INTRODUCTION: The ABX Pentra DX 120 (Pentra DX 120, ABX Diagnostics, Montpellier, France) adopted new technologies to perform differential leukocyte and erythroblast counts. The double matrix can discriminate Large Immature Cell (LIC), Immature Granulocyte (IMG), Immature Monocyte (IMM), Immature Lymphocyte (IML), and Atypical lymphocyte (ALY) in addition to a routine 5-differential count. For erythroblast (ERB), a fluorescence method is employed. In this study, we evaluated the performance of the Pentra DX 120 in the performance of differential leukocyte and erythroblast counts. METHODS: Precision was evaluated using 3-level control materials. Comparison analysis was performed on 200 samples: 100 normal and 100 abnormal samples. We evaluated the 5 part differential count, LIC, IMG, IMM, IML, ALY, and ERB. These parameters were analyzed in comparison with the results from the reference method, manual differential count. RESULTS: The coefficients of variation (CVs) of precision were <5% for neutrophils and lymphocytes, <15% for eosinophils and basophils and <7% for erythroblasts. The correlation coefficients (r) were >0.9 except monocytes and basophils. IMG, ALY and erythroblasts were also well correlated with manual count (r=0.8315, 0.5602, 0.8144, respectively). The efficiency of flagging system was 84% for LIC, 80% for ALY, and 78.0% for increased ERB (>2/100WBCs). CONCLUSIONS: The Pentra DX 120 performed reliable differential leukocyte and IMG, ALY, and ERB results demonstrated comparable performance to manual count. And, the flagging system was efficient for detecting each abnormal cell population. We expect the Pentra DX 120 double matrix and erythroblast count can reduce microscopic review rate in routine laboratory and promote laboratory efficiency.
Basophils ; Eosinophils ; Erythroblasts ; Fluorescence ; Granulocytes ; Leukocytes ; Lymphocytes ; Monocytes ; Neutrophils

We found an unusual morphology of eosinophil nucleus having longer chromatin filament in addition to a single narrow chromatin bridge. The nucleus having two chromatin filament bridge looked like two legged eosinophil, instead of usual glasses shape. As the physiologic function of the nucleus of granulocyte segmentation and the mechanism by which the lobes are formed during differention is still unknown, we could not know the definite nature and significance of these double chromatin filament. However we could suggest that they may be a reactive change of eosinophilia. This not uncommon morphology has not been described as yet. Here we report three cases of unusual morphology of eosinophil nucleus presenting double chromatin filament bridge, one case with a band form nucleus looked like ring shape, with brief review of literatures.
Chromatin* ; Eosinophilia ; Eosinophils* ; Eyeglasses ; Glass ; Granulocytes ; Leg

PURPOSE: Fc receptors and Mac-1 play an important role in the protective response of granulocytes and monocytes against microbial infection. FcgammaRl, FcgammaRll, FcgammaRlll as well as CD11b/ CD18 have never been measured in a quantitative way during hemopoiesis. Thus we quantified the expression of Fc Rl, Fc Rll, Fc Rlll, and CD11b/CD18 during hematopoietic differentiation using HL-60 cells, which was induced to differentiate by DMSO, or PMA. METHODS: HL-60 cells (ATCC CCL-240) were induced to differentiate by adding 1.0% DMSO, or 16nM PMA. On the 4th and 7th day after stimulation as well as before stimulation, phenotypic analysis was performed by flow cytometry after staining the cells with PE-conjugated anti- human CD64, CD32, CD16, CD11b, CD18, and isotype controls. And the measured fluorescent intensity was transformed into Molecules of Equivalent Soluble Fluorochromes (MESF). RESULTS: Percent positive cells and MESF of CD11b on HL-60 cells increased upon induction by DMSO, but not by PMA. Percent positive cells of CD18 on HL-60 cells was 99% regardless of differentiation. But MESF of CD18 was increased on the 4th day and decreased on the 7th day by DMSO or PMA. Percent positive cells and MESF of FcgammaRl on HL-60 cells increased upon induction by DMSO or PMA. Percent positive cells of FcgammaRll on HL-60 cells was above 90% regardless of differentiation. MESF of FcgammaRll showed no significant change by DMSO or PMA. CONCLUSION: Quantitative expression of FcgammaRl, FcgammaRll, FcgammaRlll, and CD11b/CD18 of HL-60 cells changed during induction of differentiation by DMSO or PMA. MESF of FcgammaR and CD11b/ CD18 a better indicator than percent positive cells to compare the differentiation of HL-60cells.
Dimethyl Sulfoxide ; Flow Cytometry ; Fluorescent Dyes ; Granulocytes ; HL-60 Cells* ; Humans ; Monocytes ; Receptors, Fc

BACKGROUND: Rhinovirus infection of the airways results in increased permeability of the airway vascular endothelium with the influx of plasma proteins, including lipids such as LDL. In vitro studies on the effect of oxLDL on leukocytes has shown many proinflammatory effects on multiple leukocytes. We hypothesized that oxLDL is one mechanism for recruiting granulocytes to the airways during a RV infection. Therefore, chemotaxis and transendothelial migration, in response to nLDL, was determined for these granulocytes. METHODS: nLDL was oxidized with 5mM Cu2SO4 for 20-24 hours. 3-5×10(5) cells were loaded into the Transwell filter while the chemotatic agonists were placed in the lower well for chemotaxis. Confluent monolayers on HPMEC were grown on Transwell filters for transendothelial migration. The filters were washed and eosinophils and neutrophils loaded on to the filter with the chemotatic agonist was were placed in the lower well. The wells were incubated for 3 hours. The number of migrating cells was counted on a hemocytometer. RESULTS: OxLDL, but not nLDL, is chemotatic for eosinophils and neutrophils. The level of granulocytes chemotaxis was dependent on both the concentration of LDL and its degree of oxidation. OxLDL stimulates eosinophil and neutrophils migration across HPMEC monolayers (±IL-1β preactivation) in a dose dependent manner. CONCLUSION: Increased vascular permeability during a RV infection may lead to the influx and oxidation of LDL. The resulting oxLDL. is one possible mechanism for the recruitment of neutrophils and eosinophils to the airway interstitial matrix. Once in the airways, granulocytes can further interact with oxLDL to promote airway inflammation.
Blood Proteins ; Capillary Permeability ; Chemotaxis ; Endothelium, Vascular ; Eosinophils* ; Granulocytes ; Inflammation ; Leukocytes ; Neutrophils* ; Permeability ; Rhinovirus ; Transendothelial and Transepithelial Migration

Neutrophils and eosinophils, 2 prominent granulocytes, are commonly derived from myelocytic progenitors through successive stages in the bone marrow. Our previous genome-wide transcriptomic data unexpectedly showed that genes encoding a multitude of neutrophil primary granule proteins (NPGPs) were markedly downregulated during the end period of eosinophilic terminal differentiation when cord blood (CB) cluster of differentiation (CD) 34+ cells were induced to differentiate toward the eosinophil lineage during a 24-day culture period. Accordingly, this study aimed to examine whether NPGP genes were expressed on the way to eosinophil terminal differentiation stage and to compare their expression kinetics with that of genes encoding eosinophil-specific granule proteins (ESGPs). Transcripts of all NPGP genes examined, including proteinase 3, myeloperoxidase, cathepsin G (CTSG), and neutrophil elastase, reached a peak at day 12 and sharply declined thereafter, while transcript of ESGP genes including major basic protein 1 (MBP1) attained maximum expression at days 18 or 24. Growth factor independent 1 (GFI1) and CCAAT/enhancer-binding protein α (C/EBPA), transactivators for the NPGP genes, were expressed immediately before the NPGP genes, whereas expression of C/EBPA, GATA1, and GATA2 kinetically paralleled that of eosinophil granule protein genes. The expression kinetics of NPGPs and ESGPs were duplicated upon differentiation of the eosinophilic leukemia cell line (EoL-1) immature eosinophilic cells. Importantly, confocal image analysis showed that CTSG was strongly coexpressed with MBP1 in differentiating CB eosinophils at days 12 and 18 and became barely detectable at day 24 and beyond. Our results suggest for the first time the presence of an immature stage where eosinophils coexpress NPGPs and ESGPs before final maturation.
Bone Marrow ; Cathepsin G ; Cell Line ; Eosinophils* ; Fetal Blood ; Granulocytes ; Hypereosinophilic Syndrome ; Kinetics ; Leukocyte Elastase ; Myeloblastin ; Neutrophils* ; Peroxidase ; Trans-Activators

BACKGROUND: Telomeres are repetitive DNA fragments at the termini of chromosomes functioning as stabilizing elements of the DNA. A ribonucleoprotein polymerase, called telomerase, is responsible for the synthesis of such telomeric repeats in embryo and germ cells. During ontogenesis of most normal human somatic cells, there exists a physiological telomerase repressing mechanism. In contrast, malignant cells are characterized by an unlimited progressive potential. Certain physiological agents, such as all-trans retinoic acid (ATRA), 13-cis retinoic acid (13-cisRA), 1alpha-25 dihydroxy vitamin D3 (VD3) and cytosine arabinoside (Ara-C), promote further differentiation of leukemic cells into mature granulocytes and monocytes and subsequently undergo apoptosis. METHODS: To determine if a potential linkage is present between telomerase regulation and the differentiation of malignant hematopoietic cells, the changes in telomerase activity during the maturation of HL-60 cells induced by ATRA, 13-cisRA, VD3 and Ara-C were investigated. RESULTS: Differentiating agents induce HL-60 cells to differentiate into CD11b+ granulocytes and monocyte/macrophages, respectively. Approximately 98% of HL-60 cells acquired the expression of CD11b+ antigen after ATRA, 13-cisRA or Ara-C treatment for 5 days. After 1 day treatment with differentiating agents, no significant difference in telomerase activity was shown between untreated and treated HL-60 cells. A dramatic inhibition of telomerase activity occurred at 3 days treatment of ATRA compared to untreated HL-60 cells. Longer treatment for 5 days with differentiating agents resulted in further decrease of telomerase activity. However, telomerase activity in HL-60 cells was decreased slightly by the VD3 or Ara-C treatment, even though for 5 days. No evidence of differentiation and slight decrease of telomerase activity were observed in ATRA-treated K-562 cells for 5 days. These decrease of telomerase activity were dependent on the incubation time and dose. CONCLUSION: These data clearly show the role of telomerase activity during the differentiation of HL-60 cells. This in vitro model can be useful for studies of the mechanisms controlling telomerase activity and in the search for physiological telomerase modulators.
Apoptosis ; Cholecalciferol ; Cytarabine ; DNA ; Embryonic Structures ; Germ Cells ; Granulocytes ; HL-60 Cells* ; Humans ; Monocytes ; Ribonucleoproteins ; Telomerase* ; Telomere ; Tretinoin

PURPOSE: Delta neutrophil index (DNI) represents the immature granulocytes count associated with neutrophil-consumption. We investigated whether DNI might be associated with Birmingham vasculitis activity score (BVAS) at diagnosis and could predict relapse during the follow-up in patients with antineutrophil cytoplasmic antibody-associated vasculitis (AAV). MATERIALS AND METHODS: We reviewed the medical records of 97 patients having DNI results. Twenty patients had granulomatosis with polyangiitis (GPA), 58 had microscopic polyangiitis (MPA), and 19 had eosinophilic GPA (EGPA). We collected clinical and laboratory data including BVAS, five factor score (FFS), and DNI. The correlation coefficient and cumulative relapse free survival rate were obtained. The optimal cut-off of DNI was extrapolated by calculating the area under the receiver operator characteristic curve. RESULTS: DNI was significantly related to cross-sectional BVAS. Furthermore, among continuous variables, only DNI could reflect BVAS of GPA and MPA, but not EGPA. Severe AAV was defined as BVAS ≥20 (the highest quartile). At diagnosis, patients having DNI ≥0.65% had a significantly higher risk of severe GPA and MPA than those having not (relative risk 4.255) at diagnosis. During the follow-up, DNI ≥0.65% could predict the higher relapse rate. CONCLUSION: DNI could reflect BVAS at diagnosis and furthermore, DNI ≥0.65% could not only identify severe AAV at diagnosis, but also predict relapse during the follow-up in patients with GPA and MPA.
Anti-Neutrophil Cytoplasmic Antibody-Associated Vasculitis* ; Cytoplasm ; Diagnosis ; Eosinophils ; Follow-Up Studies ; Granulocytes ; Granulomatosis with Polyangiitis ; Humans ; Medical Records ; Microscopic Polyangiitis ; Neutrophils* ; Recurrence* ; Survival Rate ; Vasculitis*

No abstract available.
Dermatitis* ; Granulocytes*

BACKGROUND: We evaluated the performance of leukocyte differential counting and clinical usefulness of the morphologic flags of the SE-000, and set optimal criteria for selecting and reviewing the specimens with increased abnormal cells. METHODS: From the results of SE-000 and manual leukocyte differential counting in 100 healthy control and 520 patient specimens we evaluated the correlations on the leukocyte fractions as well as the frequency, sensitivity and false positivity of the flags. After determination of the review criteria we calculated total review rate from the 3,403 consecutive CBC specimens. RESULTS: In both control and patient groups the correlation between two methods was high with the exception of monocytes and basophils. Regarding the morphologic flags, Blast was sensitive (86.9%) however could not detect mature looking lymphoblasts. Immature granulocyte showed high sensitivity (93.7%). Left shift showed the highest frequency (34.6%) and false positive rate (82.8%). Atypical lymphocytes and NRBC showed relatively low sensitivity (63.6%, and 50.5%, respectively). We determined to review the slide when 1) All morphologic flags except Left shift are marked, 2) WBC <3,000/microliter or >20,000/microliter, Hb <8.0 g/dL or 18.0 g/dL, Platelet <100,000/microliter or >600,000/microliter, 3) Severe deviation of leukocyte fractions or 4) Specially requested by physician. As a result, total review rate was 25.0% while 14 abnormal cases with no flags could be additionally detected. CONCLUSIONS: A new review criteria determined from the results of CBC and leukocyte differential together with morphologic flags could reduce the review rate without skipping the abnormal cases.
Basophils ; Blood Platelets ; Granulocytes ; Humans ; Leukocytes ; Lymphocytes ; Monocytes

BACKGROUND: We evaluated the performance of leukocyte differential counting and clinical usefulness of the morphologic flags of the SE-000, and set optimal criteria for selecting and reviewing the specimens with increased abnormal cells. METHODS: From the results of SE-000 and manual leukocyte differential counting in 100 healthy control and 520 patient specimens we evaluated the correlations on the leukocyte fractions as well as the frequency, sensitivity and false positivity of the flags. After determination of the review criteria we calculated total review rate from the 3,403 consecutive CBC specimens. RESULTS: In both control and patient groups the correlation between two methods was high with the exception of monocytes and basophils. Regarding the morphologic flags, Blast was sensitive (86.9%) however could not detect mature looking lymphoblasts. Immature granulocyte showed high sensitivity (93.7%). Left shift showed the highest frequency (34.6%) and false positive rate (82.8%). Atypical lymphocytes and NRBC showed relatively low sensitivity (63.6%, and 50.5%, respectively). We determined to review the slide when 1) All morphologic flags except Left shift are marked, 2) WBC <3,000/microliter or >20,000/microliter, Hb <8.0 g/dL or 18.0 g/dL, Platelet <100,000/microliter or >600,000/microliter, 3) Severe deviation of leukocyte fractions or 4) Specially requested by physician. As a result, total review rate was 25.0% while 14 abnormal cases with no flags could be additionally detected. CONCLUSIONS: A new review criteria determined from the results of CBC and leukocyte differential together with morphologic flags could reduce the review rate without skipping the abnormal cases.
Basophils ; Blood Platelets ; Granulocytes ; Humans ; Leukocytes ; Lymphocytes ; Monocytes