Higher Total Nucleated Cell Count Research Articles

Initial and follow-up evaluations on cerebrospinal fluid involvement by hematologic malignancy.

Central nervous system (CNS) involvement is a serious complication in hematologic malignancy, and early detection and management of CNS involvement in these cases significantly impact the prognosis. Currently, there is no consensus on the use of multiparametric flow cytometry (MFC) and conventional cytology (CC) testing for initial and follow-up cerebrospinal fluid (CSF) specimens to diagnose CNS involvement by hematologic malignancy. In our institution, after initial MFC and CC, two subsequent negative MFCs are required before discontinuing MFC. The aim of this study is to evaluate the outcome of this approach. CSF cytology and MFC reports were retrieved from Laboratory Information System, and data was reviewed. Between January 2020 and December 2021, 1789 CSF samples from 280 patients were submitted for CSF analysis. For those 517 CSF samples tested by both MFC and CC, 97 cases tested positive by both MFC and CC with 95% concordance. Eighteen cases were MFC + /CC - and 7 were MFC - /CC + . Thirty-six cases had initially positive MFCs followed by more than one MFC evaluation. Among those 36 cases, 22 cases (61.1%) converted to negative after the second follow-up sample, 9 cases (25%) were continuously positive for at least three samples, and 5 cases (13.9%) exhibited negative to positive conversion. Compared to negative CSF cases, positive CSFs had higher total nucleated cell count and higher total protein levels while red blood cells, glucose, and lactate dehydrogenase levels remained at comparable levels. The concordance between MFC and CC was excellent. The high incidence of positive MFCs on two or more follow-up samples and the high frequency of negative MFC to positive conversion indicate the necessity of repeated negative MFCs before discontinuing MFC. The fact that more than half of the positive cases converted to negative after the second CSF specimen and most follow-up positive cases can be detected by CC alone suggests it is adequate to use CC alone for follow-up CSF study after two consecutive negative MFCs.

Comparison of Fluid Analysis and Cytologic Findings of Cerebrospinal Fluid Between Three Collection Sites in Adult Equids With Neurological Disease.

Cerebrospinal fluid (CSF) is routinely collected from three sites in the horse, the atlanto-occipital (AO), atlantoaxial (AA), and lumbosacral (LS) space. A comparison between fluid analysis parameters [total protein, total nucleated cell count (TNCC), red blood cell (RBC) count, and morphologic analysis] from samples obtained at each of the three sites has not previously been performed. A retrospective analysis was performed to evaluate the differences in fluid analysis of CSF between the AO, AA, and LS sites in equids presented to a referral service for evaluation of suspected neurological disease. A total of 113 equids aged ≥1 year that underwent CSF collection between 2008 and 2020 were included. Total nucleated cell count, RBC concentration, total protein (TP), and morphologic evaluation between CSF samples obtained from the three sites were compared. When comparing all samples, LS centesis was associated with higher RBC compared to other sites (p < 0.05); TP was lower in the AA group than in the LS group (p < 0.05). Within a subset of cytologically unremarkable samples, RBC concentration was highest in LS samples (p < 0.01); TP was higher in LS samples compared to AA samples (p < 0.05) and TNCC was higher (p < 0.01) in AA and LS groups compared to the AO. In cytologically abnormal samples, there were no significant differences between sites in any parameter. Abnormal cytology was correlated with non-survival (p = 0.0002). Non-survival was associated with higher TNCC (p < 0.01). The receiver operating characteristic (ROC) curve for TNCC had an area under the curve of 0.67 (95% CI, 0.55–0.79) and indicated that a cutoff value of 24 cells/μL maximized specificity (72%) and sensitivity (54%) to predict non-survival in all horses. Positive predictive value was 45%; negative predictive value was 78%. The concentration of RBC was higher in samples from the LS site. This has clinical implications due to the importance of comparative diagnostics and its potential impact on cytologic evaluation. There were minimal differences in multiple other parameters between sites, which are likely clinically insignificant.

Cytological Findings in Bronchoalveolar Lavage Fluid of Foals With Pneumonia Caused by Rhodococcus equi and Other Bacteria

The distinction between lower respiratory tract infections caused by Rhodococcus equi and those caused by other pathogens is difficult. The aim of this retrospective study was to describe cytological findings in bronchoalveolar lavage fluid (BALF) of foals with pneumonia caused by R. equi or other bacteria. Nineteen foals aged from 3 weeks to 6 months with evidence of respiratory disease between 2015 and 2016 were selected from the database of the Veterinary Teaching Hospital “Mario Modenato” of the University of Pisa. Eight foals of 19 (42.1%) had R. equi pneumonia, whereas 11 of 19 (57.9%) had another bacterial pneumonia. R. equi–positive foals had statistically significant higher total nucleated cell count (P = .02) and neutrophils percentage (P = .002) than R. equi–negative ones. Macrophages proportion (P = .01) was lower in R. equi–positive than in R. equi–negative foals. Bronchoalveolar lavage is a quite easy procedure that can be performed in the field with minimal equipment required. Here we reported significant differences in the cellular composition of BALF that can be used to differentiate foals with R. equi bronchopneumonia from those with other bacterial pneumonias, whereas waiting for culture results.

Targeting Cord Blood Units with Higher Total Nucleated Cell Count for Inclusion in a Public Cord Blood Bank: Impact on Inventory Diversity and Self-Sustainability

Targeting Cord Blood Units with Higher Total Nucleated Cell Count for Inclusion in a Public Cord Blood Bank: Impact on Inventory Diversity and Self-Sustainability

Relationship between automated total nucleated cell count and enumeration of cells on direct smears of canine synovial fluid

Canine osteoarthritis is a common disorder seen in veterinary clinical practice and causes considerable morbidity in dogs as they age. Synovial fluid analysis is an important tool for diagnosis and treatment of canine joint disease and obtaining a total nucleated cell count (TNCC) is particularly important. However, the low sample volumes obtained during arthrocentesis are often insufficient for performing an automated TNCC, thereby limiting diagnostic interpretation. The aim of the present study was to investigate whether estimation of TNCC in canine synovial fluid could be achieved by performing manual cell counts on direct smears of fluid.Fifty-eight synovial fluid samples, taken by arthrocentesis from 48 dogs, were included in the study. Direct smears of synovial fluid were prepared, and hyaluronidase added before cell counts were obtained using a commercial laser-based instrument. A protocol was established to count nucleated cells in a specific region of the smear, using a serpentine counting pattern; the mean number of nucleated cells per 400 × field was then calculated. There was a positive correlation between the automated TNCC and mean manual cell count, with more variability at higher TNCC. Regression analysis was performed to estimate TNCC from manual counts. By this method, 78% of the samples were correctly predicted to fall into one of three categories (within the reference interval, mildly to moderately increased, or markedly increased) relative to the automated TNCC. Intra-observer and inter-observer agreement was good to excellent. The results of the study suggest that interpretation of canine synovial fluid samples of low volume can be aided by methodical manual counting of cells on direct smears.

FEASIBILITY OF COLLECTING UMBILICAL CORD BLOOD IN JORDAN AND THE EFFECT OF MATERNAL AND NEONATAL FACTORS ON HEMATOPOIETIC STEM CELL CONTENT

BackgroundCord blood transplant is an accepted treatment for many malignant and non-malignant diseases. We sought to determine the feasibility of collecting cord blood in Jordan and the effect of maternal and fetal factors on the quality of the cord blood units.MethodsA total of 124 cord blood units were collected, and 75 (60%) cord blood units were included in this analysis. Cord blood volume, total nucleated cell (TNC) count, cell viability and CD34+ content were measured, and clonogenic assay was performed.ResultsThe mean volume of the collected units was 68.9 ml (range 40–115) with mean nucleated cell count of 6.5 x 108 (range 1–23.0). Our results showed a positive correlation between the volume of cord blood and TNC count (p=0.008), cell viability (p=0.001), CD34+ content (p=0.034) and the length of the umbilical cord (p=0.011). In addition, our results showed an inverse relation between the Colony Forming Unit-Granulocyte Macrophage (CFU-GM) concentration and the gestation duration (p=0.038).ConclusionWe conclude that it is feasible to collect cord blood units in Jordan with excellent TNC and CD34+ cell content. The volume of cord blood collected was associated with higher TNC count and CD34+ count. Efforts toward establishing public cord blood banks in our area are warranted.

Comparison of bronchoalveolar lavage fluid obtained by manual aspiration with a handheld syringe with that obtained by automated suction pump aspiration from healthy dogs

To compare bronchoalveolar lavage (BAL) fluid obtained by manual aspiration (MA) with a handheld syringe with that obtained by suction pump aspiration (SPA) in healthy dogs. 13 adult Beagles. Each dog was anesthetized and bronchoscopic BAL was performed. The MA technique was accomplished with a 35-mL syringe attached to the bronchoscope biopsy channel. The SPA technique was achieved with negative pressure (5 kPa) applied to the bronchoscope suction valve with a disposable suction trap. Both aspiration techniques were performed in each dog in randomized order on opposite caudal lung lobes. Two 1 mL/kg aliquots of warm saline (0.9% NaCl) solution were infused per site. For each BAL fluid sample, the percentage of retrieved fluid was calculated, the total nucleated cell count (TNCC) and differential cell count were determined, and semiquantitative assessment of slide quality was performed. Comparisons were made between MA and SPA techniques for each outcome. 1 dog was removed from the study because of illness. The mean percentage of fluid retrieved (mean difference, 23%) and median TNCC (median distribution of differences, 100 cells/μL) for samples obtained by SPA were significantly greater than those for samples obtained by MA. In healthy dogs, BAL by SPA resulted in a significantly higher percentage of fluid retrieval and samples with a higher TNCC than did MA. Further evaluation of aspiration techniques in dogs with respiratory tract disease is required to assess whether SPA improves the diagnostic yield of BAL samples.

Evaluation of Cerebrospinal Fluid in Cavalier King Charles Spaniel Dogs Diagnosed with Chiari-Like Malformation with or without Concurrent Syringomyelia

Syringomyelia is a common clinical problem in the Cavalier King Charles Spaniel dog population. The underlying pathophysiology of the development and progression of syringes is currently unknown. The primary aim of the current study was to determine whether development of syringomyelia is accompanied by alterations in cerebrospinal fluid composition. Patient records of Cavalier King Charles Spaniels with Chiari-like malformations, with or without concurrent syringomyelia, that had magnetic resonance imaging and cerebrospinal fluid analysis between December 2004 and December 2009, were retrospectively reviewed. Total nucleated cell count per microliter, with differential count, red blood cell count per microliter, and protein concentration (g/l), were reviewed. Data were analyzed with the Mann-Whitney U-test. Spearman rank test was used to test association between cerebrospinal fluid composition and syrinx size, and the Fisher exact test was used to compare the presence of macrophages, reactive lymphocytes, and myelin. A P value of ≤ 0.05 was considered significant for statistical tests. Dogs with syringomyelia had a higher total nucleated cell count (syringomyelia: 4/µl [range, 0-15/µl] vs. without syringomyelia: 2/µl [0-8/µl]; P = 0.0047), higher protein concentration (syringomyelia: 0.26 g/l [0.07-0.42 g/l] vs. without syringomyelia: 0.2 g/l [0.12-0.39 g/l]; P = 0.039), and an increased neutrophil percentage (syringomyelia: 0% [0-37%] vs. without syringomyelia: 0% [0-21%]; P = 0.0203) than those with the Chiari-like malformation alone. There was a positive correlation between total nucleated cell count and syrinx size (r = 0.51; P = 0.0068). Development of syringomyelia is accompanied by alterations in cerebrospinal fluid composition. The pathogenesis of syringomyelia has not been fully elucidated; therefore, the exact origin of cerebrospinal fluid changes remains unclear.

Platelet-Activating Factor and Evidence of Oxidative Stress in the Bronchoalveolar Fluid of Thoroughbred Colts during Race Training

Inflammatory airway disease (IAD) is prevalent in young racehorses during training, being the 2nd most commonly diagnosed ailment interrupting training of 2-year-old Thoroughbred racehorses. That stabling and exercise cause oxidative stress, release of platelet-activating factor (PAF) and inflammation in airways of Thoroughbred colts. Colts in breeding farms (NC, n = 45), stabled for 30 days (EC, n = 40), and race trained (EX, n = 34). Cytological profile and parameters of bronchoalveolar lavage fluid (BALF) related to oxidative stress, bioactivity of the proinflammatory mediator PAF, catalase activity, and alveolar macrophage function. Percentages of neutrophils and eosinophils in the BALF of the EX group were higher (5.4 +/- 6.4% versus 0.9 +/- 1.2%) than the upper limits for normal horses (3-5%). BALF from the EX group (45.6 +/- 2.8 cells/microL of BALF) also displayed significantly (P = .017) higher total nucleated cell count. PAF bioactivity and the total protein concentration in the BALF were higher in the EX group (0.0683 +/- 0.076 versus 0.0056 +/- 0.007 340 : 380 nm ratio P = .0039, 0.36 +/- 0.30 versus 0.14 +/- 0.15 mg of proteins/mL of BALF P < .001). Concentration of BALF hydroperoxides was higher in the EC group (104.7 +/- 80.0 versus 35.2 +/- 28.0 nmol/mg of proteins, P = .013) and catalase activity was higher in the EX group (0.24 +/- 0.16 versus 0.06 +/- 0.02 micromol H2O2/min/mg of proteins, P = .0021). Alveolar macrophage phagocytosis (P = .048) as well as production of superoxide anion (P = .0014) and hydrogen peroxide (P = .0011) were significantly lower in EX group. Further studies should be performed to elucidate the role of PAF in the pathophysiology of IAD. Its presence in bronchoalveolar fluid of young athletic horses makes it a potential therapeutic target to be investigated.

Engraftment Rate and Kinetics Following Umbilical Cord Blood Transplantation Is Critically Affected by CD34+ Progenitor Cell Dose and CD8+ T-Cell Dose Complementary.

<Background> The risk of rejection after umbilical cord blood (CB) transplantation (CBT) is substantially higher than that after allogeneic BMT or PBSCT. In CBT, higher total nucleated cell count and CD34+ cell dose are shown to be associated with significant rapid and proper engraftment. Although CD8+ or other accessory cells are shown to have positive association with engraftment in many experimental and clinical BMT, potential contribution of these cells to engraftment after CBT has not been examined comprehensively.<Patients and methods> We examined CB subpopulation doses, such as CD34+, CD3+, CD4+, CD8+, CD14+ and CD16/56+ cells, in cryopreserved CB unit by flowcytometry, and analyzed their influence on the risks of primary graft-failure, GVHD, non-relapse mortality and the probability of survival. Data on 146 patients [median age 15.0 years old (range, 0.2–74.0), median body weight 41.0 kg (range, 4.8–85.0)] who received CBT as their first transplant between 1995 and 2002 for the treatment of malignant (n=137), including advanced disease patients (n=100), and nonmalignant (n=9) diseases were evaluated. The HLA-disparity between CB and patient was 0/6: 35 (24.0%), 1/6: 65 (44.5%), 2/6: 45 (30.8%), and 3/6: 1 (0.7%).<Results> In a multivariate analysis, CD34+ cell dose at higher than the median (>1.4 × 105/kg) and CD8+ cell dose at higher than the median (>15.7 × 105/kg) were significantly associated with successful engraftment [Hazard ratio (HR), 0.56; 95% confidence interval (CI), 0.37–0.85, and HR, 0.65; 95% CI, 0.43–0.99, respectively]. In the subgroup of CD34+ cell higher than the median group, the median time to achieve neutrophil engraftment was comparable between CD8+ higher and lower than the median group (21 days and 22 days, P=0.11). In the subgroup analysis for patients who received lower than the median (1.4 × 105/kg) of CD34+ cells, the median time to achieve neutrophil engraftment in the patients received more CD8+ cells (>15.7 × 105/kg) was significantly faster than that in the patients received less CD8+ cells (21 days versus 25 days, P=0.0047). CD8+ cell dose in at least median and HLA-disparity more than 1 locus were significant factors associated with an increasing incidence of grade II–IV acute GVHD [HR, 2.05; 95% CI, 1.18–3.59 and HR, 2.41; 95% CI, 1.16–5.00, respectively]. We could not detect any association of higher CD34+ and CD8+ cell doses with improved survival. No significant impact of other CB subpopulations on transplant outcome was observed.<Discussion> Our findings and previous study reported by Wagner et al. (Blood. 2002; 100:1611–8) demonstrated the CD34+ cell dose in CB unit is a significant and definitive predictor of engraftment rate and kinetics after CBT, and suggested the threshold dose for successful engraftment is around 1.4–1.7 × 105/kg. Additionally, CD8+ cells may have a supportive role in engraftment following CBT with an insufficient CD34+ cell dose. To confirm the effect of these cell subpopulations on survival, further study with more monotonous background is warranted.