ discussion The HSC progenitor’s colonies from the preeclampsia group were significantly lower compared to the control. This correlates with the low UCB volume, TNC and CD34+ cells count. In addition, the UCB-enriched CD34+ population were lymphoid progenitors and capable to differentiate into natural killer cells and T-lymphocytes. Conclusion These findings should be taken into consideration when selecting UCB from preeclamptic mothers for banking and predicting successful treatment related to UCB transplant.

PurposeThe purpose was to evaluate the effect of intrauterine injection of aBMNC on the endometrial function in patients with refractory Asherman’s syndrome (AS) and/or thin and dysfunctional endometrium (TE).Study designThis is a prospective, experimental, non-controlled studyMaterial and methodsThe study was carried out between December 2018 and December 2020 on 20 patients, who were of age < 45 years and had oligo/amenorrhea and primary infertility due to refractory AS and/or TE. One hundred ml BM was extracted. aBMNC cells were separated according to generic volume reduction protocol by using the Cell Separation System SEPAX S-100 table top centrifuge system. We have evaluated CD34+, mononuclear cell (MNC), and total nucleated cell (TNC) counts. The transplantation aBMNC was performed by two intrauterine injections at an interval of one week, transvaginally into the endometrial–myometrial junction by an ovum aspiration needle. Midcyclic endometrial thickness (ET) and gestations after transplantation were evaluated.ResultsThe mean TNC, MNC, and CD34+ cells were 11.55 ± 4.7 × 108, 3.85 ± 2.01 × 108, and 7.00 ± 2.88 × 106 at first injection, respectively, and 6.85 ± 2.67 × 108, 2.04 ± 1.11 × 108, and 3.44 ± 1.31 × 106 at second injection, respectively.The maximum posttransplantation ET was significantly higher than the maximum pretransplantation ET: 2.97 ± 0.48 vs. 5.76 ± 1.19 (mean ± standard deviation, p < 0.01). Twelve patients had frozen-thaw embryo transfers after the study.In 42% (n = 5 of 12) of the patients, pregnancy was achieved. One of the five patients delivered a healthy baby at term.ConclusionsAutologous BMNC transplantation may contribute to endometrial function in patients with AS and/or TE.

Abstract Background Neonatal cell therapy applications are increasing; however, data on allogeneic cell therapy are limited. Objective To summarize evidence on allogeneic cell therapy in term and preterm neonates. Methods Cochrane Central Register of Controlled Trials, Embase, Ovid Medline, and various registries were searched for studies investigating the safety, feasibility, and efficacy of allogeneic cell therapy in neonates. Two authors independently selected the articles, extracted data, and assessed the risk of bias. Results Twelve published (153 infants) and 21 ongoing studies were included. These studies predominantly sourced allogeneic cells from umbilical cord blood (UCB). Mesenchymal stromal cells (MSCs) were the main cell type used (134 of 153 infants); others included UCB-derived total nucleated cells (TNCs) and human amnion epithelial cells (hAECs). Applications included bronchopulmonary dysplasia (BPD; 113 infants), Krabbe disease (13 infants), intraventricular haemorrhage (10 infants), perinatal arterial ischemic stroke (10 infants), hypoxic-ischaemic encephalopathy (6 infants), and necrotizing enterocolitis (1 infant). Nine out of 12 studies did not report any serious adverse events (SAEs) related to cell administration. Three studies reported SAEs, such as graft versus host disease (GVHD) in 5 infants (UCB-derived TNCs for Krabbe disease); and transient cardiorespiratory compromise in 1 infant (hAECs for BPD). Data on efficacy outcomes were limited. Conclusion The safety and feasibility of allogeneic cell therapy applications in neonates are available, mainly from the use of MSCs. Further safety data for other cell types are required, and the risk of GVHD in different settings needs to be determined. Efficacy studies are largely lacking for all cell types. Protocol Registration The protocol was registered with PROSPERO (registration number CRD42023397876), the international prospective register for systematic reviews (https://www.crd.york.ac.uk/PROSPERO). Graphical Abstract Graphical Abstract

To determine the influence of graft composition in haplo-HSCT, we summarized the long-term consequences of 251 consecutive transplantations from haploidentical donors. For donor-recipient HLA3/6-matched setting, 125 cases used G-CSF-mobilized BM and PBSCs mixtures, while 126 cases only used G-CSF-mobilized PBSCs in HLA4/6-matched transplantation. On the one hand, we wanted to explore the effect of harvests (CD34+ cells and TNCs dosages) on transplantation outcome in the context of haplo-HSCT no matter HLA4/6 or HLA3/6-matched setting. On the other hand, for patients using G-CSF-mobilized BM and PBSCs combination in HLA3/6-matched setting, we attempted to analyze whether TNCs or CD34+ cells from G-CSF-mobilized BM or G-CSF-mobilized PBSCs play the most paramount role on transplantation prognosis. Collectively, patients with hematologic malignancies receiving G-CSF-primed BM and PBSCs harvests had comparable consequences with patients only receiving G-CSF-mobilized PBSCs. Moreover, when divided all patients averagely according to the total amount of transfused nucleated cells, 3-year TRM of the intermediate group (13.06-18.05×10 8 /kg) was only 4.9%, which was remarkably reduced when compared to lower and higher groups with corresponding values 18.3%, 19.6% ( P =0.026). The 3-year probabilities of OS and DFS of this intermediate group were 72.6% and 66.5%, which were slightly improved than the lower and higher groups. Most importantly, these data suggest that the transfused nucleated cells from G-CSF-primed BM above than 5.20×10 8 /kg could achieve remarkably lower TRM in haplo-HSCT receiving G-CSF-mobilized BM and PBSCs harvests. These encouraging results suggested that we could improve the efficacy of haplo-HSCT by adjusting the component and relative ratio of transfused graft cells. Nevertheless, the above findings should be confirmed in a randomized prospective comparative research with adequate follow-up.

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has led the medical and scientific community to explore the pathogenesis and clinical manifestations of coronaviruses. In felines, a widespread coronavirus known as feline coronavirus (FCoV) can lead to feline infectious peritonitis (FIP), a highly fatal disease characterised by severe systemic inflammation. Diagnosing FCoV remains challenging due to the limited accuracy of the available methods. The present study introduces the FIP Effusion Index, a novel diagnostic method that combines the albumin-to-globulin (ALB/GLOB) ratio with the delta total nucleated cell (∆TNC) count obtained via flow cytometry using the Sysmex XN-1000V® analyser in effusions. Samples from cats (n = 50) with suspected FIP were analysed for ∆TNC, with findings showing that a ∆TNC ≥ 2.1 is highly indicative of FIP and a ∆TNC ≥ 4.9 can be considered diagnostic. The FIP Effusion Index enhanced diagnostic precision in our group of samples, achieving 96.3% sensitivity and 95.7% specificity for values ≥ 5.06, and reaching perfect specificity (100%) with 96.3% sensitivity for values ≥ 7.54. This combined approach surpasses the accuracy of individual parameters, establishing the FIP Effusion Index as a superior diagnostic tool for FIP, with potential applications in both veterinary and human medicine for related coronavirus diseases.

Background Bronchoalveolar lavage (BAL) is a diagnostic method for the assessment of the lower respiratory airway health status in horses. Differential cell count and sometimes also total nucleated cell count (TNCC) are routinely measured by time-consuming manual methods, while faster automated methods exist. The aims of this study were to compare: 1) the Sysmex XN-V body fluid (BF) mode with the manual techniques for TNCC and two-part differential into mononuclear and polymorphonuclear cells; 2) the Olympus VS200 slide scanner and software generated deep-learning-based algorithm with manual techniques for four-part differential cell count into alveolar macrophages, lymphocytes, neutrophils, and mast cells. The methods were compared in 69 clinical BAL samples. Results Incorrect gating by the Sysmex BF mode was observed on many scattergrams, therefore all samples were reanalyzed with manually set gates. For the TNCC, a proportional and systematic bias with a correlation of r  = 0.79 was seen when comparing the Sysmex BF mode with manual methods. For the two-part differential count, a mild constant and proportional bias and a very small mean difference with moderate limits of agreement with a correlation of r  = 0.84 and 0.83 were seen when comparing the Sysmex BF mode with manual methods. The Sysmex BF mode classified significantly more samples as abnormal based on the TNCC and the two-part differential compared to the manual method. When comparing the Olympus VS200 deep-learning-based algorithm with manual methods for the four-part differential cell count, a very small bias in the regression analysis and a very small mean difference in the difference plot, as well as a correlation of r  = 0.85 to 0.92 were observed for all four cell categories. The Olympus VS200 deep-learning-based algorithm also showed better precision than manual methods for the four-part differential cell count, especially with an increasing number of analyzed cells. Conclusions The Sysmex XN-V BF mode can be used for TNCC and two-part differential count measurements after reanalyzing the samples with manually set gates. The Olympus VS200 deep-learning-based algorithm correlates well with the manual methods, while showing better precision and can be used for a four-part differential cell count.

Cerebrospinal fluid analysis is an important diagnostic test when assessing a neurological canine patient. For this analysis, the total nucleated cell count and differential cell counts are routinely taken, but both involve time-consuming manual methods. To investigate faster automated methods, in this study, the Sysmex XN-V body fluid mode and the deep-learning-based algorithm generated by the Olympus VS200 slide scanner were compared with the manual methods in 161 canine cerebrospinal fluid samples for the total nucleated cell count and in 65 samples with pleocytosis for the differential counts. Following incorrect gating by the Sysmex body fluid mode, all samples were reanalyzed with manually set gates. The Sysmex body fluid mode then showed a mean bias of 15.19 cells/μL for the total nucleated cell count and mean biases of 4.95% and −4.95% for the two-part differential cell count, while the deep-learning-based algorithm showed mean biases of −7.25%, −0.03% and 7.27% for the lymphocytes, neutrophils and monocytoid cells, respectively. Based on our findings, we propose that the automated Sysmex body fluid mode be used to measure the total nucleated cell count in canine cerebrospinal fluid samples after making adjustments to the predefined settings from the manufacturer. However, the two-part differential count of the Sysmex body fluid mode and the deep-learning-based algorithm require some optimization.

Purpose The most important HLA-independent factor for the selection of cord blood units (CBU) for hematopoietic stem cell transplantation is the total nucleated cell (TNC) count over 150 × 10 7 as a surrogate marker for stem cell content. The purpose of this prospective study was to define prenatal clinical predictors for TNC count that would help to identify successful CBU donors before the onset of active labor. Methods This was a prospective analysis of 594 CBUs, collected from all eligible term singleton pregnancies at Basel University Hospital between 4/2015 and 9/2016 analyzing several maternal and fetal factors. The impact of these factors on TNC count (< 150 × 10 7 cells vs. ≥ 150 × 10 7 cells) of the CBUs was modeled in a multivariate analysis. Results A total of 114 (19.2%) CBUs had a TNC count of ≥ 150 × 10 7 . In a ROC analysis there was no significant difference between the AUC of all prenatal factors (AUC 0.62) and estimated fetal birth weight by ultrasound alone (AUC 0.62). For women planning a trial of labor a recruitment cut-off at an estimated birth weight of 3300 g would allow 72.6% of all donors with sufficient TNC count to be recruited and 22.8% of all collected CBUs would have a sufficient TNC count for banking. For women planning for elective CS a cut-off of 3400 g would allow 71.4% of all donors with sufficient TNC count to be recruited and 22.7% of all collected CBUs would have sufficient TNC count for banking. Conclusion The estimated fetal birth weight within 2 weeks of delivery by ultrasound as single parameter can be considered at the time of recruitment to estimate the chances of a successful CBU donation.

Umbilical cord blood (UCB) is used as a source of donor cells for hematopoietic stem cell (HSC) transplantation. The success of transplantation is dependent on the quality of cord blood (CB) units for maximizing the chance of engraftment. Improved outcomes following transplantation are associated with certain factors of cryopreserved CB units: total volume and total nucleated cell (TNC) count, mononuclear cell (MNC) count, and CD34+ cell count. The role of the storage period of CB units in determining the viability and counts of cells is less clear and is related to the quality of cryopreserved CB units. Herein, we demonstrate the recovery of viable TNCs and CD34+ cells, as well as the MNC viability in 20-year-old cryopreserved CB units in a CB bank (MEDIPOST Co., Ltd., Seongnam-si, Gyeonggi-do, Korea). In addition, cell populations in CB units were evaluated for future clinical applications. The stable recovery rate of the viability of cryopreserved CB that had been stored for up to 20 years suggested the possibility of uses of the long-term cryopreservation of CB units. Similar relationships were observed in the recovery of TNCs and CD34+ cells in units of cryopreserved and fresh CB. The high-viability recovery of long-term cryopreserved CB suggests that successful hematopoietic stem cell (HSC) transplantation and other clinical applications, which are suitable for treating incurable diseases, may be performed regardless of long-term storage.