BACKGROUND: Neutrophil gelatinase-associated lipocalin (NGAL) is known to be one of the ideal biomarkers for acute kidney injury providing early information on damage to the kidney. METHODS: We evaluated the performance for precision and the reportable range of the automated NGAL Test (Bioporto Diagnostics, Denmark) assay and compared the values of these tests with widely used point of care test. The reference interval of NGAL was established in Korean adults. RESULTS: Within run percent coefficient of variation (%CV) and total precision %CV for 2 levels were all within 5%. The reportable range was found to be acceptable for the range of 57.0 - 3182.0 ng/mL (r=0.999). The method comparison was made between Biosite's assay and Bioporto Diagnostics' (Passing and Bablok fit, y=1.94x - 5.29; x, Biosite; y, Bioporto; n=31; y range, 250 to 1,308 ng/mL; r2=0.959). The correlation was linear within the limit of 1,500 ng/mL, but not beyond this limit. The 2.5 and 97.5 percentile of the reference range for the samples were 43.2 ng/mL and 124.8 ng/mL, respectively. CONCLUSIONS: Since NGAL Test can be used in automated chemical analyzer, it can not only reduce the man power and time consumed in but also displayed excellent precision and linearity.
Acute Kidney Injury ; Biomarkers ; Chemistry, Clinical ; Immunoassay ; Lipocalins ; Nephelometry and Turbidimetry ; Neutrophils ; Reference Values

AIMTo study the active constituents for the treatment of rheumatoid arthritis from the ethyl acetate extracts of the roots of Lasianthus acuminatissimus Merr.

METHODSVarious chromatographic techniques were used to separate and purify the constituents. Their structures were established on the basis of 1D, 2D NMR and HRMS spectroscopic analyses and their preliminary evaluation of anti-inflammation effect on the release of beta-glucuronidase was carried out.

RESULTSEight compounds were isolated and identified as lasianthuslactone A (1), codonolactone (2), 2,5-dimethoxy-1, 4-benzoquinone (3), uncargenin A (4), nonadecyl alcohol (5), 13-docosenoic acid (6), tetracosanoic acid (7) and beta-sitosterol (8). Compound 3 showed a significant inhibitory effect on release of beta-glucuronidase rat polymorphous nuclear leukocytes activated by platelet activating factor (PAF).

CONCLUSIONCompound 1 is a new one, the others were isolated from the plant for the first time and 3 is one of active anti-inflammation compound in the plant.

Animals ; Anti-Inflammatory Agents, Non-Steroidal ; chemistry ; isolation & purification ; pharmacology ; Benzoquinones ; chemistry ; isolation & purification ; pharmacology ; Glucuronidase ; metabolism ; Molecular Conformation ; Molecular Structure ; Neutrophils ; metabolism ; Plant Roots ; chemistry ; Plants, Medicinal ; chemistry ; Rats ; Rubiaceae ; chemistry ; Sesquiterpenes ; chemistry ; isolation & purification ; pharmacology

Neutrophils play an important role in the human immune system for protection against such microorganisms as a protozoan parasite, Trichomonas vaginalis; however, the precise role of neutrophils in the pathogenesis of trichomoniasis is still unknown. Moreover, it is thought that trichomonal lysates and excretory-secretory products (ESP), as well as live T. vaginalis, could possibly interact with neutrophils in local tissues, including areas of inflammation induced by T. vaginalis in humans. The aim of this study was to investigate the influence of T. vaginalis lysate on the fate of neutrophils. We found that T. vaginalis lysate inhibits apoptosis of human neutrophils as revealed by Giemsa stain. Less altered mitochondrial membrane potential (MMP) and surface CD16 receptor expression also supported the idea that neutrophil apoptosis is delayed after T. vaginalis lysate stimulation. In contrast, ESP stimulated-neutrophils were similar in apoptotic features of untreated neutrophils. Maintained caspase-3 and myeloid cell leukemia-1 (Mcl-1) in neutrophils co-cultured with trichomonad lysate suggest that an intrinsic mitochondrial pathway of apoptosis was involved in T. vaginalis lysate-induced delayed neutrophil apoptosis; this phenomenon may contribute to local inflammation in trichomoniasis.
Animals ; *Apoptosis ; Cells, Cultured ; Female ; Humans ; Membrane Potentials ; Mitochondrial Membranes/physiology ; Neutrophils/chemistry/*immunology ; Receptors, IgG/analysis ; Trichomonas vaginalis/*immunology

We analyzed the comparative amounts of granulocyte-colony stimulating factor (G-CSFr) and granulocyte macrophage CSF (GM-CSFr) receptors expressed on neutrophils and monocytes in measles patients to investigate the role of these CSFrs in the development of leukopenia including neutropenia and monocytopenia in measles. EDTA-anticoagulated peripheral blood of 19 measles patients, 10 children with other infections showing leukopenia and 16 children with normal complete blood cell counts (CBC)s were analyzed using flow cytometry and QuantiBRITE. The leukocyte (5260 +/- 2030/uL vs. 9900 + 2680/uL, p=0.000), neutrophil (2580 +/- 960/uL vs. 4250 +/- 2750/uL, p=0.024) and the lymphocyte counts of measles patients (1810 +/- 1430/uL vs. 4530 +/- 3450/uL, p= 0.006) were lower than in the normal controls. The neutrophils of measles patients expressed similar amounts of G- CSFr (1858 +/- 355) as normal children (1764 +/- 477, p= 0.564) and leukopenic patients (1773 +/- 673, p=0.713), but lower levels of GM-CSFr (535 +/- 118) than normal children (957 +/- 344, p=0.000) and leukopenic patients (832 +/- 294, p=0.002). The monocytes of measles patients expressed similar amounts of G-CSFr (916 +/- 336) and GM-CSFr (3718 +/- 906) as normal children (1013 +/- 391 and 4125 (2645, p > 0.05) but less than leukopenic patients (1454 +/- 398 and 5388 +/- 806, p > 0.05). The neutrophil and monocyte counts of measles patients did not correlate with the amount of G-CSFr or GM-CSFr expressed on neutrophils or monocytes (p > 0.05), but in the normal children, the monocyte count correlated with the levels of GM-CSFr on monocytes (r=0.951, p=0.049). In conclusion, neutropenia is one of the more important characteristics of measles patients, which could be due to the decreased GM-CSFr expression on neutrophils. However, the monocytopenia found in measles patients is not due to the decreased expression of CSFr on the monocytes.
Human ; Leukocyte Count ; Measles/*blood ; Monocytes/*chemistry ; Neutropenia/etiology ; Neutrophils/*chemistry ; Receptors, Granulocyte Colony-Stimulating Factor/*blood ; Receptors, Granulocyte-Macrophage Colony-Stimulating Factor/*blood

OBJECTIVETo investigate the significance of CD11b expression in neutrophils and lymphocytes in children with systemic inflammatory response syndrome (SIRS).

METHODSCD11b expression in neutrophils and lymphocytes was measured using flow cytometry in 36 children with SIRS (SIRS group) and 28 children with infectious disease but without SIRS (control group). The sensitivity and specificity of neutrophil CD11b for diagnosis of SIRS were evaluated.

RESULTSDuring the acute phase, an increased CD11b expression in neutrophils (96.7+/-8.1%) was observed in the SIRS group compared with the control group (85.1+/-5.1%) (p<0.05). Using neutrophil CD11b expression >92.2% as a cut-off value for diagnosis of SIRS, the sensitivity and the specificity were 97.2 % and 92.9% respectively. Lymphocytic CD11b expression in the SIRS group (13.4+/-8.6%) was lower than that in the control group (19.2+/-6.4%) in the acute phase (p<0.05). In the SIRS group, lymphocytic CD11b expression was remarkably suppressed in the severe sepsis subgroup (7.27+/-3.04%), showing significantly decreased expression compared with the non-infectious subgroup (19.3+/-2.9%) and the sepsis subgroup (15.9+/-12.5%) (p<0.01). In the convalescence stage lymphocytic CD11b expression in the SIRS group was similar to that in the control group.

CONCLUSIONSCD11b expression in neutrophils may serve as a reliable indicator for diagnosis of SIRS. The down-regulation of lymphocytic CD11b expression might be a signal of the condition aggravation in children with SIRS.

C-Reactive Protein ; analysis ; CD11b Antigen ; blood ; Child ; Child, Preschool ; Female ; Flow Cytometry ; Humans ; Infant ; Lymphocytes ; chemistry ; Male ; Neutrophils ; chemistry ; Sensitivity and Specificity ; Systemic Inflammatory Response Syndrome ; diagnosis ; immunology

Primary ciliary dyskinesia is characterized by chronic upper and lower respiratory infections which are caused by the grossly impaired ciliary transport. Since the cilia and neutrophils both utilize microtubular system for their movement, it has been speculated that neutrophil motility such as chemotaxis might be impaired in patients with primary ciliary dyskinesia. Neutrophils were purified from whole blood from 16 patients with primary ciliary dyskinesia and from 15 healthy controls. Chemotactic responses of neutrophils to leukotriene B4 (LTB4), complement 5a (C5a), and formylmethion-ylleucylphenylalanine (fMLP) were examined using the under agarose method. The chemotactic differentials in response to LTB4, C5a, and fMLP in neutrophils from the patient group were significantly lower than the corresponding values in neutrophils from the control group (p<0.05 for all comparisons). The difference in chemotactic index between the two groups was statistically significant for LTB4 and fMLP (p<0.05 for both comparisons), but not for C5a (p=0.20). Neutrophils from patients with primary ciliary dyskinesia showed a decreased chemotactic response as compared with those from normal subjects. It is concluded that the increased frequency of respiratory tract infection in patients with primary ciliary dyskinesia is possibly due to the defective directional migration of neutrophils, as well as to the defective mucociliary clearance of the airways.
Adolescent ; Chemotactic Factors/pharmacology ; Chemotaxis* ; Child ; Cilia/ultrastructure ; Comparative Study ; Complement 5a/pharmacology ; Dose-Response Relationship, Drug ; Dynein ATPase/chemistry ; Human ; Kartagener Syndrome/blood* ; Kartagener Syndrome/classification ; Leukotriene B4/pharmacology ; Male ; N-Formylmethionine Leucyl-Phenylalanine/pharmacology ; Neutrophils/physiology* ; Neutrophils/ultrastructure

AIMTo validate the abundance of Interleukin 8 receptor beta (IL-8Rbeta) mRNA in single human neutrophil.

METHODSHuman neutrophils were isolated and purified from volunteers, total RNA was extracted and a regular RT-PCR aiming at IL-8Rbeta mRNA was performed to ascertain its expression profile in human neutrophils and optimize the reaction conditions for the following single-cell RT-PCR procedures. Subsequently, single neutrophil or the cellular content was harvested to conduct reverse transcription and two-round PCR with the same primer pairs used before. Serial dilution of single neutrophil cDNA pool was carried out at the same time with the exact two-round PCR followed. The specificity of this single-cell RT-PCR procedure was verified by the BamHI restriction endonuclease digestion on the final cDNA products.

RESULTSRegular RT-PCR indicated IL-8Rbeta mRNA expression in human neutrophils. While single-cell RT-PCR was sensitive enough to detect trace IL-8Rbeta mRNA as predicted cDNA product could be amplified from a 10 000 times diluted intracellular specimen from single neutrophil, which indicated an abundant expression of this mRNA in human neutrophil. Moreover, BamHI digestion on the final cDNA product clarified the specificity of this single-cell RT-PCR procedure.

CONCLUSIONThis simplified semi-quantitative single-cell RT-PCR procedure specifically confirmed that IL-8Rbeta mRNA was highly expressed in human neutrophil, which also provided the possibility of comparing mRNA abundance at single cell level.

Cells, Cultured ; Humans ; Neutrophils ; chemistry ; RNA, Messenger ; genetics ; Receptors, Interleukin-8B ; analysis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; methods ; Single-Cell Analysis ; methods

OBJECTIVETo investigate the effect of blood glucose instability on respiratory burst of leukocytes in patients with type 2 diabetes (T2DM).

METHODSForty-five patients with T2DM were divided into 3 groups after continuous glucose monitoring for 72 h with glucose wavy coefficient <1.5 (n=11), between 1.5 and 3.0 (n=19), and >3.0 (n=15). Peripheral blood neutrophils were isolated from the diabetic patients and normal control subjects for assay of glucose 6-phosphate dehydrogenase (G6PD) with a spectrophotometric method, detecting G6PD mRNA expression by real-time PCR, and determining reactive oxygen species level using the fluorescent probe DCFH-DA.

RESULTSCompared with the normal control group, the diabetic patients showed significantly lowered G6PD activity (F=78.739, P<0.05) and ROS level (F=384.962, P<0.05) but significantly increased G6PD mRNA expression (F=269.612, P<0.01). These changes were significantly correlated with the blood glucose wavy coefficients.

CONCLUSIONThe fluctuation of blood glucose in T2DM patients can decrease G6PD activity and lead to functional decline of the respiratory burst.

Blood Glucose ; chemistry ; Case-Control Studies ; Diabetes Mellitus, Type 2 ; metabolism ; Glucosephosphate Dehydrogenase ; metabolism ; Humans ; Neutrophils ; metabolism ; Pentose Phosphate Pathway ; Reactive Oxygen Species ; metabolism ; Respiratory Burst

OBJECTIVETo study the clinical value of the expression of neutrophil surface CD64 in the diagnosis of community acquired pneumonia in children.

METHODSNinety-eight children with community acquired pneumonia were recruited into the study and were classified into three groups according to pathogene: bacterial pneumonia (n=48), viral pneumonia (n=29) and Mycoplasmal pneumonia (n=21). Twenty healthy children were enrolled as controls. The bacterial infection group was subdivided into mild infection (n=36) and severe infection groups (n=12). The levels of peripheral blood neutrophil CD64 were measured using flow cytometry. Dynamic changes of C-reactive protein were also detected for each patient.

RESULTSThe CD64 index and CRP levels in the bacterial pneumonia group were significantly higher than in the other three groups (P<0.05). The CD64 index in the severe bacterial infection group was significantly higher than in the mild group (P<0.05). After antibiotic treatment, expression of CD64 in the severe bacterial infection group decreased significantly (P<0.05). The CD64 index was positively correlated with CRP value (r=0.545, P<0.01). ROC curve analysis showed that the threshold of CD64 and CRP was 2.8 and 8 mg/L respectively. Specificity of CD64 index (90%) was much higher than CRP (74%).

CONCLUSIONSThe determination of peripheral blood neutrophil CD64 contributes to the early diagnosis of pulmonary bacterial infection and the evaluation of anti-infection effect.

C-Reactive Protein ; analysis ; Child ; Child, Preschool ; Community-Acquired Infections ; blood ; diagnosis ; Female ; Humans ; Male ; Neutrophils ; chemistry ; Pneumonia, Bacterial ; blood ; diagnosis ; ROC Curve ; Receptors, IgG ; blood

OBJECTIVETo investigate the value of combined determination of neutrophil CD64 and procalcitonin (PCT) in the early diagnosis of neonatal bacterial infection.

METHODSAccording to discharge diagnosis, 37 neonates with bacterial infection were divided into sepsis (n=15) and ordinary infection (non-sepsis) groups (n=22). Twenty-one neonates without infection who were hospitalized during the same period of time were enrolled as the control group. Venous blood samples were collected immediately after admission. Flow cytometry was used to measure the serum level of neutrophil CD64. Chemiluminescence and immune transmission turbidimetry were used to measure the serum levels of PCT and CRP respectively.

RESULTSThe sepsis group had higher serum levels of neutrophil CD64, PCT, and CRP than the control group (P<0.01), the ordinary infection group had a higher serum level of neutrophil CD64 than the control group (P<0.01), and the sepsis group had higher serum levels of PCT and CRP than the ordinary infection group (P<0.01). The areas under the ROC curve (AUC) of neutrophil CD64, PCT, and CRP in diagnosing bacterial infection were 0.818, 0.818, and 0.704 respectively, and the AUC of combined neutrophil CD64 and PCT was 0.926. A combination of neutrophil CD64 and PCT had a sensitivity of 97.29% and an accuracy of 89.65% in the early diagnosis of neonatal bacterial infection.The sensitivity and accuracy were higher than those of a combination of CRP and neutrophil CD64 or PCT as well as neutrophil CD64, PCT, or CRP alone for the early diagnosis of neonatal bacterial infection.

CONCLUSIONSThe combined determination of neutrophil CD64 and PCT can improve the sensitivity and accuracy in the diagnosis of neonatal bacterial infection, which helps with early identification of bacterial infection.

Bacterial Infections ; blood ; diagnosis ; C-Reactive Protein ; analysis ; Calcitonin ; blood ; Early Diagnosis ; Female ; Humans ; Infant, Newborn ; Male ; Neutrophils ; chemistry ; ROC Curve ; Receptors, IgG ; blood